Induction, amplification, and propagation of diabetic retinopathy-associated inflammatory cytokines between human retinal microvascular endothelial and Müller cells and in the mouse retina

Ocular levels of IL-1 beta, TNF alpha, IL-8, and IL-6 correlate with progression of diabetic retinopathy (DR). M & uuml;ller cells (MC), which are crucial to maintaining retinal homeostasis, are targets and sources of these cytokines. We explored the relative capacities of these four DR-associated cytokines to amplify inflammatory signal expression both in and between human MC (hMC) and retinal microvascular endothelial cells (hRMEC) and in the mouse retina. Of the four cytokines, IL-1 beta was the most potent stimulus of transcriptomic alterations in hMC and hRMEC in vitro, as well as in the mouse retina after intravitreal injection in vivo. Stimulation with IL-1 beta significantly induced expression of all four transcripts in hMC and hRMEC. TNF alpha significantly induced expression of some, but not all, of the four transcripts in each cell, while neither IL-8 nor IL-6 showed significant induction in either cell. Similarly, conditioned media (CM) derived from hMC or hRMEC treated with IL-1 beta, but not TNF alpha, upregulated inflammatory cytokine transcripts in the reciprocal cell type. hRMEC responses to hMC-derived CM were dependent on IL-1R activation. In addition, we observed a correlation between cytokine expression changes following direct and CM stimulation and NF kappa B-p65 nuclear translocation in both hMC and hRMEC. Finally, in mice, intravitreal injections of IL-1 beta, but not TNF alpha, induced retinal expression of Il1b and CXCL8 homologues Cxcl1, Cxcl2, Cxcl3, and Cxcl5, encoding pro-angiogenic chemokines. Our results suggest that expression of IL-1 beta, TNF alpha, IL-8, and IL-6 may be initiated, propagated, and sustained by autocrine and paracrine signals in hRMEC and hMC through a process involving IL-1 beta and NF kappa B. Targeting these signals may help thwart inflammatory amplification, preventing progression to vision-threatening stages and preserving sight.