The Effects Of Transection Of The Cervical Sympathetic Trunk On Monocrotaline-induced Pulmonary Arterial Hypertension In Rats And Its Possible Mechanism

PART Ⅰ EXPLORATION OF THE OPTIMAL INTERVENTION TIME OF TCST ON PAH RATSObjective: Excessive activation of sympathetic nerve has been proved to be one of the reasons for pulmonary arterial hypertension(PAH).This part mainly detects whether transection of the cervical sympathetic trunk(TCST)can inhibit the increase of pulmonary artery pressure in rats,and explores the optimal intervention time.Methods: 40 male SD rats were randomly divided into 7 groups: 10 rats in monocrotaline(MCT)group and 30 rats in the other groups(each group of 5 rats).MCT was intraperitoneally injected into rats at a dose of50 mg/kg.The control group was given intraperitoneal injection of the same amount of saline.The rats in the Control + TCST group received TCST on the day of saline injection.The other four groups were treated with TCST on the day of MCT injection,and on the 3th,7th and 14 th day after MCT injection.Then the right ventricular systolic pressure(RVSP)and mean pulmonary arterial pressure(m PAP)were measured by right heartcatheterization on 21 day(D21)after MCT administration.Results: The RVSP and m PAP in MCT group were significantly higher than those in Control group(RVSP: 58.31 ±4.94 mm Hg vs.29.52±2.25 mm Hg;m PAP: 33.20 ±3.13 mm Hg vs.17.46 ±1.05 mm Hg;P < 0.01),suggesting that PAH modeling was successfully developed in rats with MCT administration.Compared with the MCT group,RVSP and m PAP was decreased in varying degrees in MCT+TCST groups,and the earlier the time of receiving TCST,the greater the decrease.In addition,there was no significant change in RVSP and m PAP in Control+TCST group compared with Control group.Conclusions: The animal model of PAH is successfully established by intraperitoneal injection of MCT in rats.The prognosis of rats with PAH was related with TCST operational time,the earlier TCST started,the better effect was.PART Ⅱ TCST DELAYS THE PROGRESSION OF PAH AND RIGHT HEART FAILURE IN RATSObjective: Studies have shown that PAH patients are often accompanied by elevated norepinephrine(NE)level in circulating blood,and NE levels are associated with prognosis.We speculate that TCST may delay the progression of PAH by blocking the sympathetic nerve innervating the pulmonary artery and then reducing the concentration of NE in lungs.In this part,the rats were divided into four groups and treated with differttent experimentation accordingly,and then the data related to hemodynamics,histomorphology and tissue protein expression were detected to verify our hypothesis.Methods: Rats were randomly divided into four groups,including a control group,an MCT group,an MCT + sham group and an MCT + TCST group.After performing haemodynamic and echocardiographic measurements,the rats were sacrificed for the histological study,and the NE concentrations and protein expression level of tyrosine hydroxylase(TH)were evaluated.The protein expression levels of extracellular signal-regulated kinase(ERK)-1/2,proliferating cell nuclear antigen(PCNA),cyclin A2 and cyclin D1 in pulmonary artery vessels were determined.Results: Compared with the MCT + sham group,TCST profoundly reduced the m PAP(22.02 ± 4.03 mm Hg vs.31.71 ± 2.94 mm Hg),RVSP(35.21 ± 5.59 mm Hg vs.48.36 ± 5.44 mm Hg),medial wall thickness of the pulmonary artery(WT%)(22.48 ± 1.75% vs.46.10 ± 3.16%),and right ventricular transverse diameter(RVTD)(3.78 ± 0.40 mm vs.4.36 ± 0.29mm)and increased the tricuspid annular plane systolic excursion(TAPSE)(2.00 ± 0.12 mm vs.1.41 ± 0.24 mm)(all P<0.05).The NE concentrations and protein expression levels of TH were increased in the PAH rats but significantly decreased after TCST.Furthermore,TCST reduced the increased protein expression of PCNA,cyclin A2 and cyclin D1 induced by MCT in vivo.Conclusions: TCST alleviates pulmonary artery remodeling and right heart failure induced by MCT in PAH rats.PART Ⅲ NE PROMOTES PASMCS PROLIFERATION VIA ERK-1/2 PATHWAYObjective: The results of second part of the experiment showed that pulmonary artery proliferation in PAH rats,accompanied by an increased concentration of NE in lung tissue,which was inhibited by TCST.Thereby it is presumed that NE may participate and promote pulmonary artery proliferation.In this part,NE and related inhibitors were used to stimulate rat pulmonary artery smooth muscle cells(PASMCs)to explore the mechanism of NE promoting the proliferation of PASMCs.Methods: Culture and identification of primary PASMCs in Rats.PASMCs were divided into four groups: Control group,NE group,NE +Pra group and NE + U0126 group.Ed U was used to detect the proliferation of cells;western blot was used to detect the expression of ERK,p-ERK,PCNA,cyclin A2 and cyclin D1 in each group.Results: The results of Ed U showed that the rate of positive cells in NE group was about twice as high as that in Control group(P < 0.05).However,the positive cell rate in NE+Pra and NE+U0126 group was significantly lower than that in NE group(P<0.05).Compared with the control group,the phosphorylation level of ERK-1/2 increased in NE group,while the expression of PCNA,cyclin A2 and cyclin D 1 increased,but decreased in NE+Pra and NE+U0126 groups(P<0.05).Conclusions: NE increase the phosphorylation level of ERK-1/2 in cytoplasm by activating the alpha-1 adrenergic receptor of PASMCs,thereby increasing the expression of PCNA,cyclin D 1 and cyclin A2 in nucleus and promoting the proliferation of PASMCs.