Abstract 382 Secreted long non-coding RNAs Gadlor1 and Gadlor2 act on intra-cardiac communication to affect cardiac remodelling during pressure overload

Clin Res Cardiol (2023). https://doi.org/10.1007/s00392-023-02180-w
Secreted long non-coding RNAs Gadlor1 and Gadlor2 act on intra-cardiac communication to affect cardiac remodelling during pressure overload
M. Keles¹, S. Grein¹, N. Froese², F. A. Trogisch¹, R. Wardman¹, S. Hemanna¹, N. Weinzierl¹, S. Lomada³, G. M. Dittrich¹, M. Szaroszyk², T. Wieland³, J. Heineke¹
¹Department of Cardiovascular Physiology, ECAS (European Center for Angioscience), Mannheim Faculty of Medicine, Heidelberg University, Mannheim; ²Kardiologie und Angiologie, Medizinische Hochschule Hannover, Hannover; ³Experimentelle Pharmakologie, Universitätsmedizin Mannheim der Universität Heidelberg, Mannheim;
Introduction. Pathological stimuli such as pressure overload can lead to the development of heart failure with maladaptive changes in the myocardium. Recent studies have shown that long non-coding RNAs (lncRNAs) contribute to the development of cardiac disease. We have identified two novel lncRNAs, Gadlor1 and Gadlor2, which are enriched in endothelial-cell (EC) derived extracellular vesicles (EVs). Methods and Results. Analysis of Gadlor1/2 expression in different cardiac cell types revealed that they are mainly expressed in ECs, followed by cardiac fibroblast (FB) and cardiomyocytes (CM). Interestingly, the abundance of Gadlors is even more pronounced in EC-derived EVs and they are mainly taken up by CMs, suggesting a possible role in cardiac cell communication. Expression levels of Gadlor1/2 were significantly increased in mouse cardiac tissue after transverse aortic constriction (TAC) and in the myocardium and serum of heart failure patients. The effect of Gadlor knock-out (Gadlor-KO) and Gadlor overexpression during cardiac pressure overload was analysed by echocardiography, histological analyses and bulk RNA sequencing from isolated cardiac cells. Systemic deletion of Gadlor1 and Gadlor2 markedly ameliorated cardiac hypertrophy following TAC as shown by a less pronounced increase in cardiomyocyte area and hypertrophy-associated genes compared to WT. Gadlor-KO mice exhibited less TAC-induced cardiac dysfunction and were protected from pulmonary congestion. Additionally, Gadlor-KO mice showed less deposition of myocardial fibrosis and higher capillarization, however unexpectedly, suffered from sudden death during prolonged pressure overload. To investigate a potential arrhythmic behaviour in KO mice, we performed contractility and calcium transient measurements. In isolated adult KO-CMs after TAC, a considerable delay in diastolic sarcoplasmic calcium re-uptake was observed compared to WT-CMs. On the contrary, Gadlor overexpression triggered cardiomyocyte hypertrophy, fibrosis and cardiac dysfunction, and faster CM relaxation. RNA sequencing from cardiac cells isolated after 2-weeks of TAC confirmed significant upregulation of angiogenesis and cell-cycle associated genes in ECs, as well as a downregulation of matrix gene expression in FBs in Gadlor-KO mice. Moreover, deletion of Gadlor1/2 downregulated mitochondrial organization-associated genes in CMs after TAC. Mechanistically, CaMKII was identified as Gadlor1/2 interaction partner in cardiomyocytes by RNA antisense purification coupled with mass-spectrometry (RAP-MS). We further investigated the phosphorylation of phospholamban (PLN) at threonine-17 (pThr-17) by CaMKII and found a significant reduction of pThr17-PLN in Gadlor-KO mice, and a considerable increase in Gadlor1/2 overexpressing mouse hearts after TAC, respectively. These findings indicate that Gadlor1/2 might promote CaMKII activation and thereby trigger aggravated myocardial remodelling and changes in excitation-contraction coupling. Conclusion. Gadlor1 and Gadlor2 are novel lncRNAs that are mainly enriched in EC-derived EVs and upregulated during cardiac pathological overload. Gadlor lncRNAs induce cardiac hypertrophy, fibrosis and alterations in the calcium homeostasis during relaxation in CMs, which mainly take up Gadlor1/2 by EVs transferred from ECs. Targeted inhibition of Gadlor1/2 in non-myocytes might therefore be considered as a therapeutic strategy for heart failure.
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