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Involvement of NF-kappaB activation in the apoptosis induced by extracellular adenosine in human hepatocellular carcinoma HepG2 cells.
Biochem Cell Biol. 2010 Aug; 88(4):705-14.BC

Abstract

Adenosine can exhibit cytotoxic activity in vivo and in vitro, though its mechanisms are still uncertain. In this study, we investigated the adenosine-mediated apoptotic signaling pathway and the role of NF-kappaB in human hepatocellular carcinoma HepG2 cells. HepG2 cells were treated with different concentrations of adenosine for 12-48 h, and the effect of adenosine on cell proliferation was evaluated by MTT assay. The cytotoxicity of adenosine alone or in combination with an NF-kappaB inhibitor, pyrrolidine dithiocarbamate (PDTC), was also evaluated by MTT assay and the mode of cell death was detected by Hoechst 33342 staining. Cell cycle progress was performed by flow cytometry with PI staining. The protein expressions of Bcl-2, p53, NF-kappaB subunit p65, and caspase-3 were assayed by Western blot. Caspase-3 activity was measured by spectrophotomteric assay. The results showed that adenosine significantly reduced the viability of HepG2 cells in a dose- and time-dependent manner, with IC 50 (24 and 48 h) of 2.52 and 1.89 mmol x L(-1), respectively. The apoptotic index (percentage of sub-G1 phase) of HepG2 cells in adenosine treatment alone for 12 and 24 h or in combination with PDTC were 8.30%, 22.32% and 20.18%, 30.89%, respectively. All of them were higher than that in the control group (0.81%, p < 0.01). The characteristic changes of cell apoptosis (chromatin condensation and sub-G1 peak) were observed under fluorescent microscopy and flow cytometry. We also found that the apoptotic process triggered by adenosine was involved in G0-G1 cell-cycle arrest, enhanced the activity of caspase-3, upregulated p53 and NF-kappaB p65 expression, and downregulated Bcl-2 expression. Inhibition of NF-kappaB by PDTC decreased NF-kappaB p65 expression, enhanced cell apoptosis ratio, and increased caspase-3 activity. NF-kappaB may play an anti-apoptosis role in adenosine-induced HepG2 cytotoxicity.

Authors+Show Affiliations

Department of Gastroenterology, Second Affiliated Hospital, Shantou University Medical College, Shantou, China. lingfeiwu@21cn.comNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

20651843

Citation

Wu, Ling-Fei, et al. "Involvement of NF-kappaB Activation in the Apoptosis Induced By Extracellular Adenosine in Human Hepatocellular Carcinoma HepG2 Cells." Biochemistry and Cell Biology = Biochimie Et Biologie Cellulaire, vol. 88, no. 4, 2010, pp. 705-14.
Wu LF, Li GP, Su JD, et al. Involvement of NF-kappaB activation in the apoptosis induced by extracellular adenosine in human hepatocellular carcinoma HepG2 cells. Biochem Cell Biol. 2010;88(4):705-14.
Wu, L. F., Li, G. P., Su, J. D., Pu, Z. J., Feng, J. L., Ye, Y. Q., & Wei, B. L. (2010). Involvement of NF-kappaB activation in the apoptosis induced by extracellular adenosine in human hepatocellular carcinoma HepG2 cells. Biochemistry and Cell Biology = Biochimie Et Biologie Cellulaire, 88(4), 705-14. https://doi.org/10.1139/O10-008
Wu LF, et al. Involvement of NF-kappaB Activation in the Apoptosis Induced By Extracellular Adenosine in Human Hepatocellular Carcinoma HepG2 Cells. Biochem Cell Biol. 2010;88(4):705-14. PubMed PMID: 20651843.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Involvement of NF-kappaB activation in the apoptosis induced by extracellular adenosine in human hepatocellular carcinoma HepG2 cells. AU - Wu,Ling-Fei, AU - Li,Guo-Ping, AU - Su,Jian-Dong, AU - Pu,Ze-Jin, AU - Feng,Jia-Lin, AU - Ye,Yan-Qing, AU - Wei,Bi-Liu, PY - 2010/7/24/entrez PY - 2010/7/24/pubmed PY - 2010/12/16/medline SP - 705 EP - 14 JF - Biochemistry and cell biology = Biochimie et biologie cellulaire JO - Biochem Cell Biol VL - 88 IS - 4 N2 - Adenosine can exhibit cytotoxic activity in vivo and in vitro, though its mechanisms are still uncertain. In this study, we investigated the adenosine-mediated apoptotic signaling pathway and the role of NF-kappaB in human hepatocellular carcinoma HepG2 cells. HepG2 cells were treated with different concentrations of adenosine for 12-48 h, and the effect of adenosine on cell proliferation was evaluated by MTT assay. The cytotoxicity of adenosine alone or in combination with an NF-kappaB inhibitor, pyrrolidine dithiocarbamate (PDTC), was also evaluated by MTT assay and the mode of cell death was detected by Hoechst 33342 staining. Cell cycle progress was performed by flow cytometry with PI staining. The protein expressions of Bcl-2, p53, NF-kappaB subunit p65, and caspase-3 were assayed by Western blot. Caspase-3 activity was measured by spectrophotomteric assay. The results showed that adenosine significantly reduced the viability of HepG2 cells in a dose- and time-dependent manner, with IC 50 (24 and 48 h) of 2.52 and 1.89 mmol x L(-1), respectively. The apoptotic index (percentage of sub-G1 phase) of HepG2 cells in adenosine treatment alone for 12 and 24 h or in combination with PDTC were 8.30%, 22.32% and 20.18%, 30.89%, respectively. All of them were higher than that in the control group (0.81%, p < 0.01). The characteristic changes of cell apoptosis (chromatin condensation and sub-G1 peak) were observed under fluorescent microscopy and flow cytometry. We also found that the apoptotic process triggered by adenosine was involved in G0-G1 cell-cycle arrest, enhanced the activity of caspase-3, upregulated p53 and NF-kappaB p65 expression, and downregulated Bcl-2 expression. Inhibition of NF-kappaB by PDTC decreased NF-kappaB p65 expression, enhanced cell apoptosis ratio, and increased caspase-3 activity. NF-kappaB may play an anti-apoptosis role in adenosine-induced HepG2 cytotoxicity. SN - 1208-6002 UR - https://www.unboundmedicine.com/medline/citation/20651843/Involvement_of_NF_kappaB_activation_in_the_apoptosis_induced_by_extracellular_adenosine_in_human_hepatocellular_carcinoma_HepG2_cells_ L2 - https://cdnsciencepub.com/doi/10.1139/O10-008?url_ver=Z39.88-2003&amp;rfr_id=ori:rid:crossref.org&amp;rfr_dat=cr_pub=pubmed DB - PRIME DP - Unbound Medicine ER -