| Project/Area Number |
11670148
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Pathological medical chemistry
|
|---|
| Research Institution | Kyorin University |
|---|
Principal Investigator |
NAGAMATSU Shinya Kyorin University School of Medicine, Professor, 医学部, 教授 (80231489)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
WATANABE Takashi Kyorin University School of Medicine, Professor, 医学部, 教授 (00191768)
ISHIDA Hitoshi Kyorin University School of Medicine, Professor, 医学部, 教授 (80212893)
|
|---|
| Project Period (FY) |
1999 – 2001
|
| Project Status |
Completed (Fiscal Year 2001)
|
| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2001: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2000: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
|
|---|
| Keywords | Insulin / GFP / Fusion / confocal microscoov / evanescent / secretory granule / diabetes mellitus / fusion / インスリン分泌 / Exocytosis / SNARE / Ca^<2+> |
|---|
| Research Abstract |
In order to elucidate the mechanism of impaired insulin release in diabetes mellitus, we firstly investigated the expression of SNAREs and its related proteins in pancreatic β cells. We revealed that MIN6 β cells expressed syntaxin 1A, syntaxin 2 (epimorphin), synaptosomal associated protein of 25 kDa (SNAP25), vesicle associated membrane protein-2 (VAMP2 or synaptobrevin), synaptotagmin I, III, IV. To examine how these proteins are involved in the etiology and progression of diabetes mellitus, we examined the expression levels of syntaxin 1A and SNAP25 in pancreatic β cells of GK rats, type 2 DM animal model. These protein levels were decreased to about a half-fold that of control. Then, we tried to recover the decreased expression levels to the normal levels using recombinant adenovirus system. Infection of Adex 1CA-syntaxin 1A and Adex 1CA-SNAP25 with GK rat islets recovered these protein levels, thereby the impaired insulin release was partly normalized. Thus, our data strongly indicated the involvement of t-SNAREs expression in the etiology and progression of type 2-DM. We then investigated how endocytosis insulin secretory granules post fusion is associated with insulin release. For this purpose, we have cloned the novel-cellubrevin related peptide (NGRP), which was localized in the early endosome. To determine the functional role of NCRP (endobrevin) in the exocytosis of insulin, wild-type and dominant-negative endobrevin were overexpressed using adenovirus system. The data revealed that endobrevin plays a physiological role in the exo-endocytosis of insulin via an interaction between endocytic vesicles and the endosome. Finally, we developed the new strategy to detect the insulin fusion per se by fusion protein between insulin and pH-sensitive GFP mutant (pHluorin).
|
