论文部分内容阅读
目的:探讨γ-分泌酶抑制剂(DAPT)抑制Notch信号通路对心肌成纤维细胞(CFs)平滑肌肌动蛋白-α(α-SMA)的表达及细胞外基质(ECM)降解平衡变化的影响。方法:采用胰酶消化法和差速贴壁法分离和纯化SD乳鼠CFs进行体外培养,将CFs分为正常组(N组,培养时不加干预药物)和尾加压素Ⅱ组(U组,加UⅡ10~(-8) mol/L)和DAPT+尾加压素Ⅱ组(D组,同时加入UⅡ10~(-8) mol/L和DAPT 75μmol/L),培养干预48h后采用RT-PCR法测基质金属蛋白酶-1(MMP-1)、组织金属蛋白酶抑制剂-1(TIMP-1)、α-SMA和Notch蛋白胞内段(Notch1)mRNA的表达;Western Blot检测α-SMA及Notch1蛋白的表达。结果:U组TIMP-1、α-SMA及Notch1的mRNA表达水平明显高于N组(均P<0.01),MMP-1mRNA的表达和MMP-1mRNA/TIMP-1mRNA比值低于N组(均P<0.01);D组TIMP-1、α-SMA、Notch1的mRNA表达以及α-SMA和Notch1蛋白的表达均较U组下调(均P<0.01),MMP-1mRNA的表达和MMP-1mRNA/TIMP-1mRNA比值较U组上调(P<0.01);但D组上述指标与N组相比差异无统计学意义。结论:DAPT可通过阻断Notch信号通路抑制尾加压素Ⅱ诱导的CFs表型转换,激活胶原降解途径,并可重调MMP-1/TIMP-1平衡,此作用有助于抑制心肌纤维化。
AIM: To investigate the effect of γ-secretase inhibitor (DAPT) on the expression of α-smooth muscle actin-α (α-SMA) and the balance of extracellular matrix (ECM) degradation in cardiac fibroblasts (CFs) induced by Notch signaling pathway. Methods: CFs of SD suckling rats were isolated and purified by trypsin digestion and differential adherent method. CFs were divided into normal group (N group, no intervention when cultured) and urotensin Ⅱ group (U The cells were treated with UⅡ10 -8 mol / L and DAPT + urotensin Ⅱ (D group, with addition of UⅡ10 -8 mol / L and DAPT 75 μmol / L) PCR was used to detect the expression of MMP-1, TIMP-1, α-SMA and Notch1 mRNA. Western Blot was used to detect the expression of α-SMA and Notch1 protein expression. Results: The mRNA expressions of TIMP-1, α-SMA and Notch1 in U group were significantly higher than those in N group (all P <0.01), while the expressions of MMP-1 mRNA and TIMP-1 mRNA in U group were lower than those in N group <0.01). The mRNA expressions of TIMP-1, α-SMA and Notch1 and the expressions of α-SMA and Notch1 in group D were significantly lower than those in group U (all P <0.01). The expressions of MMP-1mRNA and TIMP -1 mRNA was higher than that of U group (P <0.01), but there was no significant difference between the above indexes in group D and N group. Conclusion: DAPT can inhibit the phenotype transition induced by urotensin Ⅱ, activate the pathway of collagen degradation, and repress the balance of MMP-1 / TIMP-1 by blocking the Notch signaling pathway. This effect can be used to inhibit myocardial fibrosis .