维甲酸下调高氧暴露下胎鼠肺成纤维细胞及Ⅱ型肺泡上皮细胞MMP-2表达

Retinoic acid down-regulates the hyperoxia-induced expression of MMP-2 in fetal rat lung fibroblasts and type Ⅱ alveolar epithelial cell

目的探讨维甲酸(RA)对高氧暴露下胎鼠肺成纤维细胞(LFs)和肺泡Ⅱ型上皮细胞(AECⅡ)基质金属蛋白酶-2 (MMP -2 )表达的影响及调控。方法原代培养胎鼠肺LFs及AECⅡ,待其生长至亚汇合状态时,随机分为:空气组、空气+RA组、高氧组和高氧+RA组。于培养2、6、12、2 4和4 8(AECⅡ)或72h(LFs)时,采用半定量RT- PCR方法检测MMP 2和TIMP 2mRNA表达,应用Westernblot检测p38和磷酸化p38(p p38)蛋白表达水平。结果(1)与空气组比较,高氧可促进胎鼠LFs、AECⅡMMP 2mRNA和胎鼠LFsp p38表达(P <0 . 0 1或P <0. 0 5 ) ;(2 )RA能部分下调高氧诱导的胎鼠LFs、AECⅡMMP 2mRNA高表达和明显降低胎鼠LFsp p38表达;(3)高氧、RA对胎鼠LFs、AECⅡTIMP -2mRNA和胎鼠LFsp38蛋白表达无明显影响;(4)胎鼠LFsp p38与MMP 2mRNA表达呈显著性正相关(r =0. 76 7,P <0. 0 1,n =18)。结论高氧可促进胎鼠LFs、AECⅡMMP 2mRNA表达;p38通路参与高氧状态下胎鼠LFsMMP -2表达调控;RA可在转录后水平调节p38的活性状态从而实现对MMP-2的表达调控。

Objective To study the effect of retinoic acid on the hyperoxia-induced expression and regulation of matrix metalloproteinase-2 (MMP-2) in fetal rat lung fibroblasts (LFs) and typeⅡ alveolar epithelial cells (AECⅡ). Methods The primary fetal rat LFs and AECⅡ were cultured in vitro. Cells grew to subconfluence and then were randomly divided into 4 groups: I. Air group; Ⅱ. Air plus RA group; Ⅲ. Hyperoxia group; Ⅳ. Hyperoxia plus RA group. After cells were cultured for 2, 6, 12, 24 and 48 (AECⅡ) or 72 h (LFs), the levels of MMP-2 and tissue inhibitor of metalloproteinase-2(TIMP-2) mRNA were checked by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). p38 and p38 phosphorylation in LFs were determined by Western blot. Results (1) the level of MMP-2 mRNA in fetal rat LFs and AECⅡ and p-p38 in fetal rat LFs increased significantly (P<0.01 or 0.05). (2) RA down-regulated the expression of MMP-2 mRNA in fetal rat LFs and AECⅡ and p-p38 in fetal rat LFs induced by hyperoxia. (3) Hyperoxia and RA had no effect on the expression of TIMP-2 mRNA in LFs and AECⅡ and the expression of p38 in LFs. (4) The expression of MMP-2 mRNA in LFs had positive correlation with p-p38 remarkably(r=0.767, P<0.01, n=18). Conclusion Hyperoxia could up-regulate the expression of MMP-2 mRNA in hyperoxia in LFs and AECⅡ remarkably. the p38 MAPK pathway mediated hyperoxia-induced regulation of the expression of MMP-2 mRNA in LFs. RA could down-regulate the expression of MMP-2 mRNA through down-regulation of p38 activation after translation in hyperoxia.