Mouse Liver Micrornas Expression Profiling Changes In The Early Stage Of Echinococcus Multilocularis Infection And Preliminary Study On The Function Of MiR-133A-3P

Objective:To study the expression of miRNAs in host liver of Em.infected early time, in order to clarify miRNAs in Em. infection plays an important role in liver fibrosis induced by host. Methods:In application of high-throughput sequencing methods to detect Em. infected one month mice normal liver tissue and alveolar hydatid disease nearby tissue miRNAs expression profiling, and screening of differentially expressed miRNAs. The predicted target gene of miRNAs were identified by application of TargetScan and miRDB two sites, and selecting target genes associated with inflammation and fibrosis related. Real-time fluorescence quantitative PCR(qRT-PCR) to verify the sequencing results; detection of Em protein MTT method can effectively stimulate rat hepatic stellate cells (T6cells) proliferation and differentiation; the expression change of qRT-PCR detection of miRNAs Em. protein in T6cells induced by fibrosis, and detect the expression of fibrosis related gene α-SMA, CollA1, Col3A1, TGF-β, TGF-β R Ⅰ and TGF-β RⅡ changes; the changes of protein expression of Western-Blot detection of Em. protein caused by fibrosis T6fibrosis α-SMA, CollA1, Col3A1, TGF-β, TGF-β RⅡ, Smad-2/3and Smad-4. Detection of T6cells miR-133a-3p and fibrosis related genes expression after been transfected by miR-133a-3p-mimic. Results:High throughput sequencing to detect Em.infection early mouse liver miRNAs expression were screened out of20differentially expressed miRNAs,13were up-regulated and7down regulated expression of miRNAs was detected by MTT. In application of TargetScan and miRDB two websites8miRNAs were screened participated in the regulation of tissue necrosis,inflammation and fibrosis;30%Em. protein can effectively stimulate the proliferation and differentiation of rat hepatic stellate cells;30%Em. protein stimulated T6cells48h miR-133a-3p expression was down regulated; and TGF-β/Smad signaling pathway related liver fibrosis the mRNA of Col-1A1, Col-3A1, TGF-β and TGF-β R Ⅰ were high expressed; Col-1A1, α-SMA, Smad-4and TGF-β protein also increased. After transfection of miR-133a-3p-mimic to T6cells miR-133a-3p expression was significantly increased, and decreased expression of fibrosis related factors gene. Conclusion:The successful establishment of the difference of Em. infection early mouse liver miRNAs expression profiling, obvious differences in table miR-133a-3p to a key role in the occurrence and development of Em. infection induced liver fibrosis, its role may be through regulation of TGF-β/Smad signal.