Association of a Lithogenic Abcg5/Abcg8 Allele on Chromosome 17 (Lith9) with Cholesterol Gallstone Formation in PERA/EiJ Mice

Abstract

We employ quantitative trait locus (QTL) mapping to identify genomic regions that harbor cholesterol gallstone susceptibility (Lith) genes in inbred mice. In a previous intercross of gallstone susceptible PERA and resistant I/LnJ mice, we detected a Lith locus on chromosome (Chr) 17 that was inherited from strain PERA and named Lith9. QTL mapping in an intercross of strain PERA with DBA/2J mice, another gallstone resistant strain, detected only a weak association of Lith9 with gallstone formation. We re-examined the genetic determinants of cholesterol gallstone susceptibility in strain PERA in a combined QTL analysis of the datasets from the two independent crosses sharing PERA as the gallstone susceptible parental strain. In addition, a haplotype analysis for the Lith9 region was conducted in the three parental strains based on single nucleotide polymorphisms and candidate genes were sequenced. Expression of candidate genes was determined by real-time PCR. QTL analysis of the dataset derived from the combined crosses confirmed a significant shared QTL in the Lith9 region with a gallstone susceptible allele in strain PERA and resistant alleles in strains I/Ln and DBA/2. The haplotype analysis of Lith9 identified a region of 3.4 megabase pairs with shared haplotypes between strains I/Ln and DBA/2 that differed from the haplotype of strain PERA. The narrowed Lith9 interval contained the candidate genes Abcg5 and Abcg8 encoding the canalicular cholesterol transporter. Sequencing confirmed identical Abcg5 and Abcg8 alleles in strains I/Ln and DBA/2 and different alleles of both genes in strain PERA. The PERA alleles of Abcg5 and Abcg8 were associated with significantly higher hepatic mRNA expression levels compared with strains I/Ln and DBA/2 that correlated positively with higher biliary cholesterol concentration. In conclusion, our analyses confirmed the Lith9 locus with a gallstone susceptible allele in strain PERA and narrowed the Lith9 interval. Higher expression levels of Abcg5/Abcg8 in strain PERA compared with I/Ln and DBA/2 substantiate the notion of dysregulated canalicular cholesterol transport as a principal cause of cholesterol gallstone formation in inbred mice.