the japanese journal of clinical and experimental medicine

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The Japanese journal of experimental medicine

• 東京大学医科学研究所 トウキョウ ダイガク イカガク ケンキュウジョ
• 東京大学伝染病研究所 トウキョウ ダイガク デンセンビョウ ケンキュウジョ

the Institute of Medical Science, the University of Tokyo

Kinokuniya, 1928-1990

• Vol. 7, no. 1 (1928)-v. 60, no. 6 (1990)

Jpn. j. exp. med

大学図書館所蔵  件 /  全 104 件

愛知学院大学 歯学・薬学図書館情報センター 歯薬図 1951-1984

21(1-4), 22(1-5), 23-54

旭川医科大学 図書館 1928-1990

7-18, 20-25, 28-31, 33-60

麻布大学 附属学術情報センター 1949-1990

20-22, 27(5), 28(1), 46, 47(2-6), 48-60

岩手医科大学 附属図書館 図 1940-1990 400

18(6), 19(1-5), 20(2), 23-60

岩手大学 図書館 1976-1990

奥羽大学 図書館 1969-1985

39-54, 55(1)

大阪公立大学 阿倍野医学図書館 図 1980-1989

大阪公立大学 杉本図書館 図書館 1955-1990

26-28, 33-34, 45-60

大阪大学 附属図書館 生命科学図書館 生命図 1930-1990

8-18, 19(1-4), 20-60

岡山大学 附属図書館 資源植物科学研究所分館 植物研図 1955-1990 203.0/J

25-58, 59(1-3, 5-6), 60

岡山大学 附属図書館 鹿田分館 鹿田図 1930-1990

8-18, 19(1-5), 20-60

帯広畜産大学 附属図書館 図 1969-1990

鹿児島大学 附属図書館 桜ヶ丘分館 桜丘分館 1970-1990

神奈川工科大学 附属図書館 1964-1972

34-35, 36(1-2, 4, 6), 37(1-3, 5-6), 38-42

神奈川歯科大学 図書館 1979-1990

金沢大学 附属図書館 自然図自動書庫 1928-1969

金沢大学 附属図書館 医学図書館 医学図雑誌 1970-1990

鹿屋体育大学 附属図書館 図 1951-1983

21-50, 51(1-2, 4-6), 52-53

北里大学 医学図書館 1980-1990

北里大学 獣医学部図書館 1973-1990

北里大学 図書館 白金分館 1928-1990

7-17, 20-39, 41-60

九州大学 医学図書館 医分 1930-1990

8-10, 11(1-4, 6), 12-19, 21-23, 24(2-6), 25-60

九州大学 中央図書館 1928-1939

京都大学 農学部 図書室 図 1940-1990

18, 19(4-5), 20(6), 21-60

京都大学 附属図書館 BNC 1928-1941 BNC||J||031E

7-18, 19(1-5)

京都薬科大学 図書館 1987-1990 P-49||JPNJEXM

杏林大学 医学図書館 医図 1928-1990

杏林大学 井の頭図書館 井の頭図 1950-1970

20-33, 39-40

京都大学 医学図書館 医図 1938-1990 J||4

16-17, 20-60

京都府立医科大学 附属図書館 図 1928-1990

7-18, 19(1-5), 20-52, 53(1-5), 54-60

近畿大学 医学部図書館 医図 1954-1982

24-32, 33(1-2, 4-6), 34-38, 40-41, 48-50, 51(1-4, 6), 52

岐阜大学 医学図書館 医分 1960-1990

30(3-6), 31-32, 33(1-4), 34-60

熊本大学 附属図書館 医学系分館 図書館 1928-1990

7-18, 19(1-3), 20-25, 26(3-6), 27(4-6), 28-41, 42(1-2, 4-6), 43-60

久留米大学 附属図書館 医学部分館 図書館 1934-1990

12(1, 3-5), 14(1-2, 4-6), 16(1-5), 17(2-6), 34-60

群馬大学 総合情報メディアセンター 医学図書館 図書館 1949-1990

結核予防会結核研究所 図書室 1953-1990

厚生労働省 国立医薬品食品衛生研究所 安全情報部図書係 1949-1990

厚生労働省 国立感染症研究所 図書 1928-1990

厚生労働省 国立感染症研究所 村山庁舎 1964-1984

34-52, 53(1-5), 54

神戸大学 附属図書館 医学分館 1964-1988 J-75

34-55, 56(1-5), 57-58

公立大学法人 福島県立医科大学 附属学術情報センター 図 1952-1990

22(2-3, 5-6), 23(1), 28-60

国立 がん研究センター 図書館 1964-1990

国立科学博物館 書庫 1949-1990

20, 21(2-3), 22(2), 29(4), 31(1, 3-6), 32(1), 34(3-6), 35-39, 40(1-4), 45-60

国立研究開発法人 理化学研究所 図書館 1928-1990

国立保健医療科学院 1937-1990

15-16, 17(1, 3-6), 18-53, 54(1-4, 6), 55-60

埼玉医科大学 附属図書館 埼医大図 1975-1983

札幌医科大学 附属総合情報センター 図 1928-1990

滋賀医科大学 附属図書館 図 1975-1990

静岡県立大学 附属図書館 草薙図書館 1964-1990 490.5||J-8

34-58, 59(1-4, 6), 60

島根大学 附属図書館 医学図書館 1974-1979

昭和大学 図書館 1976-1990

信州大学 附属図書館 医学部図書館 1933-1990

11, 12(1, 6), 13(1-4), 14-16, 18-20, 22-60

自治医科大学 図書館 1930-1990

8(3-4, 6), 9-14, 15(4-6), 16-18, 19(4-5), 20(1), 21(4), 22(3-6), 23-35, 36(1-3, 5-6), 37-60

千葉大学 附属図書館 亥鼻分館 亥分 1928-1985

7-18, 20(2-6), 21-55

筑波大学 附属図書館 医学図書館 1979-1990 洋 J

鶴見大学 図書館 1971-1990

41-55, 56(1-5), 57-60

東京医科歯科大学 図書館 図 1928-1990

7-10, 23-60

東京医科大学 図書館分館 図 1928-1990

7-17, 18(1-5), 19(1-5), 20-60

東京工業大学 すずかけ台図書館 1979-1990

49(5-6), 50-60

東京国立博物館 1974-1984 405-32

44(1, 3-6), 45-46, 47(1, 3-6), 48(1-3, 6), 49-54

東京歯科大学 図書館 1949-1990

20(1-2, 4-6), 21, 22(1-5), 23(2-6), 24-60

東京慈恵会医科大学 学術情報センター 1976-1990

東京大学 医科学研究所 図書室 図書室 1928-1990

東京大学 医学図書館 図書 1981-1990

東京大学 柏図書館 書庫 1928-1990

7-18, 19(1-5), 20-60

東京大学 駒場図書館 自図 1952-1990

22-24, 25(1-2), 31-59, 60(1, 3-5)

東京大学 大学院人文社会系研究科・文学部 図書室 心理 1929-1940

東京大学 農学生命科学図書館 図 1987-1990

東京大学 薬学図書館 図書 1960-1984

30-38, 53-54

東邦大学 医学メディアセンター 1975-1990

東北大学 附属図書館 医学分館 図 1928-1990

東北大学 附属図書館 農学分館 図 1951-1977

徳島大学 附属図書館 蔵本分館 1940-1990

18(2, 4-5), 19(1-4), 20-52, 53(1-3, 5-6), 54-59

鳥取大学 附属図書館 医学図書館 図 1928-1990

7-19, 21(4), 22-60

富山大学 附属図書館 医薬学図書館 図 1949-1990

獨協医科大学 図書館 1941-1990

長崎大学 附属図書館 医学分館 1949-1990

名古屋大学 附属図書館 医学部分館 医分館 1928-1990

7-52, 53(2-6), 54-60

奈良県立医科大学 附属図書館 1964-1990

鳴門教育大学 附属図書館 1964-1971

34(2, 6), 37(4-6), 38(1, 4), 39(3, 5), 40(2-4), 41(2, 6)

新潟大学 附属図書館 旭町分館 旭分 1928-1991 490

日本医師会 医学図書館 1950-1990

20(4-6), 21-32, 33(1-5), 34-35, 36(1, 3-6), 37-60

日本歯科大学 生命歯学部図書館 1968-1990

38-54, 55(1), 57-60

日本獣医生命科学大学 付属図書館 1966-1990 490.76

日本生物科学研究所 図書室 1955-1990 継続中

日本大学 医学部図書館 1935-1990

13(2), 20-60

日本大学 生物資源科学部 図書館 図 1953-1990

農業・食品産業技術総合研究機構 動物衛生研究部門 図書室 1949-1990 ||||||E3224

農林水産省 農林水産研究情報総合センター 1954-1963

24(4-6), 25(1-2, 6), 26(1-2), 27(3-4, 6), 28(1, 3, 6), 29(2), 30(5), 32(1-2), 33(5)

弘前大学 附属図書館 医学部分館 1928-1990

7-11, 12(1, 3-6), 13-14, 15(1), 16-18, 19(1-5), 23(2), 28(2), 30, 31(3, 5-6), 32-60

広島大学 図書館 中央図書館 特殊 1955-1990

25-51, 52(1-3, 5-6), 53-60

福井大学 附属図書館 1968-1969

福岡大学 図書館 医学部分館 1973-1990 WB

北海道大学 医学研究科・医学部図書館 図書 1928-1990

北海道大学 大学院歯学研究院・大学院歯学院・歯学部図書室 図書 1968-1989

38-41, 42(1, 3-6), 43-54, 59(4)

防衛省防衛医科大学校 図書館 1951-1978

21-23, 24(1-3, 5), 25-26, 27(3-6), 28(2-6), 29-47, 48(1-5)

三重大学 医学部図書館 医学科 1976-1976 49

三重大学 附属図書館 図 1980-1990 49

宮崎大学 附属図書館 医学分館 図 1928-1990

7-15, 30-31, 32(1-5), 33(1-2, 4-6), 34-35, 36(1), 37(3-6), 38-55, 56(1-5), 57(3-6), 58-60

山口大学 図書館 医学部図書館 1954-1990

24(5-6), 25-60

山梨大学 附属図書館 医学分館 図 1968-1979 W

横浜市立大学 医学情報センター 1930-1990

8-17, 20-60

酪農学園大学 附属図書館 図 1922-1988

7-18, 19(1-5), 20-39, 40(1-2, 4-6), 41(1, 3-6), 42-47, 48(1-5), 49-55, 57-58

和歌山県立医科大学 図書館 紀三井寺館 図 1928-1990

7-13, 34-60

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Statement of responsibility varies: The Government Institute for Infectious Diseases of The Tokyo Imperial University

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Tokyo Imperial University

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Institute of Medical Science, University of Tokyo 1992. 3-

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PERSPECTIVE article

The current status and future direction of clinical research in japan from a regulatory perspective.

• Department of Regulatory Science, Faculty of Pharmacy, Meiji Pharmaceutical University, Kiyose, Japan

In Japan, a law called the Clinical Trials Act went into being effective on April 1, 2018, and clinical research on human subjects conducted in Japan has been undergone major changes. Those other than clinical trials for marketing approval of drugs or medical devices are broadly classified into “specific clinical trials” and others, and regulations have been tightened for each. As a result, clinical interventional study was drastically reduced, and observational clinical study increased. For the observational clinical study, the two previous ethical guidelines were merged into the “Ethical Guidelines for Medical and Biological Research Involving Human Subjects,” which was enacted in March 2021. The observational clinical study is now subjected to these ethical guidelines. In addition, changes are planned for the Act on the Protection of Personal Information, which greatly affects data collection in clinical research. Clinical research in Japan must be conducted appropriately while adapting to these various changes in the external environment and legal framework. Adapting to these changes is not an easy task, as it requires increased financial and human resources for all stakeholders.

Introduction

Advances and developments in medical technology lead to higher quality medical care and better health for people. The creation and reinforcement of evidence based on clinical research are important for the development of medicine. In spite of this, clinical research in Japan is insufficient in terms of the related systems and implementation mechanisms and has therefore fallen behind Europe and the United States ( 1 , 2 ). After the Diovan scandal, a misconduct case related to a post-marketing clinical trial of an antihypertensive agent, valsartan in 2012 and similar scandals involving the clinical research at that time ( 3 ), trust in the clinical research conducted in Japan was lost ( 4 , 5 ). Since then, to regain trust in clinical research, industries, government, and academia have been united in their efforts to ensure the reliability and scientific soundness of clinical research, improve the mechanisms used to implement research, and create and revise laws and other regulations that support these changes. Against this background, in recent years, legal measures and policies related to clinical research are being strengthened in Japan.

The legal system concerning clinical research in Japan consists mainly of two laws or guidelines. One is the Clinical Trials Act (“ Rinsho-Kenkyuu hou ” in Japanese) for interventional research ( 6 ), which was established in April 2018. The other is an ethical guideline for medical research, such as observational clinical studies. This guideline is known as the Ethical Guidelines for Medical and Biological Research Involving Human Subjects ( 9 ), which was developed by merging the existing Ethical Guidelines for Medical Research Involving Human Subjects ( 7 ) and the Ethical Guidelines for Human Genome/Analysis Research ( 8 ). The new merged guideline was announced in March 2021. The Clinical Trials Act for interventional research was established in April 2018, and over 3 years have passed since its establishment. Although stakeholders such as researchers, medical institutions, and pharmaceutical companies that conduct interventional research are required to understand and appropriately comply with this Act, it is believed that there is still room for making further improvements in the Act. As the Clinical Trials Act was originally created for purpose of restoring trust in the clinical research conducted in Japan after several scandals, it requires bigger changes and more careful handling to be carried out by stakeholders, such as medical institutions and pharmaceutical companies than those required under the regulations stipulated by the existing ethical guidelines. While these changes were appropriate in some cases, in others, they simply led to increase in paperwork and complexity. The enactment of the Clinical Trials Act has caused continuing confusion at institutions where research is conducted; however, in general, it has led to the reduction of outdated habits, changes in ways of thinking, and improvements in clinical research operations as well as in the relationship between pharmaceutical companies and medical institutions. Based on this, I believe that the Clinical Trials Act currently remains effective in improving the clinical research conducted in Japan. It has been <1 year since the establishment of the Ethical Guidelines for Medical and Biological Research Involving Human Subjects, which targets clinical research other than interventional research, such as observational clinical studies. It can be expected that issues related to the handling of these guidelines will be brought up in the future, but the issue related to the definition of “observational clinical studies” has already been pointed out as a problem. Therefore, researchers, medical institutions, and pharmaceutical companies will search for better ways to carry out the clinical research in Japan.

In this paper, we provide an overview of the history of legal regulations related to clinical research and discuss the responsibilities and roles played by various stakeholders in Japan. The objective is to point out the current issues in the legal system and guidelines related to the Japanese clinical research and discuss the future direction of clinical research in Japan.

1. The types of clinical research

Clinical research is a part of medical research that is conducted for determining the causes and treatment of diseases; making improvements for disease prevention, diagnosis, and treatment; and improving the quality of life of patients. Clinical research naturally involves human subjects. There are a variety of definitions of “clinical research” and none of them has become the established definition; however, it is believed that clinical research can be classified into the following four types:

(1) Clinical interventional study: research related to the development of medicines, treatments, therapeutic methods, and drugs.

(2) Prognostic factor clinical study: research that investigates factors that predict patient prognoses.

(3) Epidemiological clinical study: research that investigates the causes of diseases.

(4) Validity clinical study: known as a validation study, this research assesses tests and surveys.

Prospective clinical research includes interventional studies involving interventions, such as drugs; medical devices; surgery; radiation, exercise, and diet therapies as well as non-interventional studies or observational studies, which do not involve any intervention. Specially in Japan, prospective clinical research conducted for obtaining approval to manufacture and market drugs and medical devices is known as a “clinical trial for the approval of drugs or medical devices” (“ Chiken ” in Japanese). Chiken fall under the regulations of Japanese Good Clinical Practice (J-GCP) which is more stringent guideline than international guideline for GCP (ICH-GCP). As a result of the establishment of the Clinical Trials Act in 2018, clinical research that involved interventions other than Chiken and was conducted under previously existing ethical guidelines that also need to comply with the new law.

2. Legal regulations related to clinical research

In Japan, the first legal regulation related to clinical research other than Chiken consisted of guidance in the form of ethical guidelines for each type of study, i.e., observational clinical study, clinical research, and human genome/analysis research.

The first regulation was the Ethical Guidelines for Human Genome/Analysis Research ( 8 ) developed in 2001. In addition, the Ethical Guidelines for Epidemiological Clinical Study ( 10 ), which targeted observational clinical studies conducted in the field of epidemiology, was developed in June 2002. The Ethical Guidelines for Clinical Research ( 11 ) was developed in 2003 and covered clinical research other than those mentioned above. Thus, each type of clinical research was conducted in accordance with one of the above ethical guidelines. Subsequently, from around 2011, problems, such as overlapping guidelines and uncertainties regarding the guideline that should be followed when conducting research that would fall under multiple ethical guidelines were brought up. Further, in the wake of the 2012 Diovan incident ( 3 ), a review of ethical guidelines was conducted; in December 2014, the Ethical Guidelines for Epidemiological Clinical Study ( 10 ) and the Ethical Guidelines for Clinical Research ( 11 ) were merged, and the Ethical Guidelines for Medical Research Involving Human Subjects ( 7 ) was officially announced. Based on what was learned as a result of the Diovan scandal, the legal system and financial aspects were reviewed, which led to the enactment of the Clinical Trials Act for interventional research in April 2018 ( 6 ). In addition, the Ethical Guidelines for Medical Research Involving Human Subjects ( 7 ) and the Ethical Guidelines for Human Genome/Analysis Research ( 8 ) were merged, and the Ethical Guidelines for Medical and Biological Research Involving Human Subjects ( 9 ) was established in March 2021. Therefore, currently, the clinical interventional research that receives funding from a company and similar studies fall under the Clinical Trials Act ( 6 ) and all other clinical research falls under the Ethical Guidelines for Medical and Biological Research Involving Human Subjects ( 9 ).

The main changes that the legal regulations related to clinical research have undergone are shown in Figure 1 .

