癌相关成纤维细胞通过TGF-βRⅡ/smad7调控前列腺癌细胞株PC-3增殖

Cancer Associated Fibroblasts Regulate the Proliferation of Prostate Cancer Cell Line PC-3 by TGF-βRⅡ/Smad7

目的研究癌相关成纤维细胞(cancer associated fibroblasts,CAFs)通过TGF-β/Smads信号通路对前列腺癌细胞株PC3、LNCa P增殖能力的影响及通路关键蛋白TGF-βRⅡ、Smad2、Smad7在其中的作用。方法原代培养源于人前列腺癌基质的CAFs。Western blot检测CAFs、PC3和LNCa P细胞TGF-β受体II型(TGF-βRⅡ)表达。用CAFs原代培养液制备条件培养液CAFs-CM,用TGF-βRⅡ抑制剂(LY2109761)处理CAFs后制备出另一条件培养液CAFs-LY-CM,分别观察PC3和LNCa P在常规培养基、CAFs-CM、CAFs-LY-CM条件培养液中的增殖能力及差异性,以及活性形式磷酸化的Smad2(P-Smad2)、Smad7表达水平的变化。结果 CAFs、PC3细胞阳性表达TGF-βRⅡ,LNCa P细胞呈弱阳性表达。和常规培养基相比,浓度为0.75g/L CAFs-CM的条件培养基可显著提升PC3[分别为(6.36±0.65)和(2.94±0.19),P<0.05]和LNCa P细胞[分别为(5.06±0.38)和(2.86±0.39),P<0.05]的增殖能力。CAFs-CM上调PC3细胞Smad7蛋白表达水平,对LNCa P细胞P-Smad2蛋白表达几无影响。采用10^(-6)mol/L LY2109761处理CAFs后,CAFs-LY-CM可显著抑制PC3细胞增殖[分别为(4.09±0.36)和(6.36±0.65),P<0.05],并呈剂量依赖性,但对LNCa P细胞增殖无显著抑制作用。结论 CAFs通过TGF-βRⅡ/Smad7关键蛋白调控PC3增殖,是PCa潜在的治疗新靶点。CAFs通过TGF-β信号通路调控LNCaP细胞增殖的作用不显著,存在其他的分子机制。

Objective To explore the impact of cancer-associated fibroblasts( CAFs) on the growth of prostate cancer cell PC3 and LNCa P by TGF-beta R II/smads signaling pathway and the role of key protein TGF-βRⅡ,Smad2,Smad7 underling it. Methods CAFs from human prostate cancer tissue was primary cultured,and then treated with TGF-beta receptor type II( TGF-beta RII) antagonist( LY2109761). The conventionally cultured CAFs were used for control. Detection of TGF-beta receptor II expression in CAFs,PC3 and LNCa P cells by Western blotting. The prepared condition mediums were designated as CAFs-LY-CM and CAFs-CM respectively. The difference on proliferation capacity and levels of Phosphorylated Smad2( P-Smad2) 、Smad7 of PC3 and LNCa P cells in this two conditional mediums were investigated. Results CAFs and PC3 cells were positive for TGF-beta R II,while LNCa P cells were weakly positive. 0. 75g/L CAFs can significantly promote the proliferation of PC3 and LNCa P cells. CAFs-CM could up-regulate the expression of Smad7 protein in PC3 cells( 6. 36 ± 0. 65 vs 2. 94 ± 0. 19,P < 0. 05),but had no effect on the expression of P-Smad2 protein in LNCaP cells( 5. 06 ± 0. 38 vs 2. 86 ± 0. 39,P < 0. 05). After 10-6mol/L LY2109761 treatment,CAFs-LY-CM could significantly inhibit the proliferation of PC3( 4. 09 ± 0. 36 vs 6. 36 ± 0. 65,P < 0. 05),which was dose dependent,but not significantly inhibited the proliferation of LNCa P cells. Conclusions CAFs is a potential therapeutic target for PCa through the regulation of PC3 proliferation by TGF-βRII/Smad7. CAFs regulates the proliferation of LNCa P cells by TGF-β/Smads signaling pathway is not significant,there are other molecular mechanisms.