P45. Characterization of Cyp2b-Knockdown mouse reveals changes in lipid metabolism Basma Damiri and William Baldwin Biology/ Environmental Toxicology, Clemson Univeristy, Clemson, SC
The Cyp2b subfamily contains five members (Cyp2b9, Cyp2b10, Cyp2b13, Cyp2b19, and Cyp2b23) of which three (Cyp2b9, Cyp2b10, Cyp2b13) are hepatic enzymes involved in xenobiotic detoxification. We made a Cyp2b-knockdown mouse using lentiviral-promoted shRNA homologous to all five Cyp2b subfamily members to characterize Cyp2b’s role in xenobiotic detoxification. Western blots from 8-week old mice demonstrate that hepatic Cyp2b’s are repressed 5-10X by the shRNA. Unexpectedly, Cyp2b-KD mice showed significant changes in some organ weights, especially an increase in abdominal, inguinal, and renal adipose. Both male and female Cyp2b-KD mice showed significant increase in fat to body weight ratio (112.1% and 73.5% for males and females, respectively). Interestingly, associated with changes in fat to body ratios was changes in non-fasting triglycerides levels. Non-fasting serum triglyceride was increased 18-39% in 8-10 weeks old female and male mice, respectively with a statistically significant increase in males. Therefore, wild-type and Cyp2b-knockdown mice were housed for 35 weeks and necropsies performed to test whether older Cyp2b-knockdown mice show perturbed lipid utilization. Thirty-five week old Cyp2b-KD mice exhibited a 17 to 22% increase in their body weight caused by a significant increase in fat deposition compared to wild-type (FVB) mice. This was accompanied by increased triglyceride and low density lipoprotein levels. Subsequent studies were performed with 100ml of corn oil or the CAR activator and CYP-inducer TCPOBOP at 3mg/kg/ in corn oil as a carrier. Surprisingly, 8-12 weeks Cyp2b-knockdown mice showed some difficulty in dealing with the corn oil. Significant increases in total serum cholesterol (25-39%) was observed in the corn oil control Cyp2b-knockdown male and female mice, respectively, compared to the WT mice, probably because the Cyp2b-KD mice were unable to respond to the unsaturated fatty acids in the corn oil. The increases in non-fasting serum total cholesterol and triglyceride was reversed by TCPOBOP treatment in Cyp2b-knockdown male mice but not in Cyp2bknockdown female mice as TCPOBOP treatment caused a significant increase in triglyceride in Cyp2b-KD females compared to WT mice. Cyp2b-KD mice appear to have perturbations in clearing corn oil from the liver as corn oil treatment increased Oil Red O staining indicating accumulation of lipids in the livers of Cyp2b-KD mice compared to WT mice. FoxA2 and Cpt1a, involved in triglyceride homeostasis, were increased 2-6 fold, albeit not significantly, in Cyp2p-KD mice treated with corn oil. In conclusion, changes in Cyp2b expression led to perturbation in lipid metabolism in Cyp2b-KD mice. This suggests that Cyp2b is more than a detoxification enzyme, but is also involved in the metabolism of unsaturated fatty acids, as Cyp2b-KD mice have increased fat deposition and show increased serum and liver lipid levels.