目的：观察吡格列酮（ PGZ）联合5－氮杂胞苷（5－AzaC）干预对大鼠胰岛素瘤细胞增殖、凋亡及胰岛素分泌量的影响。方法2015年6月—2016年6月于河南省平顶山市第二人民医院内分泌科进行实验。将大鼠胰岛素瘤细胞（RIN－m5f）行体外原代及传代培养，将培养好的细胞分为5组：空白组（不加任何试剂）、模型组[白细胞介素－1β（IL－1β）2 ng／ml＋干扰素γ（IFN－γ）100 U／ml]、吡格列酮组（IL－1β2 ng／ml ＋IFN－γ100 U／ml＋PGZ 15μmol／L）、5－AzaC组（IL－1β2 ng／ml ＋IFN－γ100 U／ml ＋5－AzaC 1．5μmol／L）及PGZ＋5－AzaC组（IL－1β2 ng／ml ＋IFN－γ100 U／ml ＋PGZ 15μmol／L ＋5－AzaC 1．5μmol／L）。应用倒置显微镜观察RIN－m5f细胞形态学的变化，流式细胞仪测定RIN－m5f细胞凋亡情况，应用Western blot法检测B细胞淋巴瘤－2（Bcl－2）、多糖聚合酶（PGZRP）表达情况，应用ELISA法检测葡萄糖刺激胰岛素分泌能力。结果 RIN－m5f细胞培养24 h、48 h、72 h后应用流式细胞仪可观察到PGZ组、5－AzaC组、PGZ＋5－AzaC组和RIN－m5f细胞凋亡率低于模型组（ q PGZ组＝12．122、8．452、8．252， P均＜0．05；qt5－AzaC组＝12．252、8．563、7．529， P均＜0．05；q PGZ＋5－AzaC组＝8．563、7．452、8．963， P ＜0．05），PGZ＋5－AzaC组细胞凋亡率低于PGZ组与5－AzaC组（ q PGZ组＝7．022、6．986、8．523， P均＜0．05；q 5－AzaC组＝8．963、9．123、10．523， P均＜0．05），培养48 h、72 h后PGZ＋5－AzaC组细胞凋亡率较培养24 h时显著下降（ q ＝5．996、6．789， P均＜0．05）。 Bcl－2、PG－ZRP表达水平：模型组＞PGZ组＞5－AzaC组＞PGZ＋5－AzaC组（ q ＝7．896、8．233、9．102， P均＜0．05）。 PGZ＋5－AzaC组葡萄糖刺激胰岛素分泌能力大于PGZ组、5－AzaC组（ q ＝8．252、7．896， P均＜0．05），5－AzaC组葡萄糖刺激胰岛素分泌能力大于PGZ组，差异均有统计学意义（q＝8．123， P ＜0．05）。结论 PGZ联合5－AzaC能有效抑制高糖诱导大鼠胰岛素瘤凋亡，恢复胰岛素瘤分泌能力，其作用机制可能与其能下调Bcl－2、PGZRP表达水平有关。%Objective To investigate pioglitazone ( PGZ) combined with 5 azacytidine (5-AzaC) intervention on in-sulin the proliferation , apoptosis ofβcell proliferation and insulin βcell insulin secretion .Methods The experiment was car-ried out in Department of Endocrinology , Second People's Hospital of Henan province from June 2015 to June 2016 in Ping-dingshan.The rat insulinoma cell line (RIN-m5f) in vitro and cultured in vitro, the cultured cells were divided into 5 groups:control group ( without any reagent ) , model group ( IL-1β2 ng/ml+IFN-γ100 U/ml) ,pioglitazone group ( IL-1β2 ng/ml +IFN-γ100 U/ml+PGZ 15 μmol/L),5-AzaC group ( IL-1β2 ng/ml +IFN-γ100 U/ml +5-AzaC 1.5 μmol/L) and PGZ+5-AzaC group (IL-1β2 ng/ml+IFN-γ100 U/ml +PGZ 15 μmol/L +5-AzaC 1.5 μmol/L).The inverted micro-scope was used to observe the change of morphology of RIN-m5f cells,RIN-m5f-determination of cell apoptosis by flow cytome-try and the detection of B cell lymphoma by Western blot 2 (Bcl-2), polysaccharides (PGZRP) expression was detected by ELISA, glucose stimulated insulin secretion.Results The apoptotic rate of RIN-m5f cells in PGZ group, 5-AzaC group and PGZ +5-AzaC group was lower than that in the model group after 24,48,72 h( q PGZ group=12.122,8.452,8.252, P <0.05;qt5-AzaC group =12.252、8.563、7.529, P <0.05;q PGZ+5-AzaC group =8.563、7.452、8.963, P <0.05).The apoptosis rate of PGZ+5-AzaC group was significantly lower than that of PGZ group and 5-AzaC group ( q PGZ group =7.022、6.986、8.523, P <0.05;q 5-AzaC group =8.963、9.123、10.523, P <0.05),but the difference was not statistically significant ( P >0.05).The apoptosis rate in PGZ +group was significantly lower than that in PGZ +group ( P >0.05).The expression of Bcl-2 and PGZRP was decreased in model group >PGZ group>5-AzaC group>PGZ +5-AzaC group.PGZ +5-AzaC group glucose stimulation better than PGZ group and 5-AzaC group ( P <0.05).The insulin secretion ability of the 5-AzaC group was sig-nificantly higher than that of the PGZ group , 5-AzaC group ( q =8.252, q =7.896, P <0.05).Conclusion The PGZ combined with 5-AzaC can effectively inhibit the apoptosis of insulinoma induced by high glucose , and restore the ability of in-sulin secreting tumor.The mechanism may be related to the down regulation of the expression of Bcl-2 and PGZRP.
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