Department of Orthopedics, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
Department of Orthopedics, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
Biochem Biophys Res Commun. 2018 Nov 17;506(1):41-47. doi: 10.1016/j.bbrc.2018.10.043. Epub 2018 Oct 15.
Glucocorticoids (GCs)-induced osteoblast apoptosis has been identified as an important cause of GCs related osteonecrosis of the femoral head (ONFH). Glycogen synthase kinase 3β (GSK3β) has been proved to mediate dexamethasone (Dex)-induced osteoblast apoptosis. This study aimed to investigate the underlying mechanism of GSK3β in Dex-induced osteoblast apoptosis.
Osteoblast cells were transfected with lentivirus expressing GSK3β-shRNA, and a DNA microarray was performed to analyze gene expression after Dex treatment with or without GSK3β-shRNA. Some differentially expressed genes were further validated by quantitative real-time-PCR (qRT-PCR).
460 genes were up-regulated (at least 2-fold) with Dex treatment but down-regulated (at least 2-fold) with GSK3β-shRNA treatment. In addition, 315 genes were down-regulated (at least 2-fold) with Dex treatment but up-regulated (at least 2-fold) with GSK3β-shRNA treatment. Among these genes, the apoptosis-related genes Hoxb8, Kif18a, Dock8, Dlk1, Tnfsf14, Casq2, Bcl2l14 and mechanosensation-related gene Piezo2 were selected for further qRT-PCR analysis. 7 of 8 genes (Piezo2, Hoxb8, Kif18a, Dlk1, Tnfsf14, Casq2, Bcl2l14) showed the same tendency between gene chip results and qRT-PCR results. The microarray data also showed that apoptotic pathway, MAPK pathway, TGFβ pathway and Wnt pathway might be related to the mechanism of GSK3β in Dex-induced osteoblast apoptosis.
Our findings indicate that GSK3β-shRNA treatment can alter various genes expression levels and change diverse signaling pathways involved in Dex-induced osteoblast apoptosis. Furthermore, Piezo2, Hoxb8, Kif18a, Dlk1, Tnfsf14, Casq2 and Bcl2l14 genes may play an important role in the GSK3β-mediated osteoblast apoptosis process.
糖皮质激素（GCs）诱导的成骨细胞凋亡已被确定为 GCs 相关股骨头坏死（ONFH）的一个重要原因。糖原合酶激酶 3β（GSK3β）已被证明介导地塞米松（Dex）诱导的成骨细胞凋亡。本研究旨在探讨 GSK3β在 Dex 诱导的成骨细胞凋亡中的作用机制。
用表达 GSK3β-shRNA 的慢病毒转染成骨细胞，并用 DNA 微阵列分析 Dex 处理后有无 GSK3β-shRNA 表达时的基因表达。通过定量实时-PCR（qRT-PCR）进一步验证一些差异表达基因。
Dex 处理可使 460 个基因（至少 2 倍上调）上调，但 GSK3β-shRNA 处理可使这些基因（至少 2 倍下调）下调。此外，Dex 处理可使 315 个基因（至少 2 倍下调）下调，但 GSK3β-shRNA 处理可使这些基因（至少 2 倍上调）上调。在这些基因中，选择凋亡相关基因 Hoxb8、Kif18a、Dock8、Dlk1、Tnfsf14、Casq2、Bcl2l14 和机械感受相关基因 Piezo2 进行进一步的 qRT-PCR 分析。在基因芯片结果和 qRT-PCR 结果之间，8 个基因中有 7 个（Piezo2、Hoxb8、Kif18a、Dlk1、Tnfsf14、Casq2、Bcl2l14）表现出相同的趋势。微阵列数据还显示，凋亡途径、MAPK 途径、TGFβ 途径和 Wnt 途径可能与 GSK3β 在 Dex 诱导的成骨细胞凋亡中的作用机制有关。
我们的研究结果表明，GSK3β-shRNA 处理可改变各种基因的表达水平，并改变 Dex 诱导的成骨细胞凋亡中涉及的各种信号通路。此外，Piezo2、Hoxb8、Kif18a、Dlk1、Tnfsf14、Casq2 和 Bcl2l14 基因可能在 GSK3β 介导的成骨细胞凋亡过程中发挥重要作用。
