Effects of Bufalin on Human Esophageal Cancer Cells Proliferation and Mechanism Study / 中国药学杂志

ABSTRACT

OBJECTIVE: To investigate the effect of bufalin on the proliferation of esophageal cancer and its possible molecular mechanism. METHODS: The effects of bufalin on the activity of esophageal cancer cell line KYSE-70 were determined by MTT assay and LDH assay kit. Colony-forming and EdU (5-ethynyl-2-deoxyuridine) assay were performed to detect the inhibitory effect of bufalin on the proliferation of KYSE-70 cells. DAPI staining, TUNEL and flow cytometry were used to assess the effects of bufalin on the apoptosis of KYSE-70 cells. Quantitative real-time PCR (qPCR) and Western blotting were used to determine the relevant expression changes in mRNA and protein of apoptosis-related factors Bcl-2, Bax, NF-κBp65, NF-κBpp65. The mitochondrial function changes of KYSE-70 cells were studied by using JC-1 fluorescent probe kit and ATP detection kit after bufalin treatment. Finally, the effect of bufalin on esophageal cancer proliferation in vivo was studied by xenograft model in nude mice. RESULTS: The results of MTT and LDH assay shown that bufalin inhibited the activity of KYSE-70 cells. Colony-forming and EdU assay showed that bufalin significantly suppressed KYSE-70 cells proliferation. DAPI staining showed that chromatin heterogeneity, nuclear concentration and fragmentation were observed after bufalin treatment. It was found that bufalin treatment significantly promoted KYSE-70 cells apoptosis by TUNEL staining and flow cytometry assay of qPCR and Western blot showed that Bcl-2 expression was down-regulated, Bax expression was up-regulated and Bax/ Bcl-2 expression ratio was increased after bufalin treatment. The mitochondrial membrane potential and the ATP production of KYSE-70 cells were significant reduced after bufalin treatment. In vivo, the growth of xenograft tumors was significantly inhibited in bufalin group. CONCLUSION: Bufalin markedly inhibits KYSE-70 cells proliferation by promoting apoptosis, and the possible mechanism of apoptosis may be related to mitochondrial pathway. Our results indicate that bufalin may be a potential therapeutic agent for esophageal cancer.