Evaluation of the immunomodulatory potency of adipose-derived mesenchymal stem cells in response to pro-inflammatory stimuli

Abstract Mesenchymal stem cells (MSCs) have shown strong potential in regenerative medicine due to their immunomodulatory capacity through paracrine signaling. However, their therapeutic effectiveness could be enhance in respond to inflammatory environments, such as those that occur in the early stages of myocardial infarction. Selecting the most responsive cell lines is therefore key to maximizing their clinical impact. This study aimed to establish a standardized protocol to assess the immunomodulatory potential of porcine adipose-derived MSCs (pASCs) under pro-inflammatory stimulation. Four independent pASC lines (pASCs014, pASCs15, pASCs016 y pASCs017) were treated with TNF-α, IFN-γ, or their combination at concentrations of 1, 3, 10, and 30 ng/mL. After 48 hours, surface expression of PD-L1, CD106, and TNFR1 was analyzed via flow cytometry. In parallel, total RNA was extracted and analyzed by qPCR to quantify the expression of IDO-1, PTGS2, IL1RN, and TNFAIP6. Among the cell lines tested, pADCs014 showed the highest upregulation of IDO-1, TSG6, and IL1RN, especially in response to TNF-α. Flow cytometry confirmed elevated expression of PD-L1 and CD106 after TNF-α stimulation, and increased TNFR1 expression following combined cytokine exposure. Based on gene and surface marker response, pADCs014 was identified as the most suitable candidate for further therapeutic applications. This protocol allows for the pre-selection of highly responsive MSC lines and could improve consistency in regenerative strategies involving immune modulation.