| BackgroundHypertension is one of the cardiovascular diseases which threaten the health of human being. It is an important risk factor of many diseases such as coronary heart disease, cerebrovascular and renal diseases, and even cause of death. Prevention and treatment of hypertension and its complications have an important clinical significance. Hypertension is the common result of both hereditary and environmental factors, which, when accumulates to a suitable extent, cause the abnormal genetic expression and elevated blood pressure. But the mechanisms of molecular biology are still unknown for most patients with hypertension.Most of CYP enzymes are involved in the metabolism of drugs. However, it has become increasingly clear that P-450 metabolites of arachidonic acid play a crucial role in the control of cardiovascular function. Cytochrome P450 epoxgenase 2C11 (CYP2C11) and hydroaylase 4A1 (CYP4A1) are expressed in the vascular endothelial and smooth muscle (VSM) cells, myocardium and kidney, and metabolize arachidonic acid to biologically active epoxyeicosatrienoic acid (EETs) and 20-hydroxyeicosatetraenoic acid (20-HETE), which play a role in regulating vascular tone and blood pressure. EETs are endothelium-derived vasodilators thathyperpolarize VSM cells by activating Kca (Ca2+-sensitive potassium channel). EETs can be further metabolized to the corresponding dihydroxyeicosatrienoic acids (DHETs) that also can diastole vessel. 20-HETE is a vasoconstrictor produced in VSM cells that reduces the open-state probability of Kca.Statins have some effects independent of cholesterol-lowering, and researches of the mechnisms suggest that atorvastatin may have helpful effects on the control of hypertension. We hypothesized that the mechanisms by which atorvastatin affected blood pressure could be as follow: atorvastatin may up-regulate the expression of CYP 2C11 mRNA and protein and increase the production of EETs, and/or atorvastatin may down-regulate the expression of CYP 4A1 mRNA and protein in SHR, which may in turn, at least partly, improve the imbalance between vasoconstrictive and vasodilative factors and avoid the following development of hypertension.The effects of long over-loading pressure on the vascular endothelium and myocardium in spontaneously hypertensive rats (SHR) are similar to those in patients with essential hypertension. Therefore, the study of effects and mechanisms of atorvastatin on SHR will contribute to supplying useful experimental evidence for human.ObjectivesIn the present study, we investigated the effects of atorvastatin on the expression of CYP2C11 and CYP4A1 mRNA and protein in heart, liver, kidney, and aorta, on the urinary 14,15-DHETs excretion, and on the LVWI in SHR, observed renal pathological changes and explored the underlying mechanisms by which atorvastatin affects blood pressure.MethodsEighteen healthy eight-week-old male SHR with body weight 180-200g were randomly divided into three groups: SHR control group (n=6), 50 mg atorvastatin group (HATV group, 50 mg·kg-1·d-1, n=6), and 10 mg atorvastatin group (LATVgroup, 10 mg·kg-1d-1 , n=6). Six age- and weight-matched male Wistar-Kyoto (WKY) rats were selected as normal control group (WKY group, n=6). Atorvastatin was resolved in 1ml saline and was administered to HATV and LATV groups by gavage for 10 weeks. Rats in WKY group and SHR control group were given the same amount of saline by gavage for 10 weeks. Systolic blood pressure (SBP) was measured before and after treatment with atorvastatin every 2 weeks. At the end of treatment, the rats were anesthesiated by intraperitoneal injection of 2% sodium pentobarbital, 2 ml arterial blood was drawn for the determination of serum concentrations of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C). Left ventricle isolated along ventricle septum (ventricle septum included), wet weight of left ventricle to body weight ratio was calculated as left ventricle weight index (LVWI). The renal specimen were fixed with 10% formalin, embedded in paraffin wax, and stained with hematoxylin and eosin (HE). The pathologic changes of kidney were observed with the light microscope. The heart, liver, kidney, and thoracic aorta of rats were rapidly excised, weighed, sectioned and snap frozen in liquid nitrogen and then stored at - 70 ℃. The expression of CYP2C11 and CYP4A1 mRNA and protein in heart, liver, kidney, and aorta in the four groups were detected by RT-PCR and Western blot analysis, respectively. Urinary 14,15-DHETs excretion was quantified by ELISA method.Statistical analysis was performed with the Statistical Package for the Social Sciences (SPSS version 11.5). Data are presented as mean±SD. Comparisons between more than two groups were analyzed by One-Way ANOVA. A probability level of P<0.05 was considered significant.ResultsDuring the experimental period, we found that: 1. Comparison between the SHR control group and WKY group: (1) SBP in SHR control group was elevated, while WKY group had no significant changes in the same time period.