While precise mechanisms underlying cardiovascular diseases (CVDs) are still not fully understood, previous studies suggest that the innate immune system, through Toll-like receptor 4 (TLR4), plays a crucial part in the pathways leading to these diseases, mainly because of its interplay with endogenous molecules. The Heat-shock protein 70 family (HSP70-70kDa) is of particular interest in cardiovascular tissues as it may have dual effects when interacting with TLR4 pathways. Although the hypothesis of the HSP70 family members acting as TLR4 ligands is becoming widely accepted, to date no co-crystal structure of this complex is available and it is still unknown whether this process requires the co-adaptor MD2. In this study, we aimed at investigating the interplay between the TLR4/MD2 complex and HSP70 family members in the human cardiovascular system through transcriptomic data analysis and at proposing a putative interaction model between these proteins. We report compelling evidence of correlated expression levels between TLR4 and MD2 with HSP70 cognate family members, especially in heart tissue. In our molecular docking simulations, we found that HSP70 in the ATP-bound state presents a better docking score towards the TLR4/MD2 complex compared to the ADP-bound state (-22.60 vs. -10.29 kcal/mol, respectively). Additionally, we show via a proximity ligation assay for HSP70 and TLR4, that cells stimulated with ATP have higher formation of fluorescent spots and that MD2 might be required for the complexation of these proteins. The insights provided by our computational approach are potential scaffolds for future in vivo studies investigating the interplay between the TLR4/MD2 complex and HSP70 family members in the cardiovascular system.
Keywords: HSP70; TLR4/MD2 complex; cardiovascular system; computational modelling.