COST Action B17

COST Action B17

INSULIN RESISTANCE, OBESITY AND DIABETES MELLITUS IN THE ELDERLY

PROGRESS REPORT

Period: from (12-99) to (12-03)

This Report is prepared by the Management Committee of the Action and presented to the relevant Technical Committee or directly to the Committee of Senior Officials.

The report is a "cumulative" report, i.e. it is updated annually and covers the period beginning from the start date of the Action.

CONTENTS

1. Overview: action identification data

2. Objectives

3. TECHNICAL DESCRIPTION AND IMPLEMENTATION

3.1 Mode of Operation

3.2 Working Groups of COST B17

3.3 Secretarial Services

4. PARTICIPATION AND COORDINATION

4.1 Management Committee

4.2 Participating Institutions

4.3 Meetings of the Management Committee

4.4 Meetings of the Working Groups

4.5 Short-term Scientific Missions and their Results

5. RESULTS

5.1 Key Scientific Outcomes

5.2 Outstanding Scientific Publications

5.3 Joint Publications and Reports

5.4 Shared Resources and European Added Value

5.5 Participation and Awards of Young Scientists

6. DISSEMINATION OF RESULTS

6.1 Scientific Publications

6.2 Conferences and Workshops

6.3 Web-site

6.4 Scientific and Technical Cooperation

6.5 Transfer of results

6.6 Information of the Public and Influence on Politics

7. ECONOMIC DIMENSION

7.1 Total Manpower per Year

7.2 Funds received from the Commission

7.3 Costs of the Action

8. SELF EVALUATION (will be added only to the last annual progress report)

1. Overview: Action Identification Data

COST Action B17 Title INSULIN RESISTANCE, OBESITY AND DIABETES MELLITUS IN THE ELDERLY

TC Recommendation: 12/2/99	First MC meeting: (day/month/year) 10/03/00
CSO Approval: 27/5/99	Last MC meeting: (day/month/year) N.A.
Start date: (day/month/year) ⁽¹⁾ 16/12/99	Final Report: (day/month/year) ⁽²⁾ N.A.
Duration: months: 72	Evaluation Report: (day/month/year) ⁽²⁾ N.A.
Extension: months 12	TC Evaluation: (day/month/year) N.A.
End date: (day/month/year) 15/12/05

Number of signatories: (number) 23

Signatories and date of signature: (day/month/year)

Austria 5/4/2000	Greece 12/1/2000	Poland XX/XX/2003
Belgium 9/2/2000	Hungary 7/1/2000	Portugal
Bulgaria	Iceland	Romania 25/2/2000
Croatia	Ireland 26/1/2000	Slovakia 12/1/2000
Cyprus	Italy 5/1/2000	Slovenia 3/2/2000
Czech Rep. 2/2/2000	Latvia	Spain 16/12/99
Denmark 16/12/99	Lithuania 15/5/2000	Sweden 16/12/99
Estonia	Luxembourg 7/2/2001	Switzerland 15/12/99
Finland	Malta	Turkey
France 16/12/99	Netherlands 16/12/99	United Kingdom 16/12/99
Germany 16/12/99	Norway 20/12/99	Israel 19/12/2000

Institutes of non-COST countries: (list) Canada (Arend Bonen, Dept. of Human Biology and Nutritional Sciences, Univ. of Guelph; and Amira Klip, Hospital for Sick Children, Toronto)

Area: diabetes

Action Web site: http://www.webio.hu/workshops/cost/

Chairperson:

Title, name: Prof. Peter Csermely Tel.: +36-1-266-2755 extn.: 4102

Affiliation: Semmelweis Univ., Dept. Med. Chem. Fax:+36-1-266-7480

Postal Address P.O.Box 260, H-1444 Budapest 8, Hungary

E-Mail:csermely@puskin.sote.hu

TC Rapporteur:

Title, name: Dr. R. S. Burkle, Scientific Programme Manager, International

Affiliation: Medical Research Council

Tel.: +44-207-670-5223, Fax:+44-207-670-5124

Postal Address: 20 Park Crescent, UK, W1B 1AL London

External Evaluator: Title, name, affiliation, country N.A.

⁽¹⁾When 5 Signatures have been collected

⁽²⁾When the report is received by TC Secretariat
2. Objectives

General objectives – state of the art at the start of the Action

Diabetes mellitus of aged people is a considerable health burden of western societies. At present no widely established screening method is available to detect the persons who are predisposed in the population and due to our incomplete knowledge on the molecular mechanisms leading to NIDDM we have very few means to influence the onset and development of this diverse disease. The ongoing research of candidate genes as well as various differential screening methods will be coordinated with the Action.

Various steps of insulin action will be studied starting from the insulin receptor and following insulin action till it reaches the cell nucleus with special emphasis on the investigation of pathological changes of the molecular mechanism of insulin action in insulin resistance, obesity and NIDDM of the aged people. This research activity is thought to lead to the development new drug-candidates for the curing-easing of the consequences of this diverse disease.

Secondary objectives

The following steps of insulin action will be studied to gain a better understanding of the etiology of insulin resistance, obesity and NIDDM of the aged people, in hope to develop drug-candidates and a reliable screening method to detect the individuals with inherited risk for the disease.

1. Identification of diabetogenes

1.1. examination of candidate genes

1.2. utilization of various differential screening methods

1.3. possible links of diabetes-related genes to insulin signalling

1.4. search for diabetes-related obesity genes

1.5. age-dependent analysis of critical genes

2. Studies on the intracellular mechanism of insulin action in normal and pathological states

2.1. mechanism of the interactions between insulin receptor and its substrates, analysis of tissue-specificity of insulin signalling pathways

2.2. molecular characterization of glucose transport in normal and insulin-resistant states

2.3. molecular analysis of insulin signalling and action during the whole lifespan

2.4. interrelationships of insulin resistance and leptin action

2.5. development of insulin- and leptin-related signalling molecules as potential drug candidates

The list of secondary objectives is open for amendments incorporating the specific aims of new groups willing to participate in the concerted action. To join to the proposed research action an expertise in insulin (leptin, insulin-like growth factor) dependent signal transduction, insulin resistance/obesity-related physiological changes and/or in structural studies of insulin resistance/obesity-related genes is required.

3. Technical description and implementation

3.1. MODE OF OPERATION

The Action is focused on promoting collaboration and joint actions between scientists of participating countries in the field. This is performed by organizing collaborative meetings, where free discussions bring the participating laboratories, and invited guests together, by establishing working groups and by promoting a series of short term scientific missions. These areas of collaboration-promotion are extended by the web/site of the Action as well as by regular, informal exchanges of research materials as well as scientific expertise and ideas by letters, FAX-es and mostly: by email. It is a primary goal of the Action to encourage joint publications of collaborative research in top-level international scientific journals.

3.2. WORKING GROUPS OF COST B17

Keeping in mind the major areas of diabetes and ageing mechanism and research, the MC decided to set up 5 WGs. To ensure proper responsibility for the individual projects and help the inter-WG contacts only one co-ordinator per WG was nominated:

WG 1: Genetic aspects of NIDDM, obesity, insulin resistance and ageing

Co-ordinator: Ton Maassen

Objectives

· identification of population studies in Europe that can be used for genetic association studies

· what kind of clinical data are minimally required for proper genetic association studies

· techniques needing improvement (high throughput SNP analysis on large populations, what microarray systems are most suited for human and mouse systems, etc.)

2002 status

A number of cohorts became available within WG-1 for studies related to the development of insulin resistance and beta cell dysfunction in ageing populations. These cohorts are the Prague Cohort (B.Bendlova) and the Hoorn cohort (Maassen-Heine) for genetic analysis related to insulin resistance. These cohorts will be evaluated for SNP association related to newly discovered biochemical steps that modulate insulin sensitivity (Van Obberghen, Nice-France). Also the results emerging from the other Working Groups will be translated into assays that allows genetic association analysis. Part of the Hoorn cohort (Heine-‘t Hart), the Utrecht cohort (van Haeften) and the collaboration with the Stumvoll-Fritsche-Häring consortium in Tübingen-Germany also allows a detailed analysis of changes in glucose-induced insulin secretion in relation to the development of glucose intolerance. These cohorts are used to test the contribution of candidate genes that emerge from the biochemical studies within the Working Groups 2-5 to changes in diabetes-related clinical parameters.

2003 status

· With the association of Poland to the Action a novel situation arose in WG1. There three EU candidate countries (Czech Republic, Slovakia and Poland), where diabetes-related genetic studies are pursued using highly similar approaches. These three countries can now associate in WG1 and form a close consortium for further research. Their studies are greatly helped by the dutch lab of Prof. Maassen as well as by the UK lab of Ken Siddle transferring techniques expertise and sharing scientific tools as well as data with them. As an excellent occasion three STSM-s were organized at the same time from the three candidate countries to Prof. Maassen’s lab to learn genetic techniques and to share future plans. These concerted STSM-s can be set as an example to initiate a gross change in the status of research at a given area with the existing tools of COST.

· The collection of DNAs and proper phenotypic characterization of WG1-asociated cohorts of diabetic patients, offsprings and controls, patients with polycystic ovary syndrome, patients with gestational diabetes and obese patients was continued.

Results of collaborative efforts coordinated by WG1

· The frequency of the neurogenin, UCP1, beta2-adrenergic receptor, FABP2 and PPARgamma2 gene mutations and polymorphisms was studied and their possible role in the pathogenesis of diabetes and obesity was assessed in various populations.

· A collaboration has been established with WG4 on diabetes subphenotypes and pharmacological therapy.

· The reduced lipid oxidation in skeletal muscle from type 2 diabetes subjects may be of genetic origin (Gaster et al. Diabetes, in press)

WG 2: Glucose and other transport processes and ageing

Co-ordinator: Antonio Zorzano

Objectives

During the operation of COST-B17, the working group 2 is concentrating in the identification of novel components in glucose transport regulation and to understand their impact for the pathogenesis of insulin resistance and type 2 diabetes in the aged population. The specific aims are the following:

· identification of the molecular basis of glucose transport regulation in cells and tissues,

· identification of processes related to glucose transport in cells or tissues which are defective in ageing,

· determination of whether any of the regulatory proteins on glucose transport is amenable of pharmacological manipulation to prevent or treat diseases associated to ageing,

· identification of the role played by small GTP-binding proteins on glucose transport and specific mechanisms involved,

· covalent modification of glucose transporters and regulation of glucose transport in cells,

· mechanisms that mediate glucose transporter internalization,

· expression of glucose transporters in cells,

· mechanisms by which phosphatidylinositol 3-kinases and protein kinases regulate glucose transport,

· regulation of glucose transport in response to cellular stress,

· glucose transporter trafficking in adipose and muscle cells,

2002 status

Ongoing intensive collaboration in several research areas, where participating groups utilize the combination of their widely different expertise:

· GLUT4 trafficking in muscle cells: J. Eckel (D), P. Shepherd (UK), F. Giorgino (I), C. Reynet (DK), J. Jensen (N), S. Sasson (IL), A. Handberg (DK), A. Zorzano (E) and J.R. Zierath (S)

· expression of glucose transporters and of regulatory proteins of GLUT4 trafficking: J. Eckel (D), F. Giorgino (I), C. Reynet (DK), J. Jensen (N), A. Handberg (DK), A. Zorzano (E) and J.R. Zierath (S)

· collaboration with WG4 (Drugs and New Preventive Therapeutic Approaches to Improve Glucose Metabolism and Insulin Resistance)

· collaboration with WG3 (Muscle and Ageing)

2003 status

· Besides the continuation of the successful collaboration of the two previous task forces (“GLUT4 trafficking in muscle cells” and “Expression of glucose transporters and of regulatory proteins of GLUT4 trafficking”) a novel task force has been formed to study the “Signaling pathways that regulate glucose transport” with the participation of J. Eckel (D), P. Shepherd (UK), F. Giorgino (I), C. Reynet (DK), J. Jensen (N), S. Sasson (IL), A. Handberg (DK), A. Zorzano (E), J.R. Zierath (S), JA Maassen (NL), M. Ouwens (NL), M. Lorenzo (S), R. Zorec (SI).

