放散式体外冲击波对软骨细胞生物学行为的影响

Effect of radial extracorporeal shock wave on the biological behaviors of chondrocytes

目的放散式体外冲击波(radial extracorporeal shock wave,rESW)能够促进软骨缺损和骨关节炎(osteoarthritis,OA)软骨损伤的修复,但具体机制仍需进一步探讨。本研究以软骨细胞为研究对象,分析rESW对软骨细胞生物学行为的影响及相关机制,为rESW的临床应用奠定研究基础。方法采用胶原酶联合胰蛋白酶消化的方法获取大鼠软骨细胞,通过甲苯胺蓝染色对软骨细胞进行鉴定。分别在正常培养条件以及IL-1β介导的炎性条件下,给予不同能量的rESW刺激,采用CCK8法和划痕法分别检测软骨细胞的增殖和迁移。通过对软骨细胞培养上清中一氧化氮(nitric oxide,NO)的Griess法测定,检测rESW对软骨细胞NO分泌的影响。Western blot分析软骨细胞诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)的蛋白表达。结果正常环境下,rESW对软骨细胞的增殖、迁移和NO分泌均未有显著影响。IL-1β存在的炎性环境下,软骨细胞增殖和迁移受到显著抑制,rESW应用后,软骨细胞增殖能力有明显恢复,但迁移未受影响。炎性环境下,软骨细胞NO生成显著升高,且iNOS蛋白表达上调,rESW抑制NO的分泌和iNOS蛋白的表达。结论炎性环境下,rESW能够保护软骨细胞的增殖功能,且抑制软骨细胞分泌前炎症因子NO,改善关节腔微环境,因此rESW能够作为一种辅助手段应用于关节软骨损伤的治疗。

Objective Radial extracorporeal shock wave(rESW)has curative effect on osteochondral lesions and osteoarthritis(OA).In order to find out the mechanisms of this effect,we apply rESW on chondrocytes directly to observe the changes of their biological behaviors under normal and inflammatory conditions.Methods Chondrocytes from cartilage of SD rats were achieved by enzymatic method.The cells were treated with interleukin 1 beta(IL-1β)as an in vitro inflammatory model.Three different energy levels of rESW were tested for the effects on the proliferation and migration of chondrocytes,with CCK8 method and cell scratch-wound assay,respectively.Nitric oxide(NO)secretion was detected through Griess method.Western blot was used to determine the protein expression of inducible nitric oxide synthase(iNOS).Results The proliferation,migration and NO secretion were not changed under normal condition by rESW.When IL-1βwas added,the proliferation of chondrocytes decreased,but recovered significantly after cells were treated with rESW;while the migration was not significantly affected by rESW.RESW significantly downregulated the NO level and iNOS protein expression in the presence of IL-1β.Conclusions RESW helps to protect the biological behaviors of chondrocytes under inflammatory condition,and may be used as a supplementary approach clinically in OA treatment.