Figure 1 . Major steps for regulations and guidelines related to clinical research in Japan.

“Clinical research” as defined by the Clinical Trials Act is interventional research other than Chiken designed to identify the efficacy or safety of drugs and other products through the use of drugs, medical devices, etc., by people. The Clinical Trials Act defines “specific clinical trials” (“ Tokutei-Rinsho-Kenkyu ” in Japanese) as interventional trials on previously approved drugs and medical devices that receive funding from companies, clinical interventional studies on unapproved drugs and medical devices, and interventional clinical research for off-label uses. Specific clinical trials must be conducted in accordance with the Clinical Trials Act, and medical institutions must have a research system and all relevant standards established. In addition, when conducting specific clinical trials, a Certified Review Board (CRB) is required to inspect and approve the study, and the study protocol must be submitted to the Ministry of Health, Labor, and Welfare. As of July 1, 2021, 101 medical institutions have CRBs. The medical institution or institutions conducting the research and all researchers involved in the research must reveal any conflict of interest (COI) related to the financial support received from pharmaceutical companies. Prior to the enactment of the Clinical Trials Act, it was not necessary in Japan to have an established research system, obtain CRB approval after due inspection, reveal COI, or submit any paperwork to the Ministry of Health, Labor, and Welfare.

3. Implementation scheme and role allotment

The Clinical Trials Act assumes that the research initiative is conducted by either a researcher or a researcher in cooperation with a company. On the other hand, in cases of clinical research on already approved drugs and medical devices that is conducted by companies as post-marketing clinical trials or surveillance, the research must be conducted in accordance with a risk management plan (RMP) and the company must conduct the research as the sponsor in Japan. Thus, based on the research initiative, interventional clinical research in Japan is currently carried out as one of the following three types:

(1) Investigator-initiated research.

(2) Joint research with company (investigator-initiated).

(3) Joint research with company (company-initiated).

As there must be a particular format for administrative procedures and contracts, which are required during study implementation, the Japan Pharmaceutical Industry Legal Affairs Association ( Ihoken ) has established formats for contracts and other documents used in each type of clinical research ( 12 ).

The research material, labor, and financial support that companies may provide for conducting clinical research under the Clinical Trials Act are detailed in Table 1 . Regardless of the type of clinical research, there are precautions stipulating that companies cannot be involved in the selection of participating centers; execution of any tasks related to applying for the approval of Institutional Review Boards; submission of study protocols to the Ministry of Health, Labor, and Welfare; and execution of activities, such as monitoring, supervision, data management, or statistical analysis ( 13 ). Furthermore, while there are guidelines related to financial support provided by companies for clinical research that is not covered by the Clinical Trials Act, there are currently no clear guidelines on the contents and labor that companies can provide as support.

Table 1 . The involvement of companies under the Clinical Trials Act in Japan.

4. Current issues with the Clinical Trials Act

Although it has been a little over 3 years since the enactment of the Clinical Trials Act, several issues related to the implementation have been pointed out ( 14 ). Comparison of the Clinical Trials Act to the regulations stipulated by the previous ethical guidelines reveals a number of characteristic features. Examples are listed below:

- The new category of clinical research known as “specific clinical trials.”

- The establishment of CRBs, which allows centralized inspection rather than inspections at each center.

- The shift in the responsibility of the research from the director of the center to the principal investigator (researcher).

- The establishment of details regarding conflicts of interest.

A specific clinical trial is a clinical research that satisfies at least one of the following: (1) Utilizes research funding provided by the manufacturer and marketer of the drug for which the research is being conducted, and (2) Utilizes drugs that are either unapproved or are being used off-label. However, as clinical settings are complex, there are a variety of questions regarding the exact moment that a clinical trial begins. For example:

- Is a clinical trial with dose modifications for elderly or children that are common in routine practice but strictly off-label considered as a “specific clinical trial”?

- Are studies utilizing an old drug that is covered by insurance for an off-label purpose considered “specific clinical trials”?

- Is it acceptable to not classify as a “specific clinical trial” an “observational clinical study” whose funding is provided by a pharmaceutical company in cases in which testing is not performed during standard medical examinations or when a higher number of examinations and tests are performed than would be as a part of standard medical examinations?

- There are no issues on the study drug of the anticancer drugs used in the study, but if the research funding is provided by the company of the antiemetic agent used in the study, is it acceptable to exclude from the “specific clinical trial”?

In addition, there are no clear guidelines regarding rules and the allotment of responsibilities, which make it difficult to know how to handle such issues.

For example, there is no single uniform way to make judgments in cases wherein it would be better to obtain the consent of the study participants for the purpose of having a paper published by a leading journal. However, according to the ethical guidelines, patients can opt out of granting consent to participate in studies in which the methods for gathering and reporting safety information, as required for observational clinical studies, are not established or in cases in which the requirements of the principal investigator and medical institutions implementing the study are not clear. There are also cases in which the monitor conducts an excessive amount of source data verification. Finally, there are examples in which companies are still involved in the creation of protocols, selection of centers, analysis, and case investigation even though they are prohibited from doing so.

5. Current issues, future direction, and effort toward revising the Clinical Trials Act

As little time has passed since the establishment of the Ethical Guidelines for Medical and Biological Research Involving Human Subjects, the issues with these guidelines have yet to be clearly identified. However, 3 years have passed since the Clinical Trials Act has been established, and discussions on how to improve it are currently under way. Here, we will list several points of dispute regarding the revision of the Clinical Trials Act, its current state, and our opinion regarding the direction that the improvements should take.

(1) The handling of observational clinical studies

Current Status

Although observational clinical studies are not subject to the Clinical Trials Act, the definition of an observational clinical study is not clear; therefore, there are cases that should not necessarily be excluded from the regulations of the Act simply because the researcher calls their study as an “observational clinical study.” In particular, there are cases in which actions, such as additional hospital visits for the purpose of the study, the addition of measurement items, and collection of small amounts of additional blood sampling are determined not to be “the most appropriate medical care for the patient” and, as a result, the CRB determines that the study should be classified as a specific clinical trial.

Making Improvements

- The scope of application needs to clearly indicate “interventional studies that utilize drugs, etc.”

- The definition of “observational clinical studies,” which are excluded from the Act, needs to be revised.

(2) The concept of “sponsor”

The principal investigators and all centers that are involved in study implementation play the role of both a “sponsor” and an “investigator.”

- Each study should have one sponsor.

- Sponsors can be individuals, companies, research institutions, or organizations.

- Sponsors are responsible for the implementation of the study (e.g., regarding adverse event reports, it should be determined by the sponsor whether there is a causal relationship with test drugs or not, and based on adverse event reports collected from the participating investigators).

(3) CRB requirements

There are many CRBs in Japan, and there exists a wide disparity in the review standards, procedures, skills, and fees, which means that there are cases in which the appropriate review is not conducted.

Based on the fact that there are disparities in the quality of CRBs, in the future, the CRBs should be consolidated.

(4) The scope of applying the Clinical Trials Act in studies involving medical devices

“Off-label” refers to cases of usage that differs even slightly from the approved, certified, or applied for usage, efficacy, and performance. If “off-label,” then the study is subject to Chiken or the Clinical Trials Act.

With respect to the clinical research involving off-label medical devices, cases in which the medical device can be regarded as having the same level of risk as that determined when the medical device in question received certification, the status of the study should be investigated and the issue of whether the study should be subjected to Chiken or the Clinical Trials Act based on the results of that investigation should be considered.

Discussion and Conclusion

In the wake of scandals involving clinical research, such as the 2012 Diovan scandal, efforts have been under way to ensure the trustworthiness and scientific soundness of clinical research, strengthen regulations and guidelines for clinical research, and examine and adjust the regulations that support these changes to regain trust in the Japanese clinical research. Against this background, in recent years, legal measures and policies related to clinical research have been taken in succession in Japan. The Japanese medical institutions, pharmaceutical companies, and stakeholders in regulatory agency must carry out clinical research appropriately while adapting to a variety of external environment-related and legal changes. Handling these changes will not be easy as they entail increases in funding and human resources. However, currently, clinical research in Japan is undergoing major changes and working toward improvements. We expect that as a result of these improvements, the Japanese clinical research will develop further and make additional contributions toward medical progress.

Data Availability Statement

The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.

Author Contributions

The author confirms being the sole contributor of this work and has approved it for publication.

Conflict of Interest

The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Publisher's Note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

Acknowledgments

The author thank Hiroshi Asai, Nobuhiko Sato, Shizuka Watanabe, Noriko Kawajiri, and Nao Shimoda for their support and guidance in conducting this research.

1. Soteriades ES, Rosmarakis ES, Paraschakis K, Falagas ME. Research contribution of different world regions in the top 50 biomedical journals (1995-2002). FASEB J. (2006) 20:29–34. doi: 10.1096/fj.05-4711lsf

PubMed Abstract | CrossRef Full Text | Google Scholar

2. Jusue-Torres I, Mendoza JE, Brock MV, Hulbert A. The 100 Most cited papers about cancer epigenetics. Cureus . (2020) 12:e7623. doi: 10.7759/cureus.7623

3. Tanimoto T, Kami M, Shibuya K. Research misconduct and scientific integrity: a call for a global forum. Lancet. (2013) 382:940. doi: 10.1016/S0140-6736(13)61933-9

4. Sawano T, Ozaki A, Saito H, Shimada Y, Tanimoto T. Payments from pharmaceutical companies to authors involved in the valsartan scandal in Japan. JAMA Netw Open . (2019) 2:e193817. doi: 10.1001/jamanetworkopen.2019.3817

5. Maeda H. Medical affairs in pharmaceutical companies and related pharmaceutical regulations in Japan. Front Med Regulat Sci. (2021) 8:672095. doi: 10.3389/fmed.2021.672095

6. Ministry Ministry of Health Labour Welfare Japan. Clinical Trials Act . Available online at: https://hourei.net/law/429AC0000000016 (accessed November 20, 2021).

Google Scholar

7. Ministry Ministry of Health Labour Welfare Japan. Ethical Guidelines for Medical Research Involving Human Subjects . Available online at: https://www.mhlw.go.jp/file/06-Seisakujouhou-10600000-Daijinkanboukouseikagakuka/0000069410.pdf (accessed November 20, 2021).

8. Ministry Ministry of Health Labour Welfare Japan. Ethical Guidelines for Human Genome/Analysis Research . Available online at: https://www.mhlw.go.jp/general/seido/kousei/i-kenkyu/genome/0504sisin.html (accessed November 20, 2021).

9. Ministry Ministry of Health Labour Welfare Japan. Ethical Guidelines for Medical and Biological Research Involving Human Subjects . Available online at: https://www.mhlw.go.jp/content/000769923.pdf (accessed November 20, 2021).

10. Ministry Ministry of Health Labour Welfare Japan. Ethical Guidelines for Epidemiological Clinical Study . Available online at: https://www.mhlw.go.jp/general/seido/kousei/i-kenkyu/ekigaku/0504sisin.html (accessed November 20, 2021).

11. Ministry Ministry of Health Labour Welfare Japan. Ethical Guidelines for Epidemiological Clinical Study . Available online at: https://www.mhlw.go.jp/general/seido/kousei/i-kenkyu/rinri/0504sisin.html (accessed November 20, 2021).

12. Japan, Pharmaceutical Industry Legal Affairs Association. Clinical Research Contract Forms . Available online at: https://www.ihoken.or.jp/htdocs/index.php?page_id=136 (accessed November 20, 2021).

13. Asai H. For appropriate implementation of Clinical Trials Act and regulations. Jpn Pharmacol Ther. (2019) 47:s122–5.

14. Maeda H. The establishment of medical affairs functions in Japan-history, current status, and future perspective. Pharm Med Dev Reg Sci . (2020) 51:345–53.

Keywords: clinical research, clinical trial, Japan, regulation, regulatory science

Citation: Maeda H (2022) The Current Status and Future Direction of Clinical Research in Japan From a Regulatory Perspective. Front. Med. 8:816921. doi: 10.3389/fmed.2021.816921

Received: 17 November 2021; Accepted: 06 December 2021; Published: 13 January 2022.

Reviewed by:

Copyright © 2022 Maeda. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY) . The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

*Correspondence: Hideki Maeda, maeda@my-pharm.ac.jp

Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.

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• Published: 13 August 2024

Amplification of autoimmune organ damage by NKp46-activated ILC1

• Stylianos-Iason Biniaris-Georgallis 1 , 2 , 3 , 4   na1   nAff5 ,
• Tom Aschman   ORCID: orcid.org/0000-0002-2101-6146 1 , 2 , 3 , 6   na1   nAff7 ,
• Katerina Stergioula 1 , 2 , 3 , 4   na1 ,
• Frauke Schreiber 1 , 2 , 3 ,
• Vajiheh Jafari 1 , 2 , 3 ,
• Anna Taranko   ORCID: orcid.org/0000-0002-2660-7762 1 , 2 , 3 ,
• Tejal Karmalkar 1 , 2 , 3 , 4 ,
• Ana Kasapi 1 , 2 , 3 ,
• Tihana Lenac Rovis 8 ,
• Vedrana Jelencic 8 ,
• David A. Bejarano 9 ,
• Lea Fabry 1 , 2 , 3 ,
• Michail Papacharalampous 6 ,
• Irene Mattiola   ORCID: orcid.org/0000-0002-2922-9947 2 , 3 ,
• Martina Molgora 10   nAff11 ,
• Jinchao Hou 10   nAff12 ,
• Karolin W. Hublitz   ORCID: orcid.org/0000-0003-4840-4496 3 ,
• Frederik Heinrich 2 ,
• Gabriela Maria Guerra 2 ,
• Pawel Durek 2 ,
• Giannino Patone   ORCID: orcid.org/0000-0002-7242-0341 13 ,
• Eric Lars-Helge Lindberg   ORCID: orcid.org/0000-0002-2979-433X 13 ,
• Henrike Maatz 13 , 14 ,
• Oliver Hölsken   ORCID: orcid.org/0000-0001-6086-9275 2 , 3   nAff15 ,
• Gerhard Krönke   ORCID: orcid.org/0000-0002-7566-4325 1 , 2 ,
• Arthur Mortha   ORCID: orcid.org/0000-0003-2673-0485 16 ,
• Reinhard E. Voll   ORCID: orcid.org/0000-0002-5542-9133 6 ,
• Alexander J. Clarke   ORCID: orcid.org/0000-0003-2846-7860 17 ,
• Anja E. Hauser   ORCID: orcid.org/0000-0002-7725-9526 1 , 2 ,
• Marco Colonna   ORCID: orcid.org/0000-0001-5222-4987 10 ,
• Kevin Thurley 2 , 18 ,
• Andreas Schlitzer   ORCID: orcid.org/0000-0001-7662-3712 9 ,
• Christoph Schneider   ORCID: orcid.org/0000-0002-0452-2960 19 ,
• Efstathios G. Stamatiades   ORCID: orcid.org/0000-0002-2785-9005 3 ,
• Mir-Farzin Mashreghi   ORCID: orcid.org/0000-0002-8015-6907 2 , 20 ,
• Stipan Jonjic   ORCID: orcid.org/0000-0001-5003-3108 8 ,
• Norbert Hübner   ORCID: orcid.org/0000-0002-1218-6223 13 , 14 , 21 ,
• Andreas Diefenbach   ORCID: orcid.org/0000-0002-9176-9530 2 , 3   na2 ,
• Masatoshi Kanda   ORCID: orcid.org/0000-0003-3468-3586 13 , 22   na2 &
• Antigoni Triantafyllopoulou   ORCID: orcid.org/0000-0003-0123-7117 1 , 2 , 3   na2  

Nature ( 2024 ) Cite this article

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We are providing an unedited version of this manuscript to give early access to its findings. Before final publication, the manuscript will undergo further editing. Please note there may be errors present which affect the content, and all legal disclaimers apply.

• Autoimmunity
• Innate lymphoid cells
• Lupus nephritis
• Monocytes and macrophages

In systemic lupus erythematosus (SLE) loss of immune tolerance, autoantibody production and immune complex deposition are required but not sufficient for organ damage 1 . How inflammatory signals are initiated and amplified in the setting of autoimmunity remains elusive. Here, we set out to dissect layers and hierarchies of autoimmune kidney inflammation in order to identify tissue-specific cellular hubs that amplify auto-inflammatory responses. Using high-resolution single-cell profiling of kidney immune and parenchymal cells, in combination with antibody blocking and genetic deficiency, we show that tissue-resident NKp46 + innate lymphoid cells (ILC) are crucial signal amplifiers of disease-associated macrophage expansion and epithelial cell injury in lupus nephritis, downstream of autoantibody production. NKp46 signaling in a distinct subset of ILC1 instructed an unconventional immune-regulatory transcriptional program, which included the expression of the myeloid cell growth factor CSF2. CSF2 production by NKp46 + ILC promoted the population expansion of monocyte-derived macrophages. Blockade of the NKp46 receptor (using the antibody mNCR1.15 2 ) or genetic deficiency of NKp46 abrogated epithelial cell injury. The same cellular and molecular patterns were operative in human lupus nephritis. Our data support that NKp46 + ILC1 promote parenchymal cell injury by granting monocyte-derived macrophages access to epithelial cell niches. NKp46 activation in ILC1 thus constitutes a previously unrecognized, critical tissue rheostat that amplifies organ damage in autoimmune hosts, with broad implications for inflammatory pathologies and therapies.

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Author information.

