The Expression And The Role In Endoplasmic Reticulum Stress Of Calcium-Sensing Receptor In Rats With Pulmonary Hypertension Induced By High Pulmonary Blood Flow

Part ⅠEstablishment of Rat Models of Increased Blood Flow-induced Pulmonary Artery HypertensionObjective:To establish two rat models of increased blood flow-induced pulmonary artery hypertension using the left lung pneumonectomy and abdominal aorta-inferior vena cava shunting (A-V) procedures, and to compare the changes of hemodynamics and pulmonary vascular morphology in the two models so as to determine the optimal model for further studies.Methods:One hundred and sixty SD rats were randomly divided into three groups:control group (n=40),the left lung pneumonectomy group(PE,n=60) and the abdommal aorta-inferior vena cava shunting group(A-V,n=60). Each group was further divided into four subgroups based on the time points:Week1,Week 2, Week4and Week6. Ten rats were assigned in each subgroup of control group and fifteen rats were assigned in each subgroup of PE and A-V. At each time point,mean pulmonary artery pressure (mPAP) was detected; the right ventricle and left ventricle plus septum were separated and weighted for calculation the index of right ventricular hypertrophy (RVHI=RV/(LY+SP); Lung tissues were collected for HE staining.The pathomorphological changes of the small and medium-sized pulmonary arteries were observed,and the percentage of pulmonary vascular wall thickness (WT%) and the percentage of wall arae (WA%) were evaluated.Results:①The numbers that rats died during the process of modeling establishment was significantly greater in the PE group than that in the A-V group. Bleeding and heart failure were the major reasons of death for the PE group.②The trend of variations of mPAP in the A-V and PE groups were similar: quickly increased at Week1, decreased to the lowest at Week2, and gradually started to go up from Week4.Compared to the Control group, mPAP in the PE group was significantly increased at each time point (Week1, Week2, Week4and Week6)(P<0.01); mPAP in the A-V group was significantly increased at Week4and Week6(P<0.01). Compared to the A-V group, mPAP in the PE group was significantly increased at each time point (P<0.05).(3)RVHI in both the A-V and PE groups were gradually increased over time. Compared to the control group, RVHI (right ventricular hypertrophy index) in both the A-V and PE groups were significantly increased at each time point (P<0.05). Compared to the A-V group, RVHI in the PE group was significantly increased at Week1, Week2and Week4(P<0.05).(4)WT%in both the A-V and PE groups were gradually increased over time.Compared to the Control group, WT%in both the A-V and PE groups were significantly increased from Week2(P<0.01or P<0.05).Compared to the A-V group, WT%in the PE groups was significantly increased at Week2, Week4and Week6(P<0.01or P<0.05).(5)WA%in both the A-V and PE groups were gradually increased over time. Compared to the Control group, MA%in both the A-V and PE groups were significantly increased from Week2(P<0.01or P<0.05). Compared to the A-V group, WA%in the PE groups was significantly increased from Week4(P<0.01).Conclusions:①Both PE and A-V rat models demonstrated the pathophysiologic characteristics of congenital heart disease-related pulmonary hypertension, showing clinical manifestations of pulmonary hypertension and right ventricular hypertrophy, and histopathological changes of increased media membrane wall thickness and increased numbers of muscularized pulmonary small arteries. Both of the two models could be used as effective and reliable animal models for studies of congenital heart disease-related pulmonary hypertension.