P6300Endothelial cell activation by pro-inflammatory cytokines exerts novel paracrine effects on co-cultured cardiomyocytes

Autor: Justin C. Mason, I Bardi, J Fourre, Cesare M. Terracciano, R T Maughan
Rok vydání: 2019
Předmět:
Zdroj: European Heart Journal. 40
ISSN: 1522-9645
0195-668X
DOI: 10.1093/eurheartj/ehz746.0898
Popis: Background Anti-inflammatory therapies have failed to meet expectations in recent clinical trials for heart failure, despite the number of studies demonstrating pro-fibrotic, arrhythmogenic and hypertrophic effects of inflammation. To enlighten this, we sought to examine the contribution of non-myocytes in the cardiac inflammatory response. Endothelial cells (EC) can regulate cardiomyocyte (CM) function with multiple soluble factors. While many studies have shown the effects of pro-inflammatory cytokines on EC or CM separately, it is still unclear how the activation of EC can affect CM function. Purpose We studied the effect of pro-inflammatory pre-conditioning of EC on CM in indirect co-culture systems and in an ex vivo model of cardiac tissue. We hypothesised that pro-inflammatory activation of EC would alter the contractility of co-cultured CM. Methods Human cardiac microvascular EC were first pre-conditioned for 24h with Cytomix (1 ng/ml TNF-α, 1 ng/ml IL-1β, 25 ng/ml IL-6 Rα/IL-6 chimera) and co-cultured in a transwell system (pore size: 0.4μm) with adult rat ventricular CM. Co-culture supernatants were screened using a Cytokine Profiler Array. In vitro analysis of calcium handling in CM utilised the optical mapping technique and Fluo-4. Contractility of cardiac tissue was measured ex vivo using myocardial slices of 300 μm, prepared from left ventricles of adult rat and cultured for 24h with field stimulation and a fixed stretch producing a sarcomeric length of 2.2 μm. Results Treatment of EC by Cytomix prior to co-cultures induced a release of CC and CXC chemokines, G-/GM-CSF, ST2 and the adhesion molecule ICAM-1. Using published RNAseq datasets we noticed that adult CM do not constitutively express the receptors for the chemokines identified with the Profiler Array. However, duration of calcium transients in CM was significantly reduced from 454 to 322 ms in co-cultures with EC pre-conditioned by Cytomix, compared to untreated EC. Amplitude and time to peak were unchanged. In contrast, myocardial slices treated with Cytomix demonstrated a significant increase in contractility compared to control (from 3.3 to 5.3 mN/mm2) but no significant change in the duration of contraction (from 467 to 434 ms) or the rate of relaxation. Conclusions Pre-conditioned EC exert paracrine effects on the calcium handling of isolated CM, suggesting pro-inflammatory activation of proximal EC can affect CM function. In myocardial slices, pro-inflammatory stimulation provoked an inotropic response. The divergence of effect with findings in isolated cells may reflect differences in experimental design and multicellularity. Future work will aim to characterise the soluble mediators involved, and selectively target EC in slices to contextualise the effects described in co-cultures. Ultimately, this may inform the development of novel anti-inflammatory strategies for clinical use. Acknowledgement/Funding British Heart Foundation
Databáze: OpenAIRE
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