Deletion of the Murine Cytochrome P450 Cyp2j Locus by Fused BAC-Mediated Recombination Identifies a Role for Cyp2j in the Pulmonary Vascular Response to Hypoxia

Abstract

Epoxyeicosatrienoic acids (EETs) confer vasoactive and cardioprotective functions. Genetic analysis of the contributions of these short-lived mediators to pathophysiology has been confounded to date by the allelic expansion in rodents of the portion of the genome syntenic to human CYP2J2, a gene encoding one of the principle cytochrome P450 epoxygenases responsible for the formation of EETs in humans. Mice have eight potentially functional genes that could direct the synthesis of epoxygenases with properties similar to those of CYP2J2. As an initial step towards understanding the role of the murine Cyp2j locus, we have created mice bearing a 626-kb deletion spanning the entire region syntenic to CYP2J2, using a combination of homologous and site-directed recombination strategies. A mouse strain in which the locus deletion was complemented by transgenic delivery of BAC sequences encoding human CYP2J2 was also created. Systemic and pulmonary hemodynamic measurements did not differ in wild-type, null, and complemented mice at baseline. However, hypoxic pulmonary vasoconstriction (HPV) during left mainstem bronchus occlusion was impaired and associated with reduced systemic oxygenation in null mice, but not in null mice bearing the human transgene. Administration of an epoxygenase inhibitor to wild-type mice also impaired HPV. These findings demonstrate that Cyp2j gene products regulate the pulmonary vascular response to hypoxia.

Pulmonary Hypertension after Prolonged Hypoxic Exposure in Mice with a Congenital Deficiency of Cyp2j

“…The availability of novel gene-targeting techniques allowed us to determine the role of murine Cyp2j in the regulation of chronic pulmonary vascular responses to hypoxic exposure in mice. In a previous study, we demonstrated that Cyp2j-deficient mice exhibit impaired HPV (13). Based on these data, in the present study we hypothesized that Cyp2j deficiency would prevent the development of chronic PH and vascular remodeling during prolonged exposure to hypoxia.…”

“…Alternatively, it is possible that cytochrome P450 epoxygenases other than Cyp2j can contribute to the development of PH in Cyp2j 2/2 mice. We have previously reported that Cyp2c genes are expressed in the lungs of Cyp2j 2/2 mice of both genders (13). We reported that pulmonary Cyp2c29 and Cyp2c38 mRNA levels did not differ between Cyp2j 1/1 and Cyp2j 2/2 mice breathing at FI O 2 0.21 (13).…”

“…We have previously reported that Cyp2c genes are expressed in the lungs of Cyp2j 2/2 mice of both genders (13). We reported that pulmonary Cyp2c29 and Cyp2c38 mRNA levels did not differ between Cyp2j 1/1 and Cyp2j 2/2 mice breathing at FI O 2 0.21 (13). Therefore, we hypothesized that Cyp2c family members could compensate for the absence of Cyp2j in the development of murine PH induced by chronic hypoxia.…”

“…Ephx2 encodes sEH mRNA. The sequences of primers for Cyp2c38 and Cyp2c29 were listed previously (13). The forward primer for Ephx2 is 59-ACCACTCATGGAT GAAAGCTACA-39, and the reverse primer is 59-TCAGGTAGATTGGCTCCACAG-39.…”

Anti-inflammatory ω-3 endocannabinoid epoxides

“…Human CYP2J2 epoxygenates AEA and 2-arachidonoylglycerol (2-AG) (28) and is physiologically relevant, because it is the second most highly expressed P450 in the human brain (29) and is the most highly expressed P450 in human cardiomyocytes (30). However, the cross-species studies are not completely translatable, because BV-2 microglial cells are derived from mice, which have 10 subfamily CYP2J isozymes (CYP2J-5, -6, -7, -8, -9, -11, -12, -13, -14, -15) but lack a comparable CYP2J2 isozyme, making comparative species studies more difficult (31). Importantly, we show that coincubation of BV-2 microglia with a broad inhibitor of CYP enzymes (ketoconazole) significantly reduces the levels of the EEQ-EA and EDP-EA regioisomers ( Fig.…”