· As a joint effort the working group identified new components of neuregulin signaling on glucose transport in muscle cells.

· The interaction with WG3 and WG4 has been extended at the 2003 joint meeting of these WG-s in Gentofte, Danemark.

Results of collaborative efforts coordinated by WG2

· Inhibition of protein kinase CbetaII increases glucose uptake in 3T3-L1 adipocytes through elevated expression of glucose transporter 1 at the plasma membrane. (Bosch et al. Mol Endocrinol)

· Expression of myotubularin by an adenoviral vector demonstrates its function as a PtdIns(3)P phosphatase in muscle cell lines. Involvement of PtdIns(3)P in insulin-stimulated glucose transport. (Chaussade et al., Mol Endocrinol)

· Insulin action in cultured human skeletal muscle cells during differentiation: assessment of cell surface GLUT4 and GLUT1 content (Al Khalili et al.)

· GLUT11 but not GLUT8 and GLUT12, is expressed in human skeletal muscle in a fiber type-specific pattern (Gaster et al.)

· Human Glycated Albumin Affects Glucose Metabolism in L6 Skeletal Muscle Cells by Impairing Insulin-induced Insulin Receptor Substrate (IRS) Signaling through a Protein Kinase C{alpha}-mediated Mechanism (Miele et al)

· Differential regulation of the muscle-specific GLUT4 enhancer in regenerating and adult skeletal muscle. (Moreno et al)

WG 3: Muscle and ageing

Co-ordinator: Aase Handberg

Objectives

· Study the impact of age, primary and secondary insulin resistance, exercise, adrenaline, and fatty acids on the expression and function of transporting and signalling proteins important for substrate uptake and processing in skeletal muscle

· biological factors behind muscle differentiation

· combined cellular and physiological approach using different muscle-specific transgenic mouse models (AMPK, etc.).

· co-culture of human adipocytes and human skeletal muscle cells for studies on the pathogenesis of insulin resistance

2002 status

· signaling through insulin and IGF1 receptors: K. Siddle (UK), C. Reynet (Denmark), A. Handberg (Denmark), J.A. Maassen (Netherlands), A.C. Rustan (Norway)

· molecular, cellular and metabolic perspectives of insulin resistance using human skeletal muscle cells in culture: A. Handberg (Denmark), C. Reynet (Denmark), A. C. Rustan (Norway), J. Jensen (Norway), J.R. Zierath (Sweden), P. Shepherd (UK), A. Zorzano (Spain), J.Eckel (Germany)

· critical role of AMPK: J.R. Zierath (Sweden), L. Hue (Belgium), M.H. Rider (Belgium), A.C. Rustan (Norway)

· collaboration with WG2 (“Glucose Transport and Ageing”) and WG4 (“Drugs and New Preventive Therapeutic Approaches to improve Glucose metabolism and Clinical Insulin Resistance”); planned joint action with WG5

· plans to develop joint actions on interactions between muscle and fat

2003 status

Results of collaborative efforts coordinated by WG3

· The reduced lipid oxidation in skeletal muscle from type 2 diabetes subjects may be of genetic origin - evidence from cultured myotubes (Gaster et al.)

· CD36 deficiency increases insulin sensitivity in muscle, but induces insulin resistance in the liver in mice (Goudriaan et al.)

· Proteome analysis reveals phosphorylation of ATP synthase beta-subunit in human skeletal muscle and proteins with potential roles in type 2 diabetes. (Hojlund et al)

· Aberrant p38 mitogen-activated protein kinase signalling in skeletal muscle from Type 2 diabetic patients (Koistinen et al)

· Expression profiling of the {gamma}-subunit isoforms of AMP-activated protein kinase suggests a major role for {gamma}3 in white skeletal muscle (Mahlapuu et al)

· Skeletal muscle reprogramming by activation of calcineurin improves insulin action on metabolic pathways (Ryder et al)

· Insulin receptor substrate-4 is expressed in muscle tissue without acting as a substrate for the insulin receptor (Schreyer et al)

WG 4: Drugs and new preventive therapeutic approaches to improve glucose metabolism and clinical insulin resistance

Co-ordinator: Francesco Giorgino

Objectives

· Study of insulin action and insulin resistance at molecular and clinical levels; identification of critical signaling intermediates for glucose utilization; investigation of molecules that act as stimulators of glucose utilization or targets for drugs known to regulate this process; clarification of the mechanism of action of insulin-sensitizing drugs; gene therapy of insulin resistance.

· Investigation of indexes of insulin resistance in ageing people; validation of surrogate markers for in vivo insulin sensitivity; creation of transgenic animals mimicking human models of insulin resistance.

2002 status

To accomplish the above goals this WG works in cooperation with all other WG of the action (genetics, glucose transport, muscle, cardiovascular) and act to develop the therapeutic implications of the activities of each WG and provide contacts with pharmacological and biotechnology industries to pursue therapeutic outcomes. Specific programmes:

· mechanism of action of insulin sensitizers

· tissue determinants of in vivo insulin resistance

· pharmaceutical control of glucose metabolism through regulation of gene expression

2003 status

The specific programmes have been extended by the two programmes below:

· insulin signaling to glucose transport: novel glucose transport regulatory mechanisms

· non-insulin signaling related mechanisms for glucose transport stimulation

Achievements:

pharmacologically relevant novel findings based on the work of WG 4

· a novel insulin analogue, [LysB3, GluB29] insulin, been found with enhanced beta cell protective action and its role in insulin signaling was analyzed

· the action of two pharmacologically relevant mediators of insulin action, inositol phosphoglycan and prostaglandylinositol cyclic phosphate, were compared in detail

· a highly efficient small molecule, Arimoclomol was found to be protective against neurodegeneration, a common chronic consequence of diabetes and its mechanism of action (help of heat shock protein induction by prolonged binding of the transcription factor HSF-1 to DNA) was clarified

· the agent and cell-type specificity of HIV protease inhibitors was assessed in the induction of insulin resistance

· effects of a low molecular weight heparin, Enoxaparin, on the restoration of the altered vascular reactivity of resistance arteries in aged and aged-diabetic hamsters were analyzed

clinically relevant analysis of large-cohort-studies

· with the participation of the WG an evaluation of vascular risk factors and markers of endothelial function was performed as determinants of inflammatory markers in type 1 diabetes as a part of the EURODIAB prospective complications study

· early signs of diabetes were assessed in the nationwide population-based Diabetes Incidence Study in Sweden (DISS)

· a screening for metabolic syndrome in hypertensive and/or obese subjects registered in primary health care was performed in Hungary.

WG 5: Cardio-vascular diseases and ageing

Co-ordinator: Maya Simionescu

Objectives

· effects of high glucose on the signalling pathways of the cells of the vascular wall

· effects of hypercaloric diet on hamster, as an experimental model for mild diabetes

· the possible roles of leptin in cardiovascular function

· insulin glycation

2002 status

· a highly successful WG meeting was held in Bucharest in the fall of 2002 where several bi- and multilateral collaborations were established, like

· a collaboration between the Institute of Cellular Biology and Pathology "N.Simionescu" (Bucharest, Romania) and the University College of London, Department of Biochemistry (UK) on studies on the effects of lipids, glucose and leptin on the cardiovascular system

· a collaboration of the laboratory "Posttranslational modifications of proteins" from the Institute of Biochemistry, Bucharest, Romania with the University of Rhode Island, Kingston, USA, Dept. of Chemistry, on the inhibitory effects of various compounds on early or advanced glycation products formation.

· the WG plans to develop intensive interactions with WG3 and WG4

2003 status

Two books are about to be published with collaboration of the WG5 members:

1. "Cellular Dysfunction in Atherosclerosis and Diabetes-Reports form Bench to Bedside” (Maya Simionescu, Anca Sima, Doina Popov, editors). The idea of this book came during the Workshop “Cardiovascular dysfunction in hyperlipidemia and diabetes” held together with the Meeting of the Working group “Cardiovascular dysfunction” of COST B17. The contributors to this book are: the WG5 COST members: Professor Juergen Eckel (Germany), Professor Peter Shepherd (UK), Professor Nava Bashan (Israel), Professor Shlomo Sasson (Israel), Professor Dan M. Cheta, (Romania), Dr. Elena Ganea (Romania), Dr. Doina Popov (Romania), along with personalities of the cell biology scientific community in atherosclerosis and diabetes from USA (Prof. David Stern and Prof. Vasilis Zannis), Canada (Prof. Moise Bendayan), UK (Prof. Chis Packard), France (Prof. Jean-Charles Fruchart), Italy (Prof. Francesco Giorgino and Prof. Michele Maiello), Israel (Prof. Michael Aviram), Netherlands (Prof. Vicor van Hinsberg), Sweden (Prof. German Camejo, and Prof. Olov Wiklund), Finland (Prof. Petri Kovanen), Hungary (Prof. Peter Csermely). Renowned Romanian cardiovascular clinicians will also contribute with chapters to this book, such as Professor Eduard Apetrei, and Prof. Leonida Gherasim, along with the Romanian top experts in diabetes, such as Prof. Constantin Ionescu-Tirgoviste and Prof. Nicolae Hancu.

2. “Vascular Involvement in Diabetes: Clinical, Experimental and Beyond” (DM Cheta, editor) This book involves academic personalities from the main universitary centers of Romania, as well as from USA (including Harvard Medical School), UK, France, Germany, Belgium, and Hungary. The following members of WG 5 will contribute with chapters: Professor Maya Simionescu (Romania), Dr. Doina Popov (Romania), Dr Elena Ganea (Romania), Dr. Dorel Radu (Romania) - as authors, and Professor Dan Mircea Cheta (Romania) - as both author and editor. The book will approach up-date medical and biological problems, and will consist of 40 chapters. Professor Eugene Braunwald (Harvard, USA) will write to foreword to this book.

As a collaborative effort of several WG5 labs as well as outside experts the effects of high glucose on the signalling pathways of the cells of the vascular wall were studied and the effects of hypercaloric diet on hamster, as an experimental model for mild diabetes were compared in several in vitro and animal models, such as

· by measurements on signal transduction in human smooth muscle cells

· on adrenaline signaling in muscle cells

· a collaboration was established between ICBP “N.Simionescu” and the Institute “Cantacuzino”on hyperglycemia in the development of diabetic complications investigated in transgenic mouse.

3.3. SECRETARIAL SERVICES

The Secretarial Services are kindly provided by the COST Secretariat. The financial administration of short term scientific missions as well as the maintenance of the web-site is provided by Prof. Csermely’s institute. From January 2004 the administration of Short Term Scientific Missions will be provided by the COST Secratariat.

4. Participation and coordination

4.1. MANAGEMENT COMMITTEE

4.1.1. Chairperson

Prof. Peter CSERMELY

Semmelweis University,

Dept. Med. Chem.

