Study On The Phenotype And Mechanism Of Pulmonary Hypertension In NRAGE Knockout Mice

MAGE-D1(NRAGE),MAGE-G1 and NECDIN are members of MAGE II gene family.The latter two genes are involved in Prader-Willi syndrome(PWS)which includes growth retardation,gonadal dysgenesis.Some reports have shown that NRAGE can inhibit metastasis of tumor cells,NRAGE interacted with P75NTR can regulate the apoptosis of neuronal growth factor.In mice experiments,loss of NRAGE leads to a series of phenotypic changes such as obesity,fertility decline and a shortened life span.Here we work on the relationship of NRAGE and pulmonary arterial hypertension(PAH).We find that in knockout mice pulmonary artery blood pressure increases significantly and beyonds the normal level measured by echocardiography,all the same,that aortic blood pressure was significantly higher than wild type mice measured by the tail artery interventional method.Paraffin sections show that pulmonary artery and aortic artery wall become thickening and lung tissue edema occurrs in NRAGE knock out mice.Pulmonary arterial hypertension refers to elevated mean pulmonary arterial pressure>25mmHg at rest and>30mmHg during exercise.It is a very serious disease,increased pulmonary vascular resistance was caused by unnormal arterioles,and which results in sustained pulmonary artery pressure that leads to right heart failure.According to the clinical classification,PAH is further subclassified into familial(FPAH),idiopathic(IPAH)and associated(APAH)varieties.The role of BMP signaling pathway in pulmonary arterial hypertension pathogenesis has been identified.BMPR Ⅱ was believed to be main genetic basis of PAH,accounting for approximately 60%of FPAH and 10-30%of IPAH.BMPR Ⅱ interacts with BMPR Ⅰto be heterotetramer,it is a serine/threonine receptor kinase,which binds BMP proteins can phosphorylate downstream Smad 1/5/8,the abnormal function of BMPR Ⅱ makes serine/threonine receptor kinase inactivation which promotes the proliferation of pulmonary artery smooth muscle cells and finally leads to PAH.No matter in NEAGR knock out mice and in 293 cells which NRAGE has been interfered or in MEF cells isolated from mice,we find the expression of BMPR Ⅱreduced.BMP4 interacted with BMPR heterotetramer can promote BMP pathway to phosphorylate downstream Smadl/5/8 which is a kind of R-Smad.P-Smad 1/5/8 translocate to the nucleus,where,binds with other transcription factors regulate target gene expression together.In VSMC adenovirus was used to interfere NRAGE,BMP4 is used to induce phosphorylation,we find the phosphorylation of Smadl/5/8 downregulated.Moreover,Smad4 decreases more when we detect the expression of Smad proteins in MEF cells.Smad4 is a kind of Co-Smad which plays an important role in helping R-Smad transportation into the nucleus.It can not only regulate cell proliferation,differentiation and apoptosis but also be recognized as tumor suppressor genes involved in various inflammations,cancer related signaling pathways.We conclude that NRAGE is required for BMPR Ⅱ and Smad4 processing or stability.The down-regulation of BMPR Ⅱ and Smad4 leads to hypertension in NRAGE knockout mice.Thus,our research works on the relationship of NRAGE and BMPR Ⅱ,NRAGE and Smad4,and how NRAGE protect BMPR Ⅱ and Smad4 not to be degraded.By illuminating these relationships we can have a good knowledge of the role of NRAGE in PAH.So far PAH is still a problem and it is difficult to overcome for medical community.Our researches suggest that,in human,NRAGE plays an important role in PAH,moreover,we provide a new animal model in studying PAH and a strong basis for the development of new therapies.