Abstract TMP120: Autoantigen(s) Trigger a Robust Immune Response in Cerebral Cavernous Malformations

Abstract

Introduction: Previous studies have reported robust inflammatory cell infiltration, selective synthesis of IgG, B-cell clonal expansion, and deposition of immune complexes and complement within Cerebral Cavernous Malformation (CCM) lesions. Furthermore,B-cell depletion has been shown to reduce the maturation of CCM in murine models. We hypothesize that specific autoantigen(s) within the lesional milieu trigger the pathogenetic immune responses in CCMs. This study aims to identify those putative autoantigen(s) using recombinant antibodies (rAbs) derived from plasma cells found in surgical human CCM lesions. Methods: CD138 + plasma cells were laser captured from fresh frozen surgically resected human CCM lesions. Clonally expanded immunoglobulin heavy- and light-chain variable region pairs were cloned into IgG expression vectors and expressed as monoclonal antibodies. Purified rAbs were assayed by immunofluorescence with CCM lesion tissue and normal brain tissue sections. rAbs assayed by immunocytochemistry with human primary cell line were used to further define the staining pattern. The cell lysates were immunoprecipitated with rAb, after protein purification by SDS-PAGE, and analyzed by Mass spectrometry. Results: In normal brain tissue, rAbs stained endothelial cells with limited staining of glial cells. In CCM lesional tissue, rAbs stained endothelial cells, glial cells as well as structures in the acellular matrix adjacent to caverns. In cultured Human Brain Microvascular Endothelial Cells (HBMECs) and Human Astrocytes (HAs), rAbs co-localized with cytoplasmic components. After HBMEC and HA cell lysates were immunoprecipitated with rAb, a Coomassie Stain detected bands of approximately 50 kDa. Conclusions: Our results suggest that autoantigen(s) in human CCM lesions are cytoplasmic components present in lesional tissue as well as in normal brain tissue. Molecular level identification of the triggering antigen is still ongoing by mass spectrometry. Identification of the autoantigen(s) in the lesional milieu might explain the propensity of lesion development from leaky endothelium in the neuroglial parenchyma. Characterization of the autoantigen triggers will open new venues for therapy or vaccine in this disease.