P.O.Box 260

H-1444 Budapest

Tel: + 36-1-266 2755/4102

Fax: +36-1-266 7480

E-mail : csermely@puskin.sote.hu

4.1.2. Vice Chairperson

Prof. Jürgen ECKEL

Deutsches Diabetes-Forschungsinstitut

Aufm Hennekamp 65

D-40225 Düsseldorf

Tel./Fax : +49 211 3382 561

Mobile : +49 172 2045 992

E-mail : eckel@uni-duesseldorf.de

4.1.3. Secretary

Prof. Mihail PASCU

DG Research / B5 - COST Secretaria

Rue de la Loi 200

Office: B 7 - 3/27

B-1049 Brussels

Tel: +32-2-299 15 56

Fax: +32-2-296 42 89

E-mail : mihail.pascu@cec.eu.int

4.1.4. Members

AUSTRIA

Prof. Josef PATSCH

Univ. Klinik f. Innere Medizin

Universität Innsbruck

Dr. T. EGGER

Ludwig Boltzmann-Institut für Altersforschung

I. Med. Abt./Donauspital, SMZO

BELGIUM

Prof. Louis HUE

Université Catholique de Louvain

ICP Département de Biochimie et de Biologie cellulaire

CZECH REPUBLIC

Dr. Bela Bendlova

Institute of Endocrinology

Dr. Jaroslav VESELY

Institute of Pathophysilogy - Medical faculty

Palacky University

DENMARK

Dr. Aase HANDBERG

Diabetes Research Center

Odense Universitets Hospital

FRANCE

Dr. Emmanuel VAN OBBERGHEN

INSERM U 145

Dr. Eric CLAUSER

INSERM U36

GERMANY

Prof. Jürgen ECKEL

Deutsches Diabetes-Forschungsinstitut

GRAND DUCHE DE LUXEMBOURG

GREECE

Prof. George DIMITRIADIS

Athens

Dr. Effie TSILIBARI

NRCPS “Demokritos”

Institute of Biology

HUNGARY

Prof. Peter CSERMELY

Semmelweis University,

Dept. Med. Chem.

Dr. György JERMENDY

Bajcsy-Zsilinszky Hospital, 3^rd Medical Department

IRELAND

Dr. John J. NOLAN

Metabolic Research Unit

Department of Endocrinology

St James Hospital

ISRAEL

Prof. Shlomo SASSON

Department of Pharmacology

Hebrew University of Jerusalem

Prof. Nava BASHAN

Dept. Clin. Biochem.

Ben Gurion Univ. Negev

ITALY

Prof. Francesco GIORGINO

Universita di Bari

LITHUANIA

Dr. Vaidotas URBANAVICIUS

Vilnius University Hospital

Clinics of Internal Medicine - Dept of Endocrinology

Dr. Valentinas MATULEVICIUS

Institute of Endocrinology - Kaunas Medical University

NETHERLAND

Dr. J.A. MAASSEN

Leiden University Medical Center

Department of Molecular Cell Biology

NORWAY

Dr. Jorgen JENSEN

National Institute of Occupational Health

Dept of Physiology

Prof. Arild Chr. RUSTAN

University of Oslo

Dept. of Pharmacology

POLAND

Prof. Aldona DEMBINSKA-KIEC

Department of Clinican Biochemistry

Collegium Medicum Jagellonian University

ROMANIA

Prof. Maya SIMIONESCU

Institute of Cellular Biology and Pathology “Nicolae Simionescu”

Prof. Dan CHETA

Institute of Diabetes Nutrition and Metabolic Diseases

SLOVAKIA

Dr. Elena SEBOKOVA

Institute of Experimental Endocrinology

Slovak Academy of Sciences

Dr. Iwar KLIMES

Institute of Experimental Endocrinology

Slovak Academy of Sciences

SLOVENIA

Prof. Marjan KORDAS

University of Ljubljana

Institute of Pathophysiology

Prof. Robert ZOREC

Institute of Pathophysiology

University of Ljubljana

SPAIN

Prof. Antonio ZORZANO

Universidad de Barcelona

Dpto. Bioquimica y Biologia Molecular

Dr. Isabel VARELA-NIETO

Universidad Autonomica - CSIC

Instituto Investigaciones Biomedicas “Alberto Sols”

Prof. Margarita LORENZO

Departamento de Bioquimica Biol. Mol. II

Facultad de Farmacia, Universitad Complutense

SWEDEN

Dr. Juleen R. ZIERATH

Karolinska Institutet

Inst. för kirugisk vetenskap

Assoc. Prof. Jan ERIKSSON

Department of Medicine

Umea University Hospital

SWITZERLAND

Dr. Markus NIESSEN

Universitätsspital Zürich

Prof. Jean-Louis CARPENTIER

Faculty of Medicine - Dept. of Morphology

University of Geneva

UNITED KINGDOM

Dr. Peter SHEPHERD

Department of Biochemistry

University College London

Prof. Kenneth SIDDLE

University of Cambridge

Department of Clinical Biochemistry

Addenbrooke’s Hospital

4.2. Participating Institutions

Canada (Arend Bonen, Dept. of Human Biology and Nutritional Sciences, Univ. of Guelph; and Amira Klip, Hospital for Sick Children, Toronto)

4.3. Meetings of the Management Committee

1^st MC meeting: 10/03/00, Brussels, Belgium

2^nd MC meeting: 9-10/10/00, Oslo, Norway

3^rd MC meeting: 23/05/01, Kerkrade, The Netherlands

4^th MC meeting: 10/09/01, Glasgow, Scotland, UK

5^th MC meeting: 16/02/02, Praha, Czech Republic

6^th MC meeting: 17/11/02, Bari, Italy

7^th MC meeting: 14/09/03, Gentofte, Danemark

4.4. Meetings of the Working Groups

2000

· WG 5 « Cardiovascular » : Bucarest – Romania – 28 November 2000, organisers : P. Shepherd, M. Simionescu (approx. 50 participants)

· WG 2 « Glucose transport » : Barcelona – Spain – 21 December 2000 (participants : J. Eckel, A. Zorzano, A. Guma).

2001

· WG 2 « Glucose and other transport processes and ageing » Kerkrade – The Netherlands – 22-23 May 2001 (organisers: J. Eckel, H. Joost, approx. 100 participants).

2002

· WG 1 « Genetics of diabetes, obesity and ageing » Prague – Czech Republic – 16 February 2002 (organisers: B. Bendlova, T. Maassen, approx. 50 participants).

· WG 5 « Cardiovascular dysfunction » Bucharest – Romania – 10-13 October 2002 (organiser: M. Simionescu, approx. 150 participants).

· WG 2+4 « Clinical and experimental insulin resistance/glucose transporter trafficking » Bari – Italy – 15-16 November 2002 (organiser: F. Giorgino, approx. 80 participants).

2003

· WG3+4 Skeletal muscle metabolism: Regulation, Exercise, Diabetes, Ageing, Gentofte, Danemark, 12-13 September 2003 (organiser: Aase Handberg, approx. 80 participants)

4.5. Short-term scientific missions and their results

(List: dates, hosts and nationality of scientists and topics)

First round

Name of Applicant	Country	Host Institute	Date	Topics
Jorgen Jensen	Norway	Univ. Coll. London	28/6-6/7/01	PI-3-kinase measurements
Shlomo Sasson	Israel	Diab. Forschungsinst. Düsseldorf, Germany	19-25/01/01	PPAR-related competitive PCR measurements
Andrej Janez	Slovenia	Diab. Forschungsinst. Düsseldorf, Germany	*Cancelled*	N.A.
Valentinas Matulevicius	Latvian	Univ. Barcelona	5/02-4/03/01	preparation of future collaboration involving clinical subjects
Elisabet Suarez	Spain	Karolinkska Inst.	10-31/05/01	to study heregulin-stimulated glucose trp.
Doina Popov	Romania	Univ. Coll. London	16-30/7/01	MAPK/STAT measurements in vascular smooth muscle cells
Michael Gaster	Denmark	Univ. Oslo	30/7-12/8/01	Fatty acid metabolism of satellite cells
Erlend Olav Brennesvik	Norway	Univ. Coll. London	1-15/9/01	learning of PI-3-kinase techniques
Doina Popov	Romania	Univ. Coll. London	28/11-5/12/01	Leptin signaling of human aortic endothelial cells

STSM results:

· the STSM of Jorgen Jensen resulted in a long-term collaboration and several joint publications of the participating labs

1. Whitehead J. P., Soos, M. A., Aslesen, R., O’Rahilly, S., & Jensen, J. Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349:775-781, 2000.

2. Whitehead J.P., Soos, M.A., Aslesen, R., O Rahilly, S., and Jensen, J.: Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349, 775-781, 2000.

3. Soos, M. A., Jensen, J., Brown, R.A., O’Rahilly, S., Shepherd, P.R., & Whitehead, J.P. Class II phosphoinositide 3-kinase is activated by insulin but not by contraction in skeletal muscle. Arch Biochem. Biophys. 396:244-248, 2001.

4. Foukas, L.C., Daniele, N., Ktori, C., Anderson, K.E., Jensen, J., and Shepherd, P.R. Direct effects of caffeine and theophylline on the p110d and other phosphoinositiode 3-kinases; Differential effects on lipid kinase and protein kinase activities. J. Biol. Chem. 277, 37124-37130, 2002.

· the STSM of Shlomo Sasson resulted in a long-term collaboration and several joint publications of the participating labs

1. Dransfeld, O., Uphues, I., Sasson, S., Schurmann, A., Joost, H.G. and Eckel, J.: Regulation of subcellular distribution of GLUT4 in cardiomyocytes: Rab4A reduces basal glucose transport and augments insulin responsiveness. Exp. Clin. Endocrinol. Diabetes 107, 26-36, 1999.

2. Dransfeld O, Rakatzi I, Sasson S, Gruzman A, Schmitt M, Haussinger D, Eckel J. Eicosanoids participate in the regulation of cardiac glucose transport by contribution to a rearrangement of actin cytoskeletal elements. Biochem. J. 359, 47-54, 2001.

3. Dransfeld, O., Rakatzi, I., Sasson, S. and Eckel, J.: Eicosanoids and the regulation of cardiac glucose transport. Ann. N.Y. Acad. Sci. 967, 208-216, 2002.

The joint experiments resulted in a successful grant application to the German Israel Foundation (2004-2006).

