Increase In Caveolin-1 Expression Enhance The Function Of Receptor-Operated Ca²⁺ Entry On Aortic Artery Of 2K1C Hypertensive Rats

Hypertension is the most important risk factor of cardiovascular disease.There are several hypotheses about the pathogenesis of hypertension at present.Among these,the imbalance of Ca2+ homeostasis in vascular smooth muscle cells(VSMCs)is he focus of research.Caveolae is widely distributed in the vascular system,including a variety of channel proteins related to the regulation of intracellular Ca2+ concentration,and plays an important role in the signal transduction of cell surface.Caveolin is an important structural protein of Caveolae,Among of it,Caveolin-1(Cav-1)is an primary regulator of intracellular Ca2+ concentration fluctuations,which has the greatest influence on vascular tension.Previous studies have shown that up regulation of Caveolae in the aorta of rats with pulmonary hypertension can affect the function of non selective cation channels.Our study constructed HPA model rats by two kidney one clip(two-kidney one-clip 2K1C)surgery,through narrowing renal artery on the left renal.Our study compared the expression change of Cav-1 and TRPC1/6,which associated with the non selective cation channel.We observed the effect of Caveolae on the aorta contraction induced by agonist in rats by using Methyl-?-cyclodextrin(M?CD)to destroy the structure of Caveolae.We treated AoSMCs with specific enhancing polypeptide of Cav-1,and confirmed the effect of up regulation of Caveolae on agonist induced rat AoSMCs increased intracellular Ca2+ concentration further.Our study also conducted experiments in the resistance arteries,and compared with the results of the aorta.Objective to explore the role of Caveolae in the pathogenesis of HPA,and find out more experimental ideas and theoretical basis for the study of the pathogenesis and treatment of HPA.Objective: To observe the effect and mechanism of Caveolae and Cav-1 in the aortaand the third-order branches of the mesenteric arteries of HPA rats induced by 2K1 C,and to provide a new scientific basis for the study of HPA pathogenesis and targeted therapy.Methods: To establish HPA rat models by performing 2K1 C surgery on SD rats.(1)Using hemodynamics and electron microscopy examination methods to measure the variation of rats systemic systolic blood pressure and diastolic blood pressure and the number of Caveolae on AoSMCs;(2)Using Real-time Q-PCR and Western blot methods to measure the expression levels of Cav-1 and TRPC1/6 mRNA or protein in aorta and the third-order branches of the mesenteric arteries of rats;(3)Using vascular circle tone detection methods to observe the influence of various agonists-induced aorta contraction by destroying Caveolae on AoSMCs by M?CD in rats;(4)Using DMT micro vessel tension detection methods to observe the influence of various agonists-induced the third-order branches of the mesenteric arteries contraction by destroying Caveolae by M?CD in rats;(5)Using Fluo-3 fluorescence detection [Ca2+]i method to research the influence of specific enhancing polypeptide of Cav-1 on CPA/OAG-induced Ca2+ entry in AoSMCs of CON and 2K1 C rats.Results: Compared with normal group:(1)The SBP and DBP of 2K1 C model rats were both significantly increased,suggesting that the successful preparation of the 2K1 C model rats.The number of Caveolae on AoSMCs of rats were observed under the transmission electron microscope,there were more Caveolae on AoSMCs of 2K1 C rats;(2)The expression level of Cav-1 and TRPC1/6 mRNA and protein on aorta of 2K1 C model rats were significant increased,suggesting that the expression of Cav-1 was up-regulated and the non voltage dependent calcium channel function was enhancement in aorta of 2K1 C model rats.(3)The effect of KCl-induced contraction on aortic of 2K1 C rats was greater.According to our research,voltage-dependent calcium channel(VDCC)has no closed relationship with Caveolae on aorta of rats,because it had slight effect on KCl-induced aorta contraction by destroying Caveolae with M?CD;(4)The effect of ET-1-induced contraction on aortic of 2K1 C rats was greater.The inhibitory effect of M?CD on ET-1 induced aortic contraction was more obvious,suggesting that the non-voltage-dependent calcium channel was closely correlated with the Caveolae on aorta of rats;(5)Exogenous Chol was able to reverse the inhibitory effect of M?CD on agonist-induced contraction,and confirmed that M?CD destroy the Caveolae integrity by exhaust Chol;(6)OAG and CPA-induced Ca2+ resting and entry level was increased on AoSMCs of rats by using specific enhancing polypeptide of Cav-1,proved that ROCE and SOCE were significantly increased in 2K1 C rats.CPA-induced Ca2+ entry enhancement degree was increased on AoSMCs of rats by using specific enhancing polypeptide of Cav-1,suggesting that Caveolae and Cav-1 on aortic of rats not closely related to SOCE;(7)OAG-induced Ca2+ entry enhancement degree was increased on AoSMCs of rats by using specific enhancing polypeptide of Cav-1,proved that enhanced ROCE was closely related with up-regulated Caveolae in AoSMCs of 2K1 C rats from the cell level;(8)The mRNA expression level on the third-order branches of the mesenteric arteries of rats showed no significant difference between CON and 2K1 C groups,suggesting that Caveolae and Cav-1 on the third-order branches of the mesenteric artery not closely related to non voltage dependent calcium channels;(9)KCl-induced constriction on the third-order branches of the mesenteric arteries of2K1 C rats was enhanced,but not closely related to Caveolae;(10)The experiment on the third-order branches of the mesenteric arteries of rats showed no significant difference in 2K1 C groups,suggesting that Caveolae and Cav-1on the third-order branches of the mesenteric artery not closely related to non voltage dependent calcium channels in 2K1 C groups;Conclusion: Caveolae and Cav-1 are widespread in VSMCs,and the up-regulation of Cav-1 and ROCE in aortic of 2K1 C rats may be one of the pathogenesis of HPA.However,the changes of Caveolae function in the third-order branches of themesenteric arteries of rats did not have significant influence on the calcium channel.In our paper,we will reveal the role of Caveolae in 2K1 C model HPA rats and research the relationship with Ca2+ channel,which will provide a new approach for the study of the pathogenesis of hypertension and the breakthrough of therapeutic pathway.