Akt与ERK1/2在人骨关节炎软骨细胞中的表达

目的: 观察蛋白激酶B(Akt)与细胞外信号调节激酶1/2(ERK1/2)在正常和骨关节炎(OA)软骨细胞中的表达,探讨Akt与ERK1/2在OA病程中的意义。方法: 手术中取5例正常和18例OA人膝关节软骨组织,包埋制备切片,免疫组织化学技术观察 p-Akt及p-ERK1/2在正常和OA 软骨组织中的表达;培养人软骨细胞,甲苯胺蓝染色、免疫组化鉴定并观察聚集蛋白聚糖及Ⅱ型胶原在正常和OA软骨细胞中的表达; Western blotting技术检测Akt、p-Akt、ERK1/2、p-ERK1/2、磷酸化70 kD核糖体蛋白S6激酶(p-p70S6K)及增殖细胞核抗原(PCNA)蛋白在正常和OA软骨细胞中表达水平; 实时荧光定量PCR技术检测聚集蛋白聚糖及Ⅱ型胶原在正常和OA软骨细胞中mRNA表达水平。结果: 与正常软骨细胞比较,OA软骨细胞内p-Akt和p-p70S6K蛋白表达明显降低(P<0.05),且聚集蛋白聚糖和Ⅱ型胶原mRNA和蛋白在OA软骨细胞中的表达水平降低(P<0.05),而 p-ERK1/2和PCNA蛋白表达明显提高(P<0.05)。结论: Akt可能通过p-p70S6K来调控OA软骨细胞外基质聚集蛋白聚糖及II型胶原的合成,ERK1/2可能通过PCNA来调控OA软骨细胞增殖;Akt与ERK1/2可能参与了OA的病理过程。

Expression of Akt and ERK1/2 in human osteoarthritic chondrocytes

AIM: To investigate the expression of protein kinase B(Akt) and extracellular signal-regulated kinase 1/2(ERK1/2) in normal and osteoarthritic chondrocytes.METHODS: The samples of knee cartilage were obtained from the normal donors(n=5) and the patients(n=18) undergoing total knee arthroplasty with the diagnosis of osteoarthritis(OA).The expression of p-Akt and p-ERK1/2 in the normal and osteoarthritic cartilage tissues was detected by the method of immunohistochemistry.The chondrocytes were isolated and identified by toluidine blue staining and immunohistochemical method.The expression levels of Akt,p-Akt,ERK1/2,p-ERK1/2,phosphorylated 70-kD ribosomal protein S6 kinase(p-p70S6K) and proliferating cell nuclear antigen(PCNA) were tested in normal and osteoarthritic chondrocytes by Western blotting.Real-time fluorescence quantitative PCR was used to measured the expression levels of aggrecan and type II collagen gene in normal and osteoarthritic chondrocytes.RESULTS: The expression of p-Akt in normal cartilage was higher than that in OA cartilage.The expression of p-ERK1/2 in OA cartilage was higher than that in normal cartilage.Compared with the normal chondrocytes,the expression of p-Akt and p-p70S6K,and the mRNA levels of aggrecan and type II collagen were increased(P<0.05),and the expression of p-ERK1/2 and PCNA was decreased in OA chondrocytes(P<0.05).CONCLUSION: Akt might regulate aggrecan and type II collagen synthesis via p-p70S6K,and ERK1/2 might regulate OA chondrocyte proliferation through PCNA.Both Akt and ERK1/2 play important roles in the pathogenesis of OA.