Stylianos-Iason Biniaris-Georgallis

Present address: Department of Nephrology and Medical Intensive Care, Charité-Universitätsmedizin, Berlin, Campus Mitte, Berlin, Germany

Tom Aschman

Present address: Department of Neuropathology, Charité-Universitätsmedizin Berlin, Campus Mitte, Berlin, Germany

Martina Molgora

Present address: H.Lee Moffitt Cancer Center & Research Institute, Department of Immunology, 12902 USF Magnolia Drive, Tampa, FL, USA

Jinchao Hou

Present address: Department of Anesthesiology, Children’s Hospital, Zhejiang University School of Medicine, National Clinical Research Center for Child Health, Hangzhou, China

Oliver Hölsken

Present address: Department of Anesthesiology and Intensive Care Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany; BIH Academy, Clinician Scientist Program, Charitéplatz 1, Berlin, Germany

These authors contributed equally: Stylianos-Iason Biniaris-Georgallis, Tom Aschman, Katerina Stergioula

These authors jointly supervised this work: Andreas Diefenbach, Masatoshi Kanda, Antigoni Triantafyllopoulou

Authors and Affiliations

Department of Rheumatology and Clinical Immunology, Charité-Universitätsmedizin Berlin, Campus Mitte, Berlin, Germany

Stylianos-Iason Biniaris-Georgallis, Tom Aschman, Katerina Stergioula, Frauke Schreiber, Vajiheh Jafari, Anna Taranko, Tejal Karmalkar, Ana Kasapi, Lea Fabry, Gerhard Krönke, Anja E. Hauser & Antigoni Triantafyllopoulou

German Rheumatism Research Center, A Leibniz Institute, Berlin, Germany

Stylianos-Iason Biniaris-Georgallis, Tom Aschman, Katerina Stergioula, Frauke Schreiber, Vajiheh Jafari, Anna Taranko, Tejal Karmalkar, Ana Kasapi, Lea Fabry, Irene Mattiola, Frederik Heinrich, Gabriela Maria Guerra, Pawel Durek, Oliver Hölsken, Gerhard Krönke, Anja E. Hauser, Kevin Thurley, Mir-Farzin Mashreghi, Andreas Diefenbach & Antigoni Triantafyllopoulou

Institute of Microbiology, Infectious Diseases and Immunology (I-MIDI), Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany

Stylianos-Iason Biniaris-Georgallis, Tom Aschman, Katerina Stergioula, Frauke Schreiber, Vajiheh Jafari, Anna Taranko, Tejal Karmalkar, Ana Kasapi, Lea Fabry, Irene Mattiola, Karolin W. Hublitz, Oliver Hölsken, Efstathios G. Stamatiades, Andreas Diefenbach & Antigoni Triantafyllopoulou

Department of Biology, Chemistry and Pharmacy, Free University of Berlin, Berlin, Germany

Stylianos-Iason Biniaris-Georgallis, Katerina Stergioula & Tejal Karmalkar

Department of Rheumatology and Clinical Immunology, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg, Germany

Tom Aschman, Michail Papacharalampous & Reinhard E. Voll

Center for Proteomics, Faculty of Medicine, University of Rijeka, Rijeka, Croatia

Tihana Lenac Rovis, Vedrana Jelencic & Stipan Jonjic

Quantitative Systems Biology, Life and Medical Sciences (LIMES) Institute, University of Bonn, Bonn, Germany

David A. Bejarano & Andreas Schlitzer

Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, MO, USA

Martina Molgora, Jinchao Hou & Marco Colonna

Cardiovascular and Metabolic Sciences, Max Delbrück Center for Molecular Medicine in the Helmholtz Association (MDC), Berlin, Germany

Giannino Patone, Eric Lars-Helge Lindberg, Henrike Maatz, Norbert Hübner & Masatoshi Kanda

DZHK (German Centre for Cardiovascular Research), Partner Site Berlin, Berlin, Germany

Henrike Maatz & Norbert Hübner

Department of Immunology, University of Toronto, Toronto, ON, Canada

Arthur Mortha

Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK

Alexander J. Clarke

Biomathematics Division, Institute of Experimental Oncology, University Hospital Bonn, Bonn, Germany

Kevin Thurley

Institute of Physiology, University of Zurich, Zurich, Switzerland

Christoph Schneider

German Center for Child and Adolescent Health (DZKJ), partner site Berlin, Berlin, Germany

Mir-Farzin Mashreghi

Charité Universitätsmedizin, Berlin, Germany

Norbert Hübner

Department of Rheumatology and Clinical Immunology, Sapporo Medical University School of Medicine, Sapporo, Hokkaido, Japan

Masatoshi Kanda

You can also search for this author in PubMed   Google Scholar

Corresponding authors

Correspondence to Andreas Diefenbach , Masatoshi Kanda or Antigoni Triantafyllopoulou .

Supplementary information

Reporting summary, supplementary table 1.

(whole kidney scRNA-seq dataset) Whole kidneys from NZB/W F1 mice were analysed by scRNA-seq. (a) Differentially expressed genes across clusters of whole.

Supplementary Table 2

(kidney NKp46+ ILC scRNA-seq Dataset) Tissue NKp46+ ILC isolated from NZB/W F1 mice were analysed by scRNA-seq. (a) Differentially expressed genes across clusters of tissue NKp46+ ILC (related to Fig. 2a). (b) Comparison of differentially expressed genes within cluster 6 from nephritis vs. young conditions (related to Figure 2c).

Supplementary Table 3

(kidney ILC RORgtfm scRNA-seq Dataset) Tissue NKp46+ ILC isolated from ROR?t-FM mice.

Supplementary Table 4

(kidney CD45+ scRNA-seq Dataset) Kidney leukocytes.

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Biniaris-Georgallis, SI., Aschman, T., Stergioula, K. et al. Amplification of autoimmune organ damage by NKp46-activated ILC1. Nature (2024). https://doi.org/10.1038/s41586-024-07907-x

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• Published: 14 August 2024

Urine-derived podocytes from steroid resistant nephrotic syndrome patients as a model for renal-progenitor derived extracellular vesicles effect and drug screening

• Adele Tanzi 1   na1 ,
• Lola Buono 1   na1 ,
• Cristina Grange 2 ,
• Corinne Iampietro 1 ,
• Alessia Brossa 1 ,
• Fanny Oliveira Arcolino 3 , 4 ,
• Maddalena Arigoni 1 ,
• Raffaele Calogero 1 ,
• Laura Perin 5 ,
• Silvia Deaglio 2 ,
• Elena Levtchenko 3 , 6 ,
• Licia Peruzzi 7   na2 &
• Benedetta Bussolati   ORCID: orcid.org/0000-0002-3663-5134 1   na2  

Journal of Translational Medicine volume  22 , Article number:  762 ( 2024 ) Cite this article

Metrics details

Personalized disease models are crucial for evaluating how diseased cells respond to treatments, especially in case of innovative biological therapeutics. Extracellular vesicles (EVs), nanosized vesicles released by cells for intercellular communication, have gained therapeutic interest due to their ability to reprogram target cells. We here utilized urinary podocytes obtained from children affected by steroid-resistant nephrotic syndrome with characterized genetic mutations as a model to test the therapeutic potential of EVs derived from kidney progenitor cells (nKPCs).

EVs were isolated from nKPCs derived from the urine of a preterm neonate. Three lines of urinary podocytes obtained from nephrotic patients’ urine and a line of Alport syndrome patient podocytes were characterized and used to assess albumin permeability in response to nKPC-EVs or various drugs. RNA sequencing was conducted to identify commonly modulated pathways after nKPC-EV treatment. siRNA transfection was used to demonstrate the involvement of SUMO1 and SENP2 in the modulation of permeability.

Treatment with the nKPC-EVs significantly reduced permeability across all the steroid-resistant patients-derived and Alport syndrome-derived podocytes. At variance, podocytes appeared unresponsive to standard pharmacological treatments, with the exception of one line, in alignment with the patient’s clinical response at 48 months. By RNA sequencing, only two genes were commonly upregulated in nKPC-EV-treated genetically altered podocytes: small ubiquitin-related modifier 1 (SUMO1) and Sentrin-specific protease 2 (SENP2). SUMO1 and SENP2 downregulation increased podocyte permeability confirming the role of the SUMOylation pathway.

Conclusions

nKPCs emerge as a promising non-invasive source of EVs with potential therapeutic effects on podocytes with genetic dysfunction, through modulation of SUMOylation, an important pathway for the stability of podocyte slit diaphragm proteins. Our findings also suggest the feasibility of developing a non-invasive in vitro model for screening regenerative compounds on patient-derived podocytes.

Introduction

Podocytes are highly dynamic and terminally differentiated renal cells that act as the final barrier to proteins during glomerular blood filtration, thus playing a pivotal role in controlling glomerular permeability [ 1 ]. Damage to podocytes can result in primary nephrotic syndrome, a prevalent pathology in children characterized by heavy proteinuria, edemas, hypoalbuminemia and hyperlipidemia, with an annual incidence of 2–7 cases per 100,000 children. The standard treatment involves steroid therapy, but around 10% of patients are resistant (steroid-resistant nephrotic syndromes) [ 2 , 3 , 4 ] and eventually progress to end-stage kidney disease [ 5 ].

The advent of reliable genetic testing has facilitated the identification of causative genetic mutations in one-third of steroid-resistant cases [ 6 ]. Glomerular genetic variants affecting structural or secreted podocyte proteins can disrupt multiple podocyte functions. The main mutations involve genes encoding for slit diaphragm proteins (NPHS1 coding for nephrin, and NPHS2 coding for podocin), and for mesangial matrix synthesis (among which COL4A3, COL4A4 and COL4A5, mutated in Alport syndrome and in a percentage of cases of steroid-resistant nephrotic syndrome, as well as LAMB2 coding for laminin subunit beta) [ 6 , 7 ]. These disruptions can lead to high-grade proteinuria and severe nephrotic syndrome that often progress to a rapid decline in kidney function within a few years [ 8 ]. Rate of progression can be forecasted basing on the expected impact of the variant on the protein expression but still, several clinical modifiers (hypertension, response to supportive therapy, other clinical additional risk factors) can influence the individual slope of kidney function decline [ 9 ]. For these reasons, in vitro tools to evaluate the response of the single subjects to treatments are highly warranted, with the aim to avoid the multiple attempts to induce proteinuria reduction and the burden of adverse events and toxicity.

Focal detachment of podocytes is a phenomenon observed in cases of massive proteinuria in both experimental [ 10 ] and human diseases [ 11 ]. Given this, isolating podocytes from urine samples has acquired interest for understanding their pathological characteristics and identifying potential therapeutic responses [ 12 ]. In line with this, we recently isolated and characterized podocytes from the urine of patients with Alport syndrome [ 13 ].

Extracellular vesicles (EVs), cell-released vesicles involved in cell-to-cell communication, are gaining attention as biological tools with regenerative potential [ 14 ]. In renal pathology, EVs derived from stem cells of different sources exert anti-apoptotic, anti-inflammatory, and pro-angiogenic effects, possibly through the transfer of mRNAs, miRNAs, and proteins to renal cells [ 15 , 16 , 17 , 18 ]. We recent demonstrated the beneficial effect of mesenchymal stromal cell-derived EVs (MSC-EVs) on podocyte injury in an in vitro millifluidic model of the glomerular filtration barrier [ 19 ]. Similarly, EVs derived from endothelial were shown to inhibit complement-induced podocyte apoptosis, prevent nephrin shedding, and maintain permselectivity during inflammatory damage [ 20 ]. However, the potential effect of stem cell-derived EVs in treating genetically altered podocytes remains unexplored.

In this study, we hypothesized that EVs derived from neonatal kidney progenitor cells (nKPCs) could offer specific benefits to podocytes, given their renal origin. These cells, isolated from preterm neonatal urine were previously reported to express embryonic kidney transcription factors, including SIX2 and CITED1 [ 21 ], and were shown to protect renal cells from hypoxic damage in a model of kidney graft perfusion [ 22 ]. To explore this hypothesis, we used podocytes with genetic alterations obtained from the urine of children with steroid-resistant nephrotic syndrome and we evaluated the effects and potential mechanisms of nKPC-EVs. These were compared with common drugs known to modulate podocyte permeability. Additionally, we employed urinary conditionally immortalized podocytes from an Alport syndrome patient as a model for genetically diseased podocytes with altered permeability.

Ethical statement

All the enrolled subjects provided informed written consent. The study protocol was approved by the Bioethics Committee of the A.O.U. Città della Salute e della Scienza Hospital (protocol no. 0021671). The study was conducted according to the principles expressed by the Declaration of Helsinki of 1975, as revised in 2013.

Generation of podocyte cell lines

A total of three patients diagnosed with steroid-resistant nephrotic syndrome were recruited in this study. Steroid resistance was defined according to the global consensus [ 23 ] as persistence of proteinuria in nephrotic range without significant reduction from onset after a course of steroid of 4 weeks prednisone 60 mg/m2/day, followed by 3 methylprednisolone pulses of 10 mg/kg each on three consecutive days and two more weeks of observation. If proteinuria is not modified by this treatment the child is classified as “steroid resistant” and is addressed to genetic study. At the time being a panel of 70 genes associated to steroid resistant nephrotic syndrome encoding for proteins involved in podocyte functions is explored [ 23 ]. Genetic and clinical features of patients are described in Table  1 . ACMG classification refers to the time of genetic diagnosis. During follow up patients 1 and 2 did not respond to all the treatments attempted, progressed to CKD 5 (chronic kidney disease, stage 5) and were successfully transplanted without recurrence. Patient 3, which initially proved to be steroid resistant, had a late satisfactory response to steroids combined with angiotensin-converting enzyme inhibitors and one single dose of Rituximab, and after 48 months of follow up has a normal kidney function and minimal proteinuria (Table  1 ). As control, we used urine from a healthy pregnant woman, a valuable source for control podocytes due to pregnancy-associated podocyturia [ 24 ].

Urine samples (~ 50 ml) were freshly centrifuged at 200 g for 10 min. The pellet was resuspended in DMEM/F-12 (Life Technologies, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% foetal calf serum (FCS; Invitrogen), 50 IU/ml penicillin, 50 g/ml streptomycin, 5 mM glutamine, 5 g/ml insulin, 5 g/ml transferrin, and 5 mg/ml selenium (all from Sigma-Aldrich, St Louis, MO, USA). Subsequently, primary cells were grown at 37 °C up to the third passage and characterized as podocytes. Three lines of steroid resistant nephrotic syndrome podocytes (NS-POD1-3) and a control urine podocyte line (CTL-uPOD) were generated.

In addition, a conditionally immortalized podocyte cell line (AS-POD) from urine of an Alport syndrome patient, previously isolated and characterized in our laboratory, was used [ 13 ]. Finally, tissue-derived conditionally immortalized podocytes, which were obtained from a nephrectomy of a healthy subject, were kindly gifted by MA Saleem and used as control for selected experiments (CTL-tPOD) [ 25 ]. AS-POD and CTL-tPOD were maintained in culture as previously described [ 26 ]. Briefly, cells were grown at a permissive temperature of 33 °C and, 8–10 days before experiments, moved to 37 °C, to ensure growth arrest and differentiation. The culture medium for AS-POD and CTL-tPOD was the same of primary podocyte cell lines.

Generation of nKPC cell line

Urine sample (500 mL) was collected from a newborn (born at 32 gestational weeks) at day 1 after birth from a catheter bag, as described [ 21 ]. The sample was centrifugated at 200 g for 10 min and the cell pellet was resuspended in α-MEM Medium (Gibco, Thermo Fisher Scientific) supplemented with 20% Chang Medium B (Irvine Scientific, Santa Ana, California, USA) and 2% Chang Medium C (Irvine Scientific), 20% FCS (Invitrogen), 50 IU/mL penicillin, 50 g/mL streptomycin, 5 mM glutamine (all from Sigma‐Aldrich). At passage 2, 1 × 10 4 primary nKPCs were infected in DMEM F12 20% FCS with a retrovirus containing a pBABE-puro-hTERT plasmid (Addgene plasmid #1771) [ 27 ]. The day after the infection, the medium was replaced with their growth medium. From passage 3, cells were selected in their growth medium containing 1 µg/mL puromycin (Gibco) for four weeks, until passage 7.

Non-renal cell lines

Primary human umbilical vein endothelial cells (HUVEC) were purchased from ATCC (ATCC-PCS-100-010, Manassas, VA, USA). Human corneal endothelial cells were previously isolated from discarded cornea patients undergoing corneal transplantation or enucleation and characterized in our laboratory [ 28 ]. For HUVEC and corneal endothelial cells, EndoGRO-VEGF (Merck Millipore, Burlington, MA, USA) supplemented with 5% FCS (Invitrogen), was used as culture media. Breast cancer cells were previously isolated, grown in DMEM-F12 supplemented with 10 ng/mL basic fibroblast growth factor, 20 ng/mL epidermal growth factor, 5 µg/mL insulin, and 0.4% bovine serum albumin (all from Sigma-Aldrich), and characterized in our laboratory [ 29 ]. Human bone marrow-derived MSCs were purchased from Lonza (Switzerland) and grown in mesenchymal stem cells basal medium (MSCBM, Lonza) [ 28 ].

EV isolation

nKPC-EVs and MSC-EVs were obtained respectively from the supernatant of nKPCs and MSCs cultured overnight in serum free conditions (RPMI 0% FCS), as previously described [ 19 , 28 ]. Serum-EVs were obtained from a total of 100 mL of serum isolated from a blood pool of five healthy donors, as previously mentioned [ 30 ]. Informed consent was obtained by the Blood Bank of “Città della Salute e della Scienza di Torino” from all the donors. Culture supernatant and serum were centrifuged for the removal of cell debris and apoptotic bodies at 3,000 g for 20 min. EVs were then purified by a 2 h-ultracentrifugation at 100,000 g at 4 °C (Beckman Coulter, Brea, CA, USA) and used fresh or stored at − 80 °C after resuspension in RPMI without FSC and supplemented with 1% dimethyl sulfoxide. Analysis of the size distribution and particle quantification were performed using NanoSight NS300 (NanoSight Ltd, Malvern, UK) equipped with a 405 nm laser and the Nanoparticle Tracking Analysis (NTA) 2.3 software (NanoSight Ltd).

