单个核细胞趋化蛋白-1和马兜铃酸Ⅰ在诱导人类肾小管上皮细胞转分化中的协同作用

Synergistic effect of monocyte chemotactic protein-1 and aristolochic acid Ⅰ on transdifferentiation of human tubular epithelial cells in vitro

目的 探讨单个核细胞趋化蛋白 1(MCP 1)和马兜铃酸Ⅰ (aristolochicacidⅠ ,AAⅠ )及其两者的协同作用 ,对人类肾小管上皮细胞 (HKC)转分化的影响。方法 应用间接免疫荧光法测定HKC细胞波形蛋白 (vimentin)、α 平滑肌肌动蛋白 (α smoothmuscleactin ,α SMA)及角蛋白 (cytocreatin)的表达 ;应用流式细胞技术测定表达α SMA( +) HKC细胞的百分数。结果 HKC细胞经MCP 1、AAⅠ或AAⅠ与MCP 1共同作用后 ,HKC细胞角蛋白表达减弱 ,波形蛋白、α SMA表达明显增强。无血清培养的HKC细胞中有少量细胞 (0 3 %～ 7 5 % )表达α SMA ,当培养液中的MCP 1浓度为 0 0 0 1、0 0 1、0 0 5、0 1、0 2、 0 4μg/L时 ,HKC细胞表达α SMA( +) 的百分率为 2 0 0 %、2 6 2 %、2 0 3%、2 3 2 %、11 2 %和 0 4% ,与无血清对照组 (平均为 3 1% )比较差异有显著性 (P <0 0 5 ) ;当加入AAⅠ的浓度为 5、10 μg/L时 ,HKC细胞表达α SMA( +) 的百分率分别为 0 5 %和 1 4% ,与无血清对照组比较差异无显著性 (P >0 0 5 ) ,当AAⅠ为 2 0、40和 80 μg/L时 ,HKC细胞表达α SMA( +) 的百分率分别为 8 6 %、9 6 %和 13 4% ,与无血清对照组比较差异有显著性 (P <0 0 5 )。当培养液中同时加入MCP 1(0 1μg/L)和AAⅠ (5、

Objective To test the possible role of monocyte chemotactic protein-1 (MCP-1) and its synergistic effect with aristolochic acid I (AAⅠ) on tubular epithelial-myofibroblast transdifferentiation (TEMT) of human renal tubular epithelial cells (HKC) in vitro. Methods The cultured HKC cells were divided into four groups: (1) negative control (serum-free); (2) MCP-1 group; (3) AAⅠ group; (4) AAⅠ+MCP-1 group. The expression of α-smooth muscle actin (α-SMA), vimentin and cytocreatin was assessed by indirect enzyme immunohistochemiry and the percentages of α-SMA (+) HKC cells were assessed by flow cytometry. Results The expression of cytocreatin of HKC cells decreased, while the expression of α-SMA, vimentin increased when treated with MCP-1 or with AAⅠ and MCP-I concomitantly. α-SMA (+) HKC cells cultured in serum-free medium was 3 1% by flow cytometry. The percentages of α-SMA (+) HKC cells were 8 6%, 9 6%,13 4% ( P <0 05 vs control) when treated with 20, 40 ,80 μg/L of AAⅠ. The percentages of α-SMA (+) HKC cells were 0 5% and 1 4% ( P >0 05 vs control) when treated with 5,10 μg/L. The percentages of α-SMA (+) HKC cells were 20%, 26 2%, 20 3%, 23 2% ( P <0 05 vs control) when treated with 0 001、0 01, 0 05, 0 1 μg/L of MCP-1. When the HKC cells were treated with MCP-1(0 1 μg/L) and AAⅠ(5, 10, 20, 40 μg/L), the percentage of α-SMA (+) cells increased markedly to 23 2%, 98 7%, 81 5%, 65 1% ( P <0 01 vs group 1, 2, 3, respectively). Conclusions These findings suggest that : (1) MCP-1 may induce the transdifferentiation of HKC cells into myofibroblasts in vitro; (2) AAⅠ at some doses may partialy induce HKC cells transdifferentiaing. (3) MCP-1 and AAⅠ may have a synergistic effect on transdifferentiation of HKC cells in vitro.