二氧化硅诱导大鼠肺成纤维细胞转分化中基因组甲基化水平

DNA methylation levels of genome in cFb transdifferentiation induced by SiO： in rats

目的探讨二氧化硅（SiO2）诱导大鼠肺成纤维细胞转分化过程中基因组的甲基化水平。方法采用直接和间接细胞共培养方法对sD大鼠巨噬细胞、肺成纤维细胞进行原代培养，以25、50、100μg／ml的SiO2作为受试物，采用免疫细胞化学方法对肺成纤维细胞转分化为肌成纤维细胞进行鉴定，采用高效液相色谱法检测成纤维细胞的基因组甲基化水平。结果间接共培养方式下，与空白组比较，25、50、100μg／ml SiO：剂量组成纤维细胞的基因组甲基化水平分别降低19．9％、26．9％、30．3％，差异有统计学意义（P〈0．05）；与100μg／ml SiO2剂量组比较，对照组5一氮杂-2-脱氧胞苷（5-aza—dC）处理后成纤维细胞基因组甲基化水平降低22．0％，差异有统计学意义（P〈O．05）。ThinCertTM共培养方式下，与空白组比较，25、50、1013μg／ml SiO2剂量组成纤维细胞的基因组甲基化水平分别降低22．2％、30．2％、36．7％，差异有统计学意义（P〈0．05）；与100μg／ml SiO2剂量组比较，对照组5-aza-dC处理后成纤维细胞基因组甲基化水平降低20．6％，差异具有统计学意义（P〈0．05）。结论矽肺体外细胞模型共培养方式下，SiO2可引起成纤维细胞基因组甲基化水平降低，用ThinCertTM共培养方式建立的体外细胞模型在矽肺纤维化发生研究中有应用前景。

Objective To explore the DNA methylation levels of genome in cFb transdifferentiation induced by SiO2 in rats. Methods The primary macrophages and fibroeytes of SD rats were co-cultured directly and indirectly, which were exposed to SiO2 at the doses of 25, 50 and 100 g/ml. The transdifferentiation of cFb was identified with immunohistoehemieal assay. The genomie DNA methylation levels of cFb were detected with HPLC. Results Under the condition of indirect co-culture, as compared with control group, the genomic DNA methylation levels of cFb exposed to SiO2 at the doses of 25, 50 and 100 g/ml reduced by 19.9%, 26.9% and 30.3%, respectively （P〈O.05）; as compared with cFb exposed to 100 g/ml SiO2, the genomic DNA methylation levels of cFb exposed to 5-aza-dC decreased by 22.0% （P〈O.05）. Under the condition of ThinCertTM direct coculture, as compared with control group, the genomic DNA methylation levels of cFb exposed to SiO2 at the dos- es of 25, 50 and 100 g/ml reduced by 22.2%, 30.2% and 36.7%, respectively （P〈O.05）; as compared with cFb exposed to 100 g/ml SiO2, the genomic DNA methylation levels of cFb exposed to 5-aza-dC decreased by 20.6% （P〈O.05）. Conclusion Under the co-culture condition in vitro, SiO2 could reduce the genomic DNA methyla- tion levels of cFb. The ThinCertTM direct co-culture can be used to study the silicosis fibrosis.