Abstract
Background: Systemic administration of bone marrow derived stromal cells (MSCs) releases a broad range of trophic factors mediating stroke recovery. How MSCs release these factors may depend on the state of the circulation after systemic administration. We assessed whether the secretomes of MSCs, upon exposure to peripheral blood monocytes (Mo), differs depending on the temporal course when the Mo were isolated after stroke. Methods: Peripheral blood mononuclear cells (PBMCs) from stroke patients was isolated at 24h (N=10) and day 5-7 (N=7) after stroke, and from healthy controls (N=5) using Ficoll gradient. CD14+ Mo were isolated using indirect magnetic beads labelling. Contact and Trans-well co-culture assays were established using 100,000 CD14+ Mo and MSCs each in a 0.8um transwell system. After 24h, media was collected and analyzed for secretome using multiplex cytokine assay. Subtypes of Mo were evaluated using flow cytometry. Results: There was a significant increase in non-classical (CD14+CD16++) Mo and a significant decrease in classical (CD14++CD16-) Mo at Day5-7 after stroke as compared to 24h after stroke (Fig). When Day5-7 CD14+ Mo were co-cultured with MSCs, we found a significant reduction in inflammatory IL-1β and TNF-α, and a significant increase in anti-inflammatory IL-4 as compared to co-cultures of MSCs with healthy Mo. This effect was more pronounced in transwell (vs contact) co-cultures. Co-cultures of MSCs and Mo from controls or from patients 24h after stroke led to reductions in TNF-α and upregulation in IL-4 compared with Mo alone. There were no differences in effect of MSCs co-cultures with healthy vs 24h stroke Mo. Conclusion: Our study indicates that the immunomodulatory effect of MSCs may be dependent on the phenotype of Mo in the peripheral circulation of stroke patients. Hence, depending on the temporal course after stroke, the systemic circulation of stroke patients may differentially regulate MSCs to release soluble factors.
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