Endothelial GATA-6 Deficiency Promotes Pulmonary Arterial Hypertension

Abstract

Pulmonary arterial hypertension (PAH) is a chronic and progressive disease characterized by pulmonary vasculopathy with elevation of pulmonary artery pressure, often culminating in right ventricular failure. GATA-6, a member of the GATA family of zinc-finger transcription factors, is highly expressed in quiescent vasculature and is frequently lost during vascular injury. We hypothesized that endothelial GATA-6 may play a critical role in the molecular mechanisms underlying endothelial cell (EC) dysfunction in PAH. Here we report that GATA-6 is markedly reduced in pulmonary ECs lining both occluded and nonoccluded vessels in patients with idiopathic and systemic sclerosis-associated PAH. GATA-6 transcripts are also rapidly decreased in rodent PAH models. Endothelial GATA-6 is a direct transcriptional regulator of genes controlling vascular tone [endothelin-1, endothelin-1 receptor type A, and endothelial nitric oxide synthase (eNOS)], pro-inflammatory genes, CX3CL1 (fractalkine), 5-lipoxygenease-activating protein, and markers of vascular remodeling, including PAI-1 and RhoB. Mice with the genetic deletion of GATA-6 in ECs (Gata6-KO) spontaneously develop elevated pulmonary artery pressure and increased vessel muscularization, and these features are further exacerbated in response to hypoxia. Furthermore, innate immune cells including macrophages (CD11b+/F4/80+), granulocytes (Ly6G+/CD45+), and dendritic cells (CD11b+/CD11c+) are significantly increased in normoxic Gata6-KO mice. Together, our findings suggest a critical role of endothelial GATA-6 deficiency in development and disease progression in PAH. Pulmonary arterial hypertension (PAH) is a chronic and progressive disease characterized by pulmonary vasculopathy with elevation of pulmonary artery pressure, often culminating in right ventricular failure. GATA-6, a member of the GATA family of zinc-finger transcription factors, is highly expressed in quiescent vasculature and is frequently lost during vascular injury. We hypothesized that endothelial GATA-6 may play a critical role in the molecular mechanisms underlying endothelial cell (EC) dysfunction in PAH. Here we report that GATA-6 is markedly reduced in pulmonary ECs lining both occluded and nonoccluded vessels in patients with idiopathic and systemic sclerosis-associated PAH. GATA-6 transcripts are also rapidly decreased in rodent PAH models. Endothelial GATA-6 is a direct transcriptional regulator of genes controlling vascular tone [endothelin-1, endothelin-1 receptor type A, and endothelial nitric oxide synthase (eNOS)], pro-inflammatory genes, CX3CL1 (fractalkine), 5-lipoxygenease-activating protein, and markers of vascular remodeling, including PAI-1 and RhoB. Mice with the genetic deletion of GATA-6 in ECs (Gata6-KO) spontaneously develop elevated pulmonary artery pressure and increased vessel muscularization, and these features are further exacerbated in response to hypoxia. Furthermore, innate immune cells including macrophages (CD11b+/F4/80+), granulocytes (Ly6G+/CD45+), and dendritic cells (CD11b+/CD11c+) are significantly increased in normoxic Gata6-KO mice. Together, our findings suggest a critical role of endothelial GATA-6 deficiency in development and disease progression in PAH. Pulmonary arterial hypertension (PAH) is a severe vascular disorder characterized by an increase in resistance and blood pressure in the pulmonary artery (PA) or lung vasculature.1Simonneau G. Robbins I.M. Beghetti M. Channick R.N. Delcroix M. Denton C.P. Elliott C.G. Gaine S.P. Gladwin M.T. Jing Z.C. Krowka M.J. Langleben D. Nakanishi N. Souza R. Updated clinical classification of pulmonary hypertension.J Am Coll Cardiol. 2009; 54: S43-S54Abstract Full Text Full Text PDF PubMed Scopus (1847) Google Scholar The key pathological features of PAH occur in small PAs and include vasoconstriction, thrombosis, and inflammation, leading to intimal proliferation and fibrosis. Elevations in pulmonary vascular resistance and PA stiffness lead to pressure and volume overloading of the right ventricle, and eventually cause right heart failure and death. PAH is one of five groups within the pulmonary hypertension World Health Organization clinical classification system, which contains various forms including patients with idiopathic pulmonary arterial hypertension (IPAH) and familial pulmonary arterial hypertension (FPAH), and is associated with connective tissue diseases, primarily systemic sclerosis (SSc) pulmonary arterial hypertension (SSc-PAH).1Simonneau G. Robbins I.M. Beghetti M. Channick R.N. Delcroix M. Denton C.P. Elliott C.G. Gaine S.P. Gladwin M.T. Jing Z.C. Krowka M.J. Langleben D. Nakanishi N. Souza R. Updated clinical classification of pulmonary hypertension.J Am Coll Cardiol. 2009; 54: S43-S54Abstract Full Text Full Text PDF PubMed Scopus (1847) Google Scholar SSc-PAH has similar clinical and histopathological features such as IPAH; however, it is unclear whether the molecular mechanisms responsible for the pathogenesis of the two forms are similar.2Fisher M.R. Mathai S.C. Champion H.C. Girgis R.E. Housten-Harris T. Hummers L. Krishnan J.A. Wigley F. Hassoun P.M. Clinical differences between idiopathic and scleroderma-related pulmonary hypertension.Arthritis Rheum. 