Knockout Of The Ath26 Quantitative Trait Locus Candidate Gene Cyp4f13 Decreases Atherosclerosis In Dba/2 Apoe-Deficient Mice

We previously identified the atherosclerosis modifying Ath26 quantitative trait locus (QTL) on mouse chr. 17 from intercrosses of apoE-/- mice on the AKR and DBA/2 background, where the DBA/2 allele is associated with larger lesions. The aim of our study was to identify causal genes responsible for Ath26 . Here we report confirmation of the Ath26 QTL in female AKR.DBAchr17 congenic mice, where inheriting 2 DBA/2 vs. AKR alleles at this locus resulted in ~5x larger lesions (p Cyp4f13 as a candidate gene in this locus from gene expression microarray data of bone marrow derived macrophages from individual F2 mice, where Cyp4f13 expression had the strongest correlation with lesion area of all expressed genes (r=0.46, p Cyp4f13 knockout (KO) embryonic stem cells and created KO mice that we backcrossed on the DBA/2 apoE-/- background. We bred male and female DBA/2 apoE-/- mice that were wild type (WT), hemizygous (HET), or KO for the Cyp4f13 gene. Chow diet fed mice were euthanized at 16 weeks of age. There were no significant differences in body weight, plasma total cholesterol or HDL-C among the 3 genotypes in either sex. Cross section lesion areas in the aortic root were not normally distributed and analyzed with non-parametric statistics. In male mice the lesions areas were smaller in the HET and KO vs. WT mice, and since the HET and KO mice had ~ equivalent lesions, we combined these two groups and found a significant 35 % decrease in median lesion area vs. WT mice (P=0.03, two-tailed Mann-Whitney test, HET+KO n=42, WT n=11). In female mice, lesions were 50% smaller in the HET and KO vs. WT mice (P=0.001, HET+KO n=31, WT n=17). Cyp4f13 belongs to the Cyp4F gene family containing 9 members in mice, 8 of which are clustered on chr. 17. Cyp4F proteins hydroxylate the omega carbon on arachadonic acid forming 20-HETE, and also hydroxylate leukotriene B4 (LTB4). Liver microsomes from Cyp4f13 HET mice had decreased 20-HETE forming activity vs. WT mice. 20-HETE and/or LTB4 metabolism may regulate inflammation and contribute to atherogenesis. We conclude that Cyp4f13 is an atherosclerosis modifier gene that may be partially responsible for the Ath26 QTL on chr. 17.