| It is well established that insulin-dependent diabetes mellitus (IDDM)is an autoimmune disorder in which the insulin-producing beta cells are specifically destroyed. The non-obese diabetic (NOD) mouse spontaneously develops IDDM that has many immunological and pathological similarities to human type 1 diabetes. One of the most important mechanisms in type 1 diabetes is the breakdown of immunological tolerance and the appearance of autoreactivity. And multiple defects in immunological tolerance to "self predispose to immune-mediated (type 1) diabetes. Administration of insulin and B chain peptide (p9-23) can induce immunological tolerance, delay onset of diabetes, and decrease incidence of diabetes in NOD mice. The protection was obtained by promoting the shift from Th1 cells to Th2 cells response. Apoptosis is the main mode of death of β cells in NOD mice. The main peak of the apoptosis in NODmouse is at 15 weeks of age, which overlaps beginning time of clinic diabetes. Bcl-2 and related cytoplasmic proteins are key regulators of apoptosis. The Bcl-2 cohort promotes cell survival, whereas the Bax and BH3 cohorts facilitate apoptosis.Objective: Immunological tolerance was induced by intraperitoneally injecting insulin and B chain peptides (p9-23) at neonatal NOD mice. To observe their effects on the incidence of diabetes and insulitis in NOD mice, the expression of cytokines, Bcl-2 and Bax mRNA in pancreases, and Bcl-2 and Bax in islet cells. The aim of this study was to investigate the mechanisms of treatment with insulin and B chain peptides (9-23).Method: 96 female NOD mice were randomly divided into 3 groups. The mice were administered respectively insulin(0.25U)in 0.5ml PBS (A1 group), B chain peptides(50 μ g)in 0.5ml PBS (A2 group)and PBS(0.5ml) (A3 group) by intraperitoneal injection within 24h as neonates. The mice were reimmunized at 2,3,4and 9th week. 12 mice of each group were killed at 15th week. Pancreases were removed to examine insulitis scores through HE straining. And expression of Bcl-2 and Bax on islet cells were deteceted by immunohistochemical straining. Levels of cytokines,Bcl-2 and Bax mRNA of pancreases were measured by RT-PCR. 20 mice were used to evaluate incidence of diabetes. The mice were considered diabetes when urine was positive, and blood glucose was consistently >13.2mmol/L. Results:1) The onset time of diabetes was delayed both in Al and A2(at 22,23 weeks of age),compared with A3 (at 16 weeks of age). And the incidence of diabetes was decreased (32%,25%) in Al and A2 groups than A3 group (72%).2) Insulitis scores: Mean insulitis scores of Al and A2 group were significantly lower at 15 weeks of age compared with A3 group (P<0.01). There were no difference between Al and A2.3) Cytokines mRNA: Levels of pancreatic IFN- Y and TNF- a mRNA were lowered significantly in Al and A2 group than in A3 group(P<0.01).There were no statistical changes between A1 and A2 group (P>0.05). Compared with A3, the expression of IL-10 of pancreases increased markedly in A1 and A2 (P<0.01), but there was no difference between A1 and A2 (P>0.05).4) Bcl-2 , Bax mRNA: Levels of Bcl-2 mRNA of pancreases were higher in A1 and A2 compared with A3 (P<0.01). While levels ofBax mRNA were lower in A1 and A2.5) Expression of Bcl-2 and Bax on islet cells: the expression ofBcl-2 on islet cells was stronger in Al and A2 compared with A3(P<0.05). And the expression of Bax was weaker in Al and A2groups.Conclusion:1) Insulin or B chain peptides (p9-23) administered in intraperitoneal in mice as neonates can lighten insulitis seriousness, delay onset of diabetes and decrease incidence of diabetes.2) Insulin or B chain peptides (p9-23) can prevent NOD mice from diabetes by down-regulated Th1 cell response, up-regulated Th2 cell response, and prime immune deviation from predominant Thl to Th2 response.3) Insulin and B chain peptides(p9-23) can prevent from diabetes through an increased expression of Bcl-2... |
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