· the STSM-s of Michael Gaster, Erlend Brennesvick and Doina Popov resulted in joint publications, which are in preparation or appeared, like:

1. Popov, D., Simionescu, M., Shepherd, P.R. Saturated-fat diet induces moderate diabetes and severe glomerulosclerosis in hamsters. Diabetologia, in press

2. Gaster, M., Rustan, A.C., Aas, V. and Beck-Nielsen, H. The Reduced Lipid Oxidation in Skeletal Muscle from Type 2 Diabetes Subjects may be of Genetic Origin - Evidence from Cultured Myotubes. Diabetes, in press

Second round

Name of Applicant	Country	Host Institute	Date	Topics
Tetzlaff Stephan	Germany	Leiden Univ.	26-31/8/02	analysis of the 3243 mutation in mitochondrial DNA
Daniela Sramkova	Czech Republic	Univ. Cambridge, Dept. Med. Clin. Biochem.	2/10-22/11/02	site-directed mutagenesis, linkage studies, fat biopsies, RT-PCR
Jonas Buren	Sweden	Univ. Oslo, Norway	11-15/02/03	glucose uptake and signaling in muscle and fat
Marieke Ruiter	Netherlands	Diab. Forschungsinst. Düsseldorf, Germany	*postponed*
Daniela Gasperikova	Slovakia	Leiden University, the Netherlands	12/-25/10/03	Practical course on the analysis of mitochondrial mutations in diabetes
Beata Kiec-Wilk	Poland	Leiden University, the Netherlands	12/-25/10/03	Practical course on the analysis of mitochondrial mutations in diabetes
Marketa Vankova	Czech Republic	Leiden University, the Netherlands	12/-25/10/03	Practical course on the analysis of mitochondrial mutations in diabetes
Sebastio Perrini	Italy	Karolinska Institute, Sweden	22/10-17/11/03	PKC depdendent signaling of glucose transport in human skeletal muscle
Julius Benicky	Slovak Republic	University of Innsbruck, Austria	28/11-19/12/03	Measurements of the volume sensitive anion channel of beta-cells

STSM results:

· the second round of STSM-s were highly successful in continuing the traditional collaboration in various areas of signaling. Joint publications appeared so far (several others are in preparation):

1. Búren, J., Liu, H.-X., Jensen, J. & Eriksson, J.W. Dexamethasone impairs insulin signalling and glucose transport by depletion of IRS-1, PI3-kinase and Protein kinase B in primary cultured rat adipocytes. Eur.J.Endocrinol. 146 (3):419-429, 2002

· additionally in establishing the spread of expertise of genetic analysis of diabetic samples in various countries of Europe suhc as the Czech Republic, Poland and Slovakia. This knowledge transfer from the UK and the Netherlands to these candidate countries of the EU is a real asset of COST B17.

5. Results

5.1. KEY SCIENTIFIC OUTCOMES

(names and years in parentheses refer to the respective publications of section 6.1.)

At the initial phase the Action identified five different areas where an extra effort is needed to fulfill the aims of the MoU. The following major results have been achieved and published so far in the Action in these areas:

WG 1: Genetic aspects of NIDDM, obesity, insulin resistance and ageing

2000-2002

· Identification of the gene altered in Berardinelli-Seip congenital lipodystrophy on chromosome 11q13 (Magre et al., a Nature Medicine paper)

· ApoB genetic variations (Mazura et al, 2000)

· Genetic background of the PPAR-related polycystic ovary syndrome has been studied in detail (Bendlova et al, 2001; Vybrikova et al., 2000, 2001)

2003

· The reduced lipid oxidation in skeletal muscle from type 2 diabetes subjects may be of genetic origin (Gaster et al. Diabetes, in press)

WG2: Glucose and other transport processes and ageing

1999-2000

· Glucose transporters of human skeletal muscle (Gaster et al, 2000a,b; Kawano et al, 1999; Krook et al, 2000)

· GLUT4 translocation mechanism in adypocytes (Enrique-Tarancon et al, 2000)

· Rab11 and GLUT4 transport (Kessler et al, 2000)

· Surface GLUT4 (Ryder et al., 2000)

2001

· GLUT4 is reduced in slow fibers of diabetic muscle (Gaster et al, 2001)

· Molecular mechanisms of glucose transport (Dimitriadis and Newsholme, 2000; Newsholme and Dimitriadis, 2001; Till et al., 2000)

· PI-3-kinase and GLUT4 transport (Kessler et al, 2001)

· Semicarbazide sensitive amine oxidase substrates and glucose transport (Morin et al, 2001)

2002

· Ubc9 plays an essential role in insulin-stimulated glucose transport in adipocytes but not in undifferentiated fibroblasts (Giorgino, 2002)

2003

· Inhibition of protein kinase CbetaII increases glucose uptake in 3T3-L1 adipocytes through elevated expression of glucose transporter 1 at the plasma membrane. (Bosch et al. Mol Endocrinol)

· Insulin action in cultured human skeletal muscle cells during differentiation: assessment of cell surface GLUT4 and GLUT1 content (Al Khalili et al.)

· GLUT11 but not GLUT8 and GLUT12, is expressed in human skeletal muscle in a fiber type-specific pattern (Gaster et al.)

· Differential regulation of the muscle-specific GLUT4 enhancer in regenerating and adult skeletal muscle. (Moreno et al)

· Identification of the functional role of Rab11 and detection of the anti-apoptotic activity of adiponectin (J. Eckel, Germany)

· Identification of SUMOylation sites in GLUT4 (F. Giorgino, Italy)

· Identification of signaling pathways involved in DHEA stimulation of glucose transport in fat cells

· Activation of insulin-dependent, AMPK-dependent pathways, is an attractive strategy to enhance glucose transport in skeletal muscle from Type 2 diabetic patients through increased cell surface GLUT4 content (J. Zierath, Sweden)

WG 3: Muscle and ageing

2000-2001

· Unbalanced expression of the different subunits of elongation factor 1 in diabetic skeletal muscle (Reynet and Kahn, a PNAS paper)

· Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle (Whitehead et al, 2000)

2002

· fiber type dependent expression of GLUT11 in human skeletal muscle (Handberg, 2002)

· development of a relevant human skeletal muscle cell model to test and stydy mechanism of action of selected hit compounds that regulate glucose metabolism (Reynet, 2002)

2003

· The reduced lipid oxidation in skeletal muscle from type 2 diabetes subjects may be of genetic origin - evidence from cultured myotubes (Gaster et al.)

· CD36 deficiency increases insulin sensitivity in muscle, but induces insulin resistance in the liver in mice (Goudriaan et al.)

· Proteome analysis reveals phosphorylation of ATP synthase beta-subunit in human skeletal muscle and proteins with potential roles in type 2 diabetes. (Hojlund et al)

· Aberrant p38 mitogen-activated protein kinase signalling in skeletal muscle from Type 2 diabetic patients (Koistinen et al)

· Expression profiling of the {gamma}-subunit isoforms of AMP-activated protein kinase suggests a major role for {gamma}3 in white skeletal muscle (Mahlapuu et al)

· Skeletal muscle reprogramming by activation of calcineurin improves insulin action on metabolic pathways (Ryder et al)

· Insulin receptor substrate-4 is expressed in muscle tissue without acting as a substrate for the insulin receptor (Schreyer et al)

· Measurements of dynamics of plasma membrane in single human skeletal muscle fibres (Robert Zorec, Slovenia)

WG4: Drugs and new preventive therapeutic approaches to improve glucose metabolism and clinical insulin resistance

1999-2002

· Small insulin mimetic (Zhang et al, 1999 a Science paper)

· Receptor binding of insulin analogues (Kurtzhas et al, 2000)

· Benzylamine and vanadate in STZ-diabetes (Marti et al, 2001)

· Insulin secretagogues (Raptis and Dimitriadis, 2001)

· Role of molecular chaperones in diabetes and ageing (Csermely 2001a,b; Nardai et al, 2001, Soti and Csermely, 2000)

2003

· synthesis of small molecules that stimulate glucose transport in skeletal muscle cells in a non-insulin dependent manner, by activating distal regulatory protein in the insulin transduction mechanism (Shlomo Sasson, Israel, 2003)

· mechanism of the stimulatory effects of cyclooxygenase II inhibitors on the glucose transport system in skeletal muscle cells (Shlomo Sasson, Israel, 2003)

· The combination of SSAO substrates and vanadate exerts an antidiabetic effect in type 2 diabetic rats (Abella et al., Diabetes 2003)

· a novel insulin analogue, [LysB3, GluB29] insulin, been found with enhanced beta cell protective action and its role in insulin signaling was analyzed

· the action of two pharmacologically relevant mediators of insulin action, inositol phosphoglycan and prostaglandylinositol cyclic phosphate, were compared in detail

· the agent and cell-type specificity of HIV protease inhibitors was assessed in the induction of insulin resistance

· effects of a low molecular weight heparin, Enoxaparin, on the restoration of the altered vascular reactivity of resistance arteries in aged and aged-diabetic hamsters were analyzed

WG5: Cardio-vascular diseases and ageing

· AGE-formation (Georgescu and Popov, 2000)

· Characterization of endothelial dysfunction (Costache et al, 2000a,b; Dobrian et al, 2000; Jinga et al., 2000; Popov et al, 2001; Raicu et al, 2000; Simionescu, 2000, 2001; Simionescu et al, 2001)

· Administration of superoxide dismutase entrapped in long-circulating liposomes reverses endothelial dysfunction (2003, Simionescu and Popov)

Additional results have also been achieved in accordance with points of the MoU:

5.1.1. Identification of diabetogenes (see WG1)

5.1.2. Studies on the intracellular mechanism of insulin action in normal and pathological states

5.1.2.1. mechanism of the interactions between insulin receptor and its substrates, analysis of tissue-specificity of insulin signalling pathways

1999-2002

· The lipid phosphatase SHIP2 controls insulin sensitivity (Clement et al., 2001 a Nature paper)

· Exercise-induced changes in IRS-signaling (Chibalin et al, 2000, a PNAS paper; Yu et al., 2001)

· Insulin action in spinal injury (Mohr et al, 2001)

· Muscle fibre type specificity (Song et al., 1999)

· mTOR and PKB (Nave et al, 1999)

· Insulin receptor signaling occurs at least in part from the cell surface and does not require internalization of the receptor. This is true for the signaling resulting in the metabolic actions (including glucose transport) of the insulin receptor. This signaling is occuring from microvilli and apparentls also from caveolae.

2003

· adrenaline increases insulin-stimulated PKB activation via cAMP and EPAC (Jorgen Jensen, Norway)

· proteomic analysis of proteins secreted by 3T3-L1 adipocytes (Ken Siddle, UK)

· inhibitory effect of the adaptor protein Grb10 on insulin signalling (Ken Siddle, UK)

· use of RNAi to down-regulate protein expression in 3T3-L1 cells (Ken Siddle, UK)

· cross-talk between IRS and cytoskeletal proteins affects insulin-dependent activation of MAPK (Markus Niessen, Switzerland)

· overexpression of IRS-2 in pancreatic islets is sufficient to increase proliferation of b-cells and protects Human b-cells from hyperglycemia-induced apoptosis (Markus Niessen, Switzerland)

· Specific serine phosphorylation sites on IRS1 which are phosphorylated in response to oxidative stress (Nava Bashan, Israel)

5.1.2.2. molecular characterization of glucose transport in normal and insulin-resistant states (see WG2)

5.1.2.3. molecular analysis of insulin signalling and action during the whole lifespan

· Inositol lipids in insulin signalling (Jones and Varela-Nieto, 1999

· PI-3-kinase (Sebokova et al, 1999)

5.1.2.4. interrelationships of insulin resistance and leptin action

· Insulin and leptin acutely regulate cholesterol ester metabolism in macrophages by novel signalling pathways (O’Rourke and Shepherd, 2002; O’Rourke et al., 2001; 2002)

· Analysis of paradoxical observations on the association between leptin and insulin resistance. (Ceddia et al, 2002)

· Endocrine abnormalities in healthy first-degree relatives of type 2 diabetes patients - potential role of steroid hormones and leptin in the development of insulin resistance. (Janson et al, 2002)

· leptin stimulates peroxisomal, but not the mitochondrial FA oxidation pathways (Sebokova and Klimes, 2003)

· Leptin and TNFa activate the class-II phosphoinositide 3-kinase (Ktori et al, 2003)

