Molecular Biology of B-Cells of Pancreatic Islets of Langerhans : Basic Study to develop Gene Therapy for Diabetes

胰岛b细胞分子生物学研究结果如下：随机cDNA测序法结合Northern blot对培养的小鼠胰岛b细胞细胞系进行分析，获得了约20个胰岛b细胞特异性克隆。从胰岛A细胞中产生免疫抑制IL-10或tgf - β的转基因小鼠。IL-10在NOD小鼠中的转基因表达加重了自身免疫性糖尿病。迄今为止，NOD小鼠体内的两株tgf - β - tg未表现出糖尿病。在5个胰岛b细胞特异性th1淋巴细胞中，2个产生ifn - γ和IL-2, 3个产生IL-6, 10个在它们之上。多重t细胞功能是自身免疫性糖尿病发展的必要条件。缺乏CD40（对t细胞分化很重要）的NOD小鼠没有患上糖尿病，这表明需要CD40来产生自我反应的t细胞。用HSV-TK加更昔洛韦的失安全系统移植产生胰岛素原的成纤维细胞，或过继转移具有识别胰岛抗原能力的il- 10转导淋巴细胞，用于开发糖尿病基因治疗的动物模型。由于转基因小鼠的胰岛素分泌通过己糖激酶的转导而增加，而葡萄糖激酶反义mRNA的表达而减少，因此胰岛b细胞中的葡萄糖激酶起着葡萄糖敏感传感器的作用。在胰岛内分泌前体细胞中检测到激活素A。胰岛b细胞中检测到tgf - β。激活素A抑制atp敏感的钾通道，激活电压依赖性钙通道，导致胰岛素分泌增加。激活素A与β细胞合作，诱导AR42J胰腺外分泌细胞向胰岛素生成细胞分化。

Molecular biological approaches to pancreatic islet B-cells led to following results ;1.Random cDNA sequencing approach comined with Northern blot analysis to a cultured murine cell line of islet B-cells, yielded abount 20 islet B-cell-specific clones.Transgenic mice producing immunosuppressive IL-10 or TGF-beta from pancreatic islet A cells were produced.Transgenic expression of IL-10 in NOD mice aggravated autoimmune diabetes. Two strains of TGF-beta-Tg in NOD mice so far did not exhibit diabetes.Among 5 islet B-cell-specific Th1-lymphoctes, 2 produced IFN-gamma and IL-2, and 3 others produced IL-6 and 10 on top of them. Multiple T-cell functions are necessary for the development of autoimmune diabetes. NOD mice lacking CD40, which is important for T-cell Differentiation, did not become diabetic, suggesting the requirement of CD40 to produce self-reacting T-cells.Transplantaion of proinsulin-producing fibroblasts with the fail-safety systm of HSV-TK plus ganciclovir, or adoptive transfer of IL-10-transduced lymphocytes with the ability to recognize islet-antigens were used to develop animal models of gene therapy for diabetes.Because insulin secretion was increased by transduction of hexokinase, and was decreased by expression of antisense mRNA for glucokinase in transgenic mice, glucokinase in islet B-cells is functioning as a glucose-sensitive sensor.Activin A was detected in the endocrine precursor cells of islets. TGF-beta was detected in pancreatic islet B-cells. Activin A inhibited ATP-sensitive potassium channel, and activated voltage-dependent calcium channels, leading to the increased-insulin secretion. Activin A, in cooperation with betacelluin, induced differentiation of pancreatic exocrine cells of AR42J to insulin-producing cells.