Microglial process convergence following brain injury

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Project Abstract Traumatic brain injury (TBI) produces significant pathology, including post-injury inflammation (particularly within the thalamus), that can lead to long-term morbidities. Microglia, the innate immune cells of the brain, are critical mediators of neuroinflammation that can have either neurotoxic or neurotrophic effects. Progress has been made investigating TBI-induced neuroimmune responses in rodents and therapies showing great promise have moved to clinical trials but failed to translate into beneficial interventions for humans suffering TBI. However, therapies targeting processes that occur in higher order mammals, with immune responses, cytoarchitecture, and metabolic rates similar to humans, such as pigs, would be more likely to translate to the clinic successfully. The neuroinflammatory progression following brain injury in pigs, however, is not well- understood. Our preliminary data demonstrated that microglial processes converge onto injured axonal swellings (microglial process convergence; MPC) in the thalamus of micro pigs, that is not recapitulated in rats, following diffuse TBI but does appear to occur in humans. Studies in mice indicate that this microglial process convergence (MPC) requires functional P2Y12R, however, the mechanisms behind MPC in higher order mammals is currently unknown. Both our preliminary data and previous studies indicate that MPC may be an ameliorative process, promoting axonal outgrowth acutely post-injury. Therefore, the goal of this study is to assess the roles of microglial changes on pathological progression in a pig model of diffuse TBI. Studies indicate that males have greater pro-inflammatory responses and less axonal outgrowth compared to females. However, there are no known studies evaluating sex as a biological variable in MPC. Accordingly, the current study will address the following specific aims 1) to evaluate the effects of inflammatory and P2Y12R modulation on MPC, neuronal survival and axonal outgrowth/retraction following injury and 2) to assess the prevalence of MPC in human brain tissue following diffuse TBI and other CNS injuries/diseases. To address these aims we will complete quantitative 3D assessments of multiplexed immunohistological samples for microglial-axonal interactions in vitro and in pigs to determine the degree of MPC in relation to axonal outgrowth/retraction changes, P2Y12R expression, and sex-related variability. As the prevalence of MPC following brain injury in the human population is currently unknown, we will also probe for the degree of MPC onto injured axonal swellings, neuronal survival, axonal outgrowth/retraction, and spatially resolved RNA profiles in a unique cohort of postmortem brain tissue. This study is significant because understanding microglial-neuronal interactions and repair mechanisms in higher order species and humans of both sexes will translate into therapeutics strategies for the treatment of TBI.