Protein extraction and western blot

For protein analysis, different EV preparations (about 5) were pooled to obtain 10 11 particles and further ultracentrifuged. Subsequently, the EV pellet was resuspended in lysis buffer, composed as follow: 1:100 Phosphatase Inhibitor Cocktail 2, 1:100 Phosphatase Inhibitor Cocktail 3, 1:100 Phenylmethanesulfonyl fluoride (PMSF), 1:100 Protease Inhibitor Cocktail (all from Sigma-Aldrich), in RIPA buffer (20 nM Tris·HCl, 150 nM NaCl, 1% deoxycholate, 0.1% SDS 1% Triton X-100, pH 7.8, Sigma-Aldrich). After 20 min of incubation at 4 °C, the protein extract was centrifugated for 15 min at 14’000 g and the supernatant was used. Podocyte and nKPC pellets were similarly resuspended in lysis buffer for protein extraction. Protein concentration of podocyte lysate was determined by Bradford solution, according to the manufacturer’s procedures (Bio-Rad Inc, Berkeley, CA, USA). At variance, total protein concentration of nKPCs and nKPC-EVs was determined spectrophotometrically using a micro-BCA™ Protein Assay Kit (Thermo Fisher Scientific), as previously described [ 31 ]. Either 30 µg (for podocytes) or 8 µg (for nKPCs and nKPC-EVs) of proteins were electrophoresed through 4–12% Mini-Protean TGX Stain-Free Gels (Bio-Rad). Using the iBLOT2 system (Life Technologies, Carlsbad, CA, USA), gels were blotted onto PVDF membrane filters according to the manufacturer’s procedures. Each membrane was incubated with blocking solution, consisting in 5% bovine serum albumin (BSA; Sigma-Aldrich) in PBS, for 1 h before overnight incubation with primary antibodies at the indicated dilutions. After rinsing in wash buffer (0.1% Tween in PBS), horseradish peroxidase-conjugated secondary antibodies were used for 1 h at 1:3000–1:5000 dilutions. After final washings, membranes were incubated with ECL chemiluminescence reagent (Bio-Rad, Milan, Italy). Images were acquired using a ChemiDoc ™ XRS + System (Bio-Rad). For podocytes analysis, the following antibodies were used: rabbit monoclonal anti-Podocin (Cat. No. sc-21009; Santa-Cruz, Dallas, TX, USA), and mouse monoclonal anti-CD2AP (Cat. No. sc-25272, Santa-Cruz). Goat monoclonal anti-Vinculin (Cat. No. sc-7648, Santa-Cruz) and mouse monoclonal anti-PCNA (Cat. No. sc-56, Santa-Cruz) were used as housekeepings. In the case of nKPC and nKPC-EV protein analyses, mouse monoclonal anti-CD63 (Cat. No. sc-5275, Santa-Cruz), rabbit monoclonal anti-Calreticulin (Cat. No. 2891, Cell signalling, Milan, Italy) and mouse monoclonal anti-TSG101 (Cat. No. sc-7964, Santa-Cruz) antibodies were used. The protein bands were detected using either rabbit, mouse, or goat peroxidase-labeled secondary antibodies.

Immunofluorescence

Immunofluorescence on podocytes was performed as follows: cells were plated at a density of 4 × 10 4 cells/cm 2 and the next day fixed in 4% paraformaldehyde for 20 min at room temperature and permeabilized with PBS 0.1% Triton X-100 (Sigma-Aldrich) for 10 min at 4°C. PBS 1.5% BSA (Sigma-Aldrich) was used to block non-specific sites for 20 min at room temperature. Subsequently, Texas Red-X Phalloidin (Cat. No. T7471, Thermo Fisher Scientific) was incubated for 1 h. Fixed cells were washed with PBS 0.1% BSA before nuclear staining with 4.6-diamidine-2-phenylindole (DAPI, Sigma-Aldrich) for 8 min. After the final wash, coverslips were mounted with Fluoromount (Sigma-Aldrich). Images were acquired by the videoconfocal system ViCo microscope Nikon Eclipse 80i (Nikon, Japan).

Transmission electron microscopy

The transmission electron microscopy (TEM) was performed on EVs fixed in glutaraldehyde placed on 200-mesh nickel formvar carbon-coated grids (Electron Microscopy Science) for 20 min to promote adhesion. The grids were then incubated with 2.5% glutaraldehyde plus 2% sucrose. EVs were negatively stained with NanoVan (Nanoprobes, Yaphank, NY, USA) and observed using a Jeol JEM 1400 Flash electron microscope (Jeol, Tokyo, Japan) [ 32 ].

Super-resolution microscopy

Super-resolution microscopy was performed with Nanoimager S Mark II microscope from ONI (Oxford Nano-imaging, Oxford, UK) equipped with a 100x, 1.4NA oil immersion objective, an XYZ closed-loop piezo 736 stage, and triple emission channels split at 640, 555 and 488 nm on nKPC-EV. EV profiler Kit (EV-MAN-1.0, ONI) was used for the experiments following manufacturer’s protocol. The Kit contains fluorescent antibodies, anti CD9-488, CD63-568 and CD81-647, washing buffer and the imaging buffer. Images were acquired sequentially in dSTORM mode in total reflection fluorescence (TIRF). Single-molecule data was filtered using NimOS software (v.1.18.3, ONI). Data analysis was conducted using Collaborative Discovery (CODI) online analysis platform www.alto.codi.bio from ONI and the drift correction pipeline version 0.2.3 was used [ 33 ].

RNA isolation, real time PCR and RNA sequencing analysis

Total RNA of patient-derived podocytes untreated or treated for 24 h with nKPC-EVs or MSC-EVs (5 × 10 4 EVs/cell), as well as RNA of CTL-tPOD untrasfected or transfected with siRNAs, was isolated using Trizol Reagent (Ambion, Austin, TX, USA) according to the manufacturer’s protocol. At variance, RNA of nKPCs and of nKPC-EVs was extracted using miRNeasy mini kit (Qiagen GmbH, Hilden, Germany) according to the manufacturer’s protocol. RNA was then quantified spectrophotometrically (Nanodrop ND-1’000, Wilmington, NC, USA). For the gene expression analysis, quantitative real-time PCR (RT-PCR) was performed. Briefly, one-strand cDNA was produced from 200 ng of total RNA using a High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems, Waltham, MA, USA). RT-PCR experiments were performed in a 20 µL-reaction mixture containing 5 ng of cDNA template, the sequence-specific oligonucleotide primers (purchased from MWG-Biotech, Eurofins Scientific, Brussels, Belgium), and the Power SYBR Green PCR Master Mix (Applied Biosystems). GAPDH mRNA was used to normalize the RNA inputs. The fold change expression with respect to the control was calculated for all the samples. Primer list can be found in supplementary data (Supplementary table 1 ). For the RNA sequencing analysis, libraries for RNA-seq were generated using a TruSeq RNA stranded sample preparation kit v2 (Illumina Inc, San Diego, CA, USA) following the manufacturer’s instructions, using 1 µg of total RNA as input material. Libraries were pooled and sequenced with a NextSeq 500 sequencer (Illumina Inc) generating 75-bp paired-end sequences. Further analyses were performed using transcript per million (TPM) tables and genes with an average |log 2 Fold Change| ≥1 were considered for further analysis using Expression Suite and Funrich V3 Software (Bundora, Australia).

Flow cytometry

After puromycin selection, nKPCs were detached using a nonenzymatic cell dissociation solution, resuspended in PBS 0.1% BSA (Sigma-Aldrich) and incubated with antibodies. Cells were incubated with either phycoerythrin (PE)-, fluorescein isothiocyanate (FITC)-, or allophycocyanin (APC)-conjugated antibodies against CD90 (Cat. No. 130-114-859, Miltenyi Biotec, Bergisch Gladbach, Germany) CD73 (Cat. No. 550257, BD Bioscience, Franklin Lakes, NJ, USA), CD146 (Cat. No. 550315, BD Bioscience) and CD29 (Cat. No. 130-101-256, Miltenyi Biotec) CD133-1 (Cat. No. 130-090-826, Miltenyi Biotec) and appropriate isotype control. Stained cells were then analyzed using FACSCalibur machine using CellQuest software (Becton Dickinson Bioscience Pharmingen).

MACSPlex analysis

nKPC-EVs were subjected to bead-based multiplex EV analysis by flow cytometry (MACSPlex Exosome Kit, human, Miltenyi Biotec) as previously described [ 33 ]. Briefly, 5 × 10 9 EVs were diluted with a MACSPlex buffer (MPB) to a final volume of 120 µL and 15 µL of MACSPlex Exosome Capture Beads (containing 39 different antibody-coated bead subsets) were added to each sample. The samples were then incubated on an orbital shaker overnight (14–16 h) at 450 rpm at 4 °C protected from light, followed by several washings with MPB using centrifugations (3’000 g, 5 min). For EV counterstaining, 5 µL of each APC-conjugated anti-CD9, anti-CD63, and anti-CD81 detection antibodies were added to each sample and incubated on an orbital shaker at 450 rpm for 1 h at room temperature. After additional washings, samples were subjected to flow cytometric analysis using FACS Celesta (BD Biosciences, New Jersey, USA).

Permeability assay

Permeability assays were performed in a 24 well plate using cell culture inserts with a pore size of 0.4 μm (Corning, New York, USA). Four x 10 4 podocytes were plated on the insert and left adhere overnight. Subsequently, podocytes were treated for 24 h with nKPC-EVs (2–5 × 10 4 EVs/cell), methylprednisolone (Urbason, Sanofi, 40 µg/mL), cyclosporin (Sandimmun, Novartis, 25 µg/mL), tacrolimus (Prograf, Panacea Biotec, 1 ng/mL) or rituximab (Mabthera, Roche, 12 µg/mL). Subsequently, FBS-free medium (500 µL) containing or not FITC-BSA (1 mg/mL, Sigma-Aldrich) was placed in the lower compartment and upper podocyte compartments, respectively. After 6 h, BSA filtration from the lower to the upper compartment was measured, to evaluate the podocyte filtration ability in basal to apical direction. Therefore, 100 µl of medium was taken from the upper compartment and the passage of FITC-BSA was determined by fluorimetry in triplicate, using Promega™ GloMax ® Plate Reader (Promega Italia S.r.l., Milano, Italy). Data are expressed as the mean amount of filtered BSA-FITC of four different experiments using at least three inserts for each condition in each experiment.

Podocyte transfection

Transfection of CTL-tPOD was performed using Lipofectamine RNAiMAX Reagent (Invitrogen). Briefly, 4 × 10 4 CTL-tPOD/well were plated in cell culture inserts, as described for the permeability assay, left adhere overnight at 33 °C and therefore moved at 37 °C, to promote differentiation. After 8 days at 37 °C, cells were transfected with 1.2 µl of Lipofectamine and 10 picomol of the specific MISSION esiRNAs (SUMO1 and SENP2; Merck), according to the manufacturer’s instructions. The day after transfection, fresh growth medium was replaced and 48 h later the cells were used for the permeability assay. In parallel, transfected CTL-tPOD were lysed, RNA was extracted, and RT-PCR was performed to verify downregulation of SUMO1 and SENP2 mRNAs. Cells transfected with MISSION negative control (Merck) were used as control.

Statistical analysis

Data are shown as mean ± SD. Statistical analysis was carried out on Graph Pad Prism (GraphPad Software, Inc., San Diego, CA, USA) by using one-way analysis of variance (ANOVA) followed by Dunnet’s multiple comparisons test, or by unpaired t-test, where appropriate. A p value < 0.05 was considered significant.

Podocyte isolation and characterization

Podocytes were obtained from freshly collected urine derived from three different patients presenting steroid-resistant nephrotic syndrome (NS-POD). Genetically characterized mutations are described in Table 1 . Podocytes isolated from healthy pregnant woman’s urine were used as control (CTL-uPOD). Additionally, conditionally immortalized podocytes, isolated from the urine of a patient with Alport syndrome and previously characterized in our laboratory [ 13 ], were included in the study (AS-POD) (Table 1 ).

Urine-derived cells were characterized by the presence of podocyte markers such as podocin and CD2AP (Fig.  1 A). By morphology, cells showed an organized cytoskeletal structure similar to already well-characterized podocytes (Fig.  1 B) [ 13 ]. Non-renal cell appeared negative for podocytes’ markers (Supplementary Fig.  1 ).

Isolation and characterization of podocytes from urine. ( A ) Western Blot analysis (representative images and quantification) of podocytes derived from urine of three different patients (NS-POD1, NS-POD2, NS-POD3) and of a healthy pregnant woman (CTL-uPOD) positive for CD2AP and podocin. Vinculin was used as housekeeping. Data are expressed as mean of two experiments ± SD. ( B ) Representative micrographs of podocytes deriving from urine of CTL-uPOD, NS-POD1, NS-POD2 and NS-POD3 stained with phalloidin (red) and blue nuclear stain DAPI. Original magnification: X20

The podocyte phenotype was also assessed by analyzing the expression of a specific signature composed of 68 podocyte-typical genes (Supplementary Fig.  2 ), as previously outlined by Lu et al., [ 13 , 34 ]. All three podocyte lines expressed the typical podocyte genes, thereby confirming the podocyte phenotype (Supplementary Fig.  2 ) [ 13 ].

Kidney progenitor-derived EV isolation and characterization

EVs were isolated by differential centrifugations from immortalized nKPCs. These cells were originally obtained from the urine of a preterm infant and characterized as previously described [ 21 ]. The size distribution of nKPC-derived EVs was analyzed using nanosight tracking analysis (NTA), revealing a mean size distribution of approximately 142.7 nm (Fig.  2 A). Western Blot analysis confirmed the presence of the EV-specific markers CD63 and TSG101, while the absence of the cytoplasmic marker calreticulin indicated the absence of cellular contamination (Fig.  2 B). TEM image revealed the typical cup-shaped morphology of EVs (Fig.  2 C). Additionally, super-resolution microscopy demonstrated that nKPC-EVs expressed CD9, CD63, and CD81, typical EV markers, in various combinations, predominantly being CD63 and CD9 single positive (Fig.  2 D-E).

Characterization of nKPC-EVs. ( A ) Representative NTA analysis showing the EV size distribution; EV mean size is about 142.7 nm. ( B ) Representative Western Blot images showing the presence in nKPC-EVs of CD63 and TSG101, and the absence of calreticulin. ( C ) Representative micrograph of TEM of nKPC-EVs (scale bar: 200 nm). ( D ) Clustering analysis of super-resolution microscopy images shows the single, double, and triple positive EV fractions expressing the tetraspanins (CD9, CD63, CD81). The analyses were performed in 3 nKPC-EV preparations using a CODI software; the graph shows the mean ± SD of a cumulative analysis of 3 fields for each preparation. ( E ) Representative super-resolution microscopy images of nKPC-EVs showing expression of CD63 (green), CD81 (red) and CD9 (blue). The scale bars are below each EV image

The expression of mesenchymal and renal progenitor markers in nKPCs and deriving EVs was also evaluated. Notably, nKPCs exhibited a mesenchymal phenotype, expressing CD90, CD73, CD146, and CD29 (Fig.  3 A), along with the cytoplasmatic nephron progenitor marker SIX2 and stromal progenitor marker FOXD1 (Fig.  3 B), as previously reported [ 21 ]. Conversely, nKPCs were negative for markers associated with adult renal progenitor cells, such as CD133 (Fig.  3 A) [ 35 ]. Similarly, using a bead-based immunocapture assay, nKPC-EVs were found to express mesenchymal stromal markers CD146, CD29, and CD44, in addition to the typical EV markers (CD9, CD63, and CD81). At variance, no expression of CD133 or stage-specific embryonic antigen-4 (SSEA-4) was detected (Fig.  3 C). Furthermore, nKPC-EVs expressed SIX2 and FOXD1 mRNAs, as demonstrated by RT-PCR, as the originating cells (Fig.  3 D).

nKPC and nKPC-derived EV characterization of intracellular and surface markers expression. ( A ) Representative flow cytometry dot plots of nKPCs showing positive expression of CD73, CD90, CD29 and CD146; nKPCs resulted negative for CD133 marker. ( B ) RT-PCR analyses showing the expression of FOXD1 and SIX2 in nKPCs. GAPDH was used as endogenous normalizer. Data were further normalized to HUVEC cells, used as a negative control for each experiment. The graphs show the RQ average (2 −∆∆Ct ) of three independent experiments ± SD. ( C ) Quantification of the median APC fluorescence for each bead population after background correction of exosomal and mesenchymal/stem cell markers in nKPC-EVs. The fluorescence intensity of each marker was normalized to the mean fluorescence intensity of all detectable markers to 1000. Data are expressed as the average of two technical replicates ± SD. ( D ) RT-PCR analysis showing the expression of FOXD1 and SIX2 in nKPC-EVs. GAPDH was used as endogenous normalizer. Data were further normalized to HUVEC cells, used as a negative control for each experiment. The graphs show the RQ average (2 −∆∆Ct ) of two independent experiments ± SD

Permeability analysis of podocyte cultures

To evaluate whether nKPC-EVs could modulate permselectivity of the patient-derived podocytes, we used an in vitro permeability assay. Podocytes were seeded on the upper side of inserts and permeability was assessed by measuring the transit of FITC-BSA from the lower compartment to the upper podocyte compartment as described in the graphical representation (Fig.  4 ).

Schematic experimental design and graphic representation of culture set-up. Graphical representation of nKPC and patient-derived podocyte isolation and EV production. Podocytes were plated in the upper compartment of transwell inserts and treated with EVs or drugs. For the permeability assay, the BSA-FITC filtration was measured. Abbreviations NS-POD: nephrotic syndrome podocytes; AS-POD: Alport syndrome podocytes; CTL-uPOD: control urinary podocytes; M-PR: methylprednisolone; CYCLO: cyclosporin; TAC: tacrolimus; RITUX: rituximab. The figure was created with Biorender.com

To evaluate the efficacy of nKPC-EV administration in the modulation of podocyte permeability, experiments were performed using two different EV doses (2 × 10 4 and 5 × 10 4 EVs/cell). A significant reduction of permeability was observed using the 5 × 10 4 EVs/cell dose on all NS-POD lines, as well as on AS-POD (Fig.  5 A). No significant changes were observed on control urine podocytes (CTL-uPOD) (Fig.  5 A). The reduction of permeability was lacking when serum-EVs were used on AS-POD. Moreover, nKPC-EV effect was compared to that of commonly used drugs for treating renal diseases: methylprednisolone (M-PR), cyclosporin (CYCLO), tacrolimus (TAC), and rituximab (RITUX). Indeed, all these drugs have been reported to exert a direct anti-proteinuric action on podocytes, beside immunomodulation [ 36 , 37 , 38 ]. Drug treatment did not affect podocyte viability (Supplementary Fig.  3 ). No modulation of permeability by pharmacological treatments was observed in the genetically altered podocytes, except for NS-POD3 (Fig.  5 B). Interestingly, the patient generating NS-POD3 cells clinically responded to treatment with steroids, cyclosporin and rituximab, showing at 48 months a clinical remission (see methods and Table 1 ). Permeability of normal podocytes (CTL-uPOD) was significantly lower in respect to diseased podocytes (Fig.  5 B), as described [ 13 ].

