Cell counts after hMSC-PBMC co-cultures.

Allogeneic co-culture assays reveal that TLR4 priming of hMSCs (MSC1) promotes T-cell proliferation, while unprimed hMSCs and TLR3 primed hMSCs (MSC2) suppress it. Methods: T cells among the peripheral blood mononuclear cells (PBMCs) were activated with 1 µg of CD3/CD28 antibody beads prior to labeling with fluorescence label (CFSE) to monitor their activation or cell division and loaded at a 10∶1 ratio over the MSCs for 72 hr. For cell counts, an aliquot of the 72 hr spent medium was removed prior to flow cytometry for Trypan Blue staining and counting as standard. Data are representative of four independent experiments and are expressed as mean cell counts +/−SEM of 4 replicate counts on a hemocytometer after trypan blue staining. Total of 5 MSC donors and 5 PBMC donors were used in the assay. Two representative donors (d1, d2) are shown.