白藜芦醇激活PI3K/Akt信号通路对软骨细胞细胞外基质合成的影响

Influence of resveratrol on extracellular matrix synthesis of chondrocytes via activating PI3K/Akt pathway

[目的]通过白藜芦醇刺激体外培养的小鼠膝关节软骨细胞,探讨白藜芦醇通过PI3K/Akt信号通路对软骨细胞细胞外基质合成的影响。[方法]体外分离培养小鼠膝关节软骨细胞,采用HE染色、甲苯胺蓝染色和Ⅱ型胶原免疫荧光染色观察鉴定软骨细胞。根据作用药物的不同分为3组:空白对照组、白藜芦醇组(100μmol/L白藜芦醇)和LY294002组(100μmol/L白藜芦醇+25μmol/L LY294002)。采用RT-PCR检测软骨细胞中Akt1,Aggrecan、Collagen II的m RNA表达水平。[结果]荧光显微镜下可见软骨细胞胞核呈蓝色荧光,胞质呈红色荧光。显示Ⅱ型胶原染色阳性细胞数占90%以上,证实所培养细胞为软骨细胞。RT-PCR检测结果显示白藜芦醇组Akt1、Aggrecan、Collagen II m RNA表达量较空白组明显升高(P<0.05)。LY294002组Akt1、Aggrecan、Collagen II m RNA表达量较白藜芦醇组下降,但仍高于空白组(P<0.05)。[结论]白藜芦醇可以通过激活PI3K/Akt信号通路,增加软骨细胞细胞外基质合成,起到保护关节软骨的作用。

Objective] rTO investigate the effect of resveratrel on extracellular matrix synthesis of chondrocytes via PI3K/ Akt pathway. [Methods] Mouse knee chondrocytes, already isolated and cultured in vitro, were observed and identified by HE staining, toluidine blue staining and collagen type II immunofluorescence staining. The chondrocyte cultures was divided into the blank control group, the resveratrol intervention group （100μM resveratrol） and LY294002 intervention group （100μM resveratrol＋25μM LY294002） . RT PCR was used to detect the expression level of Aktt, aggrecan, collagen II mRNA in chondrocytes. [Results] The cell nucleus was blue, meanwhile, the cytoplasmic red under fluorescent microscope. Fluorescence microscopy showed that the number of positive cells of type II collagen accounted for more than 90%, confirming that the cultured cells were chondrocytes. Compared with the blank control group, the expression level of Aktl, aggrecan, collagen II mRNA in the resveratrol group was significantly higher, with a statistical difference （P〈0.05） . In addition, the expression of Aktl, aggrecan, collagen II mRNA in the LY294002 group were lower than the resveratrol group, nevertheless those still higher than the blank control group, with significant differences as well （P 〈0.05 ） . [Conclusion] Resveratrol can increase the cartilage extracellular matrix synthesis via activating the PI3K/Akt signaling pathway, which might protect the artic- ular cartilage.