基质金属蛋白酶1、3,基质金属蛋白酶组织抑制剂1和miR-29在大鼠肺纤维化中及活性维生素D3处理后的表达与作用

Effect of active vitamin D3 on expressions of matrix metalloproteinase 1 and 3,tissue inhibitor of metalloproteinase 1 and miR-29 during progression of pulmonary fibrosis of rats

目的:探讨大鼠肺纤维化发生发展中基质金属蛋白酶1(MMP1)、MMP3、基质金属蛋白酶组织抑制剂1(TIMP1)及miR-29的表达和作用以及活性维生素D3[1,25(OH)_2D_3]处理对这些基因表达的影响。方法:雄性SD大鼠随机分为预防组和治疗组。预防组分为对照组Ⅰ、模型组Ⅰ和给药组Ⅰ,治疗组分为对照组Ⅱ、模型组Ⅱ和给药组Ⅱ。模型组Ⅰ/Ⅱ和给药组Ⅰ/Ⅱ经气管注入博莱霉素,对照组Ⅰ/Ⅱ经气管注入生理盐水。预防组和治疗组分别于术后第2和14天腹腔注射给药,给药组给予活性维生素D3,模型组给予活性维生素D3溶剂,对照组给予生理盐水。预防组的各组分别于术后第14、21、28天处死大鼠取材,治疗组的各组分别于术后第21和28天处死大鼠取材。实时定量PCR检测大鼠肺组织中miR-29a及TIMP1 mRNA表达水平,免疫组织化学检测组织中MMP1、MMP3和TIMP1的表达水平。结果:模型组Ⅰ/Ⅱ和给药组Ⅰ/Ⅱ中的MMP1、MMP3和TIMP1的表达均明显高于相同时间点对照组Ⅰ/Ⅱ中的表达,而miR-29a的表达明显低于相应的对照组Ⅰ/Ⅱ;给药组Ⅰ/Ⅱ中MMP1、MMP3和TIMP1的表达均低于相应时间点的模型组Ⅰ/Ⅱ的表达,而给药组Ⅰ/Ⅱ中miR-29a的表达则高于模型组Ⅰ/Ⅱ中的表达。结论:MMP1、MMP3、TIMP1和miR-29在大鼠肺纤维化发生发展中具有重要作用,活性维生素D3可以促进miR-29表达,抑制MMP1、MMP3、TIMP1的表达;可能是通过miR-29调节包括MMP1、MMP3、TIMP1在内的多种靶基因来发挥抑制纤维化的作用。

Objective： To explore the role of active vitamin D3 in progression of pulmonary fibrosis through determining expression and function of matrix metalloproteinase 1 （MMP1）, MMP3, tissue inhibitor of metalloproteinase 1 （TIMP1） and miR-29 in rats. Methods： Male SD rats were randomly divided into prevention group and treatment group, and subdivide into control, model and medicine subgroups respectively. Animals in model and medicine subgroups were injected with bleomyein through the trachea, while rats in control subgroups were injected with sterile saline through the trachea. The active vitamin D3, solvent and sterile saline were injected intraperitoneally into rats in medicine, model and control subgroups on postoperative day 2 and 14. Samples harvested from rats in prevention group on postoperative day 14, 21 and 28, and from rats in treatment group on postoperative day 21 and 28 were submitted to detect levels of miR 29 and TIMP1 using real time qPCR and expressions of MMP1, MMP3 and TIMP1 by immunohistochemistry. Results： Considering the same postoperative time points, expressions of MMP1, MMP3 and TIMP1 in rats from model and medicine subgroups were significantly higher than those from control subgroups, while levels of miR29 in model and medicine animals were obviously lower than that from control subgroups. Administration of active vitamin D3 caused significant downregulation of MMP1, MMP3 and TIMP1 and statistical upregulation in miR-29 compared to data from model animals. Conclusion： The active vitamin D3 could prevent formation of pulmonary fibrosis by regulating expressions of MMP1, MMP3, TIMP1 and miR-29, which may important players during the progression of pulmonary fibrosis.