Session Type: ACR Poster Session C
Session Time: 9:00AM-11:00AM
Background/Purpose: Pulmonary arterial hypertension (PAH) is a progressive and fatal complication in the patients with connective tissue diseases (CTD). Elevation of endothelin-1 (ET-1) levels in pulmonary arteries is suggested as one of the pathogenesis of PAH, however, mechanism for ET-1 induction in the disease circumstances is still elusive. A driving force for ET-1 synthesis is hypoxia and ET-1 has been known as a target gene of hypoxia-inducible factors (HIFs). HIFs are heterodimeric transcription factors composed of α subunit (HIF-1α or HIF-2α or HIF-3α) and β subunit (HIF-1β), operating in gene regulation in adaptive response to hypoxic condition. We have demonstrated ablation of HIF-3α gene in mice resulted in an increase in ET-1 production by pulmonary endothelial cells and human PAH-like cardio-pulmonary phenotype such as right ventricular enlargement and pulmonary arteriole musculalizations, indicating an implication of HIF-3α in the pathogenesis of PAH. In the present study, we aimed to explore the role of genetic defect or anomalies of HIF-3α in the pathogenesis of CTD-PAH.
Methods: Using genomic DNA form patients with systemic sclerosis (SSc) without PAH, SSc complicated with PAH, and normal healthy controls, we analyzed the incidence of coding single nucleotide polymorphisms (SNPs) in HIF3A gene reported in NCBI dbSNP database by PCR-direct sequencing. The m×n χ2–test was applied to examine statistical significance of the incidence of the SNPs. Functional validations of the HIF-3α carrying the SNPs (SNP-HIF3α) were performed in the cells overexpressing SNP-HIF3α. Effect of SNP-HIF3α on ET-1 gene promoter was analyzed by reporter gene assays and chromatin immunoprecipitation assays.
Results: We found coding SNPs in HIF3A gene with significantly higher incidence in the SSc patients complicated with PAH compared to the SSc patients without PAH. Overexpression of HIF-3α carrying the SNPs (SNP-HIF3α) in the cells resulted in induction of ET-1 mRNA even under normoxic condition, indicating an implication of SNP-HIF3α in dysregulation of ET-1 production. SNP-HIF3α showed higher affinity to HIF-1β and to ET-1 gene promoter irrespectively of oxygen concentrations compared to wild type HIF-3α. Interestingly, SNP-HIF3α seemed to employ an additional cis-acting DNA sequence proximity to hypoxia response element in ET-1 gene promoter for transcriptional activation of the gene.
Conclusion: PAH-associated coding SNPs confer a novel function on HIF-3α at ET-1 gene promoter, which might contribute to the derangement of ET-1 gene regulation in patients with CTD-PAH.
To cite this abstract in AMA style:Mizumoto K, Takatori K, Takatori S, Yoshimoto R, Eguchi K, Fujishiro D, Kodama S, Kobayashi A, Okamoto K, Kawaguchi Y, Haneda M, Makino Y. Pulmonary Arterial Hypertension-Associated Single Nucleotide Polymorphisms of Hypoxia-Inducible Factor-3a Gene Cause Constitutive Activation of the Endothelin-1 Gene Promoter [abstract]. Arthritis Rheumatol. 2015; 67 (suppl 10). https://acrabstracts.org/abstract/pulmonary-arterial-hypertension-associated-single-nucleotide-polymorphisms-of-hypoxia-inducible-factor-3a-gene-cause-constitutive-activation-of-the-endothelin-1-gene-promoter/. Accessed October 16, 2019.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/pulmonary-arterial-hypertension-associated-single-nucleotide-polymorphisms-of-hypoxia-inducible-factor-3a-gene-cause-constitutive-activation-of-the-endothelin-1-gene-promoter/