Nodal induced by hypoxia exposure contributes to dacarbazine resistance and the maintenance of stemness in melanoma cancer stem‑like cells

Li,Hong; Chen,Junjie; Wang,Xiao; He,Mei; Zhang,Zhenyu; Cen,Ying

doi:10.3892/or.2018.6387

Nodal induced by hypoxia exposure contributes to dacarbazine resistance and the maintenance of stemness in melanoma cancer stem‑like cells

Authors:
- Hong Li
- Junjie Chen
- Xiao Wang
- Mei He
- Zhenyu Zhang
- Ying Cen
View Affiliations

Affiliations: Department of Burns and Plastic Surgery Unit, West China Hospital of Sichuan University, Chengdu, Sichuan 610041, P.R. China, Department of Burns and Plastic Surgery Unit, West China Hospital of Sichuan University, Chengdu, Sichuan 610041, P.R. China, Department of Burns and Plastic Surgery, The Second People's Hospital of Chengdu, Chengdu, Sichuan 610017, P.R. China
Published online on: April 23, 2018 https://doi.org/10.3892/or.2018.6387
Pages: 2855-2864

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

Nodal signaling has a critical role in the processes of embryogenesis and is necessary for maintaining cell stemness. However, its upregulation in melanoma is positively correlated with malignant potential, including cancer progression, metastasis and recurrence, and a recent report has revealed its role in promoting self‑renewal capacity in melanoma. Our study aimed to explore the effects of hypoxia exposure, which is one of the main causes of chemoresistance in melanoma, on the physiological processes of melanoma cancer stem‑like cells (CSCs) via regulating Nodal. A375 CSCs were enriched by culturing in serum‑free medium (SFM) and were analyzed for the expression levels of Nodal and its correlated proteins by semi‑quantitative western blotting. Lentiviral‑packaged Nodal coding sequence or short‑hairpin RNA (shRNA) was employed. After hypoxia exposure, the effects on glucose uptake, ATP production and O2 consumption were detected, and whether Nodal contributed to the proliferation, invasion, colony formation, self‑renewal capacity and chemoresistance was evaluated. We demonstrated that hypoxia exposure induced Nodal expression and activated the Smad2/3 pathway in A375 CSCs. Hypoxic‑induced Nodal partially promoted dacarbazine resistance, promoted invasion and self‑renewal capacity, but not proliferation which was further confirmed using Nodal knockdown. Blockage of Nodal signaling activity with the small‑molecule inhibitor SB431542 partially reversed Nodal‑induced chemoresistance. Nodal knockdown further sensitized A375 CSCs to dacarbazine after SB431542 pretreatment, indicating the involvement of proNodal in dacarbazine resistance. The introduction of mut‑proNodal induced chemoresistance further confirming the role of proNodal in this process. Taken together, our results demonstrated that Nodal induced by hypoxia exposure induced a malignant phenotype and chemoresistance in A375 CSCs, and proNodal also contributed to these processes, indicating that Nodal may be a potential therapeutic target for melanoma.