· molecules other than leptin can bind to the leptin receptor (Peter Shepherd, UK)

5.1.2.5. development of insulin- and leptin-related signalling molecules as potential drug candidates (see WG4)

Other papers on insulin, diabetes and ageing

2000

· Diffusible transcription factor in ear development (Frago et al., 2000)

2001

· Chaperone-overload is a possible contributor to “civilization diseases”: atherosclerosis, cancer, diabetes (Csermely, 2001, a Trends in Genetics hypothesis)

· Acetylcholinesterase and diabetes (Kiss et al., 2001)

· Amine oxidase subtrates and adypocytes differentiation (Fontana et al., 2001)

· Membrane changes in diabetes (Somogyi et al., 2001)

· Thiazolidindiones (Jermendy and Csermely, 2001)

· Thyroid hormones and glucose intolerance (Dimitriadis and Raptis, 2001)

2002

· a glomerular epithelial cell receptor (podocalyxin) is anti-adhesive and antagonizes the adhesive effect of integrins (Tsilibari et al, 2002)

· IGF-I deficit is associated with a progressive age-dependent lost of auditory neurons by apoptosis (Varela-Nieto et al., 2002)

2003

· the participating Romanian EU Center of Excellence started stem cell research differentiation experiments

· elucidation of the mechanism responsible for the inhibition of protein synthesis during hypoxia : inactivation of elongation factor-2 and of p70 S6 ribosomal protein kinase by the AMP-activated protein kinase (Louis Hue, Belgium)

· IGF-I regulates inner ear neurogenesis by promoting cell survival and differentiation (Isabel Varela-Nieto, Spain)

· CCAAT/enhancer binding protein alpha mediates the synergistic effect of insulin and dexamethasone in GLUT4 gene expression (Margarita Lorenzo, Spain)

· the mechanism for the induction of insulin resistance by HIV protease inhibitors (Almira Klip, Canada and Nava Bashan, Israel)

· caffeine inhibits the enzyme PI 3-kinase (Peter Shepherd, UK)

· identification of a destabilizing trans-acting protein that binds to a cis-acting element in the 3’UTR region of GLUT-1 mRNA (Shlomo Sasson, Israel)

5.2. OUTSTANDING SCIENTIFIC PUBLICATIONS (published in journals with an impact factor higher than 10)

1999

1. Zhang B, Salituro G, Szalkowski D, Li Z, Zhang Y, Royo I, Vilella D, Diez MT, Pelaez F, Ruby C, Kendall RL, Mao X, Griffin P, Calaycay J, Zierath JR, Heck JV, Smith RG, Moller DE. Discovery of a small molecule insulin mimetic with antidiabetic activity in mice. Science. 1999 284:974-977. IF: 23.9

2000

1. Chibalin AV, Yu M, Ryder JW, Song XM, Galuska D, Krook A, Wallberg-Henriksson H, Zierath JR. Exercise-induced changes in expression and activity of proteins involved in insulin signal transduction in skeletal muscle: differential effects on insulin-receptor substrates 1 and 2. Proc Natl Acad Sci U S A. 2000 97:38-43. IF: 10.8

2. Rupnik, M. M. Kreft, I., S. K. Sikdar, S. Grilc., R. Romih, G. Zupančič, T. F. J. Martin, and R. Zorec (2000) Rapid Regulated Dense-Core Vesicle Exocytosis Requires the CAPS Protein PNAS 97:5627-5632. IF: 10.8

2001

1. Clement S, Krause U, Desmedt F, Tanti JF, Behrends J, Pesesse X, Sasaki T, Penninger J, Doherty M, Malaisse W, Dumont JE, Le Marchand-Brustel Y, Erneux C, Hue L, Schurmans S. The lipid phosphatase SHIP2 controls insulin sensitivity. Nature. 2001 Jan 4;409(6816):92-7. IF: 25.8

2. Csermely, P. Chaperone-overload as a possible contributor to “civilization diseases”: atherosclerosis, cancer, diabetes. Trends in Genetics, 17, 701-704, 2001, IF: 12.9

3. Magre J, Delepine M, Khallouf E, Gedde-Dahl T, Jr., Van Maldergem L, Sobel E, Papp J, Meier M, Megarbane A, Bachy A, Verloes A, d'Abronzo FH, Seemanova E, Assan R, Baudic N, Bourut C, Czernichow P, Huet F, Grigorescu F, de Kerdanet M, Lacombe D, Labrune P, Lanza M, Loret H, Matsuda F, Navarro J, Nivelon-Chevalier A, Polak M, Robert JJ, Tric P, Tubiana-Rufi N, Vigouroux C, Weissenbach J, Savasta S, Maassen JA, Trygstad O, Bogalho P, Freitas P, Medina JL, Bonnicci F, Joffe BI, Loyson G, Panz VR, Raal FJ, O'Rahilly S, Stephenson T, Kahn CR, Lathrop M, Capeau J 2001 Identification of the gene altered in Berardinelli-Seip congenital lipodystrophy on chromosome 11q13. Nat Genet 28:365-370 IF: 30,9

4. Reynet C and Kahn CR. Unbalanced expression of the different subunits of elongation factor 1 in diabetic skeletal muscle. PNAS, 2001, 98: 3422-3427, IF: 10.8

2002

7. Almind, K., Delahaye, L., Hansen, T., VanObberghen, E., Pedersen, O., and Kahn, C.R.: Characterization of the Met3261Ie variant of phosphatidylinositol 3-kinase p85 alpha. Proc Natl. Acad. Sci. USA 99, 2124-2128, 2002. IF: 10.8

8. Ouwens DM, de Ruiter ND, van der Zon GC, Carter AP, Schouten J, van der BC, Kooistra K, Bos JL, Maassen JA, van Dam H 2002 Growth factors can activate ATF2 via a two-step mechanism: phosphorylation of Thr71 through the Ras-MEK-ERK pathway and of Thr69 through RalGDS-Src-p38. EMBO J 21:3782-3793, 2002, IF: 14,0

9. Reig N, Chillaron J, Bartoccioni P, Fernandez E, Bendahan A, Zorzano A, Kanner B, Palacin M, Bertran J. The light subunit of system b(o,+) is fully functional in the absence of the heavy subunit. EMBO J. 21, 4906-4914, 2002, IF: 14,0

2003

1. Török, Zs., Tsvetkova, N.M., Balogh, G., Horváth, I., Nagy, E., Pénzes, Z., Hargitai, J., Bensaude, O., Csermely, P., Crowe, J.H., Maresca, B. and Vígh, L. (2003) Heat shock protein co-inducers with no effect on protein denaturation specifically modulate the membrane lipid phase. Proc. Natl. Acad. Sci. USA 100, 3131-3136, IF: 10.7

5.3. JOINT PUBLICATIONS AND REPORTS

(the collaborating scientists from different countries have been set in boldface)

1999

2. Franch, J., Aslesen, R. & Jensen, J. Regulation of glycogen synthesis after contractile activity in rat skeletal muscle. Effect of adrenaline on glycogen synthesis and activation of glycogen synthase and glycogen phosphorylase. Biochem. J. 344:231-235, 1999.

3. Nave, B.T., Ouwens, M., Withers, D.J., Alessi, D.R. and Shepherd, P.R.: Mammalian target of rapamycin is a direct target for protein kinase B: identification of a convergence point for opposing effects of insulin and amino-acid deficiency on protein translation. Biochem. J. 344, 427-431, 1999.

4. Sebokova, E., Gasperikova, D., Ouwens, M., Dorrestijn, J., Eckel, J., Maasen, A. and Klimes, I.: Defect in long-term activation of phosphatidylinositol 3-kinase by insulin in vivo: studies in insulin-resistant hHTg rats. Endocr. Regul. 33, 49-54, 1999.

2000

1. Kessler, A. E. Tomŕs, D. Immler, H.E. Meyer, A. Zorzano, J. Eckel. Rab 11 is associated with GLUT4-containing vesicles and redistributes in response to insulin. Diabetologia. 43, 1518-1527, 2000.

2. Kurtzhals, P., Schaffer, L., Sorensen, A., Kristensen, C., Jonassen, I., Schmid, C., and Trub, T.: Correlations of receptor binding and metabolic and mitogenic potencies of insulin analogs designed for clinical use. Diabetes 49, 999-1005, 2000.

3. Till, M., Ouwens, D.M., Kessler, A. and Eckel, J.: Molecular mechanisms of contraction-regulated cardiac glucose transport. Biochem. J. 346, 841-847, 2000.

4. Whitehead J. P., Soos, M. A., Aslesen, R., O’Rahilly, S., & Jensen, J. Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349:775-781, 2000.

5. Whitehead J.P., Soos, M.A., Aslesen, R., O Rahilly, S., and Jensen, J.: Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349, 775-781, 2000.

2001

1. Aslesen, R., Engebretsen, E.M.L., Franch, J. & Jensen, J. Glucose uptake and metabolic stress in rat muscles stimulated electrically with different protocols. J. Appl. Physiol. 91:1237-1244, 2001.

2. Clement S, Krause U, Desmedt F, Tanti JF, Behrends J, Pesesse X, Sasaki T, Penninger J, Doherty M, Malaisse W, Dumont JE, Le Marchand-Brustel Y, Erneux C, Hue L, Schurmans S. The lipid phosphatase SHIP2 controls insulin sensitivity. Nature. 2001 Jan 4;409(6816):92-7.

3. Dransfeld O, Rakatzi I, Sasson S, Gruzman A, Schmitt M, Haussinger D, Eckel J. Eicosanoids participate in the regulation of cardiac glucose transport by contribution to a rearrangement of actin cytoskeletal elements. Biochem. J. 359, 47-54, 2001.

4. Červenková K., Belejová M., Veselý J., Chmela Z., Rypka M., Ulrichová J., Modrianský M.,Maurel P.: Cell suspensions, cell cultures, and tissue slices – important metabolic in vitro systems. Biomed. Papers, 145, 57-60, 2001.

5. Kessler, A., Uphues, I., Ouwens, D.M., Till, M. and Eckel, J.: Diversification of cardiac insulin signaling involves the p85alpha/beta subunits of phosphatidylinositol 3-kinase. Am. J. Physiol. 280, E65-E74, 2001.

6. Soos, M. A., Jensen, J., Brown, R.A., O’Rahilly, S., Shepherd, P.R., & Whitehead, J.P. Class II phosphoinositide 3-kinase is activated by insulin but not by contraction in skeletal muscle. Arch Biochem. Biophys. 396:244-248, 2001.

2002

1. Aas, V., Torblĺ, S., Andersen, M.H., Jensen, J. and Rustan, A.C.: Electrical stimulation improves insulin responses in a human skeletal muscle cell model of insulin resistance. Ann. N.Y. Acad. Sci. 967, 506-515, 2002.

2. Búren, J., Liu, H.-X., Jensen, J. and Eriksson, J.W. Dexamethasone impairs insulin signalling and glucose transport by depletion of IRS-1, PI3-kinase and Protein kinase B in primary cultured rat adipocytes. Eur.J.Endocrinol. 146 (3):419-429, 2002

3. Dransfeld, O., Rakatzi, I., Sasson, S. and Eckel, J.: Eicosanoids and the regulation of cardiac glucose transport. Ann. N.Y. Acad. Sci. 967, 208-216, 2002.

5. Franch, J., Andersen, J.L., Jensen, J., Pedersen, P.K., & Knudsen, J. Acyl-CoenzymeA binding protein is expressed fibre type specific in rat skeletal muscles but not affected by moderate endurance training. Pflügers Arch. 443:387-393, 2002.

