UH Faculty, Staff, and Student Works
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The collection gathers research products generated by University of Houston faculty, staff, and students
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Browsing UH Faculty, Staff, and Student Works by Author "Agarwal, Sandeep K."
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Item Beyond Autoantibodies: Biologic Roles of Human Autoreactive B Cells in Rheumatoid Arthritis Revealed by RNA?Sequencing(Arthritis and Rheumatism, 2017-06) Mahendra, Ankit; Yang, Xingyu; Abnouf, Shaza; Adolacion, Jay R.T.; Park, Daechan; Soomro, Sanam; Roszik, Jason; Coarfa, Cristian; Romain, Gabrielle; Wanzeck, Keith; Bridges, Louis Jr. S.; Aggarwal, Amita; Qiu, Peng; Agarwal, Sandeep K.; Mohan, Chandra; Varadarajan, NavinObjective:To obtain the comprehensive transcriptome profile of human citrulline?specific B cells from patients with rheumatoid arthritis (RA). Methods:Citrulline? and hemagglutinin?specific B cells were sorted by flow cytometry using peptide–streptavidin conjugates from the peripheral blood of RA patients and healthy individuals. The transcriptome profile of the sorted cells was obtained by RNA?sequencing, and expression of key protein molecules was evaluated by aptamer?based SOMAscan assay and flow cytometry. The ability of these proteins to effect differentiation of osteoclasts and proliferation and migration of synoviocytes was examined by in vitro functional assays. Results:Citrulline?specific B cells, in comparison to citrulline?negative B cells, from patients with RA differentially expressed the interleukin?15 receptor ? (IL?15R?) gene as well as genes related to protein citrullination and cyclic AMP signaling. In analyses of an independent cohort of cyclic citrullinated peptide–seropositive RA patients, the expression of IL?15R? protein was enriched in citrulline?specific B cells from the patients’ peripheral blood, and surprisingly, all B cells from RA patients were capable of producing the epidermal growth factor ligand amphiregulin (AREG). Production of AREG directly led to increased migration and proliferation of fibroblast?like synoviocytes, and, in combination with anti–citrullinated protein antibodies, led to the increased differentiation of osteoclasts. Conclusion:To the best of our knowledge, this is the first study to document the whole transcriptome profile of autoreactive B cells in any autoimmune disease. These data identify several genes and pathways that may be targeted by repurposing several US Food and Drug Administration–approved drugs, and could serve as the foundation for the comparative assessment of B cell profiles in other autoimmune diseases.Item Detection and isolation of auto-reactive human antibodies from primary B cells(Methods, 12/1/2014) Sendra, Victor G.; Lie, Anthony; Romain, Gabrielle; Agarwal, Sandeep K.; Varadarajan, NavinThe isolation of human monoclonal antibodies (hmAb) has emerged as a versatile platform in a wide variety of contexts ranging from vaccinology to therapeutics. In particular, the presence of high titers of circulating auto-antibodies is implicated in the pathology and outcome of autoimmune diseases. Therefore, the molecular characterization of these hmAb provides an avenue to understanding the pathogenesis of autoimmune diseases. Additionally, the phenotype of the auto-reactive B cells may have direct relevance for therapeutic intervention. In this report, we describe a high-throughput single-cell assay, microengraving, for the screening, characterization and isolation of anti-citrullinated protein antibodies (ACPA) from peripheral blood mononuclear cells (PBMC) of rheumatoid arthritis (RA) patients. Stimulated B cells are profiled at the single-cell level in a large array of sub-nanoliter nanowells (?105), assessing both the phenotype of the cells and their ability to secrete cyclic-citrullinated peptide (CCP)-specific antibodies. Single B cells secreting ACPA are retrieved by automated micromanipulation, and amplification of the immunoglobulin (Ig) heavy and light chains is performed prior to recombinant expression. The methodology offers a simple, rapid and low-cost platform for isolation of auto-reactive antibodies from low numbers of input cells and can be easily adapted for isolation and characterization of auto-reactive antibodies in other autoimmune diseases.