低强度脉冲超声对人骨性关节炎软骨细胞的影响  

Effect of low-intensity pulsed ultrasound on human osteoarthritis chondrocytes in vitro

作　　者：杜登悝 易刚[1] 唐赢[1] 陈世荣[1] 于廷和[2] 

机构地区：[1]重庆医科大学附属第二医院骨科,重庆400010 [2]重庆医科大学附属第二医院妇产科,重庆400010

出　　处：《第三军医大学学报》2016年第21期2320-2325,共6页Journal of Third Military Medical University

基　　金：重庆市基础科学与前沿技术研究一般项目(CSTC2013jcyA10048)~~

摘　　要：目的探讨低强度脉冲超声(low-intensity pulsed ultrasound,LIPUS)对体外原代培养的人骨性关节炎(osteoarthritis,OA)软骨细胞凋亡和细胞外基质(extracellular matrix,ECM)的影响。方法取人膝关节置换术后废弃软骨分离培养软骨细胞,经甲苯胺蓝染色与Ⅱ型胶原细胞免疫化学染色鉴定,分别用流式细胞仪、q PCR技术及Western blot技术检测不同超声强度(0、30、60、90 m W/cm^2)条件下软骨细胞的凋亡率和Ⅱ型胶原(COL2α1)、蛋白聚糖(aggrecan)、基质金属酶13(matrix metalloproteases 13,MMP13)在mRNA及蛋白水平的表达差异。结果流式细胞学检测显示,30、60、90 m W/cm2组的细胞凋亡率[(7.75±0.79)%、(8.52±0.91)%、(11.38±0.85)%]均较对照组低[(18.13±0.83)%,P<0.05];30、60、90 m W/cm2组细胞的COL2α1和蛋白聚糖在mRNA及蛋白水平均高于对照组(P<0.05),MMP13 mRNA及蛋白表达水平低于对照组(P<0.05),但各处理组之间不呈强度依赖性。结论LIPUS能够抑制体外单层培养的OA软骨细胞的凋亡,促进软骨ECM的合成,抑制软骨ECM的分解,可用于OA的治疗并延缓其发展。Objective To determine the effect of low-intensity pulsed ultrasound（ LIPUS） on the cell apoptosis and extracellular matrix（ ECM） of primarily cultured human osteoarthritis（ OA） chondrocytes in vitro. Methods Human OA chondrocytes were isolated from abandoned articular cartilage tissues and cultured. After the cells were identified by Toluidine Blue staining and immunochemical staining for type Ⅱcollagen,the cells at passage 2 were stimulated by LIPUS at different intensities（ 0,30,60 and 90 m W/cm） 20 min per day for 1 week. Then the apoptotic rate was evaluated by flow cytometry. The mRNA and protein expression levels of collagen type Ⅱ α1（ COL2α1）,aggrecan and matrix metalloproteases 13（ MMP13） were detected by q PCR and Western blotting. Results The apoptotic rates were（ 7. 75 ± 0. 79） %,（ 8. 52 ±0. 91） % and（ 11. 38 ± 0. 85） % respectively in the cells stimulated by 30,60 and 90 m W/cm-2 LIPUS,which were significantly lower than that in the control group [（ 18. 13 ± 0. 83） %,P〈0. 05]. The mRNA and protein expression levels of COL2α1 and aggrecan were significantly higher（ P〈0. 05）,but those of MMP13 were obviously lower（ P〈0. 05） in the cells stimulated with different intensity LIPUS than in the control cells,but no intensity-dependent manner was found in the stimulated groups. Conclusion LIPUS can reduce the rate of apoptosis of human osteoarthritis chondrocytes primarily cultured in vitro. Further LIPUS can promote the synthesis and inhibit the decomposition of ECM. It might be an useful treatment for OA and delay the progression of OA.

关 键 词：低强度脉冲超声 骨性关节炎 软骨细胞 细胞凋亡 细胞外基质 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学] R454.3[医药卫生—基础医学]