Permeability assay in podocyte cultures treated with nKPC-EVs and different drugs. ( A ) Podocytes were treated with different doses of nKPC-EVs (20 K: 2 × 10 4 EVs/cell, 50 K: 5 × 10 4 EVs/cell). Serum-EVs were used as control. Untreated cell condition was used as control for each experiment, set as BSA filtration rate of 100%. Data are expressed as the mean amount of filtered BSA-FITC of four different experiments using at least three inserts for each condition in each experiment ± SD. One-way ANOVA with Dunnett’s multiple comparisons test was performed after the normalization of each experiment to untreated podocytes; * p  < 0.05, ** p  < 0.01, *** p  < 0.001, **** p  < 0.0001, vs. the respective untreated condition. ( B ) Podocytes were treated with nKPC-EVs (5 × 10 4 EVs/cell) or the different drugs methylprednisolone (M-PR, 40 µg/mL), cyclosporin (CYCLO, 25 µg/mL), tacrolimus (TAC 1 ng/mL) and rituximab (RITUX 12 µg/mL). Untreated cell condition was used as control for each experiment, set as BSA filtration rate of 100%. Data are expressed as the mean amount of filtered BSA-FITC of four different experiments using at least three inserts for each condition in each experiment ± SD. One-way ANOVA with Dunnett’s multiple comparisons test was performed after the normalization of each experiment to untreated podocytes; * p  < 0.05, ** p  < 0.01, *** p  < 0.001, **** p  < 0.0001, vs. the respective untreated condition

Transcriptomic analysis of urine-derived podocytes treated with EVs

To explore the mechanism involved in the observed effect on permeability, we analyzed the change in the RNA profile of the NS-POD and AS-POD after treatment with 5 × 10 4 nKPC-EVs/cell. The transcripts per kilobase million (TPM) normalization method was employed to compare changes in the gene expression profile between the samples. Our analysis revealed a total of 2876 upregulated genes (log 2 fold change > 1) and 2574 downregulated genes (log 2 fold change <-1) in response to EV treatment (Fig.  6 A-B). Gene Ontology analysis highlighted cell-to-cell adhesion as the primary upregulated biological activity in treated podocytes (Fig.  6 C-D). Only two genes, SUMO1 and SENP2, resulted to be commonly upregulated in all four samples post nKPC-EV treatment (Fig.  6 F), showing a log 2 fold change ranging between 3.07 and 3.42 (Fig.  6 E-F). At variance, no gene was consistently downregulated in all patient-derived podocyte cell lines (not shown). Interaction network analysis demonstrated a direct interaction between these two genes in the SUMOylation pathway [ 39 , 40 ].

Cross-analysis for the identification of regulated genes common to the four patient-derived podocytes. ( A ) Pie chart representing the sum of all the genes differentially expressed in the four different podocyte lines after the treatment with nKPC-EVs (in blue), compared with the non-differentially expressed genes (grey). ( B ) Pie chart representation of up-regulated (green) and down-regulated (red) differentially expressed transcripts in the four different podocyte lines untreated and treated with EVs. C and D . Gene Ontology analysis of differentially expressed genes in the four different podocyte lines untreated and treated with EVs. In each table, the identification (ID) number, the name, and the P value associated with the GO are given. E . Representative Venn diagram showing the numbers of the genes that resulted up-regulated after the treatment of the podocytes with nKPC-EVs by total RNA sequencing analysis. Data were analyzed using Expression Suite and Funrich V3 Software. F . Heatmap showing the levels of expression regulation of the two genes which were upregulated in all the four podocyte lines

The nKPC-EV-induced up-regulation of both SUMO1 and SENP2 in urine-derived podocytes was confirmed by RT-PCR, where the up-regulation reached significance in three podocytes lines from nephrotic syndrome patients (Fig.  7 A-B). No effect on SUMO1 or SENP2 modulation was observed using MSC-EVs (not shown). Subsequently, we aimed at confirming the relevance of SUMO1 and/or SENP2 on podocyte function. For this purpose, we performed the permeability assay using CTL-tPOD that had been downregulated for either SUMO1 or SENP2 genes by transfection with siRNA (Fig.  7 C). Permeability assay revealed an increased BSA-FITC permeability in CTL-tPOD transfected with SENP2 siRNA either alone or combined with SUMO1 siRNA (Fig.  7 D).

Role of SUMO1 and SENP2 in podocyte permeability. A - B . Validation of differentially expressed genes in urine-derived podocytes treated with EVs. mRNA expression of SUMO1 ( A ) and SENP2 ( B ) genes in urine-derived podocytes treated or not with nKPC-EVs. Data are shown as relative quantification, normalized to GAPDH and to each untreated control respectively set as 1. The graphs show the RQ average (2 −∆∆Ct ) of at least three independent experiments ± SD. Unpaired t-test was performed after the normalization of each experiment to its untreated condition (AS-POD, NS-POD1, NS-POD 2, NS-POD3, respectively); * p  < 0.05, ** p  < 0.001, *** p  < 0.0001 vs. AS-POD, NS-POD1, NS-POD 2, NS-POD3, respectively. C . RT-PCR analysis of SUMO1 and SENP2 genes in CTL-tPOD after siRNA transfection. Data are shown as relative quantification, normalized to GAPDH and to the untransfected control (CTL), set as 1. The graphs show the RQ average (2 −∆∆Ct ) of two independent experiments ± SD. One-way ANOVA with Dunnett’s multiple comparisons test was performed; * p  < 0.05, ** p  < 0.01, *** p  < 0.001, vs. SCR. D . Permeability assay using CTL-tPOD transfected with siRNA targeting SUMO1 and/or SENP2 genes. Data are expressed as the mean amount of filtered BSA-FITC of at least six different experiments for each condition. One-way ANOVA with Dunnett’s multiple comparisons test was performed after the normalization of each experiment to untransfected podocytes (CTL); * p  < 0.05, ** p  < 0.01, *** p  < 0.001, **** p  < 0.0001, vs. SCR. Abbreviations CTL: untransfected CTL-tPOD; LP: CTL-tPOD treated only with Lipofectamine reagent; SCR: CTL-tPOD transfected with a scramble sequence; SUMO1 siRNA: CTL-tPOD transfected with siRNA targeting SUMO1; SENP2 siRNA: CTL-tPOD transfected with siRNA targeting SENP2; SUMO1 + SENP2 siRNA: CTL-tPOD transfected with the combination of siRNA targeting SUMO1 and siRNA targeting SENP2

The current study provides a novel approach to directly assess the potential of regenerative therapies on patient-derived podocytes with genetic alterations. We demonstrated the efficacy of EVs derived from neonatal renal progenitor cells in improving permeability in podocytes obtained from patients with steroid-resistant nephrotic syndrome. Furthermore, we identified the SUMOylation pathway as a possible common mechanism in all patient-derived cell lines treated with nKPC-EVs.

The availability of human podocytes that accurately replicate human pathologies in culture is currently limited. Recent advancements have allowed the derivation of podocyte-like cells from induced pluripotent stem cells from patients, offering a promising avenue [ 41 ]. However, obtaining a purely homogeneous population of podocytes is challenging; typically, only 30–50% of cells in induced pluripotent stem cell cultures exhibit podocyte-like morphology [ 42 ]. Another potential source for modeling diseases is the differentiation of urinary renal progenitor cells into podocytes, although this process may involve cell culture and differentiation stimuli [ 43 ].

Previous studies have utilized urine as a non-invasive and valuable source to obtain podocytes [ 11 ]. In patients with active glomerular diseases, podocytes are shed from the glomerulus in response to local environmental factors. Urine patient-derived podocytes have been demonstrated to be positive for podocyte markers, viable, and capable of growth in culture [ 11 ]. Conversely, due to their limited number and limited replication potential in culture, podocytes cannot be obtained from normal subjects’ urine [ 11 ]. Of interest, increased podocyturia is observed in the urine of uncomplicated pregnant women [ 24 ], making it a valuable source for obtaining healthy control podocytes.

In this context, our in vitro model of glomerular permeability utilizing podocytes directly obtained from patients’ urine holds significant promise for personalized medicine. This study includes podocytes with mutations affecting a slit diaphragm protein (podocin), matrix synthesis components (collagen IV and laminin) and a calcium balance regulator (Phospholipase C epsilon 1). We here showed the feasibility of a patient-specific permeability model that could be used in personalized medicine to evaluate the impact of drugs commonly administered to patients with nephrotic syndrome. Notably, podocytes derived from steroid resistant patients, unresponsive to therapies, also did not respond to the drugs previously reported to modulate permeability [ 36 , 37 , 38 ]. An effect on permeability was only observed on the podocyte line from a patient who, despite initially clinically steroid-resistant, exhibited subsequent clinical improvement with minimal proteinuria after 48 months (NS-POD3). In the future, a personalized approach could potentially set the stage for tests capable of predicting patient responses, guiding clinicians in treatment decisions, and avoiding unnecessary immunosuppressive interventions in the management of nephrotic syndrome. It must be however underlined that the short survival of those primary lines only allows a restricted number of tests.

Another potential application of the primary model of the glomerular filtration barrier is the evaluation of innovative biological therapeutics. We here assessed the effect of nKPS-EVs on genetically mutated patient-derived podocytes. Our findings indicate that nKPCs could serve as a valuable source of EVs with regenerative potential. Interestingly, no effect on healthy podocytes was observed, further supporting the importance of disease models to assess therapeutic approaches. Moreover, in consideration to the variability of single line response, the use of different primary cell lines is instrumental to identify a possible nKPC-EV specific mechanism of action.

EVs, in particular those obtained from MSCs, have been previously studied in in vitro and in vivo experimental models of kidney disease [ 19 , 44 , 45 , 46 , 47 , 48 ]. Using a millifluidic model of glomerular filtration barrier, we previously demonstrated that MSC-EVs could traverse the basal membrane, reach podocytes and transfer their RNA cargo, resulting in protection against doxorubicin-induced injury [ 19 ]. Other studies have underscored the protective roles of MSC-EVs on mouse podocyte lines using a model of diabetic nephropathy [ 47 , 48 ]. However, the identification of specific mechanisms involved in the effect of EVs on target cells appears quite challenging, in consideration to the multitude of EV cargoes and to the patient-derived cell heterogeneity.

In our approach, we searched for common nKPC-EV-regulated genes at the transcriptomic level. Of interest, SUMO1 and SENP2, its modulator, were identified as the only common genes concordantly regulated in all patient-derived podocyte lines. At variance, no SUMO or SENP2 modulation was detected using MSC-EV treatment. Although we did not identify the factors involved in the specific effect of nKPS-EVs on podocytes due to the complexity of EV cargo, it might be speculated that factors linked to their renal progenitor origin, such as SIX2 expression, could promote SUMO1 and SENP2 expression, known to be relevant in epithelial cell differentiation [ 22 ]. This could be supported by data obtained using the EV originating cells, nKSPCs, administered to human kidneys discarded for transplantation. In the study, the de novo expression of SIX2 in proximal tubular cells and upregulated regenerative markers, including SOX9, was observed [ 22 ].

SUMOylation pathway is considered to be involved in slit diaphragm protein stabilization, thus playing a possible role in the control of glomerular permeability [ 49 ]. Further studies will be necessary to better elucidate this point. The activity of SUMO, or SUMOylation, consists in a post-translational modification that alters the function of target proteins and modulates cell processes such as protein stability, localization, and activity [ 49 , 50 ]. Since SENP2 is a known regulator of SUMO1 [ 38 ], it may be indirectly involved in slit diaphragm stabilization as well. This was also confirmed by experiments of SUMO1 and SENP2 downregulation, demonstrating their involvement on podocyte permeability in our in vitro model. Concordantly, Gene Ontology analysis highlighted cell-to-cell adhesion as the primary upregulated biological activity in nKPC-EV treated podocytes. Therefore, the potential effect of EVs derived from kidney progenitors on genetically altered podocytes represents a promising strategy for addressing genetic pathologies currently lacking a therapeutic solution. While this strategy was tested on three podocyte lines in this study, further validation and expansion to a larger patient cohort are warranted.

In conclusion, we here set up a human in vitro functional model for the analysis of drug response on albumin permeability, thanks to combined effort of clinical, genetic and basic science. Our findings demonstrated the positive impact of nKPC-EVs in improving the function of genetically altered podocytes and identified the effect of EVs on the regulation of SUMOylation, an important pathway for stabilizing podocyte slit diaphragm proteins.

Our data may pave the road for establishing a standard non-invasive in vitro model for the screening of regenerative compounds directly on patient-derived podocytes.

Data availability

RNA sequencing data are available in the supplementary data.

Abbreviations

Extracellular vesicles

Neonatal kidney progenitor cells

Nephrotic syndrome podocytes

Alport syndrome podocytes

Control urine podocytes

Control tissue podocytes

Foetal calf serum

Bovine serum albumin

Nanoparticle tracking analysis

Real-time PCR

Phycoerythrin

Fluorescein isothiocyanate

Allophycocyanin

MACSPlex buffer

Chronic kidney disease

Stage-specific embryonic antigen-4

Methylprednisolone

Cyclosporin

Mesenchymal stromal cells

Small ubiquitin-related modifier 1

Sentrin-specific protease 2

Scramble sequence

Lipofectamine

Fukasawa H, Bornheimer S, Kudlicka K, Farquhar MG. Slit diaphragms contain tight junction proteins. J Am Soc Nephrol. 2009;20(7):1491–503.

Article   CAS   PubMed   PubMed Central   Google Scholar  

Bierzynska A, Saleem M. Recent advances in understanding and treating nephrotic syndrome. F1000Res. 2017;6:121.

Article   PubMed   PubMed Central   Google Scholar  

Kanasaki K, Kanda Y, Palmsten K, Tanjore H, Lee SB, Lebleu VS, et al. Integrin beta1-mediated matrix assembly and signaling are critical for the normal development and function of the kidney glomerulus. Dev Biol. 2008;313(2):584–93.

Article   CAS   PubMed   Google Scholar  

Noone DG, Iijima K, Parekh R. Idiopathic nephrotic syndrome in children. Lancet. 2018;392(10141):61–74.

Article   PubMed   Google Scholar  

Tullus K, Webb H, Bagga A. Management of steroid-resistant nephrotic syndrome in children and adolescents. Lancet Child Adolesc Health. 2018;2(12):880–90.

Nagata M. Podocyte injury and its consequences. Kidney Int. 2016;89(6):1221–30.

Sadowski CE, Lovric S, Ashraf S, Pabst WL, Gee HY, Kohl S, et al. A single-gene cause in 29.5% of cases of steroid-resistant nephrotic syndrome. J Am Soc Nephrol. 2015;26(6):1279–89.

Williams AE, Esezobor CI, Lane BM, Gbadegesin RA. Hiding in plain sight: genetics of childhood steroid-resistant nephrotic syndrome in Sub-saharan Africa. Pediatr Nephrol. 2023;38(7):2003–12.

Hejazian SM, Zununi Vahed S, Moghaddas Sani H, Nariman-Saleh-Fam Z, Bastami M, et al. Steroid-resistant nephrotic syndrome: pharmacogenetics and epigenetic points and views. Expert Rev Clin Pharmacol. 2020;13(2):147–56.

Kanwar YS, Rosenzweig LJ. Altered glomerular permeability as a result of focal detachment of the visceral epithelium. Kidney Int. 1982;21(4):565–74.

Vogelmann SU, Nelson WJ, Myers BD, Lemley KV. Urinary excretion of viable podocytes in health and renal disease. Am J Physiol Ren Physiol. 2003;285(1):F40–48.

Article   CAS   Google Scholar  

Oliveira Arcolino F, Tort Piella A, Papadimitriou E, Bussolati B, Antonie DJ, Murray P, et al. Human urine as a noninvasive source of kidney cells. Stem Cells Int. 2015;2015:362562.

Iampietro C, Bellucci L, Arcolino FO, Arigoni M, Alessandri L, Gomez Y, et al. Molecular and functional characterization of urine-derived podocytes from patients with Alport syndrome. J Pathol. 2020;252(1):88–100.

Harrell CR, Jovicic N, Djonov V, Arsenijevic N, Volarevic V. Mesenchymal stem cell-derived exosomes and other Extracellular vesicles as new remedies in the Therapy of Inflammatory diseases. Cells. 2019;8(12):E1605.

Article   Google Scholar  

Biancone L, Bruno S, Deregibus MC, Tetta C, Camussi G. Therapeutic potential of mesenchymal stem cell-derived microvesicles. Nephrol Dial Transpl. 2012;27(8):3037–42.

Bianchi F, Sala E, Donadei C, Capelli I, La Manna G. Potential advantages of acute kidney injury management by mesenchymal stem cells. World J Stem Cells. 2014;6(5):644–50.

Camussi G, Deregibus MC, Bruno S, Cantaluppi V, Biancone L. Exosomes/microvesicles as a mechanism of cell-to-cell communication. Kidney Int. 2010;78(9):838–48.

Ju Gqun, Cheng J, Zhong L, Wu S, Zou X, yu, Zhang G et al. yuan,. Microvesicles derived from human umbilical cord mesenchymal stem cells facilitate tubular epithelial cell dedifferentiation and growth via hepatocyte growth factor induction. PLoS One. 2015;10(3):e0121534.

Bellucci L, Montini G, Collino F, Bussolati B. Mesenchymal stromal cell-derived extracellular vesicles pass through the Filtration Barrier and protect podocytes in a 3D glomerular model under continuous perfusion. Tissue Eng Regen Med. 2021;18(4):549–60.

Medica D, Franzin R, Stasi A, Castellano G, Migliori M, Panichi V, et al. Extracellular vesicles derived from endothelial progenitor cells protect human glomerular endothelial cells and podocytes from complement- and cytokine-mediated Injury. Cells. 2021;10(7):1675.

