Betulinic acid induces apoptosis by regulating PI3K/Akt signaling and mitochondrial pathways in human cervical cancer cells

Xu,Tao; Pang,Qiuying; Wang,Yang; Yan,Xiufeng

doi:10.3892/ijmm.2017.3163

Betulinic acid induces apoptosis by regulating PI3K/Akt signaling and mitochondrial pathways in human cervical cancer cells

Authors:
- Tao Xu
- Qiuying Pang
- Yang Wang
- Xiufeng Yan
View Affiliations

Affiliations: Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Harbin, Heilongjiang 150040, P.R. China
Published online on: September 28, 2017 https://doi.org/10.3892/ijmm.2017.3163
Pages: 1669-1678
Copyright: © Xu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Betulinic acid (BA), a potential anticancer compound, could induce apoptosis in human cervical cancer (HeLa) cells, but its mechanism has yet to be fully elucidated. The present study was focused on deciphering the detailed molecular mechanism of BA-induced apoptosis. In the present study, results indicated that BA was highly effective against HeLa cells via induction of time-dependent apoptosis, and the authors demonstrated that the BA treatment acted through downregulating a phosphatidylinositol 3-kinase (PI3K) subunit and suppressing the Akt phosphorylation at Thr308 and Ser473 after increasing the generation of intracellular reactive oxygen species. Then, BA induced cell cycle arrest at the G0/G1 phase, which was consistent with the cell cycle-related protein results in which BA significantly enhanced the expression of p27Kip and p21Waf1/Cip1 in HeLa cells. This target-specific inhibition was associated with mitochondrial apoptosis, as reflected by the increased expression of Bad and caspase-9, the generation of reactive oxygen species (ROS) and the decline in mitochondrial membrane potential. Moreover, preincubation of the cells with glutathione (antioxidant) blocked the process of apoptosis, prevented the phosphorylation of downstream substrates. These results established that ROS acted as a key factor to effect apoptosis by BA treatment in HeLa cells. Therefore, these findings demonstrated that BA induced apoptosis in HeLa cells by downregulating the expression of PI3K/Akt signaling molecules via ROS, and triggering a mitochondrial pathway.

