Adipose tissue derived mesenchymal stem cells exhibit a superior immune-modulatoery effect on mature dendritic cells compared to bone marrow derived mesenchymal stem cells

Poster Abstract Presentations customized dECM which enhances cellular growth and functionality. This is achieved by culturing MSCs on hydrogels of varying stiffnesses, and assessing the bioactivity of the ECM produced during culture. Methods, Results & Conclusion: Mechanical properties of tunable stiffness polyacrylamide gels were verified using an Instron Microtester. Cells used were isolated from the decidua basalis, and ECM was sourced from decidua basalis hTERT-transfected MSCs. ECM deposition was induced by addition of ascorbic acid. Instron measurements confirm the elastic moduli of fully-hydrated polyacrylamide gels to be 4 kPa, 10 kPa, and 40 kPa, tailored to mimic the stiffness of adipose tissue, skeletal muscle tissue, and collagenous bone tissue, respectively. MSCs exhibit a spindle-shaped morphology when cultured on the hydrogels. Confocal microscopy reveals microarchitectural differences between ECM deposited on softer vs. stiffer hydrogels. We have shown that MSCs can be grown on hydrogels of varying stiffnesses, and can be induced to laydown extracellular matrix. The functionalization of the gels with collagen I to aid cell adhesion has been confirmed with immunofluorescence. The deposition of ECM has also been confirmed with confocal microscopy, and the alignment of ECM fibrils has been shown to depend on substrate stiffness. Further work will involve proteomic characterization of the dECMs, and culturing MSCs on the dECMs to assess bioactivity. We expect that the substrate stiffness will modulate the ECM bioactivity, allowing customization of tailored dECM for MSC expansion. 330 ADIPOSE TISSUE DERIVED MESENCHYMAL STEM CELLS EXHIBIT A SUPERIOR IMMUNE-MODULATOERY EFFECT ON MATURE DENDRITIC CELLS COMPARED TO BONE MARROW DERIVED MESENCHYMAL STEM CELLS R. Zaza1, F. Jamali1, M. Hassouneh2, D. Alhattab1, S. AlAlawi1 & A. Awidi1 1 Cell Therapy Center- University of Jordan, Amman, Al-Jubeiha, Jordan, 2 Department of Applied Biology, University of Sharjah- Faculty of Science, Sharjah, United Arab Emirates Background & Aim Background: The ability of mesenchymal stem cells (MSCs) to interact with immune cells and exert a modulatory effect is the core of cellular therapy approaches. Dendritic cells (DCs), the most potent of antigen presenting cells (APCs), play a key role in T-cell activation and firing immune responses. The direct interaction between MSCs and mature DCs (mDCs) results in attenuation of mDCs' activity and subsequent immune reactions. Aim: Bone marrow derived MSCs (BM-MSCs) and adipose tissue derived MSCs (AT- MSCs), are more abundant and easier to harvest compared to other types of MSCs. How BM-MSCs and AT-MSCs affect mDCs can be illustrated with changes in surface marker expression and mDCs secreted cytokines, a focus is needed on changes in mDC gene expression which may help in a better understanding of the interaction process. Methods, Results & Conclusion Methods: MSCs were isolated from two adult sources bone marrow and adipose tissue. Monocytes were isolated from peripheral blood to generate mDCs in recommended culture conditions. All cells were characterized by flowcytometry. Multiplex immunoassay was carried out to evaluate the inflammatory cytokines levels. PCR arrays were performed to demonstrate the gene expression profile of mDCs. All tests were performed before and after direct contact co-culture of mDCs with BM-MSCs, and with AT-MSCs. Results: The immune-inhibitory effect of both types of MSCs on mDCs was manifested by a decrease in the levels of inflammatory cytokines, and the downregulation of 84 investigated genes responsible for mDC activity in both co-cultures. The fold changes, and fold change ratios from PCR array data clearly demonstrate the superiority of AT-MSCs in inhibiting mDCs immune awakening functions. This finding is supported by previous microarray analysis of the inherent immune-modulatory potential of both types of MSCs, which revealed an advantage for AT-MSCs over BM-MSCs, in terms of the expression magnitude of immune-regulatory genes. Functional annotation clustering and gene ontology demonstrated that all investigated genes participate in response to stimulus (p- value= 4.3 E-26), and that 79.8% (67 out of 84 genes) are significant players in positive regulation of response to stimulus (pvalue = 4.3 E-45). Conclusion: These findings highlight the outcome of MSCs’ interaction with mDCs on the molecular level. Our data confirm the immune-modulatory properties of MSCs and suggest AT-MSCs as a more effective suppressor of mDCs immune activity than BM-MSCs.

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Protein-protein interactions as generated from PCR array data.

331 ISCT SURVEY REPONSES TO PROCESS AND PRODUCT DEVELOPMENT COMMITTEE'S COST-OF-GOODS QUESTIONNAIRE J. Fink, O. Karnieli & D. Clarke PPD, ISCT, Westborough, Massachusetts, United States Background & Aim: The ISCT Process and Product Development (PPD) committee conducted a questionnaire regarding Cost of Goods (COG) topics and activities in Cell & Gene therapy in 2017. They survey's questions were around the topics of Product type, Manufacturing, Container, Dose, Storage, Transport, Temperature and Administration. Methods, Results & Conclusion: The ISCT membership was sent the voluntary questionnaire. 51 members filled out the 45-question survey. This poster will release the results and summarize the data from the PPDs COGs questionnaire. A sample of the responses are: respondents were 33% academic vs. private, 93% are working with a therapeutic product, 65% autologous vs. 32% allogenic, T-Cells and MSC represent 68% of cell type used, 54% are seeking US FDA regulation approval and 41% seeking Europe EMEA approval, 31% plan to manufacture at the clinic/hospital, 43% at CMO and 56% internally, 54% plan to use a bag for product delivery and 48% a vial, 44% expect one dose per patient and 31% say it will vary per patient, 56% will store their product at cryogenic temperatures vs 22% fresh, 80% plan to use a DMSO containing cryoprotectant, 34% expect to ship the final product internationally, 74% require thawing of the final product, 56% require a dilution and/or wash, 78% will let the physician determine protocol if any adverse events occur and 50% will offer access to a sponsor review team. The PPD hopes the results of this survey will help further improvements to planning and cost reduction for Cell and Gene therapies. 332 REAL-TIME QUALITY CONTROL AND FUNCTIONAL ASSESSMENT OF MESENCHYMAL STEM CELLS FOR CELLULAR THERAPIES Y.A. Abassi ACEA Biosciences, San Diego, California, United States Background & Aim: One of the biggest hurdles in manufacturing of cellular therapies using mesenchymal stem cells (MSCs) is the lack of quantitative tools that simultaneously assess quality and potency of MSCs, which display big variations among donors and over limited passages. Methods, Results & Conclusion: We developed a real-time cell-based assay that generates impedance signals reflecting integrated changes in cell number, attachment, and morphology. We obtained bone marrow derived MSCs from 3 different donors and cultured them under standard conditions using xenofree media. Multiple real-time quantitative parameters are derived from the