Arcolino FO, Zia S, Held K, Papadimitriou E, Theunis K, Bussolati B, et al. Urine of Preterm neonates as a Novel source of kidney progenitor cells. J Am Soc Nephrol. 2016;27(9):2762–70.

Arcolino FO, Hosgood S, Akalay S, Jordan N, Herman J, Elliott T, et al. De novo SIX2 activation in human kidneys treated with neonatal kidney stem/progenitor cells. Am J Transpl. 2022;22(12):2791–803.

Trautmann A, Vivarelli M, Samuel S, Gipson D, Sinha A, Schaefer F, et al. IPNA clinical practice recommendations for the diagnosis and management of children with steroid-resistant nephrotic syndrome. Pediatr Nephrol. 2020;35(8):1529–61.

Furuta I, Zhai T, Umazume T, Ishikawa S, Nakagawa K, Akaishi R, et al. Increased podocyturia in pregnant women compared to non-pregnant women. J Obstet Gynaecol Res. 2017;43(5):873–9.

Saleem MA, O’Hare MJ, Reiser J, Coward RJ, Inward CD, Farren T, et al. A conditionally immortalized human podocyte cell line demonstrating nephrin and podocin expression. J Am Soc Nephrol. 2002;13(3):630–8.

Ivanova EA, Arcolino FO, Elmonem MA, Rastaldi MP, Giardino L, Cornelissen EM, van den Heuvel LP, Levtchenko EN. Cystinosin deficiency causes podocyte damage and loss associated with increased cell motility. Kidney Int. 2016;89(5):1037–48.

Counter CM, Hahn WC, Wei W, Caddle SD, Beijersbergen RL, Lansdorp PM, et al. Dissociation among in vitro telomerase activity, telomere maintenance, and cellular immortalization. Proc Natl Acad Sci U S A. 1998;95(25):14723–8.

Buono L, Scalabrin S, De Iuliis M, Tanzi A, Grange C, Tapparo M, Nuzzi R, Bussolati B. Mesenchymal stem cell-derived extracellular vesicles protect human corneal endothelial cells from endoplasmic reticulum stress-mediated apoptosis. Int J Mol Sci. 2021;22(9):4930.

Brossa A, Grange C, Mancuso L, Annaratone L, Satolli MA, Mazzone M, et al. Sunitinib but not VEGF blockade inhibits cancer stem cell endothelial differentiation. Oncotarget. 2015;6(13):11295–309.

Cavallari C, Figliolini F, Tapparo M, Cedrino M, Trevisan A, Positello L, et al. miR-130a and Tgfβ content in Extracellular vesicles derived from the serum of subjects at High Cardiovascular Risk predicts their In-Vivo angiogenic potential. Sci Rep. 2020;10(1):706.

Rampino T, Gregorini M, Guidetti C, Broggini M, Marchini S, Bonomi R, et al. KCNA1 and TRPC6 ion channels and NHE1 exchanger operate the biological outcome of HGF/scatter factor in renal tubular cells. Growth Factors. 2007;25(6):382–91.

Verta R, Grange C, Skovronova R, Tanzi A, Peruzzi L, Deregibus MC, et al. Generation of spike-extracellular vesicles (S-EVs) as a Tool to mimic SARS-CoV-2 Interaction with host cells. Cells. 2022;11(1):146.

Skovronova R, Grange C, Dimuccio V, Deregibus MC, Camussi G, Bussolati B. Surface marker expression in small and Medium/Large mesenchymal stromal cell-derived extracellular vesicles in naive or apoptotic Condition using Orthogonal techniques. Cells. 2021;10(11):2948.

Lu Y, Ye Y, Bao W, Yang Q, Wang J, Liu Z, et al. Genome-wide identification of genes essential for podocyte cytoskeletons based on single-cell RNA sequencing. Kidney Int. 2017;92(5):1119–29.

Bussolati B, Bruno S, Grange C, Buttiglieri S, Deregibus MC, Cantino D, et al. Isolation of renal progenitor cells from adult human kidney. Am J Pathol. 2005;166(2):545–55.

Faul C, Donnelly M, Merscher-Gomez S, Chang YH, Franz S, Delfgaauw J, et al. The actin cytoskeleton of kidney podocytes is a direct target of the antiproteinuric effect of cyclosporine A. Nat Med. 2008;14(9):931–8.

Coppo R, Camilla R, Porcellini MG, Peruzzi L, Gianoglio B, Amore A, et al. Saquinavir in steroid-dependent and -resistant nephrotic syndrome: a pilot study. Nephrol Dial Transpl. 2012;27(5):1902–10.

Ransom RF, Lam NG, Hallett MA, Atkinson SJ, Smoyer WE. Glucocorticoids protect and enhance recovery of cultured murine podocytes via actin filament stabilization. Kidney Int. 2005;68(6):2473–83.

Maruyama T, Abe Y, Niikura T. SENP1 and SENP2 regulate SUMOylation of amyloid precursor protein. Heliyon. 2018;4(4):e00601.

Hang J, Dasso M. Association of the human SUMO-1 protease SENP2 with the nuclear pore. J Biol Chem. 2002;277(22):19961–6.

Haynes JM, Selby JN, Vandekolk TH, Abad IPL, Ho JK, Lieuw WL, et al. Induced Pluripotent Stem Cell-Derived Podocyte-Like cells as models for assessing mechanisms underlying heritable Disease phenotype: initial studies using two Alport Syndrome patient lines indicate impaired Potassium Channel activity. J Pharmacol Exp Ther. 2018;367(2):335–47.

Musah S, Dimitrakakis N, Camacho DM, Church GM, Ingber DE. Directed differentiation of human induced pluripotent stem cells into mature kidney podocytes and establishment of a Glomerulus Chip. Nat Protoc. 2018;13(7):1662–85.

Lazzeri E, Ronconi E, Angelotti ML, Peired A, Mazzinghi B, Becherucci F, et al. Human urine-derived renal progenitors for Personalized modeling of genetic kidney disorders. J Am Soc Nephrol. 2015;26(8):1961.

Grange C, Tritta S, Tapparo M, Cedrino M, Tetta C, Camussi G, et al. Stem cell-derived extracellular vesicles inhibit and revert fibrosis progression in a mouse model of diabetic nephropathy. Sci Rep. 2019;9(1):4468.

Jiang Zzhen, Liu Y, mei, Niu X, Yin J yong, Hu B, Guo S, chun et al. Exosomes secreted by human urine-derived stem cells could prevent kidney complications from type I diabetes in rats. Stem Cell Res Ther. 2016;7:24.

Sedrakyan S, Villani V, Da Sacco S, Tripuraneni N, Porta S, Achena A, et al. Amniotic fluid stem cell-derived vesicles protect from VEGF-induced endothelial damage. Sci Rep. 2017;7(1):16875.

Duan Y, Luo Q, Wang Y, Ma Y, Chen F, Zhu X, et al. Adipose mesenchymal stem cell-derived extracellular vesicles containing microRNA-26a-5p target TLR4 and protect against diabetic nephropathy. J Biol Chem. 2020;295(37):12868–84.

Zhao T, Jin Q, Kong L, Zhang D, Teng Y, Lin L, et al. microRNA-15b-5p shuttled by mesenchymal stem cell-derived extracellular vesicles protects podocytes from diabetic nephropathy via downregulation of VEGF/PDK4 axis. J Bioenerg Biomembr. 2022;54(1):17–30.

Tossidou I, Himmelseher E, Teng B, Haller H, Schiffer M. SUMOylation determines turnover and localization of nephrin at the plasma membrane. Kidney Int. 2014;86(6):1161–73.

Wu Q, Aroankins TS, Cheng L, Fenton RA. SUMOylation Landscape of renal cortical Collecting Duct cells. J Proteome Res. 2019;18(10):3640–8.

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Acknowledgements

We thank Prof. Giovanni Camussi and Dr Chiara Deregibus (University of Turin) for electron microscopy EV analyses. We thank Dr. Roberta Verta for her support in cell line maintenance and in Western Blot analysis.

SD, EL, LP and BB are members of the European Reference Network for Rare Kidney Diseases (ERKNet). The study was supported by the National Institutes of Health (NIH) grant 1R01DK123234 to BB and LP and by European Research Council Consolidator grant 101045467 – NEOGRAFT to EL.

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Adele Tanzi and Lola Buono equal contribution first.

Licia Peruzzi and Benedetta Bussolati equal contribution last.

Authors and Affiliations

Department of Molecular Biotechnology and Health Sciences, University of Turin, Via Nizza 52, Turin, 10125, Italy

Adele Tanzi, Lola Buono, Corinne Iampietro, Alessia Brossa, Maddalena Arigoni, Raffaele Calogero & Benedetta Bussolati

Department of Medical Sciences, University of Turin, Turin, Italy

Cristina Grange & Silvia Deaglio

Department of Pediatric Nephrology, Emma Children’s Hospital, Amsterdam UMC, Amsterdam, The Netherlands

Fanny Oliveira Arcolino & Elena Levtchenko

Emma Centrum of Personalized Medicine, Emma Children’s Hospital, Amsterdam UMC, Amsterdam, The Netherlands

Fanny Oliveira Arcolino

Department of Urology, Children’s Hospital Los Angeles, Los Angeles, CA, USA

Laura Perin

Department of Development and Regeneration, Cluster Woman and Child, Laboratory of Pediatric Nephrology, KU Leuven, Leuven, Belgium

Elena Levtchenko

Pediatric Nephrology, ERKNet Center, Regina Margherita Children’s Hospital, AOU Città della, Salute e della Scienza di Torino, Turin, Italy

Licia Peruzzi

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Contributions

AT, LB: experimental procedures on podocyte permeability, data analysis, draft writing, AT, CG: experimental procedures on EV characterization and analysis, CI and AB: experimental procedures on progenitor cell isolation, characterization and immortalization, FOA: experimental procedures on Alport podocyte cultures, MA and RC: experimental procedures on RNA-seq and data analyses, LPerin: conceptualization, manuscript writing, SD: genetic analyses, manuscript writing, LPeruzzi: conceptualization, experimental procedures on urinary patient podocyte collection, manuscript writing, and BB: conceptualization, data analyses, manuscript writing,

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Correspondence to Benedetta Bussolati .

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All subjects enrolled in the present study provided informed written consent. The study protocol was approved by the Bioethics Committee of the A.O.U. Città della Salute e della Scienza Hospital (protocol no. 0021671). The study was conducted according to the principles expressed by the Declaration of Helsinki of 1975, as revised in 2013.

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Tanzi, A., Buono, L., Grange, C. et al. Urine-derived podocytes from steroid resistant nephrotic syndrome patients as a model for renal-progenitor derived extracellular vesicles effect and drug screening. J Transl Med 22 , 762 (2024). https://doi.org/10.1186/s12967-024-05575-z

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The Japanese Lung Cancer Society Guideline for non-small cell lung cancer, stage IV

Affiliations.

• 1 Internal Medicine III, Wakayama Medical University, Wakayama, Japan.
• 2 Department of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
• 3 Division of Thoracic Oncology, Shizuoka Cancer Center, Shizuoka, Japan.
• 4 Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Aichi, Japan.
• 5 Osaka City General Hospital, Osaka, Japan.
• 6 Department of Thoracic Oncology, National Cancer Center Hospital, Tokyo, Japan.
• 7 Clinical Research Center, Department of Thoracic Oncology and Medicine, National Hospital Organization Shikoku Cancer Center, Ehime, Japan.
• 8 Department of Internal Medicine, Niigata Cancer Center Hospital, Niigata, Japan.
• 9 Respiratory Center, Asahikawa Medical University Hospital, Hokkaido, Japan.
• 10 National Hospital Organization Kinki-chuo Chest Medical Center, Osaka, Japan.
• 11 Department of Internal Medicine, Division of Medical Oncology and Respiratory Medicine, Shimane University Faculty of Medicine, Shimane, Japan.
• 12 Department of Thoracic Oncology, Kansai Medical University Hospital, Osaka, Japan.
• 13 Department of Thoracic Oncology, Hyogo Cancer Center, Hyogo, Japan.
• 14 Advanced Cancer Translational Research Institute, Showa University, Tokyo, Japan.
• 15 Department of Surgery, University of California San Diego, California, USA.
• 16 National Cancer Center Hospital East, Chiba, Japan.
• 17 Department of Pulmonary Medicine and Oncology, Graduate School of Medicine, Nippon Medical School, Tokyo, Japan.
• 18 First Department of Medicine, Hokkaido University Hospital, Hokkaido, Japan.
• 19 Thoracic Oncology, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan.
• 20 Department of Thoracic Oncology and Respiratory Medicine, Tokyo Metropolitan Cancer and Infectious diseases Center Komagome Hospital, Tokyo, Japan.
• 21 Kyushu University Hospital, Fukuoka, Japan.
• 22 Regional Respiratory Symptomatology, Kanazawa University Graduate School of Medical Science, Kanazawa, Japan.
• 23 Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
• 24 Department of Thoracic Oncology, National Cancer Center Hospital East, Chiba, Japan.
• 25 Department of Biostatistics, Yokohama City University School of Medicine, Kanagawa, Japan.
• 26 Japan Federation of Cancer Patient Groups, Tokyo, Japan.
• 27 Japan Lung Cancer Alliance, Tokyo, Japan.
• 28 Department of Biomedical Statistics and Bioinformatics, Kyoto University Graduate School of Medicine, Kyoto, Japan.
• 29 Shizuoka Cancer Center, Shizuoka, Japan.
• 30 Division of Pulmonary Medicine, Allergry and Rheumatology, Department of Internal Medicine, Iwate Medical University School of Medicine, Iwate, Japan.
• 31 Thoracic Oncology, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan. [email protected].
• PMID: 31049758
• PMCID: PMC6545178
• DOI: 10.1007/s10147-019-01431-z

According to rapid development of chemotherapy in advanced non-small cell lung cancer (NSCLC), the Japan Lung Cancer Society has been updated its own guideline annually since 2010. In this latest version, all of the procedure was carried out in accordance with grading of recommendations assessment, development and evaluation (GRADE) system. It includes comprehensive literature search, systematic review, and determination of the recommendation by multidisciplinary expert panel which consisted of medical doctors, pharmacists, nurses, statisticians, and patients from patient advocacy group. Recently, we have had various types of chemotherapeutic drugs like kinase inhibitors or immune-checkpoint inhibitors. Thus, the guideline proposes to categorize patients into three entities: (1) driver oncogene-positive, (2) PD-L1 ≥ 50%, and (3) others. Based on this subgroup, 31 clinical questions were described. We believe that this attempt enables clinicians to choose appropriate treatment easier. Here, we report an English version of the Japan Lung Cancer Society Guidelines 2018 for NSCLC, stages IV.

Keywords: Chemotherapy; Guideline; Kinase inhibitor; Non-small cell lung cancer; Programed cell death-1 inhibitor; Programed death-ligand 1 inhibitor.

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Conflict of interest statement

Hiroaki Akamatsu received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Ono Pharmaceutical Co. Ltd, Pfizer, and Taiho Pharmaceutical Co. Ltd. He received research funding from MSD. Kiichiro Ninomiya received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Eli Lilly, and MSD. Hirotsugu Kenmotsu received honoraria from AstraZeneca K.K., Chugai Pharmaceutical Co. Ltd, Ono Pharmaceutical Co. Ltd. Boeringer Ingelheim, Eli Lilly K.K., Kyowa Hakko Kirin Co. Ltd., Bristol-Myers Squibb, MSD, and Novartis Pharma K.K. He also received research funding from AstraZeneca K.K., Chugai Pharmaceutical Co. Ltd, and Boeringer Ingelheim. Masahiro Morise received honoraria from AstraZeneca, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Ono Pharmaceutical Co. Ltd, and Pfizer. He also received research funding from AstraZeneca K.K., Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Pfizer, Ono Pharmaceutical Co. Ltd., Merck Serono, Kissei, Novartis and Taiho Pharmaceutical Co. Ltd. Haruko Daga received honoraria from Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, and MSD. Yasushi Goto received honoraria from AstraZeneca, Eli Lilly, Chugai, Taiho Pharmaceutical, Boehringer Ingelheim, Ono Pharmaceutical Co. Ltd., Bristol Myers Squibb, Pfizer, MSD, Shionogi Pharma and Novartis. He also received research funding from Abbvie, Eli Lilly, Taiho Pharmaceutical, Bristol Myers Squibb, and Ono Pharmaceutical Co. Ltd. Toshiyuki Kozuki received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, Kyowa Hakko Kirin Co, MSD, Nippon Kayaku, Ono Pharmaceutical Co. Ltd, Pfizer and Taiho Pharmaceutical Co. Ltd. He received research funding from AstraZeneca Chugai Pharmaceutical Co. Ltd, Eli Lilly and Merck Serono. Satoru Miura received honoraria from AstraZeneca, Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Eli Lilly and MSD. Takaaki Sasaki received honoraria from AstraZeneca, Boehringer Ingelheim, Daiichi Sankyo Co. Ltd., Eli Lilly and Novartis. He received research funding from Boehringer Ingelheim and Pfizer. Akihiro Tamiya received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Ono Pharmaceutical Co. Ltd. and Eli Lilly. He received research funding from AstraZeneca, Bristol-Myers Squibb and Ono Pharmaceutical Co. Ltd. Yukari Tsubata received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Daiichi Sankyo Co. Ltd. and Kyowa Hakko Kirin. She received research funding from Daiichi Sankyo Co. Ltd. and Ono Pharmaceutical Co. Ltd. Hiroshige Yoshioka received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Novartis, Ono Pharmaceutical Co. Ltd., Pfizer, and Taiho Pharmaceutical Co. Ltd. Yoshihiro Hattori received honoraria from AstraZeneca, Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Novartis, Ono Pharmaceutical Co. Ltd., and Taiho Pharmaceutical Co. Ltd. He received research funding from MSD and Ono Pharmaceutical Co. Ltd. Hidenori Mizugaki received honoraria from AstraZeneca, Boehringer Ingelheim, and Chugai Pharmaceutical Co. Ltd. He received research funding from Boehringer Ingelheim. Kaname Nosaki received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, Kyowa Hakko Kirin, MSD, Nippon Kayaku, Novartis, Ono Pharmaceutical Co. Ltd., Pfizer, and Taiho Pharmaceutical Co. Ltd. He received research funding from MSD and Novartis. Yusuke Okuma received honoraria from Boehringer Ingelheim, AstraZeneca, MSD, Novartis, Taiho Pharmaceutical Co. Ltd, and Eli-Lilly. He received research funding from Chugai Pharmaceutical Co. Ltd and Takeda Pharmaceutical Co. Ltd. Kentaro Tanaka received honoraria from Chugai Pharmaceutical Co. Ltd. Shigeki Umemura received research funding from MSD. Takeharu Yamanaka received honoraria from Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Taiho Pharmaceutical Co. Ltd, and Takeda Pharmaceutical Co. Ltd. He received research funding from Taiho Pharmaceutical Co. Ltd and Takeda Pharmaceutical Co. Ltd. Satoshi Morita received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Ono Pharmaceutical Co. Ltd., Pfizer, and Taiho Pharmaceutical Co. Ltd. He received research funding from Boehringer Ingelheim. Makoto Maemondo received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, MSD, Novartis, Ono Pharmaceutical Co. Ltd., Pfizer, and Taiho Pharmaceutical Co. Ltd. He received research funding from Boehringer Ingelheim. Takashi Seto received honoraria from Astellas Pharma, AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Eli Lilly, Kissei Pharmaceutical Co. Ltd, MSD, Novartis, Ono Pharmaceutical Co. Ltd., Pfizer, Roche Singapore, Takeda Pharmaceutical Co. Ltd., Taiho Pharmaceutical Co. Ltd., Thermo Fischer Scientific and Yakult Honsha Co. Ltd. He received research funding from Astellas Pharma, AstraZeneca, Bayer Yakuhin, Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Daiichi Sankyo, Eisai, Eli Lilly, Kissei Pharmaceutical Co. Ltd, LOXO Oncology, Merck Serono, MSD, Novartis, Pfizer and Takeda Pharmaceutical Co. Ltd. Nobuyuki Yamamoto received honoraria from AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Chugai Pharmaceutical Co. Ltd, Daiichi Sankyo, Eli Lilly, MSD, Novartis, Ono Pharmaceutical Co. Ltd., Pfizer, Taiho Pharmaceutical Co. Ltd. and Yakult Honsha Co. Ltd. He received research funding from Boehringer Ingelheim, Chugai Pharmaceutical Co. Ltd, Eli Lilly, and MSD. Others are no conflicts of interest.

