Monitoring microglial inflammatory phenotype in diabetic retinopathy models
Steven Abcouwer   (Ann Arbor, MI)
We hypothesize that flow cytometry can be used to efficiently monitor the activation and inflammatory status of retinal microglia during diabetic retinopathy (DR). The Specific Aim is to demonstrate the feasibility of methods to measure the activation and inflammatory status of retinal microglia during the progression of DR. Microglia are resident macrophage cells in the central and peripheral nervous system, and the sole immune cell of the retina. They perform innate immune surveillance, becoming “activated” upon infection or tissue damage. During acute and chronic neurodegenerative diseases microglia become activated, exhibiting an altered phenotype that includes changes in morphology, increased phagocytosis, surface marker expression, and gene expression. Fully activated microglia produce inflammatory and toxic molecules that contribute to neuronal inflammation, neuronal toxicity and vascular dysfunction. Microglia exhibiting an activated morphology have been observed in both human diabetic retinas and experimental models of DR. Immunohistochemical co-staining for inflammatory proteins and microglial marker proteins have qualitatively cemonstrated that activated microglia are a potential source of inflammatory proteins during retinopathies. However, during some neurodegenerative diseases microglia assume an alternative activation phenotype that coincides with the expression of anti-inflammatory proteins. These microglia may actually play a beneficial adaptive role that minimizes the inflammatory response to neuronal damage. Given that inflammation is now thought to play a key role in the progression of DR, it is vital to define the activation and inflammatory status of retinal microglia. The goal is to perfect methods enabling the expression of several inflammatory proteins, including TNFa, IL-6, IL-1ß, and iNOS, as well as anti-inflammatory proteins, including TGFß, IL-1Ra, IL-27, and arginase, to be quantified in subsets of retinal microglia by flow cytometry. To accomplish this we will: 1) use cultured microglia to initially test and develop antibody combinations for use in flow cytometry, 2) further test these combinations using the retinal IR model, and 3) apply these methods to retinas of diabetic animals.
No report available yet.