Tsuchida Shinji, Arai Yuji, Takahashi Kenji A, Kishida Tsunao, Terauchi Ryu, Honjo Kuniaki, Nakagawa Shuji, Inoue Hiroaki, Ikoma Kazuya, Ueshima Keiichiro, Matsuki Tomohiro, Mazda Osam, Kubo Toshikazu
Department of Orthopaedics, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, Japan.
J Orthop Res. 2014 Aug;32(8):975-80. doi: 10.1002/jor.22623. Epub 2014 Mar 28.
We assessed whether heat shock protein 70 (HSP70) is involved in hypoxia inducible factor 1 alpha (HIF-1α)-dependent anabolic pathways in articular chondrocytes under hypoxic conditions. Primary rabbit chondrocytes were cultured under normoxia (20% oxygen condition) or hypoxia (1% oxygen condition). Alternatively, cells cultured under normoxia were treated with CoCl2 , which induces HIF-1α, to simulate hypoxia, or transfected with siRNAs targeting HIF-1α (si-HIF-1α) and HSP70 (si-HSP70) under hypoxia. HSP70 expression was enhanced by the increased expression of HIF-1α under hypoxia or simulated hypoxia, but not in the presence of si-HIF-1α. Hypoxia-induced overexpression of ECM genes was significantly suppressed by si-HIF-1α or si-HSP70. Cell viability positively correlated with hypoxia, but transfection with si-HIF-1α or si-HSP70 abrogated the chondroprotective effects of hypoxia. Although LDH release from sodium nitroprusside-treated cells and the proportion of TUNEL positive cells were decreased under hypoxia, transfection with si-HIF-1α or si-HSP70 almost completely blocked these effects. These findings indicated that HIF-1α-induced HSP70 overexpression increased the expression levels of ECM genes and cell viability, and protected chondrocytes from apoptosis. HIF-1α may regulate the anabolic effects of chondrocytes under hypoxic conditions by regulating HSP70 expression.
我们评估了热休克蛋白70（HSP70）是否参与缺氧条件下关节软骨细胞中缺氧诱导因子1α（HIF-1α）依赖性合成代谢途径。原代兔软骨细胞在常氧（20%氧气条件）或缺氧（1%氧气条件）下培养。另外，将常氧培养的细胞用诱导HIF-1α的氯化钴处理以模拟缺氧，或在缺氧条件下用靶向HIF-1α（si-HIF-1α）和HSP70（si-HSP70）的小干扰RNA转染。缺氧或模拟缺氧条件下，HIF-1α表达增加可增强HSP70表达，但在存在si-HIF-1α时则不然。si-HIF-1α或si-HSP70可显著抑制缺氧诱导的细胞外基质（ECM）基因过表达。细胞活力与缺氧呈正相关，但用si-HIF-1α或si-HSP70转染可消除缺氧的软骨保护作用。尽管缺氧条件下硝普钠处理细胞的乳酸脱氢酶（LDH）释放及TUNEL阳性细胞比例降低，但用si-HIF-1α或si-HSP70转染几乎完全阻断了这些作用。这些发现表明，HIF-1α诱导的HSP70过表达增加了ECM基因表达水平和细胞活力，并保护软骨细胞免于凋亡。HIF-1α可能通过调节HSP70表达来调节缺氧条件下软骨细胞的合成代谢作用。