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Treatment strategy of each subgroups in NSCLC, stage IV. PD-1 programed cell death-1,…

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First-line treatment of EGFR-mutated NSCLC, stage IV. EGFR epidermal growth factor receptor, TKI…

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Second-line or further treatment of EGFR-mutated NSCLC, stage IV. EGFR epidermal growth factor…

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First-line treatment of ALK-rearranged NSCLC, stage IV. Abbreviations: ALK; anaplastic lymphoma kinase, TKI;…

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The Japanese Version of the Diabetes Treatment Satisfaction Questionnaire (DTSQ): translation and clinical evaluation

Ishii, h, bradley, clare, riazi, afsane, barendse, s and yamamoto , t.

Ishii, H, Bradley, Clare, Riazi, Afsane, Barendse, S and Yamamoto , T (2000) The Japanese Version of the Diabetes Treatment Satisfaction Questionnaire (DTSQ): translation and clinical evaluation . Journal of Clinical and Experimental Medicine, 192 (7).

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Molecular identification and functional characterization of lc-pufa biosynthesis elongase ( elovl2 ) gene in chinese sturgeon ( acipenser sinensis ).

Simple Summary

1. introduction, 2. materials and methods, 2.1. experimental animals and sample collection, 2.2. molecular cloning of elovl2 cdna and qpcr, 2.3. bioinformatic analyses, 2.4. functional characterization in yeast, 2.5. fatty acid analysis, 2.6. statistical analysis, 3.1. molecular identification of chinese sturgeon elovl2, 3.2. multiple protein sequence alignments of the elovl2 among vertebrates, 3.3. synteny and gene structure comparison of the elvol2 in vertebrates, 3.4. phylogenic analysis, 3.5. spatial and temporal distribution patterns of chinese sturgeon elovl2, 3.6. functional characterization of chinese sturgeon elovl2, 3.7. effect of dietary lipid sources on the elovl2 expression in chinese sturgeon, 4. discussion, 5. conclusions, supplementary materials, author contributions, institutional review board statement, informed consent statement, data availability statement, conflicts of interest.

• Wen, Z.; Li, Y.; Bian, C.; Shi, Q.; Li, Y. Genome-wide identification of a novel elovl4 gene and its transcription in response to nutritional and osmotic regulations in rabbitfish ( Siganus canaliculatus ). Aquaculture 2020 , 529 , 735666. [ Google Scholar ] [ CrossRef ]
• Tocher, D.R. Fatty acid requirements in ontogeny of marine and freshwater fish. Aquac. Res. 2010 , 41 , 717–732. [ Google Scholar ] [ CrossRef ]
• Pereira, S.L.; Leonard, A.E.; Mukerji, P. Recent advances in the study of fatty acid desaturases from animals and lower eukaryotes. Prostaglandins Leukot. Essent. Fat. Acids 2003 , 68 , 97–106. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Castro, L.F.C.; Tocher, D.R.; Monroig, O. Long-chain polyunsaturated fatty acid biosynthesis in chordates: Insights into the evolution of Fads and Elovl gene repertoire. Prog. Lipid Res. 2016 , 62 , 25–40. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Xie, D.; Chen, C.; Dong, Y.; You, C.; Wang, S.; Monroig, Ó.; Tocher, D.R.; Li, Y. Regulation of long-chain polyunsaturated fatty acid biosynthesis in teleost fish. Prog. Lipid Res. 2021 , 82 , 101095. [ Google Scholar ] [ CrossRef ]
• Nugteren, D.H. The enzymic chain elongation of fatty acids by rat-liver microsomes. Biochim. Biophys. Acta 1965 , 106 , 280–290. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Li, Y.; Wen, Z.; You, C.; Xie, Z.; Tocher, D.R.; Zhang, Y.; Wang, S.; Li, Y. Genome wide identification and functional characterization of two LC-PUFA biosynthesis elongase ( elovl8 ) genes in rabbitfish ( Siganus canaliculatus ). Aquaculture 2020 , 522 , 735127. [ Google Scholar ] [ CrossRef ]
• Agaba, M.; Tocher, D.R.; Dickson, C.A.; Dick, J.R.; Teale, A.J. Zebrafish cDNA encoding multifunctional Fatty Acid elongase involved in production of eicosapentaenoic (20:5n-3) and docosahexaenoic (22:6n-3) acids. Mar. Biotechnol. 2004 , 6 , 251–261. [ Google Scholar ] [ CrossRef ]
• Morais, S.; Monroig, O.; Zheng, X.; Leaver, M.J.; Tocher, D.R. Highly Unsaturated Fatty Acid Synthesis in Atlantic Salmon: Characterization of ELOVL5 - and ELOVL2 -like Elongases. Mar. Biotechnol. 2009 , 11 , 627–639. [ Google Scholar ] [ CrossRef ]
• Monroig, Ó.; Wang, S.; Zhang, L.; You, C.; Tocher, D.R.; Li, Y. Elongation of long-chain fatty acids in rabbitfish ( Siganus canaliculatus ): Cloning, functional characterisation and tissue distribution of Elovl5 - and Elovl4 -like elongases. Aquaculture 2012 , 350–353 , 63–70. [ Google Scholar ] [ CrossRef ]
• Monroig, Ó.; Tocher, D.R.; Hontoria, F.; Navarro, J.C. Functional characterisation of a Fads2 fatty acyl desaturase with Δ6/Δ8 activity and an Elovl5 with C16, C18 and C20 elongase activity in the anadromous teleost meagre ( Argyrosomus regius ). Aquaculture 2013 , 412–413 , 14–22. [ Google Scholar ] [ CrossRef ]
• Wang, S.; Monroig, Ó.; Tang, G.; Zhang, L.; You, C.; Tocher, D.R.; Li, Y. Investigating long-chain polyunsaturated fatty acid biosynthesis in teleost fish: Functional characterization of fatty acyl desaturase ( Fads2 ) and Elovl5 elongase in the catadromous species, Japanese eel ( Anguilla japonica ). Aquaculture 2014 , 434 , 57–65. [ Google Scholar ] [ CrossRef ]
• Kuah, M.-K.; Jaya-Ram, A.; Shu-Chien, A.C. The capacity for long-chain polyunsaturated fatty acid synthesis in a carnivorous vertebrate: Functional characterisation and nutritional regulation of a Fads2 fatty acyl desaturase with Δ4 activity and an Elovl5 elongase in striped snakehead ( Channa striata ). Biochim. Biophys. Acta 2015 , 1851 , 248–260. [ Google Scholar ] [ PubMed ]
• Xie, D.; Chen, F.; Lin, S.; You, C.; Wang, S.; Zhang, Q.; Monroig, Ó.; Tocher, D.R.; Li, Y. Long-chain polyunsaturated fatty acid biosynthesis in the euryhaline herbivorous teleost Scatophagus argus : Functional characterization, tissue expression and nutritional regulation of two fatty acyl elongases. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 2016 , 198 , 37–45. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Zou, W.; Lin, Z.; Huang, Y.; Limbu, S.M.; Wen, X. Molecular cloning and functional characterization of elongase ( elovl5 ) and fatty acyl desaturase ( fads2 ) in sciaenid, Nibea diacanthus (Lacepède, 1802). Gene 2019 , 695 , 1–11. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Galindo, A.; Garrido, D.; Monroig, Ó.; Pérez, J.A.; Betancor, M.B.; Acosta, N.G.; Kabeya, N.; Marrero, M.A.; Bolaños, A.; Rodríguez, C. Polyunsaturated fatty acid metabolism in three fish species with different trophic level. Aquaculture 2021 , 530 , 735761. [ Google Scholar ] [ CrossRef ]
• Yan, J.; Liang, X.; Cui, Y.; Cao, X.; Gao, J. Elovl4 can effectively elongate C18 polyunsaturated fatty acids in loach Misgurnus anguillicaudatus . Biochem. Biophys. Res. Commun. 2018 , 495 , 2637–2642. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Carmona-Antoñanzas, G.; Monroig, O.; Dick, J.R.; Davie, A.; Tocher, D.R. Biosynthesis of very long-chain fatty acids (C>24) in Atlantic salmon: Cloning, functional characterisation, and tissue distribution of an Elovl4 elongase. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 2011 , 159 , 122–129. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Monroig, Ó.; Webb, K.; Ibarra-Castro, L.; Holt, G.J.; Tocher, D.R. Biosynthesis of long-chain polyunsaturated fatty acids in marine fish: Characterization of an Elovl4 -like elongase from cobia Rachycentron canadum and activation of the pathway during early life stages. Aquaculture 2011 , 312 , 145–153. [ Google Scholar ] [ CrossRef ]
• Li, S.; Monroig, Ó.; Wang, T.; Yuan, Y.; Carlos Navarro, J.; Hontoria, F.; Liao, K.; Tocher, D.R.; Mai, K.; Xu, W.; et al. Functional characterization and differential nutritional regulation of putative Elovl5 and Elovl4 elongases in large yellow croaker ( Larimichthys crocea ). Sci. Rep. 2017 , 7 , 2303. [ Google Scholar ] [ CrossRef ]
• Oboh, A.; Betancor, M.B.; Tocher, D.R.; Monroig, O. Biosynthesis of long-chain polyunsaturated fatty acids in the African catfish Clarias gariepinus : Molecular cloning and functional characterisation of fatty acyl desaturase ( fads2 ) and elongase ( elovl2 ) cDNAs. Aquaculture 2016 , 462 , 70–79. [ Google Scholar ] [ CrossRef ]
• Gregory, M.K.; James, M.J. Rainbow trout ( Oncorhynchus mykiss ) Elovl5 and Elovl2 differ in selectivity for elongation of omega-3 docosapentaenoic acid. Biochim. Biophys. Acta 2014 , 1841 , 1656–1660. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Machado, A.M.; Tørresen, O.K.; Kabeya, N.; Couto, A.; Petersen, B.; Felício, M.; Campos, P.F.; Fonseca, E.; Bandarra, N.; Lopes-Marques, M.; et al. “Out of the Can”: A Draft Genome Assembly, Liver Transcriptome, and Nutrigenomics of the European Sardine, Sardina pilchardus . Genes 2018 , 9 , 485. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Ferraz, R.B.; Kabeya, N.; Lopes-Marques, M.; Machado, A.M.; Ribeiro, R.A.; Salaro, A.L.; Ozório, R.; Castro, L.F.C.; Monroig, Ó. A complete enzymatic capacity for long-chain polyunsaturated fatty acid biosynthesis is present in the Amazonian teleost tambaqui, Colossoma macropomum . Comp. Biochem. Physiol. B Biochem. Mol. Biol. 2019 , 227 , 90–97. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Garrido, D.; Monroig, Ó.; Galindo, A.; Betancor, M.B.; Pérez, J.A.; Kabeya, N.; Marrero, M.; Rodríguez, C. Lipid metabolism in Tinca tinca and its n-3 LC-PUFA biosynthesis capacity. Aquaculture 2020 , 523 , 735147. [ Google Scholar ] [ CrossRef ]
• Janaranjani, M.; Shu-Chien, A.C. Complete repertoire of long-chain polyunsaturated fatty acids biosynthesis enzymes in a cyprinid, silver barb ( Barbonymus gonionotus ): Cloning, functional characterization and dietary regulation of Elovl2 and Elovl4 . Aquac. Nutr. 2020 , 26 , 1835–1853. [ Google Scholar ] [ CrossRef ]
• Xu, W.; Wang, S.; You, C.; Zhang, Y.; Monroig, Ó.; Tocher, D.R.; Li, Y. The catadromous teleost Anguilla japonica has a complete enzymatic repertoire for the biosynthesis of docosahexaenoic acid from α-linolenic acid: Cloning and functional characterization of an Elovl2 elongase. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 2020 , 240 , 110373. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Peng, Z.; Ludwig, A.; Wang, D.; Diogo, R.; Wei, Q.; He, S. Age and biogeography of major clades in sturgeons and paddlefishes (Pisces: Acipenseriformes ). Mol. Phylogenet. Evol. 2007 , 42 , 854–862. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Zhuang, P.; Zhao, F.; Zhang, T.; Chen, Y.; Liu, J.; Zhang, L.; Kynard, B. New evidence may support the persistence and adaptability of the near-extinct Chinese sturgeon. Biol. Conserv. 2016 , 193 , 66–69. [ Google Scholar ] [ CrossRef ]
• Huang, Z.; Wang, L. Yangtze Dams Increasingly Threaten the Survival of the Chinese Sturgeon. Curr. Biol. 2018 , 28 , 3640–3647.e3618. [ Google Scholar ] [ CrossRef ]
• Zhang, Y.; Lu, R.; Qin, C.; Nie, G. Precision nutritional regulation and aquaculture. Aquac. Rep. 2020 , 18 , 100496. [ Google Scholar ] [ CrossRef ]
• Wang, B.; Wu, B.; Liu, X.; Hu, Y.; Ming, Y.; Bai, M.; Liu, J.; Xiao, K.; Zeng, Q.; Yang, J.; et al. Whole Genome Sequencing Reveals Autooctoploidy in the Chinese Sturgeon and its Evolutionary Trajectories. Genom. Proteom. Bioinform. 2023 , 22 , qzad002. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Cheng, X.; Xiao, K.; Jiang, W.; Peng, G.; Chen, P.; Shu, T.; Huang, H.; Shi, X.; Yang, J. Selection, identification and evaluation of optimal reference genes in Chinese sturgeon ( Acipenser sinensis ) under polypropylene microplastics stress. Sci. Total Environ. 2024 , 920 , 170894. [ Google Scholar ] [ CrossRef ]
• Livak, K.J.; Schmittgen, T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 2001 , 25 , 402–408. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Hallgren, J.; Tsirigos, K.; Pedersen, M.D.; Almagro Armenteros, J.J.; Marcatili, P.; Nielsen, H.; Krogh, A.; Winther, O. DeepTMHMM predicts alpha and beta transmembrane proteins using deep neural networks. BioRxiv 2022 . [ Google Scholar ] [ CrossRef ]
• Thompson, J.D.; Higgins, D.G.; Gibson, T.J. CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994 , 22 , 4673–4680. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Hall, T. BioEdit : A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp. Ser. 1999 , 41 , 95–98. [ Google Scholar ]
• Abramson, J.; Adler, J.; Dunger, J.; Evans, R.; Green, T.; Pritzel, A.; Ronneberger, O.; Willmore, L.; Ballard, A.J.; Bambrick, J.; et al. Accurate structure prediction of biomolecular interactions with AlphaFold 3. Nature 2024 , 630 , 493–500. [ Google Scholar ] [ CrossRef ]
• Li, M.; Tang, H.; Qing, R.; Wang, Y.; Liu, J.; Wang, R.; Lyu, S.; Ma, L.; Xu, P.; Zhang, S.; et al. Design of a water-soluble transmembrane receptor kinase with intact molecular function by QTY code. Nat. Commun. 2024 , 15 , 4293. [ Google Scholar ] [ CrossRef ]
• Tamura, K.; Stecher, G.; Kumar, S. MEGA11: Molecular Evolutionary Genetics Analysis Version 11. Mol. Biol. Evol. 2021 , 38 , 3022–3027. [ Google Scholar ] [ CrossRef ]
• Zheng, X.; Ding, Z.; Xu, Y.; Monroig, O.; Morais, S.; Tocher, D.R. Physiological roles of fatty acyl desaturases and elongases in marine fish: Characterisation of cDNAs of fatty acyl Δ6 desaturase and elovl5 elongase of cobia ( Rachycentron canadum ). Aquaculture 2009 , 290 , 122–131. [ Google Scholar ] [ CrossRef ]
• Hastings, N.; Agaba, M.; Tocher, D.R.; Leaver, M.J.; Dick, J.R.; Sargent, J.R.; Teale, A.J. A vertebrate fatty acid desaturase with Delta 5 and Delta 6 activities. Proc. Natl. Acad. Sci. USA 2001 , 98 , 14304–14309. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Christie, W.W. Gas chromatography-mass spectrometry methods for structural analysis of fatty acids. Lipids 1998 , 33 , 343–353. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Monroig, Ó.; Lopes-Marques, M.; Navarro, J.C.; Hontoria, F.; Ruivo, R.; Santos, M.M.; Venkatesh, B.; Tocher, D.R.; Castro, L.F.C. Evolutionary functional elaboration of the Elovl2/5 gene family in chordates. Sci. Rep. 2016 , 6 , 20510. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Jaillon, O.; Aury, J.-M.; Brunet, F.; Petit, J.-L.; Stange-Thomann, N.; Mauceli, E.; Bouneau, L.; Fischer, C.; Ozouf-Costaz, C.; Bernot, A.; et al. Genome duplication in the teleost fish Tetraodon nigroviridis reveals the early vertebrate proto-karyotype. Nature 2004 , 431 , 946–957. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Castro, L.F.C.; Monroig, Ó.; Leaver, M.J.; Wilson, J.; Cunha, I.; Tocher, D.R. Functional Desaturase Fads1 (Δ5) and Fads2 (Δ6) Orthologues Evolved before the Origin of Jawed Vertebrates. PLoS ONE 2012 , 7 , e31950. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Glasauer, S.M.K.; Neuhauss, S.C.F. Whole-genome duplication in teleost fishes and its evolutionary consequences. Mol. Genet. Genom. 2014 , 289 , 1045–1060. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Tocher, D.R. Metabolism and Functions of Lipids and Fatty Acids in Teleost Fish. Rev. Fish. Sci. 2003 , 11 , 107–184. [ Google Scholar ] [ CrossRef ]
• Xie, D.; Fu, Z.; Wang, S.; You, C.; Monroig, Ó.; Tocher, D.R.; Li, Y. Characteristics of the Fads2 gene promoter in marine teleost epinephelus coioides and role of SP1-binding site in determining promoter activity. Sci. Rep. 2018 , 8 , 5305. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Gillard, G.; Harvey, T.N.; Gjuvsland, A.; Jin, Y.; Thomassen, M.; Lien, S.; Leaver, M.; Torgersen, J.S.; Hvidsten, T.R.; Vik, J.O.; et al. Life-stage-associated remodelling of lipid metabolism regulation in Atlantic salmon. Mol. Ecol. 2018 , 27 , 1200–1213. [ Google Scholar ] [ CrossRef ]
• Mourente, G.; Rodríguez, A.; Grau, A.; Pastor, E. Utilization of lipids by Dentex dentex L. (Osteichthyes, Sparidae) larvae during lecitotrophia and subsequent starvation. Fish Physiol. Biochem. 1999 , 21 , 45–58. [ Google Scholar ] [ CrossRef ]
• Sargent, J.R.; Tocher, D.R.; Bell, J.G. 4—The Lipids. In Fish Nutrition , 3rd ed.; Halver, J.E., Hardy, R.W., Eds.; Academic Press: San Diego, CA, USA, 2003; pp. 181–257. [ Google Scholar ]
• Monroig, O.; Rotllant, J.; Sánchez, E.; Cerdá-Reverter, J.M.; Tocher, D.R. Expression of long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis genes during zebrafish Danio rerio early embryogenesis. Biochim. Biophys. Acta 2009 , 1791 , 1093–1101. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• You, C.; Miao, S.; Lin, S.; Wang, S.; Waiho, K.; Li, Y. Expression of long-chain polyunsaturated fatty acids (LC-PUFA) biosynthesis genes and utilization of fatty acids during early development in rabbitfish Siganus canaliculatus . Aquaculture 2017 , 479 , 774–779. [ Google Scholar ] [ CrossRef ]
• Tocher, D.R.; Leaver, M.J.; Hodgson, P.A. Recent advances in the biochemistry and molecular biology of fatty acyl desaturases. Prog. Lipid Res. 1998 , 37 , 73–117. [ Google Scholar ] [ CrossRef ]
• Datsomor, A.K.; Zic, N.; Li, K.; Olsen, R.E.; Jin, Y.; Vik, J.O.; Edvardsen, R.B.; Grammes, F.; Wargelius, A.; Winge, P. CRISPR/Cas9-mediated ablation of elovl2 in Atlantic salmon ( Salmo salar L.) inhibits elongation of polyunsaturated fatty acids and induces Srebp-1 and target genes. Sci. Rep. 2019 , 9 , 7533. [ Google Scholar ] [ CrossRef ]
• Xie, D.; Wang, S.; You, C.; Chen, F.; Tocher, D.R.; Li, Y. Characteristics of LC-PUFA biosynthesis in marine herbivorous teleost Siganus canaliculatus under different ambient salinities. Aquac. Nutr. 2015 , 21 , 541–551. [ Google Scholar ] [ CrossRef ]
• Li, Y.; Hu, C.; Zheng, Y.; Xia, X.; Xu, W.; Wang, S.; Chen, W.; Sun, Z.; Huang, J. The effects of dietary fatty acids on liver fatty acid composition and Δ6-desaturase expression differ with ambient salinities in Siganus canaliculatus . Comp. Biochem. Physiol. B Biochem. Mol. Biol. 2008 , 151 , 183–190. [ Google Scholar ] [ CrossRef ]
• Nayak, M.; Giri, S.S.; Pradhan, A.; Samanta, M.; Saha, A. Effects of dietary α-linolenic acid/linoleic acid ratio on growth performance, tissue fatty acid profile, serum metabolites and Δ6 fad and elovl5 gene expression in silver barb ( Puntius gonionotus ). J. Sci. Food Agric. 2020 , 100 , 1643–1652. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Datsomor, A.K.; Gillard, G.; Jin, Y.; Olsen, R.E.; Sandve, S.R. Molecular Regulation of Biosynthesis of Long Chain Polyunsaturated Fatty Acids in Atlantic Salmon. Mar. Biotechnol. 2022 , 24 , 661–670. [ Google Scholar ] [ CrossRef ]
• Asif, M. Health effects of omega-3,6,9 fatty acids: Perilla frutescens is a good example of plant oils. Orient. Pharm. Exp. Med. 2011 , 11 , 51–59. [ Google Scholar ] [ CrossRef ]
• Orsavova, J.; Misurcova, L.; Ambrozova, J.V.; Vicha, R.; Mlcek, J. Fatty Acids Composition of Vegetable Oils and Its Contribution to Dietary Energy Intake and Dependence of Cardiovascular Mortality on Dietary Intake of Fatty Acids. Int. J. Mol. Sci. 2015 , 16 , 12871–12890. [ Google Scholar ] [ CrossRef ] [ PubMed ]
• Tocher, D.R.; Betancor, M.B.; Sprague, M.; Olsen, R.E.; Napier, J.A. Omega-3 Long-Chain Polyunsaturated Fatty Acids, EPA and DHA: Bridging the Gap between Supply and Demand. Nutrients 2019 , 11 , 89. [ Google Scholar ] [ CrossRef ] [ PubMed ]

