SIRT1 Regulation of Apoptosis of Human Chondrocytes
SIRT1 Regulation of Apoptosis of Human Chondrocytes
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DOI：
10.1002/art.24864
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发表时间：
2009-09-01
影响因子：
--
通讯作者：
Kuroda, Ryosuke
中科院分区：
文献类型：
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作者：
Takayama, Koji;Ishida, Kazunari;Kuroda, Ryosuke
Objective.已知SIRT 1抑制细胞凋亡并促进各种类型细胞的存活。然而，SIRT 1在人软骨细胞凋亡中的作用尚未见报道。我们进行了这项研究，以探讨SIRT 1与人类软骨细胞凋亡的关系，这是骨关节炎（OA）的一个特征。SIRT 1在人软骨细胞中的表达通过逆转录-聚合酶链反应、免疫印迹和人软骨样品的免疫组织学检查。通过免疫印迹法研究了SIRT 1在分解代谢、机械和营养应激下的表达。为了检测SIRT 1对细胞凋亡的影响，在一氧化氮（NO）诱导的细胞凋亡过程中，SIRT 1被小干扰RNA（siRNA）抑制，并被白藜芦醇激活。TUNEL染色和裂解的聚（ADP-核糖）聚合酶（PAR-P）免疫印迹法检测细胞凋亡。为了研究细胞凋亡的机制，我们使用免疫印迹法测定线粒体中切割的半胱天冬酶和凋亡相关的凋亡信号蛋白Bax和Bcl-2的水平。在人软骨细胞和人软骨样品中证实了SIRT 1表达。所有分解代谢，机械和营养应力抑制SIRT 1的表达。通过siRNA抑制SIRT 1增加了TUNEL阳性细胞的百分比，并增加了NO诱导的裂解的PARP和裂解的caspase 3和9的量。相反，用白藜芦醇处理降低了TUNEL阳性细胞的百分比，并降低了NO诱导的裂解的PARP和裂解的caspase 3和9的量。SIRT 1 siRNA抑制SIRT 1后，Bax表达增加，Bcl-2表达减少;白藜芦醇抑制SIRT 1后，Bax表达减少，Bcl-2表达增加。这些结果表明，SIRT 1通过调节软骨相关的凋亡信号来调节人软骨细胞的凋亡。对SIRT 1的进一步研究可能有助于阐明OA的发病机制。
Objective. SIRT1 is known to inhibit apoptosis and to promote survival of various types of cells. However, the roles of SIRT1 in apoptosis of human chondrocytes have never been reported. We undertook this study to investigate the relationship of SIRT1 to apoptosis of human chondrocytes, which is a characteristic feature of osteoarthritis (OA).Methods. The expression of SIRT1 in human chondrocytes was examined by reverse transcription-polymerase chain reaction, immunoblotting, and immunohistology of human cartilage samples. The expression of SIRT1 under catabolic, mechanical, and nutritional stresses was investigated by immunoblotting. To examine the effect of SIRT1 on apoptosis, SIRT1 was inhibited by small interfering RNA (siRNA) and activated by resveratrol during nitric oxide (NO)-induced apoptosis. TUNEL staining and immunoblotting of cleaved poly(ADP-ribose) polymerase (PAR-P) were performed to detect apoptosis. To examine the mechanisms of apoptosis, we used immunoblotting to determine the levels of cleaved caspases and mitochondria-related apoptotic signaling proteins, Bax and Bcl-2, in the mitochondrial fraction.Results. SIRT1 expression was confirmed in human chondrocytes and human cartilage samples. All catabolic, mechanical, and nutritional stresses inhibited SIRT1 expression. SIRT1 inhibition by siRNA for SIRT1 increased the percentage of TUNEL-positive cells and increased the amounts of cleaved PARP and cleaved caspases 3 and 9 induced by NO. In contrast, treatment with resveratrol decreased the percentage of TUNEL-positive cells and decreased the amounts of cleaved PARP and cleaved caspases 3 and 9 induced by NO. Furthermore, in the mitochondrial fraction, SIRT1 inhibition by siRNA for SIRT1 increased the amount of Bax but reduced the amount of Bcl-2, while resveratrol reduced the amount of Bax but increased the amount of Bcl-2.Conclusion. These results indicate that SIRT1 regulates apoptosis in human chondrocytes through the modulation of mitochondria-related apoptotic signals. Further research on SIRT1 might contribute to resolving the pathogenesis of OA.