Opposite Regulation of Cholesterol Levels by the Phosphatase and Hydrolase Domains of Soluble Epoxide Hydrolase

Abstract

Soluble epoxide hydrolase (sEH) is a bifunctional enzyme with two catalytic domains: a C-terminal epoxide hydrolase domain and an N-terminal phosphatase domain. Epidemiology and animal studies have attributed a variety of cardiovascular and anti-inflammatory effects to the C-terminal epoxide hydrolase domain. The recent association of sEH with cholesterol-related disorders, peroxisome proliferator-activated receptor activity, and the isoprenoid/cholesterol biosynthesis pathway additionally suggest a role of sEH in regulating cholesterol metabolism. Here we used sEH knock-out (sEH-KO) mice and transfected HepG2 cells to evaluate the phosphatase and hydrolase domains in regulating cholesterol levels. In sEH-KO male mice we found a ϳ25% decrease in plasma total cholesterol as compared with wild type (sEH-WT) male mice. Consistent with plasma cholesterol levels, liver expression of HMG-CoA reductase was found to be ϳ2-fold lower in sEH-KO male mice. Additionally, HepG2 cells stably expressing human sEH with phosphatase only or hydrolase only activity demonstrate independent and opposite roles of the two sEH domains. Whereas the phosphatase domain elevated cholesterol levels, the hydrolase domain lowered cholesterol levels. Hydrolase inhibitor treatment in sEH-WT male and female mice as well as HepG2 cells expressing human sEH resulted in higher cholesterol levels, thus mimicking the effect of expressing the phosphatase domain in HepG2 cells. In conclusion, we show that sEH regulates cholesterol levels in vivo and in vitro, and we propose the phosphatase domain as a potential therapeutic target in hypercholesterolemia-related disorders. Soluble epoxide hydrolase (sEH)2 is a member of the epoxide hydrolase family (EC 3.3.2.3) with broad distribution in human tissues (1). sEH has now been conclusively shown to metabolize endogenous fatty acid epoxides generated by CYP450 epoxygenases (2, 3). These fatty acid epoxides (such as epoxyeicosatrienoic acids) have been shown to possess a wide variety of biological effects, many of which are related to cardiovascular physiology (4, 5). Epoxyeicosatrienoic acids have been described as major components of the endothelium-derived hyperpolarizing factors and are generally known for their vasodilator effects (6). sEH inhibitors have been shown to lower blood pressure in several animal models and have been proposed as therapeutic options for the management of hypertension (7,8). Additionally, epidemiological studies link sEH polymorphisms with various human diseases, many of which are related to cardiovascular disorders. The R287Q polymorphism was found to be associated with subclinical atherosclerosis and coronary artery calcification in African-Americans and was described as an emerging risk factor for atherosclerosis (9, 10). Another variant (K55R) was associated with coronary heart disease in Caucasians (11). Interestingly, the R287Q variant was also found to be associated with increased levels of plasma cholesterol and triglycerides in familial hypercholesterolemia patients (12...

Soluble epoxide hydrolase deficiency alters pancreatic islet size and improves glucose homeostasis in a model of insulin resistance

“…Previous studies show that disruption of the Ephx2 gene as well as inhibition of sEH activity by selective inhibitors result in a significant shift of the epoxy-fatty acid to diol ratio (18,22,23). Physiological and biochemical insulin signaling data support our conclusion that decreasing sEH and thus increasing plasma levels of EETs provide beneficial effects on glucose regulation possibly through modulating insulin sensitivity.…”

“…Because this sEHI has high bioavailability and a long half life (21), the high doses used resulted in high blood and tissue levels of TUPS generating a chemical knockout. Previous studies showed that plasma levels of EETs are elevated in the presence of TUPS and in KO mice (22). In spite of the high doses of TUPS, no adverse effects were seen in the pathological analysis of various organs.…”

“…1D)]. These blood and tissue levels of sEHI increase epoxy-fatty acid to diol ratios in both plasma and tissue (22). Taken together, these findings indicate that KO mice exhibit efficient gene deletion and indicate that the sEH inhibitor is distributed to tissues.…”

“…Pharmacological inhibition was achieved by treating animals with selective sEH inhibitor (TUPS). This compound is an effective inhibitor of sEH (IC 50 of 5 and 3 nM for murine and human sEH, respectively) (22,23). The concentration of TUPS in blood and tissue extracts was determined at the end of the study.…”

The Role of Cytochrome P450 Epoxygenases, Soluble Epoxide Hydrolase, and Epoxyeicosatrienoic Acids in Metabolic Diseases

“…Consistent with plasma cholesterol concentrations, liver expression of b-hydroxy-b-methylglutaryl coenzyme A reductase, an enzyme integral for the production of cholesterol, was found to be ;50% lower in sEH knockout male mice. Furthermore, the sEHI AR9276 significantly lowered cholesterol in an animal model of abdominal aortic aneurysm (29).…”