Inhibition of Jak-STAT3 pathway enhances bufalin-induced apoptosis in colon cancer SW620 cells.

Abstract

BACKGROUND:

The purpose of the research is to investigate the roles of Jak-STAT3 signaling pathway in bufalin-induced apoptosis in colon cancer SW620 cells.

METHODS:

The 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.

CONCLUSIONS:

Bufalin 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.

• Authors: Zhitu Zhu, Enze Li, Yangyang Liu, Yu Gao, Hongzhi Sun, Guangyou Ma, Zhenghua Wang, Xiaomei Liu, Qingjun Wang, Xiujuan Qu, Yunpeng Liu, Yunlong Yu
• Journal: World journal of surgical oncology (World J Surg Oncol) Vol. 10 Pg. 228 (Oct 30 2012) ISSN: 1477-7819 [Electronic] England
• PMID: 23110625 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antineoplastic Agents
• Bufanolides
• Enzyme Inhibitors
• STAT3 Transcription Factor
• STAT3 protein, human
• Tyrphostins
• alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide
• JAK1 protein, human
• Janus Kinase 1
• bufalin

Topics

• Antineoplastic Agents (pharmacology)
• Antineoplastic Combined Chemotherapy Protocols
• Apoptosis (drug effects)
• Blotting, Western
• Bufanolides (pharmacology)
• Cell Cycle (drug effects)
• Cell Proliferation (drug effects)
• Colonic Neoplasms (drug therapy, metabolism, pathology)
• Enzyme Inhibitors (pharmacology)
• Humans
• Janus Kinase 1 (antagonists & inhibitors, metabolism)
• Mitotic Index
• STAT3 Transcription Factor (antagonists & inhibitors, metabolism)
• Tumor Cells, Cultured
• Tyrphostins (pharmacology)