Caspase-3 is Involved in IFN-γ- and TNF-α-Mediated MIN6 Cells Apoptosis via NF-κB/Bcl-2 Pathway
Caspase-3 is Involved in IFN-γ- and TNF-α-Mediated MIN6 Cells Apoptosis via NF-κB/Bcl-2 Pathway
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DOI：
10.1007/s12013-013-9642-4
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发表时间：
2013-12-01
影响因子：
2.6
通讯作者：
Zhang, Ke-qin
中科院分区：
文献类型：
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作者：
Cao, Zhao-hui;Yin, Wei-dong;Zhang, Ke-qin
TNF-α和IFN-γ是β细胞破坏中的主要促炎细胞因子。然而，其潜在机制仍不清楚。本研究以小鼠胰岛素瘤细胞株MIN 6为研究对象，探讨Caspase-3在细胞因子诱导的胰岛β细胞凋亡中的作用，并分析Caspase-3的激活机制。结果表明，IFN-γ和TNF-α的组合显著降低了MIN 6细胞的活力，并且观察到的细胞生长抑制是由于细胞凋亡，如通过在共聚焦激光扫描显微镜下的形态学变化和Annexin-V/7-AAD双染色的FACS测定所判断的。IFN-γ和TNF-α处理后24 h和12 h，分别观察到裂解的Caspase-3和Caspase-3的已知底物PARP，同时伴有NF-κ B激活和Bcl-2下调。Caspase-3抑制剂预处理可显著减弱IFN-γ和TNF-α诱导的细胞凋亡。抑制NF-κ B活化导致Bcl-2表达增加，Caspase-3活性显著减弱，IFN-γ和TNF-α处理的MIN 6细胞的细胞活力明显改善。综上所述，我们的研究结果表明Caspase-3在诱导MIN 6细胞凋亡中起关键作用，并且进一步证实其活化与NF-κ B介导的Bcl-2下调有关，这可能是IFN-γ和TNF-α介导的MIN 6细胞凋亡的潜在机制。
TNF-alpha and IFN-gamma are the major pro-inflammatory cytokines in the beta-cell destruction. However, the underlying mechanism remains unclear. The present study used a murine insulinoma cell line MIN6 for further investigation of the effect of Caspase-3 on the cytokines-induced pancreatic beta-cell apoptosis and analyzed the mechanisms involved in the activation of Caspase-3. It was showed that the combination of IFN-gamma and TNF-alpha significantly reduced the viability of MIN6 cells and the observed cells growth inhibition was due to cell apoptosis as judged by the morphological changes under a confocal laser scanning microscopy and FACS assay of Annexin-V/7-AAD double staining. Accompanying with NF-kappa B activation and Bcl-2 downregulation, both the cleaved Caspase-3 and PARP, a known substrate of Caspase-3 in vivo, were observed at 24 and 12 h, respectively, after cells exposure to IFN-gamma and TNF-alpha treatment. Pretreatment of Caspase-3 inhibitors remarkably attenuated IFN-gamma- and TNF-alpha-induced cells apoptosis. Inhibition of NF-kappa B activation led to the increase in Bcl-2 expression, a significant attenuation in Caspase-3 activity, and an obvious amelioration in cells viability in IFN-gamma- and TNF-alpha-treated MIN6 cells. Taken together, our results indicate that Caspase-3 is critical for the induction of MIN6 cells apoptosis and it's activation is further confirmed to be related to the NF-kappa B-mediated Bcl-2 downregulation, which may be the underlying mechanism of IFN-gamma- and TNF-alpha-mediated MIN6 cells apoptosis.