(2)Compared to the A-V model, the PE model showed several advantages:more significant changes of hemodynamics, more obvious manifestations of pulmonary vessel remodeling, and less time required for establishment.Part ⅡThe Expression Of The Calcium-Sensing Receptor In The Pulmonary Hypertension Induced By High Pulmonary Blood FlowObjective:The observation of the calcium-sensing receptor (CaSR) in the expression of high pulmonary blood flow pulmonary hypertension in rat pulmonary arterial smooth muscle, and to explore the mechanism of its role in the formation of pulmonary hypertension in high pulmonary blood flow.Methods:The rat model of pulmonary hypertension was established by resect left lung. Twenty-seven SD rats were randomly divided into3groups (n=9), the control group, the surgery group, the surgery plus calcium-sensing receptor blockers Calhex231group. Thirty-five days later, respectively, the rats mean pulmonary artery pressure (mPAP) were measured, the right ventricular/body weight (right ventricle/body weight, RV/BW) and the right ventricle/left ventricle plus septum (right ventricle/left ventricle plus septum, RV/(LV+S)) ratios were calculated too. The expression of the CaSR mRNA in the rat pulmonary artery in each group was detected by real-time quantitative PCR. The changes in the pulmonary artery structure were observed by the HE staining.Results:①Thirty-five days after surgery, mean pulmonary artery pressure (MPAP)、right ventricle/body weight(RV/BW) and the right ventricle/left ventricle plus septum ratios (right ventricle/left ventricle plus septum, RV/LV+S) was significantly higher than in the control group(P<0.05); Compared with the surgery group, MPAP、RV/BW and RV/(LV+S)was significantly lower in the surgery plus Calhex231group (P<0.05).②Compared with the control group, the CaSR mRNA expression of the pulmonary artery significantly increased in the surgery group (P<0.05); Compared with the surgery group, the CaSR mRNA expression of the pulmonary artery in the surgery+Calhex231group significantly reduced (P<0.05).Conclusions:High pulmonary blood flow increased the CaSR expression; The CaSR was involved in the process of high pulmonary blood flow in pulmonary hypertension by inducing pulmonary vascular proliferation and reconstruction.PartⅢThe Role Of Calcium-Sensing Receptor In Endoplasmic Reticulum Stress In Rats With Pulmonary Hypertension Induced By High Pulmonary Blood FlowObjective:As the important pathophysiological element in many cardiovascular diseases, endoplasmic reticulum stress (ERS) is recognized to be important in recent years. The previous studies demonstrated that the CaSR was able to increase the degree of tissue damage through activated endoplasmic reticulum stress, but the relationship and their specific mechanism between CaSR and ERS has not studied in the progression of pulmonary hypertension induced by high pulmonary blood flow. The experiment was designed to research the effect of the CaSR on ERS and its mechanisms in the rats of pulmonary hypertension induced by high pulmonary blood flow in rats.Methods:The rat model of pulmonary hypertension was established by resect left lung. Twenty-seven SD rats were randomly divided into3groups (n=9), the control group, the surgery group, the surgery plus calcium-sensing receptor blockers Calhex231group. Thirty-five days later, the apoptosis of pulmonary artery endothelium cells was detected by the TUNEL techno logy, the expression of ERS marker proteins glucose-regulated protein78(GRP78), caspase-12and the eukaryotic translation initiation factor2(p-eIF2a) were measured in the pulmonary artery by immunohistochemistry staining method, and semi-quantitative analysis the results by using the automatic image analysis system.Results:①Thirty-five days after surgery, apoptosis present in the pulmonary artery endothelium cells in every group.Compared with the control group, the apoptosis of pulmonary artery endothelium cells in the surgery group was significantly increased (P<0.05); Compared with the surgery group, the apoptosis of surgery phus Calhex231pulmonary artery endothelium cells was obviously reduced (P<0.05).②Compared with the control group, the expression of GRP78,caspasel2and p-eIF2a in the pulmonary artery were significantly higher in the surgery group (P<0.05);The expression of GRP78, caspasel2and p-eIF2a in the surgery plus Calhex231group was lower than the surgery group obviously(P<0.05).Conclusions:In rats with pulmonary hypertension induced by high pulmonary blood flow due to the left lung pneumonectomy for thirty-five days, endoplasmic reticulum stress in pulmonary artery was significantly enhanced. By activating excessive endoplasmic reticulum stress,CaSR promoted pulmonary hypertension induced by high pulmonary blood flow and pulmonary vascular remodeling formation.