The Expression Of Soluble Exposide Hydrolase In Human Coronary Atherosclerotic Lesions And Its Significance

Atherosclerosis (AS) is the most common coronary artery disease, which leads tomyocardial infarction and heart failure. The phenotypic change of smooth muscle cell (VSMC)and the consequencing cell proliferation and migration play an important role in the developmentof AS. Epoxyeicosatrienoic acids (EETs), products of arachidonic acid metabolism by CYP450enzyme, is known as an endothelium-derived hyperpolarizing factor. EETs can induce VSMCmembrane hyperpolarization and subsequent vessel dilation. They also have anti-inflammatoryeffect, thus, are considered to be endogenous protective factors on vascular diseases, such ashypertension and AS. Soluble epoxide hydrolase (sEH) is an enzyme to hydrolyze EETs to theircorresponding dihydroxyeicosatetraenoic acids and diminishes the biological activity of EETs. Itwas reported that sEH inhibitors suppressed the progression of atherosclerotic lesion in animalmodels. However, the expression and effect of sEH in human samples have not been reportedand the mechanism has not to be fully elucidated. In present study, we investigate the expressionof sEH in samples of human coronary atherosclerosis, and the underlying mechanism in vivo andin vitro.First, we utilized 97 cases of human coronary artery specimens, including 61 cases withatherosclerotic lesions and 36 control subjects. The samples were collected by the Department ofPathology, Shantou University Medical College during 2001 to 2010. All samples wereconformed by forensic autopsy and hematoxylin-eosin (HE) staining. The samples were dividedinto grade I, II, III or IV according to the degree of the lesions. The immunohistochemistrystaining with antibodies against sEH, SMα-actin, CD68, as well as Ponceau/Victoria blue (P/VB)were performed. The results showed that the positive staining of sEH was found in early stages(grade I or II) of the coronary atheromatous plaque, but not in the normal coronary artery. ThesEH positive cells were mainly located at the layers of media and thickened endothelium. Thecolocalization of sEH and VSMC marker suggested that sEH positive cells were VSMCs orderived from media layer. The high expression of sEH in coronary atherosclerotic plaque mayplay a role on promoting the development of AS, especially in the early lesions.To further define the role of sEH in the development of AS in vitro, we isolated andcultured VSMCs from SD rat thoracic aorta, the cells were treated with different concentrationsof platelet-derived growth factor BB (PDGF-BB). Results from western blot analysis showedthat sEH expression at protein level was increased in a concentration-depended manner; at themean time, the mRNA levels of the markers of VSMC differentiation, including SMα-actin,SM22αand Cnn1, were downregulated by PDGF-BB, detected by real-time quantitative PCR. This effect can be partially reversed by TUPS, a specific sEH inhibitors. Furthermore,overexpression of sEH by adenovirus (Ad-sEH) in VSMC could induce VSMC dedifferentiationand increase cell migration. Transwell cell migration assay showed that PDGF-BB treatment andAd-sEH infection could promote the migration of VSMC, which could be attenuated by thetreatment of TUPS.In vivo, we established a carotid artery injury model through wire-injury in mice. Both sEHknockout (sEH-/-) and wild-type (sEH+/+) control mice. After 2 weeks since injury, the carotidarteries on both sides were isolated and double immunostained with antibodies against sEH andmarker of VSMCs. The results revealed that the intensity of sEH expression and the thickness ofleft carotid artery wall (wire-injury side) were significantly higher than that on right side (16.64±1.24μm/11.58±0.67μm, P <0.001) in the sEH+/+ group, which was attenuated by thetreatment of sEH inhibitor. Furthermore, compared to sEH+/+ mice, the thickness of neointima ininjured carotid artery was significantly reduced in sEH-/- mice (13.67±0.86μm, P = 0.001;11.62±0.99μm, P <0.001).In summary, this study, at first time, reported that sEH protein level was high in the layersof media and neointima in human coronary atherosclerotic plaques. sEH positive cells weremainly derived from VSMC in stages of the coronary atherosclerotic plaque. sEH expressionwas also increased induced by PDGF-BB in cultured rat primary VSMC and wire-injury modelin vivo. Thus, our data suggested that high expression of sEH may be involved in smooth musclecell differentiation and migration during the development of AS. The results may provide a novelmechanism on the pathogenesis of AS, and a potential therapeutic target for the treatment of AS.