View Figures

View References

1	Miller AJ and Mihm MC: Melanoma. N Engl J Med. 355:51–65. 2006. View Article : Google Scholar : PubMed/NCBI
2	Godar DE: Worldwide increasing incidences of cutaneous malignant melanoma. J Skin Cancer. 2011:8584252011. View Article : Google Scholar : PubMed/NCBI
3	Reed KB, Brewer JD, Lohse CM, Bringe KE, Pruitt CN and Gibson LE: Increasing incidence of melanoma among young adults: An epidemiological study in Olmsted County, Minnesota. Mayo Clin Proc. 87:328–334. 2012. View Article : Google Scholar : PubMed/NCBI
4	Bedia C, Casas J, Andrieu-Abadie N, Fabriàs G and Levade T: Acid ceramidase expression modulates the sensitivity of A375 melanoma cells to decarbazine. J Biol Chem. 286:28200–28209. 2011. View Article : Google Scholar : PubMed/NCBI
5	Mouawad R, Sebert M, Michels J, Bloch J, Spano JP and Khayat D: Treatment for metastatic malignant melanoma: Old drugs and new strategies. Crit Rev Oncol Hematol. 74:27–39. 2010. View Article : Google Scholar : PubMed/NCBI
6	Brennan J, Norris DP and Robertson EJ: Nodal activity in the node governs left-right asymmetry. Genes Dev. 16:2339–2344. 2002. View Article : Google Scholar : PubMed/NCBI
7	Takenaga M, Fukumoto M and Hori Y: Regulated nodal signaling promotes differentiation of the definitive endoderm and mesoderm from ES cells. J Cell Sci. 120:2078–2090. 2007. View Article : Google Scholar : PubMed/NCBI
8	Topczewska JM, Postovit LM, Margaryan NV, Sam A, Hess AR, Wheaton WW, Nickoloff BJ, Topczewski J and Hendrix MJ: Embryonic and tumorigenic pathways converge via Nodal signaling: Role in melanoma aggressiveness. Nat Med. 12:925–932. 2006. View Article : Google Scholar : PubMed/NCBI
9	Lee CC, Jan HJ, Lai JH, Ma HI, Hueng DY, Lee YC, Cheng YY, Liu LW, Wei HW and Lee HM: Nodal promotes growth and invasion in human gliomas. Oncogene. 29:3110–3123. 2010. View Article : Google Scholar : PubMed/NCBI
10	Lawrence MG, Margaryan NV, Loessner D, Collins A, Kerr KM, Turner M, Seftor EA, Stephens CR, Lai J; APC BioResource, ; et al: Reactivation of embryonic nodal signaling is associated with tumor progression and promotes the growth of prostate cancer cells. Prostate. 71:1198–1209. 2011. View Article : Google Scholar : PubMed/NCBI
11	Quail DF, Zhang G, Walsh LA, Siegers GM, Dieters-Castator DZ, Findlay SD, Broughton H, Putman DM, Hess DA and Postovit LM: Embryonic morphogen nodal promotes breast cancer growth and progression. PLoS One. 7:e482372012. View Article : Google Scholar : PubMed/NCBI
12	Papageorgiou I, Nicholls PK, Wang F, Lackmann M, Makanji Y, Salamonsen LA, Robertson DM and Harrison CA: Expression of nodal signalling components in cycling human endometrium and in endometrial cancer. Reprod Biol Endocrinol. 7:1222009. View Article : Google Scholar : PubMed/NCBI
13	Lonardo E, Hermann PC, Mueller MT, Huber S, Balic A, Miranda-Lorenzo I, Zagorac S, Alcala S, Rodriguez-Arabaolaza I, Ramirez JC, et al: Nodal/Activin signaling drives self-renewal and tumorigenicity of pancreatic cancer stem cells and provides a target for combined drug therapy. Cell Stem Cell. 9:433–446. 2011. View Article : Google Scholar : PubMed/NCBI
14	Fuerer C, Nostro MC and Constam DB: Nodal·Gdf1 heterodimers with bound prodomains enable serum-independent nodal signaling and endoderm differentiation. J Biol Chem. 289:17854–17871. 2014. View Article : Google Scholar : PubMed/NCBI
15	Ellis PS, Burbridge S, Soubes S, Ohyama K, Ben-Haim N, Chen C, Dale K, Shen MM, Constam D and Placzek M: ProNodal acts via FGFR3 to govern duration of Shh expression in the prechordal mesoderm. Development. 142:3821–3832. 2015. View Article : Google Scholar : PubMed/NCBI
16	Cabarcas SM, Mathews LA and Farrar WL: The cancer stem cell niche-there goes the neighborhood? Int J Cancer. 129:2315–2327. 2011. View Article : Google Scholar : PubMed/NCBI
17	Hanahan D and Weinberg RA: Hallmarks of cancer: The next generation. Cell. 144:646–674. 2011. View Article : Google Scholar : PubMed/NCBI
18	Zhu H, Wang D, Liu Y, Su Z, Zhang L, Chen F, Zhou Y, Wu Y, Yu M, Zhang Z and Shao G: Role of the Hypoxia-inducible factor-1 alpha induced autophagy in the conversion of non-stem pancreatic cancer cells into CD133⁺ pancreatic cancer stem-like cells. Cancer Cell Int. 13:1192013. View Article : Google Scholar : PubMed/NCBI
19	Quail DF, Taylor MJ, Walsh LA, Dieters-Castator D, Das P, Jewer M, Zhang G and Postovit LM: Low oxygen levels induce the expression of the embryonic morphogen Nodal. Mol Biol Cell. 22:4809–4821. 2011. View Article : Google Scholar : PubMed/NCBI
20	Zhou S, Kurt-Jones EA, Cerny AM, Chan M, Bronson RT and Finberg RW: MyD88 intrinsically regulates CD4 T-cell responses. J Virol. 83:1625–1634. 2009. View Article : Google Scholar : PubMed/NCBI
