针刀干预对膝骨关节炎兔原代软骨细胞活性、凋亡及自噬的影响

Effects of acupotomy on activity,apoptosis,and autophagy of primary chondrocytes in knee osteoarthritis rabbits

目的观察针刀干预对膝骨关节炎(KOA)兔软骨细胞活性、凋亡及自噬的影响。方法选取普通健康6月龄雄性新西兰兔24只,体重(2.5±0.5)kg,采用随机数字表法将其分为空白组、模型组,针刀组,每组8只。模型组、针刀组使用改良Videman法制备KOA兔模型,固定6周,空白组不予处理。通过X线检查进行模型评价。针刀组选取常见经筋病灶点行针刀松解,每周1次,共治疗4次。空白组和模型组不予处理。干预结束后1周,提取兔左膝关节股骨内外侧髁软骨面原代软骨细胞,采用CCK-8检测细胞活性;Annexin V-FITC/PI双染流式检测细胞凋亡率;LC3B免疫荧光染色检测细胞自噬水平。结果三组组内各时间点细胞活性比较,差异均有统计学意义(P<0.05),模型组同期细胞活性低于空白组,针刀组同期细胞活性高于模型组,差异有统计学意义(P<0.05)。模型组软骨细胞凋亡率高于空白组(P<0.05),针刀组软骨细胞凋亡率较于模型组(P<0.05)。与空白组比较,模型组LC3B荧光强度下降;与模型组比较,针刀组LC3B荧光强度增强。结论基于经筋理论针刀循膝周常见经筋病灶点松解能增强KOA兔原代软骨细胞自噬水平,提高细胞活性和减少细胞凋亡。

Objective To observe the effects of acupotomy on chondrocyte activity,apoptosis,and autophagy in rabbits with knee osteoarthritis(KOA).Methods Twenty-four healthy six-month-old male New Zealand rabbits with body weight of(2.5±0.5)kg were divided into blank group,model group,and acupotomy group by random number table method,with eight rabbits in each group.Model group and acupotomy group were used to prepare KOA rabbit model by modified Videman method,and fixed for six weeks.Blank group was not processed.The model was evaluated by X-ray examination.In acupotomy group,common tendons were selected for acupotomy release,once a week,a total of four times.The blank and model groups were not processed.One week after the intervention,primary chondrocytes were extracted from the cartilage surface of the femoral medial and lateral condyles of the rabbit left knee joint,and the cell activity was detected by CCK-8.Apoptosis rate was detected by Annexin V-FITC/PI double staining flow cytometry.LC3B immunofluorescence staining was used to detect autophagy.Results The comparison of cell activity at each time point in three groups were statistically significant(P<0.05).The cell viability in the model group was lower than that in the blank group,and the cell viability in the acupotomy group was higher than that in the model group at the same time(P<0.05).The chondrocyte apoptosis rate in the model group was higher than that in the blank group(P<0.05),and the chondrocyte apoptosis rate in the acupotomy group was lower than that in the model group(P<0.05).Compared with the blank group,the fluorescence intensity of LC3B in the model group decreased;compared with the model group,the fluorescence intensity of LC3B in the needle knife group was enhanced.Conclusion Based on the balance theory of muscles and bones through the tendon system,acupotomy can enhance the autophagy level of primary chondrocytes in KOA rabbits,improve cell activity and reduce cell apoptosis.