The Study Of Therapeutic Effect And Mechanism Of STVNa On Right Ventricular Hypertrophy And Pulmonary Vascular Remodeling Induced By Transverse Aortic Constriction In Rats

Pulmonary artery hypertension is a severe cardiovascular disease characterized by remodeling of pulmonary vascular structures and progressive elevation of pulmonary arterial pressure,which eventually leads to overloading of the right ventricle and even death.The incidence of pulmonary hypertension caused by left heart disease is highest in clinic,but the existing drugs have limited therapeutic effect on them.Large-scale long-term clinical studies have demonstrated that cyclic prostaglandin analogues and endothelin receptor inhibitors don’t display a good therapeutic effect on pulmonary hypertension caused by left heart disease.Therefore,there is an urgent need to develop new therapeutic drugs for pulmonary hypertension caused by left heart disease.It has been reported that Isostereol has anti-inflammatory,anti-oxidative,anti-proliferative and other active effects as well as protective effects on cardiovascular diseases.As a sodium salt of isosteviol,our laboratory previously studied the therapeutic effect of sodium steviol sodium（STVNa）on monocrotaline-induced pulmonary hypertension in rats.Because monocrotaline causes pulmonary artery pressure to increase by destroying pulmonary vascular endothelial cells,and most patients with pulmonary hypertension in clinical origin are from left heart disease,the pathogenesis of the two is different.Therefore,it is of great significance to investigate the therapeutic effect and mechanism of STVNa on pulmonary hypertension caused by left heart disease.This study will investigate the therapeutic effects and possible mechanisms of STVNa by establishing a pulmonary hypertension model with rat transverse aortic constriction and an in vitro hypoxia-induced pulmonary artery smooth muscle cell proliferation model.The main contents are as follows:1.At present,there is no research report on the pathogenesis of pulmonary hypertension in rats caused by transverse aortic constriction at different time points.To answer this question,the hemodynamics and hypertrophy index of left and right ventricle and lung pathology analysis were performed respectively in 3,6,9 weeks after transverse aortic constriction in rats.And it will provide a standard for the establishment of animal models of pulmonary hypertension induced by left heart disease.The results showed that the mean left ventricular pressure,the left ventricular contraction rate and the left ventricular myocardial cross-sectional area increased after 3 weeks of transverse aortic constriction in rats,which indicated a compensatory increase in left ventricular function.The left ventricular end-systolic pressure and mean right ventricular pressure began to increase after 6 weeks of transverse aortic constriction,indicating that left cardiac function subjected to the right heart,and the right heart function was relatively enhanced.The Left ventricular end-systolic pressure and mean right ventricular pressure both peaked in 9weeks after transverse aortic constriction.The cross-sectional area of the right cardiac myocyte and the thickness of the middle layer of the pulmonary vascular increased significantly,which showed that the symptoms of pulmonary hypertension are typical,significant right heart hypertrophy and pulmonary vascular remodeling appear,and can be used for subsequent pharmacodynamic studies.2.There is no report on the therapeutic effect of STVNa on pulmonary hypertension induced by left heart disease.Using the animal model of rat pulmonary hypertension induced by left heart disease,this part of the experiment focuses on the efficacy of STVNa.The experimental results showed that intragastric administration of STVNa 8mg/kg/d and 4mg/kg/d in two doses showed a dose-dependent improvement in the hemodynamics of rats after 9 weeks of transverse aortic constriction.Compared with vehicle group,8mg/kg/d STVNa significantly reduced right heart hypertrophy index and significantly improved right ventricular cardiomyocyte hypertrophy and myocardial fibrosis,and also significantly reduced the middle vascular thickness and pulmonary fibrosis.In addition,8 mg/kg/d STVNa can effectively reduce the activation of mast cells and macrophages.And the mRNA and the protein expression level of tumor necrosis factor alpha（TNF alpha）and interleukin 1 beta（IL-1 beta）was significantly decreased in lung tissue compared with the vehicle group.8mg/kg/d STVNa also reduced NOX4 protein expression and hydrogen peroxide content in rat lung tissue.3.The previous section of this study has confirmed that STVNa can reduce the expression of NOX4 and hydrogen peroxide in lung tissue,but the mechanism of further inhibiting lung tissue remodeling is not clear,and pulmonary artery smooth muscle cell proliferation is important for pulmonary artery hypertension,so it is questionable whether STVNa can inhibit smooth muscle cell proliferation by inhibiting NOX4 expression.Therefore,this part of the experiment using the acute isolation of rat pulmonary artery smooth muscle cells,an in vitro hypoxia-induced cell proliferation model was estabilshed,and the inhibition of STVNa on hypoxia-induced pulmonary artery smooth muscle cell proliferation and related mechanisms was studied.The results showed that the optimal conditions of hypoxia were 24 hours in the hypoxia incubator with 94%N₂,1%O₂,and 5%CO₂.STVNa could inhibit the hypoxia-induced proliferation of pulmonary artery smooth muscle cells in a dose-dependent manner and the best dose was 50μM,which is also proved by results of the cellular immunofluorescence of proliferating cell nuclear antigen（PCNA）.STVNa may be able to reduce the hypoxia-induced increase in intracellular reactive oxygen species（ROS）by inhibiting the expression of NOX4,thereby reducing the phosphorylation of ERK1/2,which is the downstream of ROS and thus inhibiting cell proliferation.STVNa may regulate NOX4 expression by decreasing intracellular NF-κB phosphorylation.CONCLUSIONS:After 9 weeks of transverse aortic constriction,a stable left ventricular disease induced pulmonary hypertension model was obtained.It is proved that STVNa has a significant therapeutic effect on pulmonary hypertension induced by left heart disease,which may through inhibiting NF-κB phosphorylation,then reducing NOX4 expression and intracellular ROS levels,further reducing the phosphorylation of ERK1/2,and inhibiting the proliferation of pulmonary artery smooth muscle cells.