S U M M A R Y Epoxyeicosatrienoic acids (EETs) are cytochrome P450 metabolites of arachidonic acid, which function in the brain to regulate cerebral blood flow and protect against ischemic brain injury. EETs are converted by soluble epoxide hydrolase (sEH) to the corresponding inactive diol metabolites. Previous animal studies have indicated that sEH gene deletion or treatment with sEH inhibitors results in increased levels of EETs and protection against stroke-induced brain damage. To begin elucidating the underlying mechanism for these effects, we sought to determine the distribution, expression, and activity of sEH in human brain samples obtained from patients with no neurological changes/pathologies. Immunohistochemical analyses showed the distribution of sEH mainly in the neuronal cell bodies, oligodendrocytes, and scattered astrocytes. Surprisingly, in the choroid plexus, sEH was found to be highly expressed in ependymal cells. Vascular localization of sEH was evident in several regions, where it was highly expressed in the smooth muscles of the arterioles. Western blot analysis and enzyme assays confirmed the presence of sEH in the normal brain. Our results indicate differential localization of sEH in the human brain, thus suggestive of an essential role for this enzyme in the central nervous system. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials. (J Histochem Cytochem 56:551-559, 2008)

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...

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2-Amino-2-methyl-1-propanol (AMP™) is widely used as a neutralizer/pH stabilizer in personal care products (PCPs); however, the potential health implications of dermal AMP exposure remain to be fully elucidated. Consequently, an in-depth analysis was performed to determine if PCPs containing AMP pose an elevated risk in humans under the intended use conditions. Animal studies have shown that at high doses, oral AMP exposure could lead to liver steatosis; thus, this study focused on hepatotoxicity. Our assessment revealed that the derived margin of exposure (MoE) values for AMP-containing PCPs were above 100, indicating that dermal exposure to AMP is unlikely to present an elevated risk of hepatotoxicity. Further, mode of action (MOA) analysis was conducted to elucidate the potential mechanisms underlying the observed hepatotoxicity in animal studies. Our analysis proposed that AMP interferes with the CDP-choline pathway in hepatocytes via the inhibition of one or more enzymes integral to the pathway and/or the replacement of choline in the assembly of the phospholipid unit. Ultimately, these events halt the lipid export via very lowdensity lipoproteins, which can subsequently develop into fatty liver accompanied by hepatotoxicity and other pathological changes if AMP exposure persists at sufficiently high doses. MOA analysis corroborated that dermal exposure to AMP expected from use of PCPs is highly unlikely to result in toxicologically significant systemic concentrations of AMP and thus hepatotoxicity. We concluded that dermal exposure to AMP in PCPs is not anticipated to result in an increased risk of hepatotoxicity.