21	dos Santos SC, Tenreiro S, Palma M, Becker J and Sá-Correia I: Transcriptomic profiling of the Saccharomyces cerevisiae response to quinine reveals a glucose limitation response attributable to drug-induced inhibition of glucose uptake. Antimicrob Agents Chemother. 53:5213–5223. 2009. View Article : Google Scholar : PubMed/NCBI
22	Walsh MC, Smits HP, Scholte M and van Dam K: Affinity of glucose transport in Saccharomyces cerevisiae is modulated during growth on glucose. J Bacteriol. 176:953–958. 1994. View Article : Google Scholar : PubMed/NCBI
23	Constam DB and Robertson EJ: Regulation of bone morphogenetic protein activities by prodomains and proprotein convertases. J Cell Biol. 144:139–149. 1999. View Article : Google Scholar : PubMed/NCBI
24	Vo BT and Khan SA: Expression of nodal and nodal receptors in prostate stem cells and prostate cancer cells: Autocrine effects on cell proliferation and migration. Prostate. 71:1084–1096. 2011. View Article : Google Scholar : PubMed/NCBI
25	Lai JH, Jan HJ, Liu LW, Lee CC, Wang SG, Hueng DY, Cheng YY, Lee HM and Ma HI: Nodal regulates energy metabolism in glioma cells by inducing expression of hypoxia-inducible factor 1α. Neuro Oncol. 15:1330–1341. 2013. View Article : Google Scholar : PubMed/NCBI
26	Papandreou I, Cairns RA, Fontana L, Lim AL and Denko NC: HIF-1 mediates adaptation to hypoxia by actively downregulating mitochondrial oxygen consumption. Cell Metab. 3:187–197. 2006. View Article : Google Scholar : PubMed/NCBI
27	Wang F, Zhang GY, Xing T, Lu ZY, Li JH, Peng C, Liu GH and Wang NS: Renalase contributes to the renal protection of delayed ischaemic preconditioning via the regulation of hypoxia-inducible factor-1α. J Cell Mol Med. 19:1400–1409. 2015. View Article : Google Scholar : PubMed/NCBI
28	Jian SL, Chen WW, Su YC, Su YW, Chuang TH, Hsu SC and Huang LR: Glycolysis regulates the expansion of myeloid-derived suppressor cells in tumor-bearing hosts through prevention of ROS-mediated apoptosis. Cell Death Dis. 8:e27792017. View Article : Google Scholar : PubMed/NCBI
29	Elliott RL and Blobe GC: Role of transforming growth factor beta in human cancer. J Clin Oncol. 23:2078–2093. 2005. View Article : Google Scholar : PubMed/NCBI
30	Glasgow E and Mishra L: Transforming growth factor-beta signaling and ubiquitinators in cancer. Endocr Relat Cancer. 15:59–72. 2008. View Article : Google Scholar : PubMed/NCBI
31	Beck S, Le Good JA, Guzman M, Ben Haim N, Roy K, Beermann F and Constam DB: Extraembryonic proteases regulate nodal signaling during gastrulation. Nat Cell Biol. 4:981–985. 2002. View Article : Google Scholar : PubMed/NCBI
32	Ben-Haim N, Lu C, Guzman-Ayala M, Pescatore L, Mesnard D, Bischofberger M, Naef F, Robertson EJ and Constam DB: The nodal precursor acting via activing receptors induces mesoderm by maintaining a source of its convertases and BMP4. Dev Cell. 11:313–323. 2006. View Article : Google Scholar : PubMed/NCBI
33	Eimon PM and Harland RM: Effects of heterodimerization and proteolytic processing on derriere and nodal activity: Implications for mesoderm induction in xenopus. Development. 129:3089–3103. 2002.PubMed/NCBI
34	Strizzi L, Postovit LM, Margaryan NV, Seftor EA, Abbott DE, Seftor RE, Salomon DS and Hendrix MJ: Emerging roles of nodal and Cripto-1: From embryogenesis to breast cancer progression. Breast Dis. 29:91–103. 2008. View Article : Google Scholar : PubMed/NCBI
35	Shen MM: Nodal signaling: Developmental roles and regulation. Development. 134:1023–1034. 2007. View Article : Google Scholar : PubMed/NCBI
36	Duan W, Li R, Ma J, Lei J, Xu Q, Jiang Z, Nan L, Li X, Wang Z, Huo X, et al: Overexpression of nodal induces a metastatic phenotype in pancreatic cancer cells via the Smad2/3 pathway. Oncotarget. 6:1490–1506. 2015. View Article : Google Scholar : PubMed/NCBI
37	Strizzi L, Postovit LM, Margaryan NV, Lipavsky A, Gadiot J, Blank C, Seftor RE, Seftor EA and Hendrix MJ: Nodal as a biomarker for melanoma progression and a new therapeutic target for clinical intervention. Expert Rev Dermatol. 4:67–78. 2009. View Article : Google Scholar : PubMed/NCBI
38	Postovit LM, Margaryan NV, Seftor EA, Kirschmann DA, Lipavsky A, Wheaton WW, Abbott DE, Seftor RE and Hendrix MJ: Human embryonic stem cell microenvironment suppresses the tumorigenic phenotype of aggressive cancer cells. Proc Natl Acad Sci USA. 105:4329–4334. 2008. View Article : Google Scholar : PubMed/NCBI
39	Brennan J, Lu CC, Norris DP, Rodriguez TA, Beddington RS and Robertson EJ: Nodal signaling in the epiblast patterns the early mouse embryo. Nature. 411:965–969. 2001. View Article : Google Scholar : PubMed/NCBI
40	Cioffi M, Trabulo SM, Sanchez-Ripoll Y, Miranda-Lorenzo I, Lonardo E, Dorado J, Vieira Reis C, Ramirez JC, Hidalgo M, Aicher A, et al: The miR-17-92 cluster counteracts quiescence and chemoresistance in a distinct subpopulation of pancreatic cancer stem cells. Gut. 64:1936–1948. 2015. View Article : Google Scholar : PubMed/NCBI