Fractalkine-mediated neuronal protection in the diabetic retina
In the diabetic retina disruption of fractalkine (FKN) signaling induces fibrin(ogen) deposition. increased production of IL-1??? by microglia. vascular and neuronal damage.But. how FKN and its receptor. CX3CR1. regulate microglia activation and retinal pathology is unknown.In humans. two single nucleotide polymorphisms in the CX3CR1 locus (hCX3CR1-I249/M280) that show defective binding to FKN. play a key role in inflammation during diabetes.Induction of diabetes in a mouse model expressing the human CX3CR1-I249/M280 revealed accelerated neuronal loss in the retina. and systemic inflammation caused microglial clustering and upregulation of pro-inflammatory cytokines.Thus. the FKN/CX3CR1 signaling pathway plays an under-appreciated role in diabetic retinopathy (DR). perhaps via inflammatory processes.However. there is a gap in knowledge regarding the exact relationship among FKN/CX3CR1 signaling. microglial activation and cell damage (neuronal and vascular) in a diabetic host in which hyperglycemia and repeated episodes of systemic inflammation occur.The central hypothesis is that FKN is neuroprotective in DR by blocking microglia activation and the subsequent vascular permeability and inflammatory changes that are characteristics of DR.The following specific aims will test the mechanism by which sustained microglial activation by fibrin(ogen) potentiates neuronal and vascular endothelial cell damage.Utilizing experimental mouse models of diabetes combined with systemic endotoxemia the following specific aims are proposed. Specific Aim 1.Determine the role of microglia in the initiation and progression of diabetic retinopathy.This aim will test the hypothesis that FKN binds to CX3CR1 polarizing microglia towards an anti-inflammatory pathway early in disease.Therefore. a) microglia depletion in diabetic mice at acute and chronic stages of disease. and b)peripheral fibrin(ogen) depletion will be used. to demonstrate that 1) dysregulated microglia in response to fibrin(ogen) induce pro-inflammatory actions and oxidative damage that contribute to neuronal damage and 2) that regulation of microglia activation can be harnessed to prevent vision loss. Specific Aim 2.Determine the neuroprotective effects of soluble FKN to mitigate microglia activation and rescue neuronal and vascular damage.FKN is expressed on neurons as a transmembrane protein. acting as a membrane-bound adhesion molecule (mFKN) or as a soluble protein (sFKN) upon cleavage from cell surfaces.Adeno-associated viral vectors will be used to express sFKN or mFKN in neurons to test the hypothesis that pathways that enhance FKN signaling via its soluble domain are neuroprotective and will prevent vision loss.Approaches will be implemented to also determine the synergistic effect of sFKN and anti-fibrin approaches to prevent vision loss in mice lacking CX3CR1 or expressing the human CX3CR1I249/M280 variant.