rBTI Combined with Oxaliplatin Promotes Apoptosis in Colon Cancer Cells
WANG Miaomiao#, ZHU Jianyun#, LI Yuying*
In this paper, the effects of rBTI (recombinant buckwheat trypsin inhibitor) combined with Oxaliplatin on apoptosis of colon cancer cells HCT116 and DLD-1 were investigated. Firstly, the effective concentration of rBTI and Oxaliplatin on colon cancer cells were screened by cell proliferation assay, and the concentration with a survival rate of more than 80% was selected as the concentration of the combined drug. MTT, morphological observation, clonogenesis, flow cytometry, Western blot and qRT-PCR were used to detect cell proliferation, morphology, clonogenesis, apoptosis, Bcl-2, Bax, Cleaved Caspase-3 protein and mRNA expression. Furthermore, the changes of MAPK signaling pathway were analyzed by reactive oxygen species detection, molecular docking and Western blot. The results showed that both rBTI and Oxaliplatin could inhibit the proliferation of colon cancer cells in a dose-dependent manner. The proliferation of colon cancer cells was significantly inhibited by the synergistic action of the drugs. Compared with the single treatment group, the number of cells in the combined treatment group decreased, the contraction became round and the volume decreased. The cell clonal formation decreased and the number of apoptosis increased. Cleaved Caspase-3 and Bax protein and mRNA expressions increased, and Bcl-2 expression decreased. The expression of reactive oxygen species increased; both rBTI and Oxaliplatin could bind closely to the key proteins of MAPK signaling pathway, and could change the expression of key proteins. In conclusion, rBTI combined with Oxaliplatin could promote apoptosis of colon cancer HCT116 and DLD-1 cells.
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