耐受性树突状细胞过继转移建立1型糖尿病小鼠感染性免疫耐受

Establishment and mechanism study of infectious immune tolerance in type 1 diabetes mellitus by tolerogenic dendritic cells

目的探讨耐受性树突状细胞（DC）过继转移，在T1DM模型小鼠体内建立感染性免疫耐受的方法和机制。方法在链脲佐菌素（STZ）所致1型糖尿病模型（type 1 diabetes mellitus，T1DM）内采用胰岛素与IEA混合乳剂皮下注射的方式诱导免疫耐受。体外分离和纯化耐受性树突状细胞。另取BALB／c小鼠，通过体内腹腔注射STZ，尾静脉注射T1DM发病小鼠脾脏T淋巴细胞（DTE），诱导T1DM。同时将不同来源DC经腹腔注射入模型小鼠体内。每周测定血糖，第4周时处死动物，取胰腺进行病理组织学检查。MTT法测定小鼠脾淋巴细胞增殖反应，采用流式细胞术检测CD4^＋CD25^＋调节性T细胞。结果采用2次小剂量STZ加DTL联合注射的方式可在小鼠体内建立以胰腺B细胞炎性破坏为主要病理特征的T1DM。通过过继转移免疫耐受性DC，可明显降低糖尿病高血糖以及小鼠脾淋巴细胞增殖能力，与模型对照组差异有统计学意义（P〈0．01）；组织病理检查发现胰岛内炎症细胞浸润减少，组织结构完整；FACS显示CD4^＋CD25^＋T细胞亚群比例显著升高。结论过继转移耐受性Dc可以在次一级T1DM模型内诱导感染性免疫耐受并防止T1DM的发生，其机制与促进体内CD4^＋CD25^＋T细胞亚群产生相关。

Objective To establish the infectious immune tolerance in T1DM murine model by adoptive transfer of tolerogenic dendritic cells（DC） and investigate the mechanism. Methods Immune tolerance was established by subcutaneously injection of the bovine insulin （ 100 μg） in IFA （ emulsified 1 ： 1 ） in streptozotocin（STZ） induced type 1 diabetes mellitus（T1DM） model. Tolerogenic dendritic cells were isolated and purified ex vivo. The T1DM model was also established by intraperitoneally injection of STZ 40 mg/kg twice accompanied by intravenous injection of diabetogenic splenic T lymphocytes（DTL） in BALB/c mice. Simultaneously, different resources of DC were injected intraperitoneally into prediabetic mice. Mice were sacrificed after 4 weeks and pancreas were taken for histopathologic examination. Lymphocyte proliferation was assayed by MTT method and CD^4＋ CD25^＋ T cell differerntiation was measured by FACS analysis. Results Low dose of STZ injected twice combined with DTL injection in BALB/c mice can develop a stable model of T1DM, characterized as the beta cell destruction by inflammatory cells; Both of the high level blood glucose and T cell proliferation were decreased significantly by adoptive transfer of tolerogenic DC（ P 〈 0.01） ; less lymphocytes infiltration was observed in islet and pancreatic histological structure was intact; CD4^＋ CD25^＋ T cells differentiation was increased remarkably. Conclusion Infectious immune tolerance could be induced by adoptive transfer of tolerngenic DC in secondary T1DM model and confer protection to mice from T1DM, the mechanism of which suppose to be associated with promoting the production of CD4^＋ CD25^＋ T cells in vivo.