RESUMEN
We evaluated whether inhibiting gene expression by small interfering RNA (siRNA) can be used for an in vivo model using a germ cell-specific gene (Tex101) as a model target in mouse testis. We generated plasmid-based expression vectors of siRNA targeting the Tex101 gene and transfected them into postnatal day 10 mouse testes by in vivo electroporation. After optimizing the electroporation conditions using a vector transfected into the mouse testis, a combination of high- and low-voltage pulses showed excellent transfection efficiency for the vectors with minimal tissue damage, but gene suppression was transient. Gene suppression by in vivo electroporation may be helpful as an alternative approach when designing experiments to unravel the basic role of testicular molecules.
RESUMEN
BACKGROUND/AIMS: Although microRNAs are known to be post-transcriptional regulators in physiological and pathological events in the liver, their role in the obstructive jaundice liver remains unclear. METHODOLOGY: We sequenced the small RNA libraries of the bile duct ligation (BDL) mouse liver to detect the in vivo microRNA expression profiles of obstructive jaundice. We also validated the differential expression of microRNAs in the BDL liver using real-time PCR. Laser microdissection was performed to identify the origin of BDL-related microRNAs. An IL6-treated normal intrahepatic biliary epithelial cell line was used as an in vitro model of obstructive jaundice. RESULTS: We found microRNAs that were upregulated in the BDL liver (let-7a, let-7d, let-7f, let-7g, miR-21, miR-125a-5p, miR-125b-5p, miR-194, miR-199a-3p, miR-199a-5p, miR-214, miR-221, and miR-486). Furthermore, laser-microdissection analysis showed that miR-199a-5p was significantly upregulated in the intrahepatic bile duct of the BDL liver. The in vitro expression of miR-199a-5p was appreciably elevated in accordance with increased proliferation of IL6-treated cells. CONCLUSIONS: We revealed dynamic changes in microRNA expression during obstructive jaundice using the BDL model. MiR-199a-5p was likely associated with the proliferation of intrahepatic bile ducts. Our data will facilitate further study of the pathophysiological role(s) of microRNAs in the obstructive jaundice liver.