Project Grant F31AI194811

ELUCIDATING THE CD301B-TN ANTIGEN AXIS IN EFFEROCYTOSIS AND INFLAMMATION RESOLUTION - PROJECT SUMMARY/ABSTRACT MAINTAINING GUT HOMEOSTASIS REQUIRES THE INTESTINAL IMMUNE SYSTEM TO RAPIDLY RESOLVE INFLAMMATION WHILE TOLERATING CONSTANT EXPOSURE TO MICROBES AND DIETARY ANTIGENS. A CRITICAL COMPONENT OF THIS BALANCE IS EFFEROCYTOSIS, THE EFFICIENT AND NON-INFLAMMATORY CLEARANCE OF APOPTOTIC CELLS BY MUCOSAL PHAGOCYTES. FAILURE OF THIS PROCESS ALLOWS CELLS TO UNDERGO SECONDARY NECROSIS, AMPLIFYING TISSUE DAMAGE AND DRIVING CHRONIC INFLAMMATORY DISEASES SUCH AS ULCERATIVE COLITIS (UC). DESPITE EXTENSIVE KNOWLEDGE OF CANONICAL EFFEROCYTOSIS RECEPTORS, THE COLON'S GLYCAN-RICH ENVIRONMENT SUGGESTS THAT CARBOHYDRATE-BINDING PATHWAYS MAY ALSO REGULATE APOPTOTIC CELL CLEARANCE. DURING APOPTOSIS AND EPITHELIAL STRESS, O-GLYCAN PROCESSING OFTEN BECOMES DISRUPTED, RESULTING IN HYPOGLYCOSYLATION AND EXPOSURE OF TN ANTIGEN (GALNAC). THIS TRUNCATED GLYCAN IS ASSOCIATED WITH MUCIN BARRIER DISRUPTION, BACTERIAL PATHOGENS, HOST IMMUNE REGULATION. STUDIES SUGGEST TN ANTIGEN MAY BE ASSOCIATED WITH STRESSED OR DYING CELLS FOR IMMUNE RECOGNITION. BECAUSE RECOGNITION OF THESE SIGNATURES THROUGH SUGAR BINDING PROTEINS IS CRITICAL FOR MUCOSAL HOMEOSTASIS, TN EXPOSURE MAY ACT AS A GLYCAN ENCODED SIGNAL FOR APOPTOTIC CELLS. ONE SUCH SUGAR BINDING PROTEIN, CD301B (MGL2) IS A MYELOID C-TYPE LECTIN RECEPTOR THAT SELECTIVELY RECOGNIZES TN ANTIGEN AND REGULATES IMMUNE RESPONSES IN MULTIPLE TISSUES, YET CD301B+ IMMUNE CELLS IN THE COLON HAVE NEVER BEEN DEFINED, AND THE TN-CD301B AXIS HAS NEVER BEEN EXAMINED IN EFFEROCYTOSIS OR MUCOSAL INFLAMMATION. MY PRELIMINARY DATA IDENTIFY COLONIC CD301B+ CELLS AS A HETEROGENOUS POPULATION OF M2-LIKE MACROPHAGES (M2S) AND CONVENTIONAL DENDRITIC SUBSET-2 CELLS (CDC2S), DEMONSTRATE THAT CD301B DEFICIENCY LEADS TO DELAYED RECOVERY FROM DSS-INDUCED COLITIS, AND SHOW THAT APOPTOTIC NEUTROPHIL-LIKE CELLS UPREGULATE TN ANTIGEN, SUPPORTING A MODEL IN WHICH GLYCAN REMODELING DURING APOPTOSIS ENABLES LECTIN-MEDIATED CLEARANCE. THEREFORE, THIS PROPOSAL AIMS TO DEFINE THE CD301B-TN AXIS AND ELUCIDATE IF GLYCAN-DEPENDENT RECOGNITION OF APOPTOTIC CELLS REGULATES EFFEROCYTOSIS AND INFLAMMATION RESOLUTION IN THE COLON. THE CENTRAL HYPOTHESIS IS THAT TN-DEPENDENT CD301B SIGNALING ENABLES EFFICIENT APOPTOTIC CELL CLEARANCE AND DRIVES RESOLUTION IN THE COLON. I WILL TEST THIS HYPOTHESIS BY 1) DEFINING THE EFFEROCYTOTIC AND HOMEOSTATIC FUNCTIONS OF CD301B+ IMMUNE CELLS USING FLOW CYTOMETRY, IN VIVO NEUTROPHIL FATE MAPPING, CYTOKINE PROFILING, AND HISTOPATHOLOGY, AND BY 2) DETERMINING HOW THE TN-CD301B AXIS MEDIATES APOPTOTIC CELL UPTAKE AND DOWNSTREAM RESOLUTION PATHWAYS USING IN VITRO EFFEROCYTOSIS ASSAYS, GLYCAN BLOCKING APPROACHES, TRANSCRIPTOMICS, AND PROXIMITY-BASED PROTEOMICS. THIS WORK ADDRESSES A FUNDAMENTAL GAP IN OUR UNDERSTANDING OF GLYCAN-LECTIN REGULATION OF EFFEROCYTOSIS IN THE COLON. BY INTEGRATING MUCOSAL IMMUNITY, GLYCOBIOLOGY, AND SYSTEMS-LEVEL APPROACHES, THIS PROJECT INTRODUCES A CONCEPTUALLY INNOVATIVE FRAMEWORK FOR LECTIN-MEDIATED INFLAMMATION RESOLUTION. THESE FINDINGS AIM TO ESTABLISH MECHANISTIC FOUNDATIONS FOR FUTURE TRANSLATIONAL STUDIES OF DEFECTIVE MUCOSAL IMMUNITY, INCLUDING UC AND OTHER INFLAMMATORY DISEASES.