This Is A Speedy Approach To Succeed With OPHN1

As compared with mice, hamster lipoprotein metabolism more closely resembles that in humans because hamsters, like humans, secrete only apoB-100 containing VLDL from liver and because their plasma contains cholesteryl ester transfer protein (CETP) (Taghibiglou et?al., 2000). Previous studies showed that a high carbohydrate diet raises the concentration of TGs in liver and plasma of these animals (Avramoglu et?al., 2003?and?Taghibiglou et?al., 2000). To eliminate hepatic Scap mRNA in hamsters, we delivered siRNA to liver by intravenous injection of siRNA complexed with Selleckchem CCI779 lipid nanoparticles (LNP) (Akinc et?al., 2009). In preliminary?experiments, we designed a series of Scap siRNAs and screened them for their OPHN1 ability to decrease Scap mRNA in primary hamster hepatocytes. The most potent siRNA reduced Scap mRNA by more than 80%, and this was used for in?vivo studies (designated siScap). As a control, we introduced four mutations into siScap to generate a mismatch version that did not reduce Scap mRNA (designated siScap-mm). Hamsters were fed the high-sucrose diet for 7?weeks. On weeks 4 and 6 of the diet, hamsters were injected intravenously with saline or 2.5?mg/kg body weight of LNP-formulated siScap or siScap-mm. siScap reduced liver Scap mRNA by 80% (Table?S2), and Scap protein was reduced below the limit of detection by our antibody (Figure?3A). The precursor and nuclear forms of SREBP-1 and -2 were also markedly reduced (Figure?3A). Although LDLR mRNA was reduced by siScap (Figure?3B), the level of LDLR protein did not decline (Figure?3A). LDLR preservation is likely attributable to the marked reduction in the mRNA encoding PCSK9 in these animals (Figure?3B). PCSK9 is a protein that destroys hepatic LDLRs (Horton et?al., 2009). PCSK9 and LDLR are both activated by SREBP-2 ( Horton et?al., 2003). In the sucrose-fed hamsters, mRNAs for FAS and SCD-1, two targets of SREBP-1c, were elevated when compared to controls (indicated by the dashed line in Figure?3B). These mRNAs were reduced to normal by siScap. siScap also reduced the mRNA for ACC-1, but the reduction did not reach the level of significance at p?BVD-523 chemical structure by siScap (Figure?3B). Control siScap-mm had none of these effects. Plasma TGs were elevated nearly 3-fold to 570?mg/dl in sucrose-fed hamsters (Figure?4A). siScap reduced these levels nearly to normal. Liver TGs were elevated more modestly, and were reduced by siScap. siScap-mm had none of these effects. Fractionation of plasma lipoproteins by fast performance liquid chromatography (FPLC) revealed a marked reduction in VLDL-TGs and VLDL-cholesterol in siScap-treated hamsters (Figure?4C). There was no significant change in LDL- or HDL-cholesterol levels.