Project Grant F32AI194808

CYSTATIN F EXPRESSION ON MICROGLIA IS CRITICAL FOR CONSTRAINING DEMYELINATION FOLLOWING VIRAL INFECTION OF THE CENTRAL NERVOUS SYSTEM - PROJECT SUMMARY MULTIPLE SCLEROSIS (MS) IS A DEBILITATING AUTOIMMUNE DISEASE WHEREIN ACTIVATED IMMUNE CELLS MISTAKENLY ATTACK THE PROTECTIVE MYELIN COATING ON NEURONS. THIS RESULTS IN SCARRING AND THE FORMATION OF LESIONS IN THE BRAIN AND/OR SPINAL CORD THAT CONTRIBUTE TO A HOST OF SYMPTOMS THAT VARY DEPENDENT ON THE LOCATION OF THE INJURY. THE PRECISE ETIOLOGY OF MS REMAINS UNKNOWN, THOUGH RECENT STUDIES HAVE POINTED TOWARDS A COMBINATION OF GENETIC AND ENVIRONMENTAL FACTORS. INTRIGUINGLY, A STRONG CORRELATION EXISTS BETWEEN PREVIOUS EXPOSURE TO EBSTEIN-BARR VIRUS, WHICH INCREASES THE RISK OF DEVELOPING MS BY 30-FOLD. AS THIS HIGHLIGHTS A POTENTIALLY IMPORTANT ROLE THAT VIRAL INFECTION MAY PLAY IN THE INITIATION OF MS, OUR LAB UTILIZES A MURINE NEUROTROPIC JHM STRAIN OF MOUSE HEPATITIS VIRUS (JHMV) THAT RESULTS IN AN ACUTE ENCEPHALOMYELITIS THAT IS ACCOMPANIED BY GRAY MATTER INVOLVEMENT WITH INFECTION OF OLIGODENDROCYTES, ASTROCYTES, AND MICROGLIA. JHMV PERSISTS IN THE WHITE MATTER TRACTS OF THE SPINAL CORD, INDUCING DEMYELINATION RESULTING IN HIND-LIMB PARALYSIS IN MICE. WE AND OTHERS HAVE DEMONSTRATED PREVIOUSLY THAT MICROGLIA, A RESIDENT INNATE IMMUNE CELL POPULATION THAT RESIDES IN THE CENTRAL NERVOUS SYSTEM (CNS), ARE CRITICAL IN CONSTRAINING THE IMMUNE RESPONSE IN DEMYELINATING MODELS OF MS. INTRIGUINGLY, IN AREAS OF DEMYELINATION UNDERGOING REMYELINATION, THERE IS A SIGNIFICANT UPREGULATION OF CYSTATIN F (CST7), A PROTEASE INHIBITOR. WHILE CST7 IS EXPRESSED ON MANY IMMUNE CELLS, WE'VE SHOWN THAT MICROGLIA ARE THE PREDOMINANT SOURCE OF CST7 IN THE BRAIN. THUS, THIS PROPOSAL SEEKS TO DEFINE THE ROLE OF MICROGLIA-DERIVED CST7 IN THE CONTEXT OF A VIRAL INFECTION WITH JHMV. WE PROPOSE TO INFECT NOVEL TRANSGENIC CONDITIONAL KNOCKOUT MICE FOR CST7 ON MICROGLIA AND WILD-TYPE ANIMALS WITH JHMV TO DETERMINE THE IMPACT OF CST7 ON THE IMMUNE RESPONSE. A COMBINATION OF FLOW CYTOMETRY AND IMAGING MODALITIES WILL BE UTILIZED TO CHARACTERIZE DIFFERENCES IN BRAIN IMMUNE CELL INFILTRATION AND ACTIVATION. THE IMPACT OF THE CONDITIONAL ABLATION OF CST7 ON DEMYELINATION AND SUBSEQUENT REMYELINATION WILL BE ASSESSED AS WELL, USING STATE-OF-THE-ART TECHNIQUES LIKE SPATIAL TRANSCRIPTOMICS (AIM 1). BASED ON OUR PRELIMINARY AND PUBLISHED FINDINGS, WE HYPOTHESIZE THAT CST7 EXPRESSED ON MICROGLIA IS HOST PROTECTIVE. THEREFORE, WE ALSO PROPOSE TO EVALUATE CST7 AS A POTENTIAL THERAPEUTIC FOR DEMYELINATING DISEASES BY EMPLOYING AN OVEREXPRESSION MODEL. CST7 WILL BE OVEREXPRESSED IN MICROGLIA IN MICE INFECTED WITH JHMV AND THE SUBSEQUENT IMMUNE RESPONSE AND MYELINATION STATUS WILL BE EVALUATED USING SIMILAR TECHNIQUES AS DESCRIBED ABOVE (AIM 2). TOGETHER, THESE AIMS SEEK TO EVALUATE THE ROLE OF MICROGLIA-DERIVED CST7 IN AN EFFORT TO FURTHER DELINEATE MECHANISMS OF DEMYELINATION AND REMYELINATION. AS IT IS EXTREMELY RARE FOR DAMAGED NEURONS TO UNDERGO REMYELINATION IN MS, DEFINING THESE MECHANISMS FURTHER WILL PROVIDE INVALUABLE INSIGHT FOR THE ADVANCEMENT OF NEW THERAPEUTICS.