Objective To explore the mechanism and inhibition of botulinum toxin type A (BTXA) on hypertrohic scar fibroblasts.Methods The cells were treated by 0 (control),0.2,0.4,0.8 U/ml BTXA for 48 h.Cell viability was detected by MTT assay.Cell apoptosis was detected by Hoechst staining.Cell cycle was detected by flow cytometry.The level of cell cycle related protein D1 (Cyclin D1),proliferation nuclear antigen (PCNA) and activation of phosphatidylinositol 3 kinase (PI3K) / protein kinase B (AKT) signaling pathway were assayed by western blot.Results Compared with control group(0.75±0.07),0.2,0.4,0.8 U/mL BTXA(0.59 ± 0.06,0.43 ± 0.04,0.34± 0.03) inhibited hypertrohic scar fibroblasts cell viability,increased cell apoptotic rate[control group(2.38±0.24)％;BTXA(15.79±1.54)％,(27.32±2.69)％,(38.46±3.90)％],down-regulated the expression of Cyclin D1(control group 1.57±0.18;BTXA 0.93±0.07,0.42±0.04,0.35±0.03) and PCNA(control group 1.46±0.16;BTXA 0.50±0.05,0.59±0.05,0.37±0.03),inhibited the expression of PI3K(control group 0.98±0.06;BTXA 0.49±0.04,0.50±0.04,0.39±0.03) and the phosphorylation of AKT(control group 1.38±0.08;BTXA 0.97±0.06,0.60±0.04,0.29± 0.02),made cell cycle arrested in G1 phase,The difference was statistically significant (P＜0.05).Conclusion These results suggested BTXA inhibit proliferation via blocking the activation of PI3K/AKT signal pathway and down-stream related cell cycle related protein.%目的 探讨A型肉毒毒素(BTXA)对增生性瘢痕成纤维细胞的抑制作用及机制.方法 0(对照组)、0.2、0.4、0.8U/mL的BTXA作用于增生性瘢痕成纤维细胞48 h,噻唑蓝(MTT)法检测细胞活力,Hoechst染色检测细胞凋亡,流式细胞术检测细胞周期,免疫印违法分析细胞周期蛋白D1 (Cyclin D1)、增殖细胞核抗原(PCNA)、磷脂酰肌醇3激酶(PI3K)/蛋白激酶B(AKT)通路的激活状况.结果 与对照组(0,75±0.07)比较,0.2、0.4、0.8 U/mL的BTXA(0.59±0.06、0.43±0.04、0.34±0.03)可明显降低增生性成纤维细胞活力;与对照组[(2.38±0.24)％]比较,可显著提高细胞凋亡率[(15.79±1.54)％、(27.32士2.69)％、(38.46±3.90)％];下调Cyclin D1及PCNA表达,降低PI3K表达及AKT磷酸化水平,并使细胞周期阻滞在G1期,差异均具有统计学意义(P＜0.05).结论BTXA可通过阻断PI3K/AKT信号通路及细胞周期蛋白表达,进而抑制增生性细胞增殖.
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