Effect Of BMPR2 Signal Pathway In Pathogenesis Of Pulmonary Arterial Hypertension Complicated With Congenital Heart Disease

Background and ObjectiveThe incidence of congenital heart disease in live newborns was estimated from 6/1000 to 12/1000. Pulmonary arterial hypertension(PAH) is a major complication of congenital heart disease with left-to-right shunt. As determined by the level of pulmonary vascular resistance, the most severe form of Pulmonary arterial hypertension is Eisenmenger's syndrome, which is very dificult to treat. Pulmonary arterial hypertension has been widely investigated for decades. To date, the pathogenesis of Pulmonary arterial hypertension remains unclear, although many studies have focused on it. During the last few years, more and more evidences have demonstrated the role of bone morphogenetic protein receptor 2 in pathogenesis of pulmonary arterial hypertension. But previous research on BMPR2-associated signal pathway was mainly focused on idiopathic and familial pulmonary arterial hypertension, not on pulmonary arterial hypertension with congenital heart disease, which still needs to elucidate. Therefore, this study is designed to observe the changes of BMPR2 and its signal transmitting molecule BMP4, Smad1 in patients with pulmonary arterial hypertension complicated with ventricular septal defect and to investigate its relevance to apoptosis of pulmonary vascular cell and dysfunction of endothelial cell, aiming to explore the pathogenesis of pulmonary arterial hypertension and define a new target for the clinical treatment.Methods1. Sixty two patients underwent ventricular septal defect repaired were divided into 4 groups according to the mPAP measured during operation: group I with no PAH (mPAP<20mmHg, n=12), group II with slight PAH (2150mmHg, n=18).2. Protein expression and mRNA of BMP4,BMPR2 of each patient were measured by real-time fluorescence quantitative RT-PCR and Western Blot . Phosphorylated protein and mRNA of Smad1 were also measured.3. The pathomorphologic change was observed by HE stain.The changes of the vascular wall thickness/ vascular diameter (WT%)and vascular area/ total vascular area(WA%) were calculated.4.Apoptosis was measured by TUNEL and apoptosis index(AI) was calculated. The protein expression of Bax and Bcl-2 were measured with immunohistochemical stain. Survivin was measured by immunohistochemical stain and Western Blot.5. Before operation, 5mL venous blood was collected and plasma was separated. ET-1 and NO concentrations in plasma were measured with radioimmunoassay and nitric acid reductase.6. Then, sixteen patients with severe PAH were randomly divided into control group (n=8) and bosentan group(n=8) treated with bosentan, a nonselective endothelin receptor antagonist. A real-time fluorescence quantitative RT-PCR assay was applied to evaluate the effects of bosentan on the mRNA expression of BMP4, BMPR2 and Smad1.Results1. With the increasing of mPAP, the mRNA and protein expression of BMP4 increased in lung tissue of patients, while the mRNA and protein expression of BMPR2, Smad1mRNA and phosphorylated protein decreased. There were statistical significant differences between each two groups (P<0.05). BMP4, BMPR2 and Smad1 were positively correlated with mPAP (P<0.05). The results suggested the BMPR2 signal pathway in lung tissue of PAH with CHD patients was restrained.2. With the increasing of mPAP, pulmonary vascular remodelling appeared. The smooth muscle cells and endothelial cells in the tunica media of pulmonary vascular proliferated, the intima became thicken and collagen deposited in patients with CHD and PAH. Meanwhile, both WT% and WA% were gradually elevated. Statistic analysis showed that there was significant difference between each two groups(P <0.05). And there were significant correlations between BMP4,BMPR2,Smad1 and WT%, WA%(P <0.05).3.In moderate and severe PAH groups, expression of survivin and bcl-2 increased while expression of bax decreased markedly. Statistic analysis showed that there were significant differences between each two groups(P <0.05). And there were significant correlations between BMP4, BMPR2, Smad1 and survivin, bcl-2(P <0.05).4.Apoptosis was observed in the lung of all the patients. Apoptosis apparently decreased in the lung tissue of group III and group IV. The difference of AI among 4 groups was significant ( P<0.05). There were significant correlations between BMP4,BMPR2,Smad1 and AI(P <0.05).5. With the aggravation of PAH, ET-1 plasma concentration increased while NO plasma concentration decreased. Statistic analysis showed that there were significant differences among each two groups(P <0.05). There were significant correlation between ET-1 and mPAP(P <0.05) and significant negative correlation between NO and mPAP(P <0.05).6. Bosentan could inhibit the expression of BMP4mRHA in the lung tissue of patients with severe pulmonary arterial hypertension and promote the expression of BMPR2mRNA and Smad1mRNA. The difference was significant between the control group and treatment group(.P<0.05).Conclusion1.The BMPR2 signal pathway is restrained because of the decreased expression of BMPR2 and Smad1, in spite of the increased expression of BMP4 in pulmonary arterial hypertension with congenital heart disease.2.The restrained BMPR2 signal pathway might repress cell apoptosis by promoting the expression of survivin and bcl-2 and suggest a significant participation in pulmonary vascular remodeling in pulmonary arterial hypertension with congenital heart disease.3. BMPR2 signal pathway might be impacted by ET-1, Bosentan can improve the dysfunctional ET-1 and BMP signal pathways in pulmonary arterial hypertension with congenital heart disease.