(2) Compared with WKY group, LVWI in SHR control group were much higher.(3) The expression of CYP2C11 gene in heart, kidney, and aorta were significantly increased in SHR control group when compared with WKY group.(4) Compared with WKY group, the expression of CYP4A1 gene in heart, kidney, and aorta were significantly increased in SHR control group.(5) Urinary 14,15-DHETs excretion in SHR control group was much higher than that in WKY group.(6) In SHR control group, serum TC, TG, LDL-C, and HDL-C level decreased, all of which reached statistical significance when compared with WKY group.(7) The renal tissue of WKY group was normal. In SHR control group, light microscopy revealed enlargement of glomerular capsule, crenation of capillary loop, thickness of renal arterial walls and stenosis of lumina.2. Comparison of the HATV and LATV groups and the above two groups:(1) Compared with SHR control group, SBP significantly decreased in HATV group at 6, 8 and 10 weeks of treatment and in LATV group merely at 10 weeks of treatment. However, compared to WKY group, the blood pressure level in HATV and LAYV groups was still higher. Compared with LATV group, the levels of blood pressure in HATV group were significantly decreased at 8 and 10 weeks of treatment.(2) A significant reduction in LVWI was found in HATV group when compared with SHR control group. However, compared with WKY group, the LVWI in HATV group was still higher.(3) Compared with SHR control group, the expression of CYP 2C11 mRNA in heart, liver, kidney and aorta of HATV group and in heart of LATV group was markedly increased. The expression of CYP 2C11 protein in four tissues of HATV group and heart, kidney and aorta of LATV group was significant higher than that in SHR control group. The expression levels of CYP 2C11 mRNA and protein in kidney reached the statistical significance between HATV and LATV groups.(4) The expression of CYP 4A1 mRNA was significant decreased in heart, kidney and aorta of HATV group and in kidney and aorta of LATV group versus SHR control group. The expression of CYP 4A1 protein in kidney and aorta of HATV group and in kidney of LATV group was markedly lower than that in SHRcontrol group. The expression levels of CYP 4A1 mRNA and protein in kidney reached the statistical significance between HATV and LATV groups.(5) After 10 weeks of treatment, urinary 14,15-DHETs excretion in HATV group was much higher than that in SHR control group.(6) Compared with SHR control group, there was a significant descent in serum TC, TG and LDL-C concentrations in HATV group, and only a decrease in LDL-C in LATV group, and HDL-C concentrations were not significantly different in the three groups of SHR.(7) The pathological abnormalities of kidney were improved in HATV group compared to SHR control group. Light microscopy of HATV group revealed slight crenation of capillary loop of glomerulus and unobvious thickness of renal arterial walls. The above abnormalities of light microscopy in LATV group were unobviously improved compared to SHR control group.Conclusions1. Administration of atorvastatin to SHR not only reduced the serum lipids levels effectively, but also prevented the development of hyertension in SHR. Within some range, the high dose of atorvastatin affected more obviously on the blood pressure in SHR than lower dose.2. Atorvastatin significantly up-regulated the expression of CYP 2C11 gene in heart, kidney, and aorta of SHR, and increased the production of 14,15-DHETs. Also, atorvastatin down-regulated the expression of CYP 4A1 gene in kidney and aorta of SHR, which might be, at least partly, the mechanism of the favorable effects of atorvastatin in regulating blood pressure.3. Atorvastatin improved myocardial hypertrophy and renal morphological changes in SHR, which play an important role in the prevention of vicious circle in hypertention. |
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