STAT1-mediated down-regulation of Bcl-2 expression is involved in IFN-γ/TNF-α-induced apoptosis in NIT-1 cells.
STAT1-mediated down-regulation of Bcl-2 expression is involved in IFN-γ/TNF-α-induced apoptosis in NIT-1 cells.
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DOI：
10.1371/journal.pone.0120921
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Zhang KQ
中科院分区：
文献类型：
--
作者：
Cao ZH;Zheng QY;Li GQ;Hu XB;Feng SL;Xu GL;Zhang KQ
肿瘤坏死因子（TNF）-α和干扰素（IFN）-γ是参与β细胞破坏的主要促炎细胞因子。胰岛β细胞在甘氨酸诱导的内在线粒体凋亡途径中的命运由Bcl-2家族成员之间的相互作用决定。然而，β细胞凋亡的调节机制仍不清楚。在本研究中，我们用TNF-α和IFN-γ处理小鼠β细胞系NIT-1，然后研究信号转导和转录激活因子-1（STAT-1）和Bcl-2家族成员在该凋亡通路中的表达的调节。结果显示TNF-α和IFN-γ协同降低NIT-1细胞活力。此外，细胞生长的减少是由于凋亡，如通过共聚焦激光显微镜检测到的凋亡小体形成所示，以及通过流式细胞术检测到的膜联蛋白-Vup+细胞百分比的显著增加。TNF-α和IFN-γ联合处理可显著增加细胞色素c的释放，增加caspase-9和caspase-3的活化，并显著增强STAT-1的磷酸化，但可下调Bcl-2的表达。在暴露于TNF-α和IFN-γ联合处理后，在另一种鼠β细胞衍生系MIN 6细胞中也观察到STAT-1活性的增强和Bcl-2表达的下调。通过siRNA敲低STAT-1基因表达或用氟达拉滨抑制STAT-1活化可逆转Bcl-2表达下调，并导致TNF-α和IFN-γ处理的NIT-1细胞凋亡显著减少。综上所述，我们的研究结果表明STAT 1介导的Bcl-2下调参与了TNF-α和IFN-γ联合诱导的NIT-1细胞凋亡。
Tumor necrosis factor (TNF)-α and interferon (IFN)-γ are the major pro-inflammatory cytokines involved in beta-cell destruction. The fate of islet beta-cells in the cytokine-induced intrinsic mitochondrial apoptotic pathway is determined by the interaction between members of the Bcl-2 family. However, the mechanism through which beta-cell apoptosis is regulated remains unclear. In this study, we treated the murine beta-cell line NIT-1 with TNF-α and IFN-γ and then investigated the regulation of signal transducer and activator of transcription-1 (STAT-1) and expression of the members of the Bcl-2 family in this apoptotic pathway. Results showed that TNF-α and IFN-γ synergistically reduced NIT-1 cell viability. In addition, the decrease in cell growth was due to apoptosis as shown by apoptotic body formation, detected by confocal laser microscope, and a significant increase in Annexin-Vup+ cell percentage, detected by flow cytometry. Combination treatment with TNF-α and IFN-γ caused a remarkable increase in the release of cytochrome c, and in the activation of caspase-9 and caspase-3, as well as, an obvious enhancement in STAT-1 phosphorylation; the treatment, however, resulted in the down-regulation in Bcl-2 expression. The enhancement in STAT-1 activity and a down-regulation in Bcl-2 expression was also observed in MIN6 cells, another murine beta-cell derived line, after cells exposure to the combination of TNF-α and IFN-γ treatment. Knockdown of STAT-1 gene expression by siRNA or inhibition of STAT-1 activation with fludarabine reversed Bcl-2 down-expression and led to a significant decrease in apoptosis in TNF-α- and IFN-γ-treated NIT-1 cells. Taken together, our results suggest that STAT1-mediated down-regulation of Bcl-2 is involved in NIT-1 cell apoptosis induced by combination treatment with TNF-α and IFN-γ.
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