Targeting IL-6 signaling as potential angiogenic and metabolic pathway

Abstract

Purpose : Unrestrained neovascularization is a major contributor to vision loss and blindness in proliferative diabetic retinopathy (PDR) and age-related macular degeneration. The vascular endothelial growth factor (VEGF) plays a key role in this context. Since we cannot satisfactorily control disease activity in all patients using anti-VEGF therapy, other signaling pathways possibly contribute in a relevant manner.
In this study, we investigate the interleukin 6 (IL6) signaling complex, IL6 and its soluble receptor (sIL6R) and their effect on angiogenesis as well as endothelial cell metabolism.

Methods : Commercially available ELISA were used to determine levels of IL6 and sIL6R in human vitreous and plasma samples of PDR patients (N=17) and control patients with macular pucker (N=17). The spheroid sprouting-, migration- and proliferation-assay were used to assess the angiomodulatory effect of IL6, sIL6R as well as the sIL6R inhibitor tocilizumab and VEGF inhibitor aflibercept on human umbilical vein vascular endothelial cells (HUVECs). Western blotting was employed to examine STAT3, STAT1, Akt, ERK signaling. Metabolic profiles were assessed by Seahorse technology and tandem mass spectrometry.

Results : PDR patients showed significantly elevated levels of IL6 and sIL6R in the vitreous and plasma compared to control patients(*p<0.05).
In vitro, only IL6+sIL6R induced a significant increase in proliferation(*p<0.0001), migration and sprouting in vascular endothelial cells, partially even in the presence of VEGF, whereas IL6 alone showed no effect. While IL6 primarily activated the STAT3 pathway, the combination of IL6+sIL6R showed a broadened spectrum of activation in inspected pathways including AKT and ERK. Interestingly, the proangiogenic effect of IL6+sIL6R in the presence of VEGF was reversed to base sprouting by co-treatment with aflibercept+tocilizumab.
The Seahorse assay revealed an elevated oxygen consumption rate of HUVECs pretreated with IL6+sIL6R as well as a higher extracellular acidification rate in the presence of VEGF+IL6+sIL6R. Targeted metabolomics also showed enhanced lactate levels in HUVECs in response to latter treatment.

Conclusions : Our data show that the interplay between IL6 and sIL6R is decisive for the resulting angiogenic and metabolic phenotype of IL6 signaling in the retina. A concise understanding of IL6 signaling activity in disease progression is necessary to successfully use it as therapeutic target.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.