Research on regulating the blood ammoniathrough silencing Rhc glycoprotein by small interfering RNA

SUN Chen; ZHAO Ji-yue; LU Zai-ming

doi:10.16048/j.issn.2095-5561.2019.01.05

Abstract：Objective To investigate the effect of small interfering RNA on regulating blood ammonia by silencing Rhcglycoprotein, and to provide a basis for targeted drug treatment of hepatic encephalopathy.Methods The original generation of small mouse brain microvascular endothelial cells was cultivated, using cationic liposome small interfering RNA sequence Rhcg-Mus-455, Rhcg-Mus-607and Rhcg-Mus-883 transfected into small mouse brain microvascular endothelial cells, the protein and RNA cells were extraction and transfected, protein western blot experiment and agarose gel electrophoresis experiment were conducted to observe the expression of intracellular ammonia transporter protein and its mRNA, so as to observe the silencing effect and silencing efficiency of small interfering RNA on Rhcg gene of cells, and to screen out the small interfering RNA sequence that can effectively inhibit the ammonia transporter gene in rat brain microvascular endothelial cells.Results The Rhcg protein levels of Rhcg-Mus-455, Rhcg-Mus-607 and Rhcg-Mus-883 of 3 small interfering RNA sequences were compared in the Western blot experiment, and the differences were not statistically significant (P＞0.05). The Rhcg protein levels of Rhcg-Mus-455, Rhcg-Mus-607 and Rhcg-Mus-883 were all lower than those of normal saline and negative control, and the difference was statistically significant (P＜0.05). Rhcg mRNA levels of Rhcg-Mus-455, RhcgMus-607 and Rhcg-Mus-883 were compared in the agarose gel electrophoresis experiment, and the differences were not statistically significant (P＞0.05). The Rhcg mRNA level of Rhcg-Mus-455 was lower than that of normal saline and negative control (P＜0.05). Conclusion All of the 3 small interfering RNA sequences have the effect of silencing the expression of ammonia transporters in the mouse cerebrovascular endothelial cells, the small interfering RNA sequence 455 may have obvious silencing efficiency.Therefore, small interfering RNA that can specifically bind ammonia transporters can be found in vitro through gene silencing technology to inhibit blood ammonia transport and thereby improve hepatic encephalopathy.