View Figures

View References

1	Pisha E, Chai H, Lee IS, Chagwedera TE, Farnsworth NR, Cordell GA, Beecher CW, Fong HH, Kinghorn AD, Brown DM, et al: Discovery of betulinic acid as a selective inhibitor of human melanoma that functions by induction of apoptosis. Nat Med. 1:1046–1051. 1995. View Article : Google Scholar : PubMed/NCBI
2	Zuco V, Supino R, Righetti SC, Cleris L, Marchesi E, Gambacorti-Passerini C and Formelli F: Selective cytotoxicity of betulinic acid on tumor cell lines, but not on normal cells. Cancer Lett. 175:17–25. 2002. View Article : Google Scholar
3	Kroemer G, Galluzzi L and Brenner C: Mitochondrial membrane permeabilization in cell death. Physiol Rev. 87:99–163. 2007. View Article : Google Scholar : PubMed/NCBI
4	Rabi T, Shukla S and Gupta S: Betulinic acid suppresses constitutive and TNFalpha-induced NF-kappaB activation and induces apoptosis in human prostate carcinoma PC-3 cells. Mol Carcinog. 47:964–973. 2008. View Article : Google Scholar : PubMed/NCBI
5	Eichenmüller M, von Schweinitz D and Kappler R: Betulinic acid treatment promotes apoptosis in hepatoblastoma cells. Int J Oncol. 35:873–879. 2009.PubMed/NCBI
6	Tan DF, Li Q, Rammath N, Beck A, Wiseman S, Anderson T, al-Salameh A, Brooks J and Bepler G: Prognostic significance of expression of p53 oncoprotein in primary (stage I–IIIa) non-small cell lung cancer. Anticancer Res. 23:1665–1672. 2003.PubMed/NCBI
7	Mullauer FB, Kessler JH and Medema JP: Betulinic acid induces cytochrome c release and apoptosis in a Bax/Bak-independent, permeability transition pore dependent fashion. Apoptosis. 14:191–202. 2009. View Article : Google Scholar
8	Eichenmüller M, Hemmerlein B, von Schweinitz D and Kappler R: Betulinic acid induces apoptosis and inhibits hedgehog signalling in rhabdomyosarcoma. Br J Cancer. 103:43–51. 2010. View Article : Google Scholar : PubMed/NCBI
9	Cheng L, Xia TS, Wang YF, Zhou W, Liang XQ, Xue JQ, Shi L, Wang Y and Ding Q: The apoptotic effect of D Rhamnose β-hederin, a novel oleanane-type triterpenoid saponin on breast cancer cells. PLoS One. 9:e908482014. View Article : Google Scholar
10	Kim JA, Åberg C, de Cárcer G, Malumbres M, Salvati A and Dawson KA: Low dose of amino-modified nanoparticles induces cell cycle arrest. ACS Nano. 7:7483–7494. 2013. View Article : Google Scholar : PubMed/NCBI
11	Liang QH, Liu Y, Wu SS, Cui RR, Yuan LQ and Liao EY: Ghrelin inhibits the apoptosis of MC3T3-E1 cells through ERK and AKT signaling pathway. Toxicol Appl Pharmacol. 272:591–597. 2013. View Article : Google Scholar : PubMed/NCBI
12	Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y and Greenberg ME: Akt phosphorylation of BAD couples survival signals to the cell-intrinsic death machinery. Cell. 91:231–241. 1997. View Article : Google Scholar : PubMed/NCBI
13	Roy B, Pattanaik AK, Das J, Bhutia SK, Behera B, Singh P and Maiti TK: Role of PI3K/Akt/mTOR and MEK/ERK pathway in Concanavalin A induced autophagy in HeLa cells. Chem Biol Interact. 210:96–102. 2014. View Article : Google Scholar : PubMed/NCBI
14	Xu T, Pang Q, Zhou D, Zhang A, Luo S, Wang Y and Yan X: Proteomic investigation into betulinic acid-induced apoptosis of human cervical cancer HeLa cells. PLoS One. 9:e1057682014. View Article : Google Scholar : PubMed/NCBI
15	Meinig JM and Peterson BR: Anticancer/antiviral agent Akt inhibitor-IV massively accumulates in mitochondria and potently disrupts cellular bioenergetics. ACS Chem Biol. 10:570–576. 2015. View Article : Google Scholar :
16	Ma K, Liu Y, Zhu Q, Liu CH, Duan JL, Tan BK and Zhu YZ: H2S donor, S-propargyl-cysteine, increases CSE in SGC-7901 and cancer-induced mice: Evidence for a novel anti-cancer effect of endogenous H2S? PLoS One. 6:e205252011. View Article : Google Scholar : PubMed/NCBI
17	Willis SN, Chen L, Dewson G, Wei A, Naik E, Fletcher JI, Adams JM and Huang DC: Proapoptotic Bak is sequestered by Mcl-1 and Bcl-xL, but not Bcl-2, until displaced by BH3-only proteins. Genes Dev. 19:1294–1305. 2005. View Article : Google Scholar : PubMed/NCBI
18	Tian F, Ding D and Li D: Fangchinoline targets PI3K and suppresses PI3K/AKT signaling pathway in SGC7901 cells. Int J Oncol. 46:2355–2363. 2015. View Article : Google Scholar : PubMed/NCBI
