Funktionelle Bedeutung eines Einzelnukleotid-Polymorphismus im Gen der endothelialen Stickstoffmonoxid-Synthase
Kardiologie, Angiologie
Rheumatologie
Zusammenfassung der Projektergebnisse
As part of this project, we could establish that a member of the signal transducers and activator of transcription (STAT) family is the long-sought shear stress-dependent transcription factor for maintaining NOS3 expression. It binds to the promoter inside two polymorphic sites, i.e. the T- 786C and the A-922G SNP (rs 1800779). Presence of the mutant -786CC- and -922GG-genotypes, which are in a near perfect linkage disequilibrium, slows down NOS3 expression in response to unidirectional shear stress (USS) and, consequently, the ability of the endothelial cells to produce NO in sufficient quantities. We presume that activating histone marks differ between the - 786CC- and -786TT-genotype while presence of the -922GG- instead of the -922AA-genotype enables tight binding of Ying Yang 1 (YY1) to the promoter thereby recruiting factors introducing repressive histone marks. In addition, there is a third SNP (A-1468T, rs 1800783) that is in a strong linkage disequilibrium with the other two. Bioinformatic analyses suggest that nuclear factor-1A preferentially binds to the promoter of -1468AA-genotype individuals but there is no working hypothesis yet what the consequences of this may be. In patients with coronary heart disease (CHD) the -786CC-genotype but not the -922GG- or the -1468AA-genotype is significantly associated with an increased risk. In patients with rheumatoid arthritis (RA) but not psoriatic arthritis and spondyloarthritis, the odds ratio is significantly and similarly increased with all 3 genotypes. This suggests that all 3 SNPs have an impact on (shear stress dependent) NOS3 expression in human endothelial cells. Since all efforts to establish a suitable mouse model reflecting the impact of these SNPs, which only exist in the human genome, on the susceptibility to contract CHD or RA were in vain, we focused on potential mechanisms/mediators that may compensate for this insufficient endothelial NO synthesis. In addition to the shear stress-dependent increase in manganese superoxide dismutase (SOD2) expression we could establish 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) as a potent anti-inflammatory eicosanoid that is released by NO-insufficient -786CC-genotype endothelial cells (in response to USS). We characterised its mechanism of action in human monocytes/macrophages through activation of the transcription factor Nrf2 and in in-vitro differentiated type 1 T-helper cells through an Nrf2-independent mechanism. Patients with CHD, RA and related chronic inflammatory diseases present with significantly increased plasma levels of 15d-PGJ2 that is highest in patients with CHD or type 2 diabetes followed by psoriatic arthritis, spondyloarthritis and RA. This increased release of 15d-PGJ2 is neither genotype-dependent nor linked to diseases significantly associated with at least the T-786C SNP. We therefore propose that it represents a possible general defence mechanism in patients with chronic inflammatory diseases and/or a decreased bioavailability of the potent anti-inflammatory mediator NO. Finally, we performed bulk RNA-sequencing analyses to identify additional compensatory mechanisms in individuals harbouring the -786CC-genotype. Despite some interesting leads that we follow up currently, we noticed that this approach does not achieve the fine resolution needed to characterize what truly differentiates the -786CC-/-922GG-/-1468AA- from the -786TT-/-922AA-/-1468TT- genotype at baseline and in response to USS. We therefore contemplate to extend this approach to single-cell RNA-sequencing.
Projektbezogene Publikationen (Auswahl)
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T-786C single nucleotide polymorphism of the endothelial nitric oxide synthase gene as a risk factor for endothelial dysfunction in polymyalgia rheumatica. Clin Exp Rheumatol. 2015 Sep-Oct;33(5):726-30
Löffers C., Heilig B. & Hecker M.
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Sensitive mass spectrometric assay for determination of 15-deoxy-Δ12,14-prostaglandin J2 and its application in human plasma samples of patients with diabetes. Analytical and Bioanalytical Chemistry, 410(2), 521-528.
Morgenstern, Jakob; Fleming, Thomas; Kadiyska, Ivelina; Brings, Sebastian; Groener, Jan Benedikt; Nawroth, Peter; Hecker, Markus & Brune, Maik
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15-Deoxy-Δ12,14-Prostaglandin J2Reinforces the Anti-Inflammatory Capacity of Endothelial Cells With a Genetically Determined NO Deficit. Circulation Research, 125(3), 282-294.
Urban, Ivelina; Turinsky, Martin; Gehrmann, Sviatlana; Morgenstern, Jakob; Brune, Maik; Milewski, Moritz R.; Wagner, Andreas H.; Rumig, Cordula; Fleming, Thomas; Leuschner, Florian; Gleissner, Christian A. & Hecker, Markus
