Background and purpose

Idiopathic pulmonary fibrosis (IPF) is a life-threatening progressive disease characterized by aberrant fibroblast activation. This study aims to explore the role of the circ0044226 on fibroblast-to-myofibroblast transition (FMT).

Methods

Bleomycin and TGF-β1 were respectively used to induce the IPF mice model and human lung fibroblasts to myofibroblast differentiation. The mRNA and protein levels were examined by qRT-PCR and western blot. Localization of α-SMA was evaluated by immunofluorescence staining. Cell viability and proliferation were evaluated by CCK8 and EDU test. Dual-luciferase reporter assay was used to analyze the interaction between miR-7 and circ0044226 or sp1. Fluorescence in situ hybridization (FISH) assay was used for the identification of sub-location of circ0044226 and miR-7 in cells. The IPF model mice received intratracheal injection of AAV-sh-NC and AAV-sh- circ0044226, and lung fibrosis was detected by HE staining, Masson staining and immunohistochemistry assay.

Results

The circ0044226 was upregulated while miR-7 was downregulated in IPF mice model and FMT-derived myofibroblasts. miR-7 was a target of circ0044226 and sp1 was a target of miR-7. circ0044226 was distributed mostly in the cytoplasm and functioned as a miR-7 sponge to positively regulate the expression of sp1. Intervention of circ0044226 could ameliorate FMT and suppress fibroblast viability and proliferation by functioning as an endogenous miR-7 sponge.

Conclusion

Circ0044226 knockdown alleviates fibroblast proliferation and FMT by functioning as a competing endogenous RNA, which may represent a promising therapy for pulmonary fibrosis.

中文翻译：

---

肺成纤维细胞的转化和纤维化受circ0044226调控。

背景和目的

特发性肺纤维化（IPF）是威胁生命的进行性疾病，其特征在于成纤维细胞活化异常。这项研究旨在探讨circ0044226在成纤维细胞向成肌纤维细胞转化（FMT）中的作用。

方法

博莱霉素和TGF-β1分别用于诱导IPF小鼠模型和人肺成纤维细胞分化为成肌纤维细胞。通过qRT-PCR和蛋白质印迹检查mRNA和蛋白质水平。通过免疫荧光染色评估α-SMA的定位。通过CCK8和EDU测试评估细胞活力和增殖。双荧光素酶报告基因分析用于分析miR-7与circ0044226或sp1之间的相互作用。荧光原位杂交（FISH）分析用于鉴定circ0044226和miR-7在细胞中的亚位。IPF模型小鼠接受气管内注射AAV-sh-NC和AAV-sh-circ0044226，并通过HE染色，Masson染色和免疫组化分析检测到肺纤维化。

结果

在IPF小鼠模型和FMT衍生的成肌纤维细胞中，miR-7被下调，而circ0044226被上调。miR-7是circ0044226的靶标，而sp1是miR-7的靶标。circ0044226主要分布在细胞质中，并起miR-7海绵的作用，正向调节sp1的表达。circ0044226的干预可起到内源性miR-7海绵的作用，可改善FMT并抑制成纤维细胞的活力和增殖。

结论

Circ0044226组合式发挥竞争性内源性RNA的作用，减轻了成纤维细胞的增殖和FMT，这可能是一种有前途的肺纤维化治疗方法。