Construction of shRNA Expressing Plasmid Targeted Rat NogoB and Observation of the Effection aboutNogoB on Hepatic Stellate Cells

Objective To construct shRNA expressing plasmid inhibiting rat NogoB and to observe its possible effect on rat primary hepatic stellate cells (HSCs) contraction. Methods Three pairs of shRNAs targeting different sequence of rat NogoB were designed and constructed into pSuper plasmid by DNA recombination technique. Culture-activated HSCs were transfected with NogoB-shRNA plasmids to scan the effective plasmid which could inhibit NogoB gene expression by Real-time PCR. And this depressant effect was also confirmed with Western blot. After NogoB was knocked-down effectively, ETA and ETB mRNA expression were assessed by Real-time PCR. Results Among the three pairs of recombinant plasmids, NogoB-shRNA2 plasmid could inhibit NogoB expression specifically. In HSCs, NogoB knockdown decreased the ratio of ETA and ETB. Conclusion We constructed specific NogoB-shRNA expression plasmid successfully which might be involved in contraction of HSCs.