LncRNA-n334788/miR-369-3p/CAZPA1轴调控特发性肺纤维化作用机制

Idiopathic pulmonary fibrosis (IPF) is a chronically progressive scarring lung disease of unclear cause. The prognosis of IPF is still poor because effective therapies are unavailable. Emerging evidence suggests that the long noncoding RNAs (lncRNAs) are involved in IPF. However, their mechanisms have not been fully elucidated. Our previous study found that the lncRNA-n334788 levels of plasma or lung were significantly higher in patients with IPF than those in healthy controls. Its target gene CAPZA1 was also increased. LncRNA-n334788 localized predominantly in the cytoplasmic in lung by performing RNA fluorescence in situ hybridization. Bioinformatics analysis showed lncRNA-n334788 could increase CAPZA1 via sponging miR-369-3p. Pervious data showed overexpression of miR-369-3p could decrease CAPZA1 level, and induce mesenchymal-epithelial transition. Accordingly, we hypothesized that lncRNA-n334788 might function as competing endogenous RNA (ceRNA) for miR-369-3p, which could promote epithelial-mesenchymal transition, and aggravate lung fibrosis by regulating CAPZA1. The study aims to explore the mechanism underlying lncRNA-n334788/miR-369-3p/CAZPA1 axis in cells’ or animals’ model of IPF. This study will provide evidence for new therapeutic target in IPF.

特发性肺纤维化（IPF）病因不明，预后极差，至今无有效治疗手段。研究报道长链非编码RNA（lncRNA）可能参与IPF，但作用和机制均不明确。我们前期发现IPF患者lncRNA-n334788及其靶基因CAPZA1明显增高，荧光原位杂交显示lncRNA-n334788定位于胞浆。生物信息分析lncRNA-n334788可能靶向miR-369-3p，进而调控CAPZA1。进一步实验发现过表达miR-369-3p降低CAPZA1，诱导间质上皮细胞转化。据此我们假设：胞浆中lncRNA-n334788可能竞争结合miR-369-3p，调控CAPZA1，诱导上皮细胞间质转化，加重肺纤维化。本课题利用细胞和动物造模等分子生物学手段，围绕上述科学假设，研究lncRNA-n334788/miR-369-3p/CAZPA1轴在IPF中的调控作用及机制，为寻找IPF的发病机制及治疗靶点提供依据。

特发性肺纤维化（idiopathic pulmonary fibrosis, IPF）是一种慢性、进行性、纤维化性肺疾病，是间质性肺炎最常见和预后最差的类型。IPF病因不明，预后极差，至今无有效治疗手段。研究表明长链非编码RNA（long noncoding RNA，lncRNA）可能参与IPF，但其确切作用和具体作用机制尚不明确。本研究结果显示，lncRNA-334788无蛋白编码功能，大部分位于细胞浆内，在TGF-β1诱导的细胞模型和博来霉素诱导的大鼠肺纤维化模型中，lncRNA-334788表达明显增高。在TGF-β1诱导的RLE-6TN细胞模型中，lncRNA-n334788过表达促进肺泡上皮间质转化（epithelial-mesenchymal transition，EMT），诱导细胞增殖，抑制细胞凋亡。反之，lncRNA-n334788敲降抑制EMT，抑制细胞增殖，促进细胞凋亡，说明lncRNA-n334788具有促纤维化的作用。通过双荧光素酶报告基因、RNA pull down、RIP等方法发现，lncRNA-n334788通过“分子海绵”的作用，竞争性结合miR-369-3p，促进CAPZA1表达，从而促进EMT，诱导细胞增殖，抑制细胞凋亡。在博来霉素诱导大鼠肺纤维化模型中使用sh-lncRNA-n334788干预后，大鼠肺纤维化程度改善。本研究首次揭示lncRNA-n334788在IPF发病过程中的分子机制，对lncRNA和lncRNA-miRNA-mRNA调控网络的深入探索，将有助加深对IPF发病及进展的理解。同时，通过对在IPF过程中起重要作用的lncRNA基因进行相应的表观遗传学修饰，使之成为可能的治疗靶点，将为IPF的治疗开辟新途径，具有良好的转化前景。