19	Goo CK, Lim HY, Ho QS, Too HP, Clement MV and Wong KP: PTEN/Akt signaling controls mitochondrial respiratory capacity through 4E-BP1. PLoS One. 7:e458062012. View Article : Google Scholar : PubMed/NCBI
20	Cantley LC: The phosphoinositide 3-kinase pathway. Science. 296:1655–1657. 2002. View Article : Google Scholar : PubMed/NCBI
21	Thorpe LM, Yuzugullu H and Zhao JJ: PI3K in cancer: Divergent roles of isoforms, modes of activation and therapeutic targeting. Nat Rev Cancer. 15:7–24. 2015. View Article : Google Scholar :
22	Brazil DP and Hemmings BA: Ten years of protein kinase B signalling: A hard Akt to follow. Trends Biochem Sci. 26:657–664. 2001. View Article : Google Scholar : PubMed/NCBI
23	del Peso L, González-García M, Page C, Herrera R and Nuñez G: Interleukin-3-induced phosphorylation of BAD through the protein kinase Akt. Science. 278:687–689. 1997. View Article : Google Scholar : PubMed/NCBI
24	Reed SI: Keeping p27(Kip1) in the cytoplasm: A second front in cancer's war on p27. Cell Cycle. 1:389–390. 2002. View Article : Google Scholar
25	Nakayama KI and Nakayama K: Ubiquitin ligases: Cell-cycle control and cancer. Nat Rev Cancer. 6:369–381. 2006. View Article : Google Scholar : PubMed/NCBI
26	Ortega S, Malumbres M and Barbacid M: Cyclin D-dependent kinases, INK4 inhibitors and cancer. Biochim Biophys Acta. 1602:73–87. 2002.PubMed/NCBI
27	Lee SK, Zhang W and Sanderson BJ: Selective growth inhibition of human leukemia and human lymphoblastoid cells by resveratrol via cell cycle arrest and apoptosis induction. J Agric Food Chem. 56:7572–7577. 2008. View Article : Google Scholar : PubMed/NCBI
28	Kang S, Dong SM, Kim BR, Park MS, Trink B, Byun HJ and Rho SB: Thioridazine induces apoptosis by targeting the PI3K/Akt/mTOR pathway in cervical and endometrial cancer cells. Apoptosis. 17:989–997. 2012. View Article : Google Scholar : PubMed/NCBI
29	Rzeski W, Stepulak A, Szymański M, Sifringer M, Kaczor J, Wejksza K, Zdzisińska B and Kandefer-Szerszeń M: Betulinic acid decreases expression of bcl-2 and cyclin D1, inhibits proliferation, migration and induces apoptosis in cancer cells. Naunyn Schmiedebergs Arch Pharmacol. 374:11–20. 2006. View Article : Google Scholar : PubMed/NCBI
30	Zhang X, Lu H, Wang Y, Liu C, Zhu W, Zheng S and Wan F: Taurine induces the apoptosis of breast cancer cells by regulating apoptosis-related proteins of mitochondria. Int J Mol Med. 35:218–226. 2015. View Article : Google Scholar
31	Zou Y, Li Q, Jiang L, Guo C, Li Y, Yu Y, Li Y, Duan J and Sun Z: DNA hypermethylation of CREB3L1 and Bcl-2 associated with the mitochondrial-mediated apoptosis via PI3K/Akt pathway in human BEAS-2B cells exposure to silica nanoparticles. PLoS One. 11:e01584752016. View Article : Google Scholar : PubMed/NCBI
32	Khan I, Guru SK, Rath SK, Chinthakindi PK, Singh B, Koul S, Bhushan S and Sangwan PL: A novel triazole derivative of betulinic acid induces extrinsic and intrinsic apoptosis in human leukemia HL-60 cells. Eur J Med Chem. 108:104–116. 2016. View Article : Google Scholar
33	Hamanaka RB and Chandel NS: Mitochondrial reactive oxygen species regulate cellular signaling and dictate biological outcomes. Trends Biochem Sci. 35:505–513. 2010. View Article : Google Scholar : PubMed/NCBI
34	Behrend L, Henderson G and Zwacka RM: Reactive oxygen species in oncogenic transformation. Biochem Soc Trans. 31:1441–1444. 2003. View Article : Google Scholar : PubMed/NCBI
35	Deeb D, Gao X, Jiang H, Janic B, Arbab AS, Rojanasakul Y, Dulchavsky SA and Gautam SC: Oleanane triterpenoid CDDO-Me inhibits growth and induces apoptosis in prostate cancer cells through a ROS-dependent mechanism. Biochem Pharmacol. 79:350–360. 2010. View Article : Google Scholar
36	Chandel NS: Mitochondrial complex III: An essential component of universal oxygen sensing machinery? Respir Physiol Neurobiol. 174:175–181. 2010. View Article : Google Scholar : PubMed/NCBI
37	Chandel NS: Mitochondrial regulation of oxygen sensing. Adv Exp Med Biol. 661:339–354. 2010. View Article : Google Scholar : PubMed/NCBI