6. Franch, J., Knudsen, J., Ellis, B.A., Pedersen, P.K., Cooney, G.J. & Jensen, J. Acyl-CoA binding protein expression is fiber type- specific and elevated in muscles from the obese insulin-resistant Zucker rat. Diabetes. 51(2):449-454, 2002.

7. Madsen, L., Guerre-Millo, M., Flindt, E.N., Berge, K., Tronstad, K.J., Bergene E., Sebokova, E., Rustan, A.C., Jensen, J., Mandrup, S., Kristiansen, K., Klimes, I., Staels, B. and Berge, R.K.: Tetradecylthioacetic acid prevents high fat diet induced adiposity and insulin resistance. J. Lipid Res. 43, 742-750, 2002.

8. Magre J, Delepine M, Khallouf E, Gedde-Dahl T, Jr., Van Maldergem L, Sobel E, Papp J, Meier M, Megarbane A, Bachy A, Verloes A, d'Abronzo FH, Seemanova E, Assan R, Baudic N, Bourut C, Czernichow P, Huet F, Grigorescu F, de Kerdanet M, Lacombe D, Labrune P, Lanza M, Loret H, Matsuda F, Navarro J, Nivelon-Chevalier A, Polak M, Robert JJ, Tric P, Tubiana-Rufi N, Vigouroux C, Weissenbach J, Savasta S, Maassen JA, Trygstad O, Bogalho P, Freitas P, Medina JL, Bonnicci F, Joffe BI, Loyson G, Panz VR, Raal FJ, O'Rahilly S, Stephenson T, Kahn CR, Lathrop M, Capeau J 2001 Identification of the gene altered in Berardinelli-Seip congenital lipodystrophy on chromosome 11q13. Nat Genet 28:365-370

2003

1. Aas V., Torblå S., Andersen M.H., Jensen J. and A.C. Rustan, Electrical Stimulation Improve Insulin Responses in a Human Skeletal Muscle Cell Model of Insulin Resistance. Annals of the New York Academy of Sciences, in press

2. Bach D, Pich S, Soriano FX, Vega N, Baumgartner B, Oriola J, Daugaard JR, Lloberas J, Camps M, Zierath JR, Rabasa-Lhoret R, Wallberg-Henriksson H, Laville M, Palacin M, Vidal H, Rivera F, Brand M, Zorzano A. Mitofusin-2 determines mitochondrial network architecture and mitochondrial metabolism. A novel regulatory mechanism altered in obesity. J Biol Chem. 2003 May 9;278(19):17190-7.

3. Ben Romano, R., Rudich, A., Torok, D., Vanounou, S., Riesenberg, K., Schlaeffer, F., Klip, A., and Bashan N. Agent and cell-type specificity in the induction of insulin resistance by HIV protease inhibitors. AIDS 17,23-32,2003.

4. Gaster, M., A. Handberg, A. Schürmann, H.-G. Joost, H. Beck-Nielsen and H.D. Schroeder. GLUT11 but not GLUT8 and GLUT12, is Expressed in Human Skeletal Muscle in a Fiber Type-specific Pattern. Accepted by JCEM, nov. 2003.

5. Gaster, M., Rustan, A.C., Aas, V. and Beck-Nielsen, H. The Reduced Lipid Oxidation in Skeletal Muscle from Type 2 Diabetes Subjects may be of Genetic Origin - Evidence from Cultured Myotubes. Diabetes, in press

6. Hojlund K, Wrzesinski K, Larsen PM,`Fey SJ, Roepstorff P, Handberg A, Dela F, Vinten J, McCormack JG, Reynet C, Beck-Nielsen H. Proteome analysis reveals phosphorylation of ATP synthase beta -subunit in human skeletal muscle and proteins with potential roles in type 2 diabetes. (2003) J. Biol. Chem. 278, 10436-10442.

7. Magre J, Delepine M, Van Maldergem L, Robert JJ, Maassen JA, Meier M, Panz VR, Kim CA, Tubiana-Rufi N, Czernichow P, Seemanova E, Buchanan CR, Lacombe D, Vigouroux C, Lascols O, Kahn CR, Capeau J, Lathrop M. Prevalence of mutations in AGPAT2 among human lipodystrophies. Diabetes. 2003 Jun; 52 (6):1573-8.

8. Miele C, Riboulet A, Maitan MA, Oriente F, Romano C, Formisano P, Giudicelli J, Beguinot F, Van Obberghen E. Human Glycated Albumin Affects Glucose Metabolism in L6 Skeletal Muscle Cells by Impairing Insulin-induced Insulin Receptor Substrate (IRS) Signaling through a Protein Kinase C{alpha}-mediated Mechanism. J Biol Chem. 2003 Nov 28;278(48):47376-47387.

9. Popov, D., Simionescu, M., Shepherd, P.R. Saturated-fat diet induces moderate diabetes and severe glomerulosclerosis in hamsters. Diabetologia, 46, 1408-1418

10. Rakatzi, I., Gromke, T., Siddle, K. and Eckel J.: Differential phosphorylation of insulin receptor substrate-1 and –2 by insulin- and IGF-I-receptors in human skeletal myoblasts. Biochemical Journal, in press

11. Ukropec, J, Reseland, JE, Gasperikova, D, Demcakova, E, Madsen, L, Berge, RK, Rustan, AC, Klimes, I, Drevonb CA, Sebokova, E. The Hypotriglyceridemic Effect of Dietary n-3 Fatty Acids is Associated with Increased b-oxidation and Reduced Leptin Expression. Lipids 2003 38 (10), 1-7, 2003.

12. Urso B, Ilondo MM, Holst PA, Christoffersen CT, Ouwens M, Giorgetti S, Van Obberghen E, Naor D, Tornqvist H, De Meyts P. IRS-4 mediated mitogenic signalling by insulin and growth hormone in LB cells, a murine T-cell lymphoma devoid of IGF-I receptors. Cell Signal. 2003 Apr;15(4):385-94.

5.4. SHARED RESOURCES AND EUROPEAN ADDED VALUE

5.4.1. Shared resources to the Network

· B (Louis Hue) AMPK-related techniques, antibodies, etc., AMPK-alpha-2 deficient transgenic mice

· CZ (Bela Bendlova) DNAs of Czech gestational diabetics, positive controls for some MODY gene mutations (GCK, HNF1alpha)

· DK (Aase Handberg) determination of CD36 in rat muscles

· F (Emmanuel van Obberghen) antibodies against various insulin signaling molecules

· G (Jurgen Eckel) techniques to study glucose transport and to culture cardiac myocytes

· GR (Effie Tsilibary) diabetic nephropathy models

· HU (Peter Csermely) protein network bioinformatics approaches, transgenic mice overexpressing damaged proteins in their liver (a possible novel diabetes model), stress protein assays and reagents

· N (Arild Rustan) lipid analysis (Jorgen Jensen) muscle exercise techniques

· NL (Ton Maassen) mitochondrial diabetes genomics

· RO (Maya Simionescu and Doina Popov) atherosclerotic model systems, stem cell research

· SE (Juleen Zierath) human skeletal muscle biopsies and techniques (Jan Eriksson) human in vivo experiments (euglycemic clamp, adrenaline infusions), human visceral and subcutaneous adipocytes obtained at abdominal surgery

· SK (Elena Sebokova and Iwar Klimes) lipid analysis, and animal models

· SL (Robert Zorec) measurements of dynamics of plasma membrane in single human skeletal muscle fibres

· SP (Antonio Zorzano) glucose transport assays

· UK (Ken Siddle) proteomic analysis of adipocytes, RNAi technique, antibodies (Peter Shepherd) immortalized human fat and muscle cell lines, PI-3-kinase antibodies and essays

5.4.2. European added value

Several WG-s initiated networks having results not been achieved without a COST Action

· WG1 initiated a joint effort of CZ, PL and SK supported by G, NL and UK to assess genetic markers in diabetes in these 3 candidate countries.

· A wide network of glucose transport and insulin signalling in muscle and adipose tissues have been established by WG-s 2 and 3 resulting in dozens of high level joint publications.

· WG5 was electrified under the leadership of the Simionescu Insitute, an EU Center of Excellence in Bucharest. Besides a wide range of bi- and multilateral collaborations they also offer several courses, such as the 4^th Advanced Study Course “From cellular and molecular biology to the 21^st century medicine” in June 2003.

· All WG-s mobilized their members to provide a large number of shared resources to the action listed under 5.4.1., which is a great asset for joint programs and provides a European research potential in the field of molecular diabetes research.

· The collaborations withing COST B17 made possible the entrance of participating groups to several 6^th Framework Programmes listed under chapter 6.4.

5.5. PARTICIPATION AND AWARDS OF YOUNG SCIENTISTS

The Action makes always a special effort tomobilize extra funds to help the participation of young scientists in the WG meetings and workshops of the Action. The second round of Short Term Scientific Missions was almost exclusively accomplished by young scientists. Young scientists are regular first- and co-authors of the joint publications listed in points 5.3. and 6.1.

The following international awards have been received by young scientists belonging to participating labs:

2002

· Best Poster Award of the EMBO course on molecular chaperones in Warsaw, Poland (Judit Hargitai in Peter Csermely’s lab, October 2002)

· Prize of European Life Scientist Organization June 2002, Nice, France (A. Burlacu and M. Voinea in Maya Simionescu’s institute)

2003

· Manuela Voinea and Adriana Georgescu, Ph.D. students (31 years old) got a prize at European Life Scientist Organization (ELSO) Meeting, Nisa (2002) that allowed them to attend the ELSO Conference in Dresden, Germany (September 2003).

· Madalina Cojocaru, Ph D student (28 years old) got the 3^rd prize at International Congress of Medical Sciences (Sofia) for the oral presentation “Particularities of ophthalmic manifestations in rheumatic diseases” at International Congress of Medical Sciences, Sofia, Bulgaria, May 2003.

· Alexandra Dobrin, Ph.D student (23 years old) got the “Student award” at the Functional Genomics and Disease Congress, 14-17 May 2003, Prague, Czech Republic, and the second prize at the 29^th Romanian Congress of Diabetes, Nutrition and Metabolic Diseases (Craiova, Romania, May 2003).

· Simona Balan, Ph.D student (26 years old) received the “Student award” at the Functional Genomics and Disease Congress, 14-17 May 2003, Prague, Czech Republic.

· Support of EUR 8,000.00 of the German Diabetes Association to Dr. Irini Rakatzi (29)

· participation of Csaba Soti (33, HU) in EU 6^th Framework programme ZINCAGE as project leader and his election to a member of the EU Board of Aging Studies

· Dr. Daniela Gasperíková, PhD was a recipient of the Guoth´s Award of the Slovak Medical Society in the year 2003

· Arie Gruzman, a Ph.D. student in my lab received two awards in 2003 for the development of novel antihyperglycemic compounds: The 2003 Bern-Schlanger award from the Hebrew University Center from Diabetes Research and the 2003 Kaye award for students from the Hebrew University of Jerusalem

6. Dissemination of results

6.1. SCIENTIFIC PUBLICATIONS

Total number of COST B17-related publications

1999: 8

2000: 34

2001: 65

2002: 133

2003: >212

2004: N.A.

2005: N.A.

