Aberrant Gene Expression in Dogs with Portosystemic Shunts

Abstract

Congenital portosystemic shunts are developmental anomalies of the splanchnic vascular system that cause portal blood to bypass the liver. Large-breed dogs are predisposed for intrahepatic portosystemic shunts (IHPSS) and small-breed dogs for extrahepatic portosystemic shunts (EHPSS). While the phenotype resulting from portal bypass of the liver of the two types of shunt is identical, the genotype and molecular pathways involved are probably different. The aim of this study was to gain insight into the pathways involved in the different types of portosystemic shunting. Microarray analysis of mRNA expression in liver tissue from dogs with EHPSS and IHPSS revealed that the expression of 26 genes was altered in either IHPSS or EHPSS samples compared with that in liver samples from control dogs. Quantitative real-time PCR of these genes in 14 IHPSS, 17 EHPSS, and 8 control liver samples revealed a significant differential expression of ACBP, CCBL1, GPC3, HAMP, PALLD, VCAM1, and WEE1. Immunohistochemistry and Western blotting confirmed an increased expression of VCAM1 in IHPSS but its absence in EHPSS, an increased WEE1 expression in IHPSS but not in EHPSS, and a decreased expression of CCBL1 in both shunt types. Regarding their physiologic functions, these findings may indicate a causative role for VCAM1 in IHPSS and WEE1 for IHPSS. CCBL1 could be an interesting candidate to study not yet elucidated aspects in the pathophysiology of hepatic encephalopathy.

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“…Gene expression differences of the selected genes were confirmed using RT‐qPCR. RNA isolation, cDNA synthesis, and gene expression profiling using RT‐qPCR was performed as described previously with a maximum of 40 cycles.…”

Lysosome-mediated degradation of a distinct pool of lipid droplets during hepatic stellate cell activation

“…An iScript cDNA Synthesis Kit (Bio-Rad) was used to synthesize cDNA. Primer design and quantitative PCR (qPCR) conditions were as described previously (41). Briefly, qPCR reactions were performed in duplicate using a Bio-Rad detection system.…”

ATGL and DGAT1 are involved in the turnover of newly synthesized triacylglycerols in hepatic stellate cells

“…An iScript cDNA synthesis kit (Bio-Rad, Veenendaal, The Netherlands) was used to synthesize cDNA. Primer design and quantitative (q) PCR conditions were as described previously (17). Briefly, qPCR reactions were performed in duplicate using the Bio-Rad detection system.…”