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纸质出版日期:2018,
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地塞米松对兔胆道成纤维细胞P311/TGF-β1/α-SMA通路表达的影响[J]. 重庆医科大学学报, 2018,43(1):26.
Effect of dexamethasone on P311/TGF-β1/α-SMA signaling pathway in rabbit biliary fibroblasts[J]. Journal of Chongqing Medical University, 2018,43(1):26.
目的:研究地塞米松(dexamethasone,DEX)对兔良性胆道狭窄(benign biliary stricture,BBS)模型胆管成纤维细胞(model fibroblast,MF)中P311/转化生长因子-β1(transforming growth factor-β1,TGF-β1)/α-平滑肌动蛋白(α-smooth muscle actin,α-SMA)通路表达的影响。方法:取2只家兔正常胆管培养正常胆管成纤维细胞(normal bile duct fibroblasts,NF),对2只家兔采用切开再吻合方法建立BBS模型并获取MF,进行细胞鉴定后分组为:NF、MF、MF+不同浓度的DEX(0.02,0.1及0.5 mg/mL)组,各组给予对应浓度的DEX干预48 h,CCK-8细胞计数法测定各组细胞增殖水平;qRT-PCR检测各组细胞P311、TGF-β1及α-SMA 基因mRNA表达水平;Western blot检测各组细胞TGF-β1及α-SMA蛋白表达水平。结果:①各组细胞A值:NF为0.331±0.002,MF组为0.533±0.005,MF+DEX 0.02组为0.487±0.003,MF+DEX 0.1组为0.434±0.004,MF+DEX 0.5组为0.381±0.004。②各组细胞P311 mRNA值:NF组为1.000 0±0.024 8,MF组为4.146 3±0.037 1,MF+DEX 0.02组为3.789 9±0.056 8,MF+DEX 0.1组为3.566 1±0.148 1,MF+DEX 0.5组为2.945 8±0.165 4。③各组细胞TGF-β1 mRNA值NF组为1.000 0±0.034 6,MF组为2.647 9±0.048 5,MF+DEX 0.02组为1.929 7±0.054 8,MF+DEX 0.1组为1.678 2±0.080 2,MF+DEX 0.5组为1.676 2±0.036 9。④各组细胞α-SMA mRNA值:NF组为1.000 0±0.056 0,MF组为4.025 7±0.065 9,MF+DEX 0.02组为3.644 9±0.196 4,MF+DEX 0.1组为2.712 9±0.102 4,MF+DEX 0.5组为2.320 7±0.031 2。⑤各组细胞TGF-β1蛋白值:NF组为0.999 6±0.046 3,MF组为2.096 3±0.029 3,MF+DEX 0.02组为1.781 8±0.040 4,MF+DEX 0.1组为1.531 4±0.032 5,MF+DEX 0.5组为1.384 0±0.035 7。⑥各组细胞α-SMA蛋白值:NF组为1.000 8±0.051 4,MF组为3.231 4±0.116 0,MF+DEX 0.02组为2.875 3±0.078 0,MF+DEX 0.1组为2.262 8±0.059 8,MF+DEX 0.5组为1.940 8±0.093 7。在DEX干预48 h后,MF增殖明显受到抑制,细胞P311、TGF-β1及α-SMA基因mRNA及蛋白表达明显下调(均P<0.05,0.1~0.5 mg/mL),呈现浓度依耐性。结论: DEX抑制BBS形成的作用机制之一可能与它对MF中P311/TGF-β1/α-SMA通路的调控有关。
Objective:To investigate effects of dexamethasone(DEX) on P311/transforming growth factor-β1(TGF-β1)/α-smooth muscle actin(α-SMA) signaling pathway in benign biliary stricture(BBS) model fibroblast(MF). Methods:Two normal rabbits were treated with normal bile duct fibroblasts(NF),BBS model was established via discission and anastomosis of two rabbits and MF was acquired;then they were divided into five groups:NF,MF,MF plus different concentration of DEX(0.02,0.1 and 0.5 mg/mL) groups. After respectively incubated with 48 h,cell proliferation was investigated by CCK-8;Relative mRNA expression of P311,TGF-β1 and α-SMA were assessed by real-time PCR;Relative protein expression of TGF-β1 and α-SMA were investigated by western blot. Results:①optical density(A) in each group:0.331±0.002 in the NF group,0.533±0.005 in the MF group,0.487±0.003 in the MF+DEX 0.02 group,0.434±0.004 in the MF+DEX 0.1 group,0.381±0.004 in the MF+DEX 0.5 group;②P311 mRNA in each group:1.000 0±0.024 8 in the NF group,4.146 3±0.037 1 in the MF group,3.789 9±0.056 8 in the MF+DEX 0.02 group,3.566 1±0.148 1 in the MF+DEX 0.1 group,2.945 8±0.654 0 in the MF+DEX 0.5 group;③TGF-β1 mRNA in each group:1.000 0±0.034 6 in the NF group,2.647 9±0.048 5 in MF group,1.929 7±0.054 8 in the MF+DEX 0.02 group,1.678 2±0.080 2 in MF+DEX 0.1 group and 1.676 2±0.036 9 in the MF+DEX 0.5 group;④cells α-SMA mRNA in each group:1.000 0±0.056 0 in the NF group,4.025 7±0.065 9 in the MF group,3.644 9±0.196 4 in the MF+DEX 0.02 group,2.712 9±0.102 4 in the MF+DEX 0.1 group and 2.320 7±0.031 2 in the MF+DEX 0.5 group;⑤TGF-β1 protein values in each group:0.999 6±0.046 3 in the NF group,2.096 3±0.029 3 in the MF group,1.781 8±0.040 4 in the MF+DEX 0.02 group,1.531 4±0.032 5 in the MF+DEX 0.1 group,and 1.384 0±0.035 7 in the MF+DEX 0.5 group;⑥The number of α-SMA protein in each group:1.000 8±0.051 4 in the NF group,3.2314±0.1160 in the MF group,2.875 3±0.078 0 in the MF+DEX 0.02 group,2.262 8±0.059 8 in the MF+DEX 0.1 group and 1.940 8±0.093 7 in the MF+DEX 0.5 group. After 48 h of DEX intervention,cell proliferation was significantly inhibited and both the mRNA and protein expression of P311,TGF-β1 and α-SMA(all P<0.05,0.1~0.5 mg/mL) were significantly regulated down,with a dose-dependent manner. Conclusion:Mechanism of DEX on anti-BBS may be because it is relevant to modulation of P311/TGF-β1/α-SMA signaling pathway.
良性胆道狭窄成纤维细胞P311/转化生长因子-β1/α-平滑肌动蛋白地塞米松兔
benign biliary stricturefibroblastsP311/transforming growth factor-β1/α-smooth muscle actindexamethasonerabbit
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