Conclusion, EX ead can capture exosomes from biofluid samples with no ultracentrifugation and exosomes is often successfully eluted in the beads.IP.Activity assays for evaluation of clinical grade MSC-EV antiinflammatory properties for use in remedy of drug-resistant epilepsy in children Alessandra Fierabracci1, Valeria La Marca1, Kelly Van Wemmel2, Sally Snyman2, Silvia Balosso3, Laura Papetti4, Maurizio Muraca5, Annamaria Vezzani6, Federico Vigevano7 and Marcin Jurga1 Children’s Hospital Bambino Ges Infectivology and Clinical Trials Location, Sort 1 Diabetes Centre; 2Esperite Cell Factory; 3Dept of Neuroscience, IRCCS Ist. Ricerche Farmacologiche Mario Negri; 4Dept of Neurosciences, Children’s Hospital Bambino Gesu’; 5Department of Women’s and Children’s Well being, University of Padua, Padua, Italy; 6Dept of Neuroscience, IRCCS Ist Ricerche Farmacologiche Mario Negri; 7Children’s Hospital Bambino Ges Dept of NeuroscienceIntroduction: MSCs exert their biological effects via secretion of extracellular vesicles (EV). We previously showed that MSC-EV have substantial immunomodulatory properties. MSC-EV inhibit B cells proliferation/differentiation upon PBMC CpG stimulation, similarly to parent MSCs. Moreover, MSC-EV induce Treg proliferation/apoptosis and IL-10 secretion, following antiCD3/CD28 PBMC stimulus. In this study we show that clinical grade (CG) EV exert similarScientific Ubiquitin-Specific Protease 1 Proteins supplier program ISEVimmunomodulation to research grade (RG) counterparts. CG EV may be created with higher PTPN3 Proteins manufacturer efficiency if in comparison to RG EV and MSCs manufacturing. Presently our group is preparing MSC-derived EV for clinical tests in therapy of epilepsy, a disorder resistant to anti-epileptic drugs in 40 of kids resulting from neuroinflammation. A novel antiinflammatory technique, determined by CG EV, is proposed. Approaches: A strategy of CG EV production is according to human umbilical cord derived (UC) MSCs cultured within a closed, scalable stirred-tank bioreactor program in totally defined GMP culture media. EVs are purified by sequential filtration/sterilization. The final item is analyzed by NTA to evaluate size and quantity, and EVs are characterized by MACSPlex immunophenotyping (FACS), to recognize certain CD markers. The immuno-modulatory activity in the CG product is evaluated in comparison with RG EVs and MSCs by precise in vitro B and T cells assays. Outcomes: The CG EV isolation process, has been optimized to obtain at the very least 1.5 10^9 EV/mL in 24 h from 0.1 10^6 MSCs. EV diameter cut off is 300 nm. MACSPlex exosome assay revealed that EV are CD9, CD63 and CD81 positive, but HLA-ABC and HLA-DRPQ negative. T and B potency assays, performed on PBMC, indicate immunosuppression by CG EV, similarly for the RG EV obtained from the identical MSCs. This effect is revealed by Treg enhance, counteracting T eff, upon T cells activation, and by reduction of B cells proliferation and plasma cell differentiation, following B cells activation. Summary/Conclusion: We’ve developed and standardized a reproducible approach for the production, quantification and immunophenotyping of CG EV, beginning from human UC MSCs, with similar immunomodulation if when compared with RG EV counterparts. Our data indicate that the use of CG EV could possibly be helpful in the treatment of a wide selection of immunological diseases, and deliver a more accessible option for allogenic MSCs. Funding: Esperite (B)IP.Urine exosome proteins CXCL9 and CXCL10 are predictors of kidney transplant rejection Christine M. Coticchia1, Ja.