1999

1. Brown R. A., Domin, J., Arcaro, A., Waterfield M.D., and Shepherd P.R. (1999) Insulin activates the alpha isoform of class-II phosphoinositide 3-kinase Journal of Biological Chemistry 274: 14529-14532

3. Jones, D.R. and Varela-Nieto, I. Diabetes and the role of inositol-containing lipids in insulin signalling. Mol. Medicine 5, 505-514. 1999. (Review).

4. Kawano Y, Rincon J, Soler A, Ryder JW, Nolte LA, Zierath JR, Wallberg-Henriksson H. Changes in glucose transport and protein kinase Cbeta(2) in rat skeletal muscle induced by hyperglycaemia. Diabetologia. 1999 42:1071-1079.

5. Nave, B.T., Ouwens, M., Withers, D.J., Alessi, D.R. and Shepherd, P.R.: Mammalian target of rapamycin is a direct target for protein kinase B: identification of a convergence point for opposing effects of insulin and amino-acid deficiency on protein translation. Biochem. J. 344, 427-431, 1999.

6. Sebokova, E., Gasperikova, D., Ouwens, M., Dorrestijn, J., Eckel, J., Msen, A. and Klimes, I.: Defect in long-term activation of phosphatidylinositol 3-kinase by insulin in vivo: studies in insulin-resistant hHTg rats. Endocr. Regul. 33, 49-54, 1999.

7. Song XM, Ryder JW, Kawano Y, Chibalin AV, Krook A, Zierath JR. Muscle fiber type specificity in insulin signal transduction. Am J Physiol. 1999 277:R1690-1696

8. Zhang B, Salituro G, Szalkowski D, Li Z, Zhang Y, Royo I, Vilella D, Diez MT, Pelaez F, Ruby C, Kendall RL, Mao X, Griffin P, Calaycay J, Zierath JR, Heck JV, Smith RG, Moller DE. Discovery of a small molecule insulin mimetic with antidiabetic activity in mice.Science. 1999 284:974-977.

2000

1. Baynes K.C.R, Beeton C.A., Panayotou G., Stein R., Simpson H., Soos M., Hansen T., O'Rahilly S., Shepherd P.R. and Whitehead J.P..(2000) A novel missense mutation in the p85a phosphoinositide 3-kinase gene found in a pedigree with familial insulin resistance impairs insulin stimulated PI 3-kinase activity Diabetologia 43: 321-331

2. Beeton, C.A., Chance E.M. Foukas, L. & Shepherd P.R. (2000) Comparison of the kinetic properties of the lipid and protein kinase activities of the p110a and p110b catalytic subunits of class Ia PI 3-kinases Biochemical Journal 350:353-359

3. Chibalin AV, Yu M, Ryder JW, Song XM, Galuska D, Krook A, Wallberg-Henriksson H, Zierath JR. Exercise-induced changes in expression and activity of proteins involved in insulin signal transduction in skeletal muscle: differential effects on insulin-receptor substrates 1 and 2. Proc Natl Acad Sci U S A. 2000 97:38-43.

4. Constantinescu, D. Alexandru, V. Alexandru, M. Raicu, M. Simionescu. Endothelial cell-derived foam cells fail to express adhesion molecules (ICAM-1 and VCAM-1) for monocytes. J Submicrosc Cytol Pathol. 32 (2), 2000.

5. Costache, G. D. Popov, A. Georgescu, M. Cenuse, V.V. Jinga, M. Simionescu. The effects of simultaneous hyperlipemia-hyperglycemia on the mesenteric resistance arteries, myocardium and kidney glomeruli. J. Submicrosc. Cytol. Pathol., 32(1), 47-58, 2000a.

6. Costache, G. D. Popov, A. Georgescu, M. Simionescu. Functional-structural alterations of the resistance arteries in experimental hyperlipemia or hyperglycemia. Proc. Rom. Acad. Ser.B, 1, 31-37, 2000b.

7. Dimitriadis, G. and Newsholme, E.: Integration of some biochemical and physiologic effects of insulin that may play a role in the control of blood glucose concentration. In: LeRoith, Taylor, Olefsky (Editors) "Diabetes mellitus, a fundamental and clinical text", Lippincott Williams and Wilkins, 2nd Edition, pp 161-176, 2000.

8. Dobrian, V. Lazar, C. Sinescu, D. Mincu, M. Simionescu. Diabetic state induces lipid loading and altered expression of lipoprotein lipase in culture monocyte-derived macrophages. Atherosclerosis, 153: 191-201, 2000.

9. Dransfeld, O., Uphues, I., Sasson, S., Schurmann, A., Joost, H.G. and Eckel, J.: Regulation of subcellular distribution of GLUT4 in cardiomyocytes: Rab4A reduces basal glucose transport and augments insulin responsiveness. Exp. Clin. Endocrinol. Diabetes 108, 26-36, 2000.

10. Enrique-Tarancón, G. Y. Castan, N. Morin, L. Marti, A. Abella, M. Camps, R. Casamitjana, M. Palacín, X. Testar, E. Degerman, C. Carpéné, A. Zorzano. Substrates of semicarbazide-sensitive amine oxidase cooperate with vanadate to stimulate tyrosine phosphorylation of IRS proteins, phosphatidylinositol 3-kinase activity and GLUT4 translocation in adipose cells. Biochem. J. 350, 171-180, 2000.

11. Frago L.M., Camarero G., Cańón S., Pańeda C., Sanz C., León Y., Giraldez F. and Varela-Nieto I. Role of diffusible and transcription factors in inner ear development: implications in regeneration. Histol. Histopathol. 15, 657-666. 2000. (Review).

12. Gaster M, Handberg A, Beck-Nielsen H, Schroder HD. Glucose transporter expression in human skeletal muscle fibers. Am J Physiol Endocrinol Metab 2000a Sep;279(3):E529-E538

13. Gaster M, Poulsen P, Handberg A, Schroder HD, Beck-Nielsen H. Direct evidence of fiber type-dependent GLUT-4 expression in human skeletalmuscle. Am J Physiol Endocrinol Metab 2000b May;278(5):E910-E916

14. Georgescu, A. D. Popov. Age-dependent accumulation of advanced glycation endproducts is accelerated in combined hyperlipemia and hyperglycemia, a process attenuated by L-Arginine. J. Am. Aging Assoc. 23:33-40, 2000.

15. Glavan, G., Zorec, R., Babic. K., Sket, D., Zivin, M. (2000) Dopaminergic regulation of Synaptotagmin I and IV mRNAs in Hemiparkinsonian Rats Neuroreport, 11: 1-5.

16. Jinga, A. Gafencu, F. Antohe, E. Constantinescu, C. Heltianu, M. Raicu, I. Manolescu, W. Hunziker, M. Simionescu. Establishment of a pure microvascular endothelial cell line from human placenta. Placenta, 21(4), 325-336, 2000.

17. Kessler, A. E. Tomŕs, D. Immler, H.E. Meyer, A. Zorzano, J. Eckel. Rab 11 is associated with GLUT4-containing vesicles and redistributes in response to insulin. Diabetologia. 43, 1518-1527, 2000.

18. Krook A, Bjornholm M, Galuska D, Jiang XJ, Fahlman R, Myers MG Jr, Wallberg-Henriksson H, Zierath JR. Characterization of signal transduction and glucose transport in skeletal muscle from type 2 diabetic patients. Diabetes. 2000 49:284-292.

19. Kurtzhals, P., Schaffer, L., Sorensen, A., Kristensen, C., Jonassen, I., Schmid, C., and Trub, T.: Correlations of receptor binding and metabolic and mitogenic potencies of insulin analogs designed for clinical use. Diabetes 49, 999-1005, 2000.

20. Mazura I., Bendlová B., Vaňková M., Včelák J., Perušičová J., Svatoš J., Štefek M., Zvárová J. (2000): Genetic variation at the apo B 3´VNTR in Czech General population and in Czech diabetes mellitus type II patient group. Journal of Human Ecology 12 (1): 19-24.

21. Mohr T, Dela F, Handberg A, Biering-Sorensen F, Galbo H, Kjaer M.: Insulin action and long-term electrically induced training in individuals withspinal cord injuries. Med Sci Sports Exerc 2001 Aug;33(8):1247-1252

22. Morin, N. J.M. Lizcano, E. Fontana, L. Marti, F. Smih, P. Rouet, D. Prevot, A. Zorzano, M. Unzeta, C. Carpéné. Semicarbazide-sensitive amine oxidase substrates stimulate glucose transport and inhibit lipólisis in human adipocytes. Journal of Pharmacology and Experimental Therapeutics 297, 563-572, 2001.

23. Pimentel, B., Sanz, C., Rapp, U., Varela-Nieto, I., de Pablo, F. and de la Rosa, E. C-Raf regulates cell survival and retinal ganglion cell morphogenesis during neurogenesis. J. Neurosci. 20, 3254-3262. 2000.

24. Raicu, S. Florea, G. Costache, D. Popov, M. Simionescu. Clotrimazole inhibits smooth muscle cells proliferation and has a vasodilator effect on resistance arteries. Fundamental and Clinical Pharmacology 14: 477-485, 2000.

25. Rupnik, M. M. Kreft, I., S. K. Sikdar, S. Grilc., R. Romih, G. Zupančič, T. F. J. Martin, and R. Zorec (2000) Rapid Regulated Dense-Core Vesicle Exocytosis Requires the CAPS Protein PNAS 97:5627-5632.

26. Rupnik, M., Kreft, M., Limpel Kržan, M. & Zorec, R. (2000) Millisecond measurements of Ca²⁺ and exocytosis in neuroendocrine and glial cells. Acta Biologica Slovenica 43: 57-61.

27. Ryder JW, Yang J, Galuska D, Rincon J, Bjornholm M, Krook A, Lund S, Pedersen O, Wallberg-Henriksson H, Zierath JR, Holman GD. Use of a novel impermeable biotinylated photolabeling reagent to assess insulin- and hypoxia-stimulated cell surface GLUT4 content in skeletal muscle from type 2 diabetic patients. Diabetes. 2000 49:647-654.

28. Simionescu. M. Structural, biochemical and functional differentiation of the vascular endothelium. In: “Morphogenesis of the endothelium”, W.Risau (ed.), Harwood Academic Publishers, p.1-23, 2000;

29. Sőti Cs. and Csermely, P. (2000) Molecular chaperones and the aging process. Biogerontology, 1, 225-233

30. Till, M., Ouwens, D.M., Kessler, A. and Eckel, J.: Molecular mechanisms of contraction-regulated cardiac glucose transport. Biochem. J. 346, 841-847, 2000.

31. Vrbíková J., Hill M., Stárka L., Vondra K., Šulcová J., Šnajderová M., Cibula D., Pobišová Z., Bendlová B. (2000): An analysis of the relationship between insulin resistance and the activity of steroid C17,20-lyase and 3b-hydroxysteroiddehydrogenase in ovaries and adrenals in women with polycystic ovary syndrome. Exp Clin Endocrinol Diab, 108, 455-462.

32. Whitehead J. P., Soos, M. A., Aslesen, R., O’Rahilly, S., & Jensen, J. Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349:775-781, 2000.

33. Whitehead J.P., Soos, M.A., Aslesen, R., O Rahilly, S., and Jensen, J.: Contraction inhibits insulin-stimulated insulin receptor substrate-1/2 associated PI 3-kinase activity, but not PKB activation or glucose uptake in rat muscle. Biochem. J. 349, 775-781, 2000.

