PODXL参与瘢痕疙瘩成纤维细胞的过度增殖

PODXL participates in the excessive proliferation of keloid fibroblasts

目的探讨podocalyxin样蛋白(PODXL)基因对瘢痕疙瘩成纤维细胞增殖、凋亡的影响。方法RT-qPCR检测正常皮肤成纤维细胞(hDF)和瘢痕疙瘩成纤维细胞(hKF)中PODXL mRNA的表达量;用重组PODXL基因的shRNA慢病毒表达载体感染瘢痕疙瘩成纤维细胞下调PODXL基因的表达量;EdU法检测细胞增殖;流式细胞仪检测细胞的周期和凋亡率;蛋白质印迹(Western blot)检测细胞中磷脂酰肌醇-3激酶(PI3K)、丝苏氨酸蛋白激酶(AKT)总蛋白和磷酸化蛋白。结果与正常皮肤成纤维细胞相比,PODXL mRNA在瘢痕疙瘩成纤维细胞中的表达量显著增加(P<0.05);下调PODXL基因表达显著抑制瘢痕疙瘩成纤维细胞增殖(P<0.05),阻滞细胞周期于S期,促进细胞的凋亡(P<0.05),降低PI3K和AKT磷酸化蛋白水平(P<0.05)。结论PODXL在瘢痕疙瘩成纤维细胞中的表达量显著增加;下调PODXL基因表达可能通过抑制PI3K/AKT信号通路激活,从而将细胞周期阻滞在S期,抑制细胞的增殖和诱导凋亡。

Objective To investigate the effect of podocalyxin-like protein(PODXL)on the proliferation and apoptosis of keloid fibroblasts.Methods RT-qPCR was used to detect the expression of PODXL mRNA in human dermal fibroblasts and keloid fibroblasts.The expression of PODXL gene in keloid fibroblasts was inhibited by recombinant PODXL gene shRNA lentiviral expression vector infection.The proliferation of cells was measured by EdU assay.The cell cycle and apoptosis rate were analyzed by flow cytometry.The level of phosphatidylinositol 3-kinase(PI3K),tyrosine kinase(AKT)total protein and phosphorylated protein in cells were detected by Western blot.Results Compared with normal skin fibroblasts,the expression of PODXL mRNA in keloid fibroblasts was significantly increased(P<0.05).Down-regulation of PODXL gene expression significantly inhibited the proliferation of keloid fibroblasts(P<0.05),arrested the cell cycle in S phase and promoted apoptosis(P<0.05),decreased PI3K,AKT phosphorylation protein levels(P<0.05).Conclusions The expression of PODXL in keloid fibroblasts is significantly increased.Down-regulation of PODXL gene expression may arrest cell cycle in S phase,inhibit proliferation and induce apoptosis by inhibiting the activation of PI3K/AKT signaling pathway.