CYP2C44, a New Murine CYP2C That Metabolizes Arachidonic Acid to Unique Stereospecific Products

Abstract

The human CYP2Cs have been studied extensively with respect to the metabolism of clinically important drugs and endogenous chemicals such as arachidonic acid (AA). Five members of the mouse CYP2C family have previously been described that metabolize arachidonic acid into regio-and stereospecific epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetraenoic acids, which have many important physiological roles. Herein, we describe the cloning and characterization of a new mouse cytochrome P450 (P450), CYP2C44, which has the lowest homology with other known mouse CYP2Cs. Western blotting and real-time polymerase chain reaction detected CYP2C44 mRNA and protein in liver Ͼ Ͼ kidney Ͼ adrenals. Kidney contained approximately 10% of the CYP2C44 mRNA content of liver. CYP2C44 metabolized AA to unique stereospecific products, 11R,12S-EET and 8R, 9S-EET, which are similar to those produced by rat CYP2C23. CY2C23 is highly expressed in rat kidney and has been suggested to be important in producing compensatory renal artery vasodilation in response to salt-loading in this species. Immunohistochemistry showed the presence of CYP2C44 in hepatocytes, biliary cells of the liver, and the proximal tubules of the kidney. Unlike mouse CYP2C29, CYP2C38, and CYP2C39, CYP2C44 did not metabolize the common CYP2C substrate tolbutamide. CYP2C44 was not induced by phenobarbital or pregnenolone-16␣-carbonitrile, two prototypical inducers of hepatic P450s. The presence of CYP2C44 in mouse liver, kidney, and adrenals and the unique stereospecificity of its arachidonic acid metabolites are consistent with the possibility that it may have unique physiological roles within these tissues, such as modulation of electrolyte transport or vascular tone.

High Potassium Intake Enhances the Inhibitory Effect of 11,12-EET on ENaC

“…Moreover, the major product generated by CYP2C44-dependent AA metabolism is 11R,12S-EET. 8 In this regard, we have observed that 11R,12S-EET is a potent inhibitor of ENaC, whereas 11S,12R-EET at the same concentration failed to inhibit ENaC, suggesting 11R,12S-EET is responsible for mediating the effect of AA on ENaC.…”

“…8 Thus, we used patchclamp technique to examine the effect of 11S,12R-or 11R,12S-EET on ENaC in the CCD. Figure 1C is a typical channel recording made in a cell-attached patch demonstrating that application of 100 nM 11R,12S-EET (directly adding EET into the bath) completely inhibited ENaC (n ϭ 5).…”

“…15 CYP2C23 has been shown to be the major homologue of CYP epoxygenase in the rat kidney and is mainly responsible for forming 11,12-EET 8,20,21 Mouse CYP2C44 is an ortholog of rat CYP2C23 and shares 84% identical amino acid sequence. 8 We previously showed that CYP2C44 is expressed in the CCD 1 and that the expression is upregulated by either high-Na intake or HK intake. 2,7 The recombinant CYP2C44 expressed in Escherichia coli was able to metabolize AA to EETs.…”

The Cyp2c44 Epoxygenase Regulates Epithelial Sodium Channel Activity and the Blood Pressure Responses to Increased Dietary Salt

“…The murine Cyp2c44 epoxygenase and its rat homologue, CYP2C23 (polypeptides 23 and 44), share 88% amino acid sequence identity (20,32) and metabolize AA to predominantly 11,12-and 14,15-EET (20,32). Roles for these enzymes in BP regulation were indicated by the demonstration that their kidney expression was dietary salt-sensitive (21,26,33) and that hypertensive salt-sensitive Dahl rats showed reduced renal CYP2C23 epoxygenase expression and EET biosynthesis (33).…”

The Nuclear Receptors Constitutive Active/Androstane Receptor and Pregnane X Receptor Activate theCyp2c55Gene in Mouse Liver

“…However, the transcriptional regulation of murine Cyp2c genes is poorly understood at the present time. Fifteen murine Cyp2c genes have been identified (Luo et al, 1998;DeLozier et al, 2004;Nelson et al, 2004;Wang et al, 2004), including Cyp2c29, Cyp2c36, Cyp2c37, Cyp2c38, Cyp2C39, Cyp2c40, Cyp2c44, and Cyp2c55. The expression of the Cyp2c genes is regulated differentially (DeLozier et al, 2004;Jackson et al, 2004Jackson et al, , 2006Goetz et al, 2006).…”