Therefore, the e pression of these MSC markers was evaluated in isolated hUCMSCs by immunostaining assay. http://www.selleckchem.com/products/XL184.html As shown in Figure 1C, OCT4 and NANOG, which represent the pluripotent embryonic stem cell phenotype, were e pressed in hUCMSCs. UCMSCs have multiple lineages potential to adipogenic and osteogenic differentiation. To characterize the isolated hUCMSCs in our system, they were cultured in the adipogenic and osteogenic complete media. Ten days after induction, osteogenic differentiation of hUCMSCs was verified as brownish orange red for e tracellular calcium deposits by Alizarin Red staining. In addition, accumulation of lipid vacuoles from the hUCMSCs as the indicator of adipogenic differentiation of MSCs was detected as bright red color by Oil red staining, implying that isolated hUCMSCs in this study had stem cell potential.
hUCMSCs inhibited the proliferation of PC 3 cancer cells To determine the antitumor effect of hUCMSCs on hu man prostate cancer cells, PC 3 prostate cancer cells were cocultured with the densities of 3. 33 104, 2 104, and 1 104 of UCMSCs. First, we determined the viability of PC 3 cells by MTT assay. The viability of PC 3 cells cocultured with UCMSCs was significantly decreased, whereas UCMSCs PC 3 did not show the difference compared with PC 3 cells cultured without hUCMSCs. In addition, we determined the proliferation of PC 3 cells cocultured with hUCMSCs by BrdU assay. The growth of PC 3 cells cocultured with hUCMSCs was decreased to 44%, 49%, and 69% of control in the presence of UCMSCs with various numbers of 3.
33 104, 2 104, and 1 104, re spectively, compared with untreated control. As shown in Figure 2C, when PC 3 cells were cocul tured in the presence of hUCMSCs, the number of PC 3 cells was rarely observed com pared with untreated control. hUCMSCs induced apoptosis and attenuated survival genes in PC 3 cells To determine whether apoptosis is induced in PC 3 cells cocultured with hUCMSCs, Western blotting was per formed. PARP cleavage, cleaved caspase 3, Ba , and phosphorylation of JNK were detected in the lysates of PC 3 cocultured with hUCMSCs. To verify whether this apoptotic event is dependent on JNK pathway, the JNK specific inhibitor SP600125 was treated in PC 3 cells cocultured with hUCMSCs.
Con versely, the apoptotic features such as PARP cleavage, cleaved caspase 3, and phosphorylation of JNK in PC 3 cells by hUCMSCs were efficiently masked by JNK in hibitor SP600125 with Western blotting and Dacomitinib immunofluorescence assay. Also, as shown in Figure 4A, PI3K and phosphorylation of AKT and ERK were attenuated in PC 3 cells by hUCMSC cells. Furthermore, the e pression of survival genes such as Bcl 2, Bcl L, Survivin, Mcl 1, and cIAP 1 was attenu ated in PC 3 cells by Western blotting. The homing of hUCMSCs and apoptosis induction in PC 3 cells in nude mouse Ne t, we investigated the homing of hUCMSCs to PC 3 cells in mice.