Schistosomiasis is one of the world's major public health problems in terms of morbidity and mortality, causing granulomatous inflammation and cumulative fibrosis. This study explored in vivo and vitro effects of miR-29b-3p in granulomatous liver fibrosis by targeting COL1A1 and COL3A1 in Schistosoma japonicum infection. Thirty male Balb/c mice were assigned to normal control and model (percutaneous infection of cercariae of S. japonicum) groups. NIH-3T3 mouse embryonic fibroblasts were designated into blank, NC, miR-29b-3p mimic, TGF-β1, TGF-β1 + NC, and TGF-β1 + miR-29b-3p mimic groups. HE and Masson staining were employed to observe the pathological changes and collagenous fibrosis. The expression of α-SMA, COL1A1, COL3A1, TIMP-1 was determined by immunohistochemistry. The RT-qPCR, Western blotting and immunofluorescence staining were conducted to determine expression of miR-29b-3p, COL1A1, and COL3A1. CCK-8 assay and flow cytometry were performed to evaluate viability and apoptosis. The relative expression of miR-29b-3p decreased in the model group. The model group showed marked fibrosis in liver tissues. The expression of α-SMA, COL1A1, COL3A1, TIMP-1 was higher in the model group than that in the normal control group. Dual luciferase reporter gene assay revealed that miR-29b-3p directly targeted COL1A1 and COL3A1. Compared with the blank, NC, TGF-β1 and TGF-β1 + NC groups, the miR-29b-3p mimic group exhibited up-regulated expression of miR-29b-3p and MMP-9 but down-regulated expression of TIMP-1, HSP47, α-SMA, COL1A1, and COL3A1; while lower cell viability but higher apoptosis rate showed. It indicated that miR-29b-3p prevents S. japonicum-induced liver fibrosis by inhibiting COL1A1 and COL3A1.

The study aimed to investigate the impact of miR-182 and FOXO1 on S. japonica-induced hepatic fibrosis. Microarray analysis was performed to screen out differential expressed miRNAs and mRNAs. Rat hepatic fibrosis model and human hepatocellular cell line LX-2 were used to study the effect of miR-182 and FOXO1. qRT-PCR and Western blot were used to detect the expression of miR-182, FOXO1 or other fibrosis markers. The targeting relationship between FOXO1 and miR-182 was verified by luciferase reporter assay. Immunohistochemistry or immunofluorescence staining was conducted to detect FOXO1 or α-SMA in rat hepatic tissues. Cell viability and apoptosis were detected by MTT assay and flow cytometry. The expression of PI3K/AKT pathway-related proteins was detected by Western blot. miR-182 was highly expressed in liver fibrosis samples, and FOXO1 expression was negatively correlated with miR-182 expression. After transfection of miR-182, FOXO1 expression was down-regulated, with the results of LX-2 cells proliferation inhibition and apoptosis induction, as well as the aggravation of rat hepatic fibrosis. The expression of p-AKT/AKT and p-S6/S6 was increased, meaning that the PI3K/AKT signal pathway was activated. The results were reversed when treated with Wortmannin (PI3K inhibitor). After transfection of miR-182 inhibitor, FOXO1 expression was up-regulated, LX-2 cell proliferation was inhibited, and apoptosis rate was increased. High-expressed miR-182 and low-expressed FOXO1 promoted proliferation and inhibiting apoptosis on liver fibrosis cells, stimulating the development of S. japonica-induced hepatic fibrosis through feeding back to PI3K/AKT signaling pathway.

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