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Ding, H.; Shi, X.; Wen, Z.; Zhu, X.; Chen, P.; Hu, Y.; Xiao, K.; Yang, J.; Tian, T.; Zhang, D.; et al. Molecular Identification and Functional Characterization of LC-PUFA Biosynthesis Elongase ( elovl2 ) Gene in Chinese Sturgeon ( Acipenser sinensis ). Animals 2024 , 14 , 2343. https://doi.org/10.3390/ani14162343

Ding H, Shi X, Wen Z, Zhu X, Chen P, Hu Y, Xiao K, Yang J, Tian T, Zhang D, et al. Molecular Identification and Functional Characterization of LC-PUFA Biosynthesis Elongase ( elovl2 ) Gene in Chinese Sturgeon ( Acipenser sinensis ). Animals . 2024; 14(16):2343. https://doi.org/10.3390/ani14162343

Ding, Haoze, Xuetao Shi, Zhengyong Wen, Xin Zhu, Pei Chen, Yacheng Hu, Kan Xiao, Jing Yang, Tian Tian, Dezhi Zhang, and et al. 2024. "Molecular Identification and Functional Characterization of LC-PUFA Biosynthesis Elongase ( elovl2 ) Gene in Chinese Sturgeon ( Acipenser sinensis )" Animals 14, no. 16: 2343. https://doi.org/10.3390/ani14162343

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Clinical and Experimental Medicine

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1 1897 May 1 1897 Jul 1 1897 Sep 1 1897 Nov 1 1896 Jan 1 1896 Apr 1 1896 Jul 1 1896 Nov 1 222 IMAGES The Japanese journal of experimental medicine (1922 edition) Japanese Journal Of Clinical Oncology-审稿速度 一般,3-8周 -首页 Journal Home Japanese Journal of Clinical and Medical Case Reports Japanese Journal of Medical Science Japanese Journal of Clinical Neurophysiology COMMENTS The Japanese Society of Lymphoma Research The Japanese Society of Lymphoma Research is dedicated to elucidating the pathology of lymphoma and related diseases, and to improving the diagnostic accuracy, treatment outcome, and quality of life of patients. ... The official journal of the society was changed from Japanese to English and is named the Journal of Clinical and Experimental ... 413557 The Japanese journal of clinical and experimental medicine. Publication Start Year: 1924 Publication End Year: Frequency: Monthly Country of Publication: Japan Publisher: Fukuoka-shi : Daidō Gakkan Shuppanbu Description: v. illus. Language: Japanese ISSN: 0021-4965(Print); 0021-4965(Linking) Coden: RIKEAZ MeSH: Medicine*; Research* Publication ... CiNii 雑誌 The Japanese journal of experimental medicine the Institute of Medical Science, the University of Tokyo Kinokuniya, 1928-1990 Vol. 7, no. 1 (1928)-v. 60, no. 6 (1990) Safety and effectiveness of tofogliflozin in Japanese people with type Journal of Diabetes Investigation is a clinical and experimental diabetes research journal publishing basic science, clinical practice, and epidemiology in diabetes. ABSTRACT Aims/Introduction Sodium-glucose cotransporter 2 (SGLT2) inhibitors effectively and safely reduce fasting and postprandial hyperglycemia while promoting weight loss. Japanese Clinical Medicine Journal portfolios in each of our subject areas. Links to Books and Digital Library content from across Sage. VIEW DISCIPLINE HUBS. Information for. Authors ; ... Japanese Clinical Medicine ISSN: 1179-6707; Online ISSN: 1179-6707; About Sage; Contact us; CCPA - Do not sell my personal information; Submission guidelines Keep lettering consistently sized throughout your final-sized artwork, usually about 2-3 mm (8-12 pt). Variance of type size within an illustration should be minimal, e.g., do not use 8-pt type on an axis and 20-pt type for the axis label. Avoid effects such as shading, outline letters, etc. The Current Status and Future Direction of Clinical Research in Japan "Clinical research" as defined by the Clinical Trials Act is interventional research other than Chiken designed to identify the efficacy or safety of drugs and other products through the use of drugs, medical devices, etc., by people. The Clinical Trials Act defines "specific clinical trials" ("Tokutei-Rinsho-Kenkyu" in Japanese) as interventional trials on previously approved ... Amplification of autoimmune organ damage by NKp46-activated ILC1 Present address: Department of Anesthesiology, Children's Hospital, Zhejiang University School of Medicine, National Clinical Research Center for Child Health, Hangzhou, China Oliver Hölsken The Tohoku Journal of Experimental Medicine Access full-text academic articles: J-STAGE is an online platform for Japanese academic journals. The Japanese journal of experimental medicine The Japanese journal of experimental medicine. Antibacterial properties of compounds having tricarbonylmethane group in their structure; antibacterial properties of 1,1-dimethyl-4 ... The Tohoku Journal of Experimental Medicine About this Journal The Tohoku Journal of Experimental Medicine (TJEM) was founded in 1920 by professors of Tohoku Imperial University, Medical School. The TJEM has been published continuously, except for the year of 1946 just after the World War II. The TJEM is open to original articles in all branches of medical sciences. Menin Inhibition With Revumenib for KMT2A-Rearranged Relapsed or Revumenib is a potent, oral, small molecule inhibitor of the menin-KMT2A interaction. In a phase I clinical study, treatment with revumenib had an acceptable safety profile and preliminary evidence of efficacy in patients with relapsed or refractory (R/R) KMT2Ar or NPM1-mutated acute leukemias and showed hallmarks of hematopoietic differentiation. 4 Here, we report the results of the pivotal ... Longevity of a Brain-Computer Interface for Amyotrophic Lateral The participant was a woman who had received a diagnosis of ALS in 2008; she was 58 years of age at study inclusion. She was in a locked-in state, received invasive ventilation through a ... Urine-derived podocytes from steroid resistant nephrotic syndrome Treatment with the nKPC-EVs significantly reduced permeability across all the steroid-resistant patients-derived and Alport syndrome-derived podocytes. At variance, podocytes appeared unresponsive to standard pharmacological treatments, with the exception of one line, in alignment with the patient's clinical response at 48 months. The Japanese Lung Cancer Society Guideline for non-small cell lung According to rapid development of chemotherapy in advanced non-small cell lung cancer (NSCLC), the Japan Lung Cancer Society has been updated its own guideline annually since 2010. In this latest version, all of the procedure was carried out in accordance with grading of recommendations assessment, development and evaluation (GRADE) system. Time-resolved scRNA-seq reveals transcription dynamics of polarized Bin Cao 1 Department of Pulmonary and Critical Care Medicine, National Center for Respiratory Medicine, National Clinical Research Center for Respiratory Diseases, Center of Respiratory Medicine, China-Japan Friendship Hospital, Beijing100029, China;2 Institute of Respiratory Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing100730, China;3 Graduate School ... Clinical medicine journals Genetics in Medicine (GIM) is an official journal of the American College of Medical Genetics and Genomics. The journal's mission is to enhance the knowledge, understanding, and practice of medical genetics and genomics through publications in clinical and laboratory genetics and genomics, including ethical, legal, and social issues as well as public health. The Japanese Version of the Diabetes Treatment Satisfaction Ishii, H, Bradley, Clare, Riazi, Afsane, Barendse, S and Yamamoto , T (2000) The Japanese Version of the Diabetes Treatment Satisfaction Questionnaire (DTSQ): translation and clinical evaluation. Journal of Clinical and Experimental Medicine, 192 (7). Animals The elongation of very-long-chain fatty acids gene 2 (elovl2) of Chinese sturgeon was identified in acipenseriformes species for the first time.Elovl2 was highly conserved in molecular evolution among vertebrates. Functional characterization in yeast demonstrated that Chinese sturgeon Elovl2 could efficiently elongate C 20 (20:4n-6 and 20:5n-3) and C 22 (22:4n-6 and 22:5n-3) substrates ... Home Overview. CEN Case Reports is a peer-reviewed online-only English-language journal that publishes original case reports in nephrology and related areas. The official journal of the Japanese Society of Nephrology. Presents clinical case reports in nephrology and related areas. A sister journal of Clinical and Experimental Nephrology. Tohoku Journal of Experimental Medicine Our mission is to publish peer-reviewed papers in all branches of medical sciences including basic medicine, social medicine, clinical medicine, nursing sciences and disaster-prevention science, and to present new information of exceptional novelty, importance and interest to a broad readership of the TJEM. The TJEM is open to original articles ... Japanese Clinical Medicine: Sage Journals A Case-Control Study of Esomeprazole plus Rebamipide vs. Omeprazole plus Rebamipide on Post-ESD Gastric Ulcers. Japanese Clinical Medicine is an international, peer reviewed, open access journal that aims to provide a dissemination point for high-quality research on clin... Viral defense protein speeds up female stem cell production A viral defense mechanism can be used to accelerate the creation of female stem cell lines in mice. The findings can boost efforts in medical research, drug testing, and regenerative therapies ... Clinical and Experimental Medicine Clinical and Experimental Medicine (CEM) is a multidisciplinary journal that aims to be a forum of scientific excellence and information exchange in relation to the basic and clinical features of the following fields: hematology, onco-hematology, oncology, virology, immunology, and rheumatology. The journal publishes reviews and editorials ... Home Clinical and Experimental Medicine is a multidisciplinary journal that aims to be a forum of scientific excellence and information exchange in relation to clinical and preclinical research in the following fields: hematology, oncology, virology, immunology and rheumatology.. Fully open access journal from January 2024. It publishes reviews and editorials, clinical and experimental studies ... The Tohoku Journal of Experimental Medicine The Tohoku Journal of Experimental Medicine. The Tohoku Journal of Experimental Medicine. Published by Tohoku University Medical Press. 11,312 registered articles (updated on June 06, 2023) Online ISSN : 1349-3329 Print ISSN : 0040-8727 ISSN-L : 0040-8727. Journal home. Advances in Clinical and Experimental Medicine A peer-reviewed, open access journal in medicine, medical sciences, public health, clinical medicine, molecular biology & genetics. ... Advances in Clinical and Experimental Medicine 1899-5276 (Print) / 2451-2680 (Online) Website ISSN Portal ... Journal of Clinical and Experimental Hematopathology Clinical Medicine; Other relevant information Title Journal of Clinical and Experimental Hematopathology; Publisher The Japanese Society for Lymphoreticular Tissue Research; Address 65 Tsurumai-cho,Showa-ku; ... Journal of the Japan Society of the Reticuloendothelial System. The Japanese Journal of Clinical and Experimental Medicine The Japanese Journal of Clinical and Experimental Medicine Publication Title - Original Language 臨牀と研究 ISSN 0021-4965 Material Format PDF Delivery Time (Days) 1-7 Raw Data Available FALSE Subjects Clinical Medicine Note Full text is usually delivered within a few hours for electronic issues and within 1-7 working days for print issues. Archive of "The Journal of Experimental Medicine". Articles from this journal are generally available in PMC after a 6-month delay (embargo); however, the delay may vary at the discretion of the publisher. ... Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press. Follow NCBI. Connect with NLM National Library of Medicine 8600 ... essay homework paper phd project resume review speech thesis