Similarly to what was accomplished together with the leukemia cell lines, to study the effects of BMSCs on CD34 cells, gene ex pression profiles from CD34 cell mono cultures and co cultures with BMSCs have been analyzed by microarrays. We discovered that 2075 genes had been differentially expressed in CD34 cell co cultures compared with mono cultures. Among probably the most up regulated genes have been SOCS3, REN and CXCL6, all with a fold transform five. Ingenu ity pathway analysis from the differentially expressed genes re vealed that one of the most represented canonical pathways were cAMP mediated signaling, VDR RXR activation and vehicle diac B adrenergic signaling. CCL2 and IL eight are enhanced in supernatants from BMSCs co cultured with leukemia cells Gene expression analysis revealed that most of the genes up regulated in BMSCs co cultured with leukemia cells were involved in IL 17 signaling.
To assess the things created by co cultured cells, we screened selelck kinase inhibitor the superna tants from co culture and mono culture samples at 48 h for cytokine production by R D Human Cytokine panel A. We chose this panel for the reason that amongst the 36 cytokines within the panel were CXCL1, sICAM 1, IL 1B, IL 8, CCL2 and Serpin E1 all of which were found to become up regulated at the gene level in co cultured BMSCs. Furthermore, with panel A we were in a position to measure the relative levels of IFN?, IL six and IL 23 that are IL 17 signaling associated cytokines. Having said that, the majority of the 36 cytokines within the panel have been undetectable in our samples and also the levels of cyto kines CXCL1, ICAM 1, IL 23, IL six, MIF and Serpin E1 weren’t substantially changed involving co culture and mono culture conditions.
Nevertheless, the levels of CCL2 and IL eight have been greater in supernatants from BMSCs co cultured with leukemia cells, however the benefits were variable amongst BMSCs from distinct subjects. The levels of IFN? p38 MAPK Inhibitors and CD40L have been higher in co culture compared with mono culture supernatants, however the differ ence was not statistically considerable. The analysis of cyto kines in the supernatant of cultured BMSCs and leukemia cells was performed in three series of experiments with BMSCs from three wholesome donors, and we discovered diverse responses amongst the unique BMSC donors. We identified increased levels of IFN? and CD40L only in the supernatants from BMSC003 co cultured with TF 1 and TF1. The levels of IL 8 have been improved within the supernatants from BMSC003, BMSC006 and, to a lesser extent, in BMSC002. The amount of CCL2 was measurable only in supernatants from BMSC003, BMSC002 and BMSC006 co cultured with TF 1 and K562 leukemia cells and within the supernatant from BMSC006 co cultured with TF 1. To confirm the improved levels of CCL2 and IL eight inside the supernatants from BMSCs co cultured with leukemia cells at 48 h, we measured the levels in the two cytokines working with ELISA assays.