压应力对破骨细胞活化的影响

目的建立3D水凝胶细胞模型,同时采用不同强度、频率、时间压应力作用于破骨细胞,观察压应力对破骨细胞分化的影响,探讨抑制破骨细胞分化的适宜压应力方案。方法 M-CSF和RANKL诱导骨髓单核细胞成破骨细胞,3D细胞水凝胶种板后,次日进行压应力干预,设计不同强度、频率压应力梯度方案对细胞水凝胶进行干预,对照组细胞不进行干预。分组如下:G0(对照组)、G1(1%,0.5 Hz,4 h)、G2(2%,0.5 Hz,4 h)、G3(3%,0.5 Hz,4 h)、G4(1%,1.0 Hz,4 h)、G5(2%,1.0 Hz,4 h)、G6(3%,1.0 Hz,4 h)。在此基础上,通过统计学计算出有效的强度和频率压应力方案后,按照不同干预时间施加压应力,分组如下:D1(4 h)、D2(8 h)、D3(12 h)、D4(16 h),每组两个孔。加压干预结束后即刻收集细胞,将RNA反转录为c DNA,再对样本中的Ctsk mRNA、NFATc1 mRNA、TRACP mRNA、M-CSF mRNA和RANK mRNA进行定量检测。结果RANK表达水平显著依赖于压应力的强度和频率(P0.01);TRACP的表达水平显著依赖于压应力的强度(P0.01),且强度和频率两者对TRACP的表达量出现交互作用(P0.01);Ctsk的表达量水平显著依赖于压应力的强度(P0.05)和频率(P0.01),且两者之间出现交互作用(P0.01)。加压8 h M-CSF的表达量比加压12 h(P0.01)和16 h(P0.05)的低。加压8 h RANK的表达量比加压12 h(P0.05)和16 h(P0.01)的低。加压16 h Ctsk和NFATc1的表达量比加压4 h和8 h的高(P0.05)。结论在3D水凝胶模型中,以1%强度、0.5 Hz频率、8 h的压应力干预方案能够抑制破骨细胞的分化。研究结果为通过适宜的运动预防骨质疏松、提高峰值骨量奠定理论基础。

Effects of Compressive Stress on Osteoclast Activation

Objective To establish the 3D hydrogel cell model and apply compressive stress with different intensities, frequencies and durations on osteoclasts, so as to observe the effect of compressive stress on osteoclast differentiation and investigate the appropriate compressive stress solution for inhibiting osteoclast differentiation. Methods M-CSF and RANKL were used to induce bone marrow mononuclear cells into osteoclasts. After the 3D cell-agarose mixture was seeded in compression culture plate, compressive stress was applied on osteoclasts with different intensities, frequencies and durations the next day. The cells in control group were not interfered. The cells were divided as following: G0 (control group), G1 (1%, 0.5 Hz, 4 h), G2 (2%, 0.5 Hz, 4 h), G3 (3%, 0.5 Hz, 4 h), G4 (1%, 1.0 Hz, 4 h), G5 (2%, 1.0 Hz, 4 h), G6 (3%, 1.0 Hz, 4 h). After the loading plan with the most effective intensity and frequency was calculated by statistical analysis, compressive stresses were applied on cells with different durations as following: D1(4 h), D2(8 h), D3(12 h), D4(16 h), and each group had two samples. Once compressive loading was finished, the total RNA extraction from cell-gel constructs were performed and Ctsk mRNA, NFATc1 mRNA, TRACP mRNA, M-CSF mRNA and RANK mRNA were measured by quantitative testing. Results RANK and TRACP mRNA expression significantly depended on intensities and frequencies of the compressive stress (P<0.01), and Ctsk mRNA significantly depended on intensities(P<0.01) while it differed notably with different frequencies (P<0.01). M-CSF mRNA expression with 8 h was much lower than that with 12 h (P<0.01) and 16 h (P<0.05). RANK mRNA expression with 8 h was lower than that with 12 h (P<0.05) and 16 h (P<0.01). In addition, Ctsk and NFATc1 mRNA expression with 16 h was higher than that with 4 h and 8 h (P<0.05). Conclusions In the 3D hydrogel model, 1% intensity, frequency of 0.5 Hz, cyclic compression intervention with 8 h can suppress the differentiation of osteoclasts. The research findings provide the theoretical basis for preventing osteoporosis and improving the peak bone mass by appropriate exercise.