虫草多糖对转化生长因子β1诱导人近端肾小管上皮细胞间充质转分化的影响  

Chinese herb Cordyceps polysaccharide inhibits transforming growth factor β1-induced epithelial-to-mesenchymal transition in proximal tubular epithelial cells

作　　者：宫壮[1] 张晓良[1] 高君[1] 李青[1] 刘必成[1] 

机构地区：[1]东南大学附属中大医院肾脏病研究所,南京210009

出　　处：《中华肾脏病杂志》2008年第9期651-656,共6页Chinese Journal of Nephrology

基　　金：国家自然科学基金（30471732）;江苏省自然科学基金（BK2006099）

摘　　要：目的探讨虫草多糖（Cp）对转化生长因子β1（TGF—B1）诱导人近端肾小管上皮细胞间充质转分化的影响。方法用四甲基偶氮唑盐（MTT）比色法检测不同浓度的Cp（0、0．01、0．1、1、5、10g／L）对人近端肾小管上皮细胞（HK-2）增殖的作用。用不同浓度的TGF-B1（0、0．1、0．5、1、5、10μg/L）作用HK-2细胞48h及5μg／L的TGF—β1作用不同时间（0、12、24、48和72h），以实时定量PCR和Western印迹方法检测仅平滑肌肌动蛋白（α-SMA）、钙黏蛋白（E-cadherin）、纤连蛋白（FN）的表达。用递增浓度的Cp（1、5、10g／L）预处理HK-2细胞24h后，观察其对TGF-β1效应的干预作用。结果Cp以剂量依赖方式促进HK-2细胞增殖（P〈0．05）。TGF—β1无论在转录水平还是蛋白水平皆能诱导HK-2细胞α-SMA、FN表达上调，而下调E—cadherin表达。分别用1、5、10g／LCp预干预24h后，同单独5μg／LTGF—β1刺激组相比，上述因子表达被不同程度逆转。在转录水平，α-SMA mRNA表达被抑制率分别为37．98％、68．08％、84．36％；FN mRNA表达被抑制率分别为46．97％、63．82％、81．85％；E—cadherin mRNA表达分别增加0．67倍、2．69倍、5．43倍（均P〈0．05）。在蛋白水平，α-SMA蛋白表达被抑制率分别为33．40％、47．75％、68．50％；FN蛋白表达抑制率分别为16．26％、65．92％、80．30％；E—cadherin蛋白表达分别增加1．33倍、3．19倍、4．29倍（均P〈0．05）。Cp能明显拮抗TGF—β1诱导的HK-2细胞的表型改变。结论Cp能有效阻止TGF-β1诱导的人近端。肾小管上皮细胞发生转分化。Objective To investigate the influence of Cordyceps polysaccharide （Cp） on epithelial-to-mesenchymal transition （EMT） induced by transforming growth factor-β1 （TGF-β1）in proximal tubular epithelial cells （PTEC）. Methods HK-2 cell proliferation was determined by MTT assay. After incubation of HK2 cells with increasing concentrations of TGF-β1 （0, 0.1, 0.5, 1, 5, 10μg/L） at 48 h and with TGF-β1 （5μg/L） at different time points, E-cadherin, α-SMA, FN expression at transcriptional and protein levels were detected by real-time PCR and Western blotting respectively. The cells were pretreated with 1, 5, 10 g/L Cp respectively for 24 h before adding TGF-β1 （5 μg/L）, then the cells were incubated for additional 48 h, mRNA and protein expression of above 3 cytokines was examined by real-time PCR and Western blotting as well. Results CP alone （0.01, 0.1, 1, 5, 10 g/L） induced HK-2 cell proliferation in a dosedependent manner. TGF-β1 enhanced α-SMA, FN expression while inhibited E-cadherin expression at both transcriptional and protein level in HK-2 cell. At transcriptional level, compared to single TGF-β1 （5 μg/L） stimulating group, after Cp （1, 5, 10 g/L） pretreatment for 24 h, the inhibition rate of α-SMA mRNA was 37.98%, 68.08% and 84.36%, respectively; FN mRNA was 46.97%, 63.82% and 81.85%, respectively; E-cadherin was up-regulated by 0.67 fold, 2.69 folds and 5.43 folds, respectively （P〈0.05）. At protein level, the inhibition rate of α-SMA was 33.40%, 47.75% and 68.50%, respectively; FN was 16.26%, 65.92% and 80.30%, respectively; E-cadherin was up-regulated by 1.33 folds, 3.19 folds and 4.29 folds, respectively （all P〈0.05）. Under Light microscopy, the Cp reversed cell shape from spindle-shape induced by TGF-β1 to nearly normal shape. Conclusion Cp may exert its inhibitive effects on TGF-β1-induced EMT.

关 键 词：纤维化 转化生长因子Β 上皮细胞 肾小管 近端 虫草多糖 HK-2细胞 上皮细胞间充质转分化 

分 类 号：R285.5[医药卫生—中药学]