Rap1A Modulates Store-Operated Calcium Entry in the Lung Endothelium: A Novel Mechanism Controlling NFAT-Mediated Vascular Inflammation and Permeability.

Autor:	Kosuru R; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.)., Romito O; Department of Pharmacology and Chemical Biology (O.R., M.T.), University of Pittsburgh School of Medicine, PA., Sharma GP; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.)., Ferraresso F; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.)., Ghadrdoost Nakhchi B; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.)., Yang K; Data Science Institute (K.Y.), Medical College of Wisconsin, Milwaukee., Mammoto T; Department of Pediatrics (T.M., A.M.), Medical College of Wisconsin, Milwaukee., Mammoto A; Department of Pediatrics (T.M., A.M.), Medical College of Wisconsin, Milwaukee., Kastrup CJ; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.)., Zhang DX; Department of Medicine (D.X.Z.), Medical College of Wisconsin, Milwaukee., Goldspink PH; Department of Physiology and Biophysics, University of Illinois Chicago (P.H.G.)., Trebak M; Department of Pharmacology and Chemical Biology (O.R., M.T.), University of Pittsburgh School of Medicine, PA.; Vascular Medicine Institute (M.T.), University of Pittsburgh School of Medicine, PA.; UPMC Hillman Cancer Center (M.T.), University of Pittsburgh School of Medicine, PA., Chrzanowska M; Versiti Blood Research Institute, Milwaukee, WI (R.K., G.P.S., F.F., B.G.N., C.J.K., M.C.).; Department of Pharmacology and Toxicology (M.C.), Medical College of Wisconsin, Milwaukee.; Cardiovascular Center (M.C.), Medical College of Wisconsin, Milwaukee.
Jazyk:	angličtina
Zdroj:	Arteriosclerosis, thrombosis, and vascular biology [Arterioscler Thromb Vasc Biol] 2024 Nov; Vol. 44 (11), pp. 2271-2287. Date of Electronic Publication: 2024 Sep 26.
DOI:	10.1161/ATVBAHA.124.321458
Abstrakt:	Background: Store-operated calcium entry mediated by STIM (stromal interaction molecule)-1-Orai1 (calcium release-activated calcium modulator 1) is essential in endothelial cell (EC) functions, affecting signaling, NFAT (nuclear factor for activated T cells)-induced transcription, and metabolic programs. While the small GTPase Rap1 (Ras-proximate-1) isoforms, including the predominant Rap1B, are known for their role in cadherin-mediated adhesion, EC deletion of Rap1A after birth uniquely disrupts lung endothelial barrier function. Here, we elucidate the specific mechanisms by which Rap1A modulates lung vascular integrity and inflammation. Methods: The role of EC Rap1A in lung inflammation and permeability was examined using in vitro and in vivo approaches. Results: We explored Ca 2+ signaling in human ECs following siRNA-mediated knockdown of Rap1A or Rap1B. Rap1A knockdown, unlike Rap1B, significantly increased store-operated calcium entry in response to a GPCR (G-protein-coupled receptor) agonist, ATP (500 µmol/L), or thapsigargin (250 nmol/L). This enhancement was attenuated by Orai1 channel blockers 10 μmol/L BTP2 (N-[4-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]phenyl]-4-methyl-1,2,3-thiadiazole-5-carboxamide), 10 μmol/L GSK-7975A, and 5 μmol/L Gd 3+ . Whole-cell patch clamp measurements revealed enhanced Ca 2+ release-activated Ca 2+ current density in siRap1A ECs. Rap1A depletion in ECs led to increased NFAT1 nuclear translocation and activity and elevated levels of proinflammatory cytokines (CXCL1 [C-X-C motif chemokine ligand 1], CXCL11 [C-X-C motif chemokine 11], CCL5 [chemokine (C-C motif) ligand 5], and IL-6 [interleukin-6]). Notably, reducing Orai1 expression in siRap1A ECs normalized store-operated calcium entry, NFAT activity, and endothelial hyperpermeability in vitro. EC-specific Rap1A knockout (Rap1A iΔEC ) mice displayed an inflammatory lung phenotype with increased lung permeability and inflammation markers, along with higher Orai1 expression. Delivery of siRNA against Orai1 to lung endothelium using lipid nanoparticles effectively normalized Orai1 levels in lung ECs, consequently reducing hyperpermeability and inflammation in Rap1A iΔEC mice. Conclusions: Our findings uncover a novel role of Rap1A in regulating Orai1-mediated Ca 2+ entry and expression, crucial for NFAT-mediated transcription and endothelial inflammation. This study distinguishes the unique function of Rap1A from that of the predominant Rap1B isoform and highlights the importance of normalizing Orai1 expression in maintaining lung vascular integrity and modulating endothelial functions. Competing Interests: None.
Databáze:	MEDLINE
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