Abstract
BackgroundThe purpose of the research is to investigate the roles of Jak-STAT3 signaling pathway in bufalin-induced apoptosis in colon cancer SW620 cells.MethodsThe inhibitory effects of bufalin on cell proliferation were determined by MTT (Methyl thiazolyltetrazolium) assay. The morphological changes of cells were measured by Wright-Giemsa staining. The cell cycle arrest and apoptosis were tested by flow cytometry analysis. Western Blot was used to determine the protein expression of the apoptosis inhibitors livin and caspase-3, the apoptosis-related proteins Bax and Bcl-2, as well as the key protein kinases in the Jak-stat3 signaling pathway, stat3 and p-stat3.Results(1) Bufalin inhibited the proliferation of SW620 cells. IC50 at 24 h, 48 h and 72 h were 76.72 ± 6.21 nmol/L, 34.05 ± 4.21 nmol/L and 16.7 ± 6.37 nmol/L. (2) Bufalin induced SW620 cell cycle arrest and apoptosis, indicated by the appearance of apoptotic bodies; (3) The results from flow cytometry demonstrated that there was cell cycle G2/M phase arrest in 20 nmol/L bufalin treatment group (36.29 ± 2.11% vs 18.39 ± 1.74%, P<0.01); there was a sub-diploid apoptosis peak in 80 nmol/L bufalin treatment group (19.69 ± 1.63% vs 0.99 ± 0.23%, P <0.01). The apoptosis rate was 34.63 ± 2.57% (vs 19.69 ± 1.63%, P = 0.002) in JAK kinase inhibitor AG490 plus bufalin treatment group. (4) During the process of bufalin-induced apoptosis in SW620 cells, transient activation of p-stat3 inhibited the activation of stat3, up-regulated Bax expression, down-regulated livin and Bcl-2 expression (P<0.01), and activated caspase-3. Inhibition of Jak-stat3 signaling pathway by pre-treatment with AG490 significantly enhanced the bufalin-induced apoptosis (P<0.01), further up-regulated Bax protein expression, down-regulated livin and Bcl-2 protein expression and enhanced caspase-3 activation.ConclusionsBufalin not only inhibited the growth of colon cancer SW620 cells, but also induced apoptosis of SW620 cells. Activation of caspase-3, up-regulation of Bax, down-regulation of livin and Bcl-2, as well as inhibition of Jak-stat3 signaling pathway might be the important mechanisms for the bufalin-induced apoptosis.
Highlights
The purpose of the research is to investigate the roles of Janus family tyrosine kinase (Jak)-signal transducers and activators of transcription 3 (STAT3) signaling pathway in bufalin-induced apoptosis in colon cancer SW620 cells
Bufalin inhibited the growth of colon cancer SW620 cells, and induced apoptosis of SW620 cells
Activation of caspase-3, up-regulation of Bax, down-regulation of livin and Bcl-2, as well as inhibition of Jak-stat3 signaling pathway might be the important mechanisms for the bufalin-induced apoptosis
Summary
The purpose of the research is to investigate the roles of Jak-STAT3 signaling pathway in bufalin-induced apoptosis in colon cancer SW620 cells. Colon cancer is a common digestive system cancer. It is estimated that there were 106,100 new cases and 49,920 deaths from colon cancer in the US in 2009 [1]. Due to the change in people’s diet, the incidence of colon cancer is rising and becoming one of the most common malignancies and the leading cause of death from cancer. With the development and application of a new generation of chemotherapy and molecular-targeted drugs, the effects of colorectal cancer treatment have been improved, but are still unsatisfactory. Researchers are engaged in discovering effective and low toxicity drugs for colon cancer treatment to improve the cure rate and reduce mortality of colon cancer
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