Effect of TNF-α on the biological characteristics of BMSCs in tumor microenvironment

Objective 　To investigate the effect of tumor necrosis factor-α (TNF-α) on the biological characteristics of rat's bone marrow mesenchymal stem cells (BMSCs) in the microenvironment of C6 glioma cells. Methods 　BMSCs were non-contact indirectly co-cultured with C6 glioma cells for 7 days. The expressions of free TNF-α in supernatants of BMSCs and co-cultured MSCs cells were tested by ELISA. Cells were divided into four groups: BMSCs group, indirectly co-cultured MSCs group, co-cultured MSCs+0.5ng/ml TNF-α group, and co-cultured MSCs+5ng/ml TNF-α group. Morphology of cells was observed by inverted microscopy. The proliferation ability was detected by CCK8, and cell cycle was detected by flow cytometry. The migration ability was tested by Transwell. The mRNA expressions of CyclinD1 and c-Myc were measured by real-time PCR. The protein expressions of NF-κB/P65, CyclinD1 and c-Myc were tested by Western blotting. Results 　The expression of TNF-α was obviously lower in co-cultured MSCs' supernatant than in BMSCs group (P<0.05). The BMSCs were spindle like fibroblast cells, and the morphology of co-cultured MSCs+5ng/ml TNF-α group was consistent with that in BMSCs group, but the apoptotic cells increased, while the morphology changed a lot in co-cultured MSCs and co-cultured MSCs+0.5ng/ml TNF-α group, showing smaller size and larger ratio of nucleus/cytoplasm. The proliferation ability, G2+S phase ratio and migration ability were significantly higher in co-cultured MSCs group than in BMSCs group (P<0.05), and the expression levels of NF-κB/P65, CyclinD1 and c-Myc mRNA and protein in co-cultured MSCs were higher than those in BMSCs group (P<0.05). While the proliferation ability, G2+S phase ratio, and migration ability in co-cultured MSCs+5ng/ml TNF-α group were markedly lower than those in co-cultured MSCs and co-cultured MSCs+0.5ng/ml TNF-α group (P<0.05), the expression levels of NF-κB/P65, CyclinD1 and c-Myc mRNA and protein were significantly lower in co-cultured MSCs+5ng/ml TNF-α group than those in co-cultured MSCs and co-cultured MSCs+0.5ng/ ml TNF-α group (P<0.05). Conclusions 　The biological characteristics of BMSCs have been changed in the microenvironment of C6 glioma, the decline of TNF-α expression after co-culture may be an important factor influencing the changes of biological characteristics. The addition of high concentration of TNF-α may be important for maintaining the stability of stem cell biological characteristics. DOI: 10.11855/j.issn.0577-7402.2018.12.02