Microbiome-dependent modulation of ocular immunity and the implications for infectious

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PROJECT SUMMARY Disruption of ocular immune homeostasis in ocular mucosa, which includes the conjunctiva, leads to disease and potential impaired vision. The conjunctiva is a mucosal tissue of the ocular surface that is rich in immune cells. These immune cells can prevent disease by detecting antigen, producing cytokines, and secreting anti- microbials. Inflammatory cells and cytokines within the conjunctiva can regulate the outgrowth and subsequent pathogenesis associated with infectious ocular disease. It has been suggested that the ocular microbiome may augment this process, thus playing a prominent role in ocular disease. Clearly defining how ocular microbiota mediate ocular immunity will be crucial for the development of more effective therapies for ocular diseases. My preliminary data suggest that commensals expand a subset of γδ T cells within ocular mucosa, which facilitate the migration of inflammatory monocytes into ocular mucosa and the release of anti-microbial molecules within the tears. The overall goals of the proposed studies are to define how ocular commensals induce γδ T cells and to determine if this specific interaction truly limits infectious ocular disease. The overall hypothesis is that ocular γδ T cells are generated and maintained by commensals in an antigen-specific manner, which in turn protects the ocular surface from infection by pathogenic methicillin-resistant Staphylococcus aureus (MRSA) infection. Rationale for the research planned is such that at its completion, new insights between the ocular microbiome and mucosal immune system will be revealed, which can then be exploited for the alleviation of disease. The K99 PHASE of this plan and Specific Aim 1 will use cell culture and fluorescence microscopy to identify how ocular γδ T cells are stimulated by ocular microbiota. Specific Aim 2 will use in vitro and in vivo techniques to assess whether innate pattern recognition receptors (PRRs) contribute to the generation and/or maintenance of commensal-directed γδ T cells. Lastly, during the INDEPENDENT R00 PHASE, in Specific Aim 3, I will use an in vivo model of ocular MRSA infection to show whether the described commensal/γδ T cell axis protects the ocular environment from pathogenic infection. The training phase of this plan will be conducted mainly in the laboratory of Dr. Rachel Caspi (mentor). TRAINING OBJECTIVES include receiving advanced training in fluorescence microscopy by Dr. Robert Fariss (co-mentor) and the NEI biologic imaging core for the purpose of imaging T cell and antigen presenting cell (APC) interactions in situ and in cell culture. I will also receive training from Dr. Sandip Datta (co-mentor) and his laboratory for the purpose of understanding pathogenesis associated with bacterial infections. Data from the proposed studies will help define how commensal microbiota modulate ocular immunity, which may aid in the treatment of ocular disease while also providing me a foundation for a life-long career as an independent researcher in the field of ocular immunology.