Second-hit and sexual dimorphism effects in PAH

PAH 的二次打击和性别二态性效应

• 批准号: 8516587
• 负责人: PRAVIN B SEHGAL
• 金额: $ 7.66万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8516587
• 关键词: Binding; Biological Assay; Blood Vessels; Cell Nucleus; Cell Size; Cell membrane; Cells; Characteristics; Chronic; Collection; Cytoplasm; Cytoplasmic Organelle; Data; Defect; Development; Dilatation - action; Disease; Endoplasmic Reticulum; Endothelial Cells; Estrogen Receptors; Estrogens; Evaluation; Family; Functional disorder; Gender; Genes; Golgi Apparatus; Guanosine Triphosphate Phosphohydrolases; Human; Hypoxia; Image; Incidence; Lesion; Lung; Lung diseases; Mediating; Membrane; Membrane Protein Traffic; Mitochondria; Morbidity - disease rate; Movement; Mus; Mutation; Nuclear; Oils; Paraffin; Pathogenesis; Patients; Penetrance; Pharmaceutical Preparations; Phenotype; Predisposition; Progressive Disease; Protein Isoforms; Proteins; Regulation; STAT5A gene; STAT5a Transcription Factor; Semilunar Bone; Sex Characteristics; Small Interfering RNA; Smooth Muscle Myocytes; Structural Protein; Structure of parenchyma of lung; Subcellular structure; Therapeutic; Thinking; Transfection; VWF gene; Vesicular stomatitis Indiana virus; Woman; abstracting; base; combinatorial; gene therapy; human RTN4 protein; insight; man; men; mitochondrial dysfunction; mortality; mutant; novel; overexpression; protein transport; pulmonary arterial hypertension; reconstruction; response; sexual dimorphism; trafficking

DESCRIPTION (provided by applicant): Pulmonary arterial hypertension (PAH) is a progressive disease with high morbidity and mortality. Mutations and/or haploinsufficiency of BMPRII is considered to underlie familial pulmonary arterial hypertension (FPAH) as well as many instances of idiopathic PAH (IPAH). However, disease development, even in FPAH with known mutations in BMPRII, has low (10-15%) penetrance leading to searches for candidate "second-hit" genes that might be involved. Additionally, there is a sexual dimorphism in this disease with the incidence approximately 2.5-fold higher in women as in men. The underlying bases for low penetrance and sexual dimorphism remain unknown. At the subcellular level, endothelial and smooth muscle cells in the lung vascular lesions of PAH have an increase in cell size ("megalocytosis"), characteristic cytoplasmic organellar changes (Golgi apparatus fragmentation, cystic dilatation of the endoplasmic reticulum) and an inability to handle intracellular trafficking of proteins. The present project is based on our unexpected discovery that a well-known transcription factor STAT5a "nongenomically" associates with the Golgi membranes in the cytoplasm of pulmonary vascular cells. siRNA mediated knockdown of STAT5a/b in primary HPAECs and HPASMCs in culture produced Golgi fragmentation, cystic dilatation of endoplasmic reticulum (ER), increased Nogo-B/RTN4 and atlastin-3 (ATL3)(respectively an ER structural protein and an ER-resident GTPase), lunate distortion of nuclei, reduced membrane trafficking and secondary mitochondrial dysfunction and fragmentation - all changes that we and others (e.g. Smith & Heath, 1977, 1979) have shown to occur in PAH - changes that we show can be blocked by overexpression of the ER/Golgi-resident GTPase atlastin-1. Also, double knockdown of STAT5a and BMPRII combinatorially inhibited ER to plasma membrane trafficking in ECs suggesting "second-hit" effects between the two. Remarkably, it is well known that STAT5 species are estrogen-responsive and are known to mediate a sexual dimorphism phenotype. In Specific Aim I we propose to investigate the occurrence of the subcellular STAT5a/b knockdown signature in cells in vascular lesions in PAH lung tissues, identify the particular cells involved, and investigate gender-based differences in these changes. In Specific Aim II we propose to investigate endothelial and smooth muscle cells in culture derived from PAH patients for the occurrence of the subcellular STAT5a/b knockdown signature and potential gender-based differences, the combinatorial effects of STAT5a/b knockdown and BMPRII changes, and the therapeutic potential of restoring wt STAT5a and atlastin isoforms in such cells. This project represents evaluation of a new paradigm in PAH with a focus on the newly discovered cytoplasmic organellar functions of STAT5a/b as potential "second-hit" targets with underlying sexual dimorphism contributing to the development of this disease.

描述（由申请人提供）：肺动脉高压（PAH）是一种高发病率和死亡率的进行性疾病。BMPRII的突变和/或单倍性不足被认为是家族性肺动脉高压（FPAH）以及许多特发性肺动脉高压（IPAH）病例的基础。然而，即使在BMPRII已知突变的FPAH中，疾病的发展也具有较低的外显率（10-15%），导致搜索可能涉及的候选“第二击”基因。此外，这种疾病还存在性别二态性，女性的发病率约为男性的2.5倍。低外显率和两性二态现象的潜在基础尚不清楚。在亚细胞水平上，PAH肺血管病变中的内皮和平滑肌细胞细胞大小增加（“巨细胞增多症”），细胞质细胞器改变（高尔基体破碎，内质网囊性扩张），无法处理细胞内蛋白质运输。目前的项目是基于我们意想不到的发现，一个众所周知的转录因子STAT5a“非基因组”与肺血管细胞细胞质中的高尔基膜相关。siRNA介导的原代hpaec和HPASMCs中STAT5a/b的敲低产生高尔基断裂，内质网（ER）囊性扩张，Nogo-B/RTN4和atlastin-3 (ATL3)（分别是内质网结构蛋白和内质网内GTPase）的增加，细胞核的月状扭曲，膜运输减少，继发性线粒体功能障碍和断裂-所有这些变化，我们和其他人（例如Smith & Heath， 1977）。1979年)的研究表明，这种变化可以通过ER/高尔基GTPase atlastin-1的过度表达而被阻断。此外，STAT5a和BMPRII的双敲低联合抑制内质网在ECs中的质膜运输，这表明两者之间存在“二次打击”效应。值得注意的是，众所周知，STAT5物种是雌激素反应性的，并且已知介导两性二态表型。