Department of Orthopedic Surgery, The Affiliated Hospital of Hebei University of Engineering, Handan, Hebei 056002, P.R. China.
Department of Dermatology, The Affiliated Hospital of Hebei University of Engineering, Handan, Hebei 056002, P.R. China.
Mol Med Rep. 2018 Sep;18(3):2681-2688. doi: 10.3892/mmr.2018.9229. Epub 2018 Jun 27.
Osteoporosis is closely associated with intervertebral disc degeneration. While parathyroid hormone (PTH) 1‑34, which is an established drug used to treatosteoporosis, is thought to inhibit the disc degeneration associated with osteoporosis, the precise mechanism involved remains unclear. In the present study, primary Sprague‑Dawley rat nucleus pulposus cells (NPCs) were cultured, phenotyped and then treated with dexamethasone (DXM) for 48 h. Cell area analysis and β‑galactosidase staining were used to investigate the effect of DXM on the senescence of NPCs. In addition, the protein levels of LC3‑II, Beclin‑1, P62, p‑mTOR and p‑p70S6k were determined by western blotting and analyzing the regulatory effect of PTH upon autophagy and the mTOR signaling pathway in cells treated with DXM. Following autophagic inhibition induced by ATG5 siRNA transfection, the regulatory effect of PTH on senescence in NPCs were investigated in addition to the potential role of autophagy. As the concentration of DXM increased, the size of the NPCs was significantly enlarged and the proportion of cells with positive β‑galactosidase staining increased significantly (P<0.05). In terms of protein expression, PTH treatment led to an increase in LC3‑II and Beclin‑1 proteins, a reduction in P62 protein, and inhibited p‑mTOR and p‑p70S6k protein expression in DXM‑treated NPCs (P<0.05). PTH attenuated the effect of DXM according to the cell size and percentage of β‑galactosidase‑positive cells. However, the inhibition of autophagy via ATG5 siRNA transfection reversed the protective effect of PTH on cell senescence (P<0.05). Collectively, the present findings suggest that PTH may inhibit the senescence of NPCs induced by DXM by activating autophagy via the mTOR pathway.
骨质疏松症与椎间盘退变密切相关。甲状旁腺激素(PTH)1-34 是一种已被证实用于治疗骨质疏松症的药物,据认为它可以抑制与骨质疏松症相关的椎间盘退变,但具体机制尚不清楚。本研究中,原代 Sprague-Dawley 大鼠髓核细胞(NPC)培养、表型鉴定后,用地塞米松(DXM)处理 48 h。细胞面积分析和β-半乳糖苷酶染色用于研究 DXM 对 NPC 衰老的影响。此外,通过 Western blot 分析和分析 DXM 处理细胞中自噬和 mTOR 信号通路的调节作用,测定 LC3-II、Beclin-1、P62、p-mTOR 和 p-p70S6k 的蛋白水平。用 ATG5 siRNA 转染抑制自噬后,研究了 PTH 对 NPC 衰老的调节作用以及自噬的潜在作用。随着 DXM 浓度的增加,NPC 的大小显著增大,β-半乳糖苷酶染色阳性细胞的比例显著增加(P<0.05)。在蛋白表达方面,PTH 处理导致 LC3-II 和 Beclin-1 蛋白增加,P62 蛋白减少,并抑制 DXM 处理 NPC 中 p-mTOR 和 p-p70S6k 蛋白表达(P<0.05)。PTH 根据细胞大小和β-半乳糖苷酶阳性细胞的百分比减弱了 DXM 的作用。然而,通过 ATG5 siRNA 转染抑制自噬逆转了 PTH 对细胞衰老的保护作用(P<0.05)。综上所述,本研究结果表明,PTH 通过激活 mTOR 通路诱导自噬可能抑制 DXM 诱导的 NPC 衰老。