2006; 54: 3043-3050Crossref PubMed Scopus (273) Google Scholar The cellular and molecular processes underpinning pathological vascular remodeling in PAH are complex and involve phenotypic alterations in different cell types within the vascular wall with further contributions from the circulating immune and progenitor cells.3Chan S.Y. Loscalzo J. Pathogenic mechanisms of pulmonary arterial hypertension.J Mol Cell Cardiol. 2008; 44: 14-30Abstract Full Text Full Text PDF PubMed Scopus (201) Google Scholar Although the inciting events remain poorly defined, current theories suggest that PAH is initiated by the disruption of EC homeostasis leading to an imbalance of vasoactive factors and production of prothrombotic and pro-inflammatory mediators. Genetic studies have linked dysregulated bone morphogenetic protein signaling to the pathogenesis of FPAH and IPAH. Heterozygous mutations of the bone morphogenetic protein receptor type II (BMPR2) were found in 50% to 70% of FPAH cases and 11% to 40% sporadic of IPAH cases.4Lane K.B. Machado R.D. Pauciulo M.W. Thomson J.R. Phillips 3rd, J.A. Loyd J.E. Nichols W.C. Trembath R.C. The International PPH ConsortiumHeterozygous germline mutations in BMPR2, encoding a TGF-beta receptor, cause familial primary pulmonary hypertension.Nat Genet. 2000; 26: 81-84Crossref PubMed Scopus (1215) Google Scholar Furthermore, a subset of patients with hereditary hemorrhagic telangiectasia carrying mutations in activin receptor-like kinase or endoglin genes also develops PAH.5Mahmoud M. Borthwick G.M. Hislop A.A. Arthur H.M. Endoglin and activin receptor-like-kinase 1 are co-expressed in the distal vessels of the lung: implications for two familial vascular dysplasias, HHT and PAH.Lab Invest. 2009; 89: 15-25Crossref PubMed Scopus (42) Google Scholar Consistently, mice with the genetic ablation of Bmpr2 gene are predisposed to develop PAH.6Hong K.H. Lee Y.J. Lee E. Park S.O. Han C. Beppu H. Li E. Raizada M.K. Bloch K.D. Oh S.P. Genetic ablation of the BMPR2 gene in pulmonary endothelium is sufficient to predispose to pulmonary arterial hypertension.Circulation. 2008; 118: 722-730Crossref PubMed Scopus (174) Google Scholar In contrast to FPAH and IPAH, BMPR2 and activin receptor-like kinase 1 mutations, so far, were not found in patients with SSc-PAH, suggesting that other molecular mechanisms may underlie the disease process in SSc-PAH.7Morse J. Barst R. Horn E. Cuervo N. Deng Z. Knowles J. Pulmonary hypertension in scleroderma spectrum of disease: lack of bone morphogenetic protein receptor 2 mutations.J Rheumatol. 2002; 29: 2379-2381PubMed Google Scholar, 8Selva-O'Callaghan E. C.P. J. of kinase 1 are not found in patients with pulmonary hypertension and underlying connective tissue Rheumatol. 26: PubMed Scopus Google Scholar patients with SSc-PAH have disease and are to to patients with J. M. L. Hassoun P.M. sclerosis-associated pulmonary arterial PubMed Scopus Google Scholar In to in systemic and inflammation, from an of the to severe in patients with SSc-PAH, and are to to J. M. L. Hassoun P.M. sclerosis-associated pulmonary arterial PubMed Scopus Google Scholar gene that lung from patients with IPAH and SSc-PAH have further the role of in both forms of E. H. J. in patients with systemic sclerosis have gene to pulmonary and pulmonary hypertension.Arthritis Rheum. PubMed Scopus Google Scholar The has also and gene in in the of the cellular processes to the development of the role of ECs in the and progression of PAH poorly GATA-6 is one of GATA factors that are highly transcription factors with two that with other transcriptional and the The transcription factors and expressed of gene 2000; PubMed Scopus Google Scholar GATA-6 is expressed in a of and small it is to the of the cells within these E. T. J. L. J.R. The GATA-6 and of by and PubMed Scopus Google Scholar GATA-6 is expressed in quiescent vascular cells and may to the of the of M. M. of response and the GATA and the transcription of and 2002; PubMed Scopus Google Scholar, H. T. T. T. protein as a of GATA-6 in the transcription of the 2000; PubMed Scopus Google Scholar Furthermore, GATA-6 is rapidly decreased in in E. T. J. L. J.R. The GATA-6 and of by and PubMed Scopus Google Scholar and in T. Z. 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The molecular mechanisms underlying the remodeling process and in the EC remain we that of GATA-6 in ECs a key pathological during development of PAH. GATA-6 is reduced in as as cells of both occluded and nonoccluded pulmonary vessels of both IPAH and SSc-PAH in suggesting that of GATA-6 is not a of vessel remodeling, vessel and may an of EC dysfunction during PAH. is by from the two of the and the chronic that of the GATA-6 in the that are in GATA-6 remain to of Gata6-KO mice that GATA-6 is for pulmonary a of GATA-6 sufficient to PA pressure, not systemic blood may in to by in the of of vascular tone and The mice develop PAH normoxic and severe PAH R.C. The pulmonary of or mice is to Invest. 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