Hydrostatic Pressure Regulates MicroRNA Expression Levels in Osteoarthritic Chondrocyte Cultures via the Wnt/β-Catenin Pathway.
Hydrostatic Pressure Regulates MicroRNA Expression Levels in Osteoarthritic Chondrocyte Cultures via the Wnt/β-Catenin Pathway.
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DOI：
10.3390/ijms18010133
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发表时间：
2017-01-12
影响因子：
5.6
通讯作者：
Fioravanti A
中科院分区：
文献类型：
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作者：
Cheleschi S;De Palma A;Pecorelli A;Pascarelli NA;Valacchi G;Belmonte G;Carta S;Galeazzi M;Fioravanti A
机械负荷和静水压力（HP）调节软骨细胞的代谢;然而，机械刺激如何起作用仍不清楚。微小RNA（microRNAs，miRNAs）在软骨稳态、力学信号转导和骨关节炎（osteoarthritis，OA）的发病机制中起着重要作用。本研究调查了正常和OA人软骨细胞中循环HP（1-5 MPa）对miR-27 a/B、miR-140、miR-146 a/B和miR-365及其靶基因（MMP-13、ADAMTS-5、IGFBP-5和HDAC-4）表达的影响。此外，我们评估了Wnt/β-catenin通路在HP应答中的可能参与。将软骨细胞暴露于HP 3 h，并在加压后立即以及12、24和48 h后进行评价。提取总RNA并用于实时PCR。β-catenin的检测采用Western blotting和免疫荧光法。在OA软骨细胞中，HP在所有分析时间点诱导miR-27 a/B、miR-140和miR-146 a的表达水平显著增加（p < 0.01），并且miR-365的表达水平显著降低（p <0.01）。MMP-13、ADAMTS-5和HDAC-4在HP后显著下调，而IGFBP-5没有显著改变。在基础条件下，OA软骨细胞中β-连环蛋白水平显著增加（p < 0.001），而HP显著降低（p < 0.01）。加压并没有引起任何显着的修改，在正常细胞。总之，在OA软骨细胞中，HP恢复了一些miRNA的表达水平，下调了MMP-13、ADAMTS-5和HDAC-4，并调节了Wnt/β-catenin通路的激活。
Mechanical loading and hydrostatic pressure (HP) regulate chondrocytes’ metabolism; however, how mechanical stimulation acts remain unclear. MicroRNAs (miRNAs) play an important role in cartilage homeostasis, mechanotransduction, and in the pathogenesis of osteoarthritis (OA). This study investigated the effects of a cyclic HP (1–5 MPa), in both normal and OA human chondrocytes, on the expression of miR-27a/b, miR-140, miR-146a/b, and miR-365, and of their target genes (MMP-13, ADAMTS-5, IGFBP-5, and HDAC-4). Furthermore, we assessed the possible involvement of Wnt/β-catenin pathway in response to HP. Chondrocytes were exposed to HP for 3h and the evaluations were performed immediately after pressurization, and following 12, 24, and 48 h. Total RNA was extracted and used for real-time PCR. β-catenin was detected by Western blotting analysis and immunofluorescence. In OA chondrocytes, HP induced a significant increase (p < 0.01) of the expression levels of miR-27a/b, miR-140, and miR-146a, and a significant reduction (p < 0.01) of miR-365 at all analyzed time points. MMP-13, ADAMTS-5, and HDAC-4 were significantly downregulated following HP, while no significant modification was found for IGFBP-5. β-catenin levels were significantly increased (p < 0.001) in OA chondrocytes at basal conditions and significantly reduced (p < 0.01) by HP. Pressurization did not cause any significant modification in normal cells. In conclusion, in OA chondrocytes, HP restores the expression levels of some miRNAs, downregulates MMP-13, ADAMTS-5, and HDAC-4, and modulates the Wnt/β-catenin pathway activation.