34. Wiese R, Homan U, M. Kreft, R. Zorec, G. Thiel (2000) Transient and irreversible exocytosis of vesicles in Zea mays coleoptile protoplasts measured in the cell-attached configuration. J. Membrane Biol. 174:15-20.

2001

2. Beauloye C, Bertrand L, Krause U, Marsin AS, Dresselaers T, Vanstapel F, Vanoverschelde JL, Hue L. No-flow ischemia inhibits insulin signaling in heart by decreasing intracellular pH. Circ Res. 2001 Mar 16;88(5):513-9.

3. Beauloye C, Marsin AS, Bertrand L, Krause U, Hardie DG, Vanoverschelde JL, Hue L. Insulin antagonizes AMP-activated protein kinase activation by ischemia or anoxia in rat hearts, without affecting total adenine nucleotides. FEBS Lett. 2001 Sep 21;505(3):348-52.

4. Becker C, Sevilla L, Tomas E, Palacin M, Zorzano A, Fischer Y. The endosomal compartment is an insulin-sensitive recruitment site for GLUT4 and GLUT1 glucose transporters in cardiac myocytes. Endocrinology. 2001 Dec;142(12):5267-76.

5. Bendlová B., Vrbíková J., Šaffková J., Stárka L. (2001): Genetické pozadí syndromu polycystických ovárií. Diabetes, metabolism , endocrinology and nutrition (DMEV) 4 (1): 58-62. )

6. Brown R. A. and Shepherd P.R. (2001) Growth factor regulation of the novel class-II PI 3-kinases Biochemical Society Transactions 29 535-537

7. Brumback, A.C., Zorec, R. & Betz, W. J. (2001) Integrins: the missing link. J. Physiol. (Lond) 530.1:1.

8. Burlacu, A. V.Jinga, A.V.Gafencu, M.Simionescu. The severity of the oxidative stress generates different mechanisms of endothelial cell death. Cell and Tissue Research. 306: 409-416, 2001

9. Cervenková K., Belejová M., Veselý J., Chmela Z., Rypka M., Ulrichová J., Modrianský M.,Maurel P.: Cell suspensions, cell cultures, and tissue slices – important metabolic in vitro systems. Biomed. Papers, 145, 57-60, 2001.

10. Chibalin AV, Kovalenko MV, Ryder JW, Feraille E, Wallberg-Henriksson H, Zierath JR. Insulin- and glucose-induced phosphorylation of the Na(+),K(+)-adenosine triphosphatase alpha-subunits in rat skeletal muscle. Endocrinology. 2001 Aug;142(8):3474-82.

11. Chmela Z., Veselý J., Lemr K., Rypka M., Hanuš J., Havlíček L., Kryštof V., Michnová L., Fuksová K., Lukeš J.: In vivo metabolism of 2,6,9-trisubstituted purine-derived cyclin-dependent kinase inhibitor bohemine in mice: glucosidation as the principal metabolic route. Drug Metab. Dispos. 29, 326-334, 2001.

12. Cinek, O., Drevinek, P., Sumnik, Z., Bendlova, B., Kolouskova, S., Snajderova, M. and Vavrinec, J.: The CTLA4 +49 A/G dimorphism is not associated with type 1 diabetes in Czech children. Eur. J. Immunogenet. 29, 219-222, 2001.

13. Clement S, Krause U, Desmedt F, Tanti JF, Behrends J, Pesesse X, Sasaki T, Penninger J, Doherty M, Malaisse W, Dumont JE, Le Marchand-Brustel Y, Erneux C, Hue L, Schurmans S. The lipid phosphatase SHIP2 controls insulin sensitivity. Nature. 2001 Jan 4;409(6816):92-7.

14. Conejo, R. and Lorenzo, M. (2001) Insulin signaling leading to proliferation, survival, and membrane ruffling in C2C12 myoblasts. J. Cell. Physiol. 187, 96-108

15. Conejo, R., Valverde, A.M., Benito, M. and Lorenzo, M. (2001) Insulin produces myogenesis in C2C12 myoblasts by induction of NF-KB and down-regulation of AP-1 activities.J. Cell. Physiol. 186, 82-94

16. Csermely, P. (2001a) A nonconventional role of molecular chaperones: involvement in the cytoarchitecture. News in Physiological Sciences. 15, 123-126, IF: 2.3

17. Csermely, P. (2001b) Chaperone-overload as a possible contributor to “civilization diseases”: atherosclerosis, cancer, diabetes. Trends in Genetics, 17, 701-704, IF: 12.9

18. Dimitriadis, G. and Raptis, S.: Thyroid hormone excess and glucose intolerance. Exp. Clin. Endocrinol. Diabetes 109 (Suppl 2), 225-239, 2001.

19. Dostálová, Mačáková J., Riegrová D., Červenková K., Veselý J.: Inhibition of excessive protein phosphorylation in neurones - new hopes for pharmacotherapy of Alzheimer dementia and other neurodegenerative diseases. Homeostasis, 41, 46-48, 2001.

20. Dransfeld O, Rakatzi I, Sasson S, Gruzman A, Schmitt M, Haussinger D, Eckel J. Eicosanoids participate in the regulation of cardiac glucose transport by contribution to a rearrangement of actin cytoskeletal elements. Biochem J. 2001 Oct 1;359(Pt 1):47-54.

21. Emanuelli, B., Peraldi, P., Filloux, C., Chavey, C., Freidinger, K., Hilton, D.J., Hotamisligil, G.S. and VanObberghen, E.: SOCS-3 inhibits insulin signaling and is up-regulated in response to tumor necrosis factor-alpha in the adipose tissue of obese mice. J. Biol. Chem. 276, 47944-47949, 2001

22. Fiedler M, Zierath JR, Selen G, Wallberg-Henriksson H, Liang Y, Sakariassen KS. 5-aminoimidazole-4-carboxy-amide-1-beta-D-ribofuranoside treatment ameliorates hyperglycaemia and hyperinsulinaemia but not dyslipidaemia in KKAy-CETP mice. Diabetologia. 2001 Dec;44(12):2180-6.

23. Fontana, E. J. Boucher, L. Marti, J.M. Lizcano, X. Testar, A. Zorzano, C. Carpéné. Amine oxidase substrates mimic several of the insulin effects on adipocyte differentiation in 3T3 F442A cells.Biochem.J. 356, 769-7717, 2001.

24. Ganea, E. "Chaperone-like Activity of alfa-Cristallin and Other Small Heat Shock Proteins", Current Protein & Peptide Science,2, 205, 2001.

25. Gaster M, Schroder HD, Handberg A, Beck-Nielsen H. The basal kinetics parameters of glycogen synthase in human myotubes cultures are not affected by chronic high insulin exposure. Biochemica et Biophysica Acta-Molecular Basis of Disease, 2001, 1537: 211-221

26. Gaster M, Staehr P, Beck-Nielsen H, Schroder HD, Handberg A. GLUT4 is reduced in slow muscle fibers of type 2 diabetic patients: is insulinresistance in type 2 diabetes a slow, type 1 fiber disease? Diabetes 2001 Jun;50(6):1324-1329

27. Georgescu, A. D. Popov, M. Simionescu. Mechanisms of decreased bradykinin-induced vasodilation in experimental hyperlipemia-hyperglycemia: contribution of nitric oxide and Ca²⁺ activated K⁺channels. Fundam. Clin. Pharmacol. 15,335-342, 2001.

28. Georgescu, A. D. Popov. Vascular reactivity of the resistance arteries to potassium in combined hyperlipemia-hyperglycemia. Proc. Rom. Acad. 2,111-115, 2001

29. Halse R, Bonavaud SM, Armstrong JL, McCormack JG, Yeaman SJ. Control of glycogen synthesis by glucose, glycogen, and insulin in cultured human muscle cells. Diabetes, 2001, 50: 720-726

30. Halse R, Pearson SL, McCormack JG, Yeaman SJ, Taylor R. Effect of tumor necrosis factor-alpha on insulin action in cultured human muscle cells. Diabetes, 2001, 50: 1102-1109

31. Halvorsen, B., A.C. Rustan, L. Madsen, J. Reseland, R.K. Berge, P. Sletnes, and E.N. Christiansen, Effect of long-chain n-3 and mono-unsaturated fatty acids on fatty acid oxidation and lipid composition in rats. Annals Nutrition & Metabolism, 2001, 45(1): p.

32. Hernandez, R., Teruel, T. and Lorenzo, M. (2001). Akt mediates insulin induction of glucose uptake and up-regulation of GLUT4 gene expression in brown adipocytes. FEBS Lett 494, 225-231

33. Jermendy, Gy. and Csermely, P. (2001) Thiazolidindiones - a new group of oral antidiabetica. Medical Weekly (Hung.), 142, 1547-1554, the publication received the Lajos Markusovszky Award for the best publication of the journal in 2001

34. Kawano Y, Ryder JW, Rincon J, Zierath JR, Krook A, Wallberg-Henriksson H. Evidence against high glucose as a mediator of ERK1/2 or p38 MAPK phosphorylation in rat skeletal muscle. Am J Physiol Endocrinol Metab. 2001 Dec;281(6):E1255-9.

35. Kessler, A., Uphues, I., Ouwens, D.M., Till, M. and Eckel, J.: Diversification of cardiac insulin signaling involves the p85alpha/beta subunits of phosphatidylinositol 3-kinase. Am. J. Physiol. 280, E65-E74, 2001.

36. Kiss, G., Heidegger, M., Somogyi, J., Csermely, P. and Ver, A. (2001) Streptozotocin-induced diabetes alters the oligomerization pattern of acetylcholinesterase in rat skeletal muscle. Diabetologia, 44, 220-223, IF: 5,2

37. Košmelj, K., A. Cedilnik, P. Veranič, G. Zupančič, M. Rupnik,L. K.Bobanovicand R. Zorec(2001) Intergranule Fusion In Rat Pars Intermedia. Image Analysis and Stereology 20:79-85.

38. Laviola L, Belsanti G, Davalli A, Napoli R, Perrini S, Weir GC, Giorgino R, Giorgino F. Effects of streptozotocin diabetes and diabetes treatment by islet transplantation on in vivo insuling signaling in diabetic heart. Diabetes 50:2709-2720, 2001.

39. Magre J, Delepine M, Khallouf E, Gedde-Dahl T, Jr., Van Maldergem L, Sobel E, Papp J, Meier M, Megarbane A, Bachy A, Verloes A, d'Abronzo FH, Seemanova E, Assan R, Baudic N, Bourut C, Czernichow P, Huet F, Grigorescu F, de Kerdanet M, Lacombe D, Labrune P, Lanza M, Loret H, Matsuda F, Navarro J, Nivelon-Chevalier A, Polak M, Robert JJ, Tric P, Tubiana-Rufi N, Vigouroux C, Weissenbach J, Savasta S, Maassen JA, Trygstad O, Bogalho P, Freitas P, Medina JL, Bonnicci F, Joffe BI, Loyson G, Panz VR, Raal FJ, O'Rahilly S, Stephenson T, Kahn CR, Lathrop M, Capeau J 2001 Identification of the gene altered in Berardinelli-Seip congenital lipodystrophy on chromosome 11q13. Nat Genet 28:365-370

40. Marti, A. Abella, C. Carpéné, M. Palacín, X. Testar, A. Zorzano. Combined treatment with benzylamine and low doses of vanadate enhances glucose tolerance and reduces hyperglycemia in streptozotocin-induced diabetic rats. Diabetes 50, 2061-2068, 2001.

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