Akt phosphorylation in human chondrocytes is regulated by p53R2 in response to mechanical stress
Akt phosphorylation in human chondrocytes is regulated by p53R2 in response to mechanical stress
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DOI：
10.1016/j.joca.2012.08.022
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发表时间：
2012-12-01
影响因子：
7
通讯作者：
Kurosaka, M.
中科院分区：
文献类型：
--
作者：
Kawakita, K.;Nishiyama, T.;Kurosaka, M.
目的：P53肿瘤抑制蛋白p53R2被激活，以响应作用于细胞信号转导的各种应激源。当DNA被损伤时，p53在其15位序列上的磷酸化诱导p53R2的产生。P53R2在软骨细胞中的作用仍然知之甚少。在这项研究中，我们评估了软骨细胞中p53R2的表达及其对机械应力的调节作用。方法：采用免疫组织化学、Western blotting和实时定量聚合酶链式反应(RT-PCR)检测p53R2在人工全膝关节置换术和正常股骨颈骨折软骨中的表达。使用FX-2000 Flexpercell系统对骨性关节炎软骨细胞施加高强度的周期性拉伸应变。接下来，对这些细胞中的硫酸糖胺聚糖(SGAG)的产量进行了量化。Western blotting检测p53R2蛋白表达及Akt、p38MAPK、ERK1/2和JNK蛋白的磷酸化。结果：5%的张应力作用于骨性关节炎软骨细胞后，软骨细胞中p53R2的表达显著增加。然而。分离株后骨关节炎软骨细胞AKT磷酸化水平下调，p53R2基因转染后AKT磷酸化水平上调。结论：p53R2可通过Akt磷酸化途径调节软骨细胞机械转导过程中基质合成。下调p53R2的表达可能成为治疗骨性关节炎的新途径。(C)2012年国际骨性关节炎研究会。爱思唯尔有限公司出版。保留所有权利。
Objective: The p53 tumor-suppressor protein p53R2 is activated in response to various stressors that act on cell signaling. When DNA is damaged, phosphorylation of p53 at its Ser 15 residue induces p53R2 production. The role of p53R2 in chondrocytes remains poorly understood. In this study, we evaluated in chondrocytes, p53R2 expression and its regulation in response to mechanical stress. Furthermore, we investigated the function of p53R2 in relation to mechanotransduction.Methods: Osteoarthritis (OA) cartilage obtained from total knee replacements and normal cartilage obtained from femoral neck fractures was used to measure p53R2 expression by using immunohistochemistry, western blotting, and real-time polymerase chain reaction (PCR). The OA chondrocytes were subjected to a high magnitude of cyclical tensile strain by using an FX-2000 Flexercell system. Next, sulfated glycosaminoglycan (sGAG) production was quantified in these cells. Protein expression of p53R2, and phosphorylation of Akt, p38MAPK, ERK1/2, and JNK was also detected using western blotting. Moreover, Akt phosphorylation was detected after transfecting the cells with p53R2-specific small interfering RNA (siRNA).Results: Expression of p53R2 was significantly increased in OA chondrocytes and in chondrocytes after applying 5% tensile strain to the cells. However. Akt phosphorylation was down-regulated in OA chondrocytes after the strain, and was up-regulated after transfection of p53R2. sGAG protein as well as collagen type II and aggrecan mRNA was increased following transfection of p53R2-specific siRNA after 5% tensile strain.Conclusions: p53R2 could regulate matrix synthesis via Akt phosphorylation during chondrocyte mechanotransduction. Down-regulation of p53R2 may be a new therapeutic approach in OA therapy. (C) 2012 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.