To Örebro University

Background and Aims: We aimed to investigate the role of IL-6 trans-signaling in regulating the release of endothelial inflammatory mediators and endothelial basement mebrane proteins, laminins. We also aimed to investigate the laminin composition in atherosclerotic plaques in relation to the immune cell content.

Methods: HUVECs were cultured for in vitro experiments. The BiKE cohort was used to assess expression of laminins in atherosclerotic plaques and healthy vessels. Gene and protein expression was analysed using Microarray/qPCR and proximity extension assay, immunostaining or immunoblotting techniques respectively.

Results: The release of 20 inflammatory proteins from ECs was significantly altered (3 downregulated, 17 upregulated) in response to stimulation with IL-6 and sIL-6R (p-value <0.05, FDR 5%). Ingenuity pathway analyses predicted that these proteins enhance lymphocyte binding and transmigration while inhibiting transmigration of granulocytes. ECs treated with combination of IL-6 and sIL-6R upregulated expression of LAMA5 while downregulating LAMA4. Transcriptomic and proteomic analyses showed that both endothelial LAMA4 and LAMA5 were significantly lower in atherosclerotic plaques compared to healthy vessels. In addition, expression of endothelial LAMA5 was significantly lower in plaques from symptomatic patients compared to those from asymptomatic patients. We also showed that endothelial laminins are negatively correlated with immune cell markers.

Conclusions: Our findings suggest that IL-6 trans-signaling regulates endothelial inflammatory proteins and laminin production that enhance binding and trans-migration of lymphocytes. In the context of atherosclerosis, expression of laminin alpha chains is altered and also appeared to be associated with plaque stability. Our data also revealed a relationship between laminin chains and immune cell content in atherosclerotic plaques.