Serum starvation-induces down-regulation of Bcl-2/Bax confers apoptosis in tongue coating-related cells in vitro

Huang,Yanhua; Fu,Ziyi; Dong,Wei; Zhang,Zhenming; Mu,Jinquan; Zhang,Junfeng

doi:10.3892/mmr.2018.8512

Serum starvation-induces down-regulation of Bcl-2/Bax confers apoptosis in tongue coating-related cells in vitro

Authors:
- Yanhua Huang
- Ziyi Fu
- Wei Dong
- Zhenming Zhang
- Jinquan Mu
- Junfeng Zhang
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Affiliations: Department of Stomatology, Nanjing Maternity and Child Health Medical Institute, Obstetrics and Gynecology Hospital Affiliated to Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China, Department of Pathogen and Immunology, Discipline of Chinese and Western Integrative Medicine, College of Basic Medicine, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, P.R. China, Department of Stomatology, Ningbo Dental Hospital, Ningbo, Zhejiang 315100, P.R. China
Published online on: January 29, 2018 https://doi.org/10.3892/mmr.2018.8512
Pages: 5057-5064
Copyright: © Huang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Tongue squamous epithelial cells are the main component of tongue coating, with proliferation, differentiation and apoptosis being the root cause of the formation and maintenance of tongue coating. The present study aimed to explore the molecular mechanism by which serum influences tongue coating, to enable a better understanding for future investigations. Tongue carcinoma squamous cells were exposed to serum‑starvation in vitro. Cellular proliferation and apoptosis were observed by using 3‑[4,5‑dimethyl‑2‑thiazolyl]‑2,5‑diphenyl‑2‑H‑tetrazolium bromide (MTT) assay, flow cytometry, Hoechst staining, scanning electron microscope (SEM), transmission electron microscope (TEM), and by measuring the expression ratio of B‑cell lymphoma 2 apoptosis regulator (Bcl‑2)/Bcl‑2 associated protein X apoptosis regulator (Bax) mRNA and protein by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blotting, respectively. MTT assays revealed that serum‑starvation results in suppression of cellular proliferation, while flow cytometry data revealed that serum‑starvation induces cell cycle arrest at G1 phase and increases apoptosis. In addition, chromatin condensation and membrane blebbing were observed through Hoechst staining, TEM and SEM. The Bcl‑2/Bax ratio was found to be significantly decreased in cells that had undergone serum‑starvation by both RT‑qPCR and western blotting analysis, further indicating that serum‑starvation induces apoptosis. Therefore, tongue carcinoma squamous cells in a serum‑free medium can simulate apoptosis related to the formation of tongue coating, which may offer guidance for future investigations about other factors.

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1	Jiang B, Liang X, Chen Y, Ma T, Liu L, Li J, Jiang R, Chen T, Zhang X and Li S: Integrating next-generation sequencing and traditional tongue diagnosis to determine tongue coating microbiome. Sci Rep. 2:9362012. View Article : Google Scholar : PubMed/NCBI
2	Zhao Y, Gou XJ, Dai JY, Peng JH, Feng Q, Sun SJ, Cao HJ, Zheng NN, Fang JW, Jiang J, et al: Differences in metabolites of different tongue coatings in patients with chronic hepatitis B. Evid Based Complement Alternat Med. 2013:2049082013. View Article : Google Scholar : PubMed/NCBI
3	Singha KB, Konar S, Mondal MK and Das J: Scanning electron microscopic study of the human fungiform papillae. J Anat Soc India. 59:154–157. 2010. View Article : Google Scholar
4	Jackowiak H and Godynicki S: The scanning electron microscopic study of lingual papillae in the silver fox (Vulpes vulpes fulva, Desmarest, 1820). Ann Anat. 186:179–183. 2004. View Article : Google Scholar : PubMed/NCBI
5	Kawasaki K, Porntaveetus T, Oommen S, Ghafoor S, Kawasaki M, Otsuka-Tanaka Y, Blackburn J, Kessler JA, Sharpe PT and Ohazama A: Bmp signalling in filiform tongue papillae development. Arch Oral Biol. 57:805–813. 2012. View Article : Google Scholar : PubMed/NCBI
6	Da Silva AE, Rados PV, Da Silva Lauxen I, Gedoz L, Villarinho EA and Fontanella V: Nuclear changes in tongue epithelial cells following panoramic radiography. Mutat Res. 632:121–125. 2007. View Article : Google Scholar : PubMed/NCBI
7	Gonçalves AS, Arantes DA, Bernardes VF, Jaeger F, Silva JM, Silva TA, Aguiar MC and Batista AC: Immunosuppressive mediators of oral squamous cell carcinoma in tumour samples and saliva. Hum Immunol. 76:52–58. 2015. View Article : Google Scholar : PubMed/NCBI
8	Zhang A, Sun H, Wang P and Wang X: Salivary proteomics in biomedical research. Clin Chim Acta. 415:261–265. 2013. View Article : Google Scholar : PubMed/NCBI
9	Sun ZM, Zhao J, Qian P, Wang YQ, Zhang WF, Guo CR, Pang XY, Wang SC, Li FF and Li Q: Metabolic markers and microecological characteristics of tongue coating in patients with chronic gastritis. BMC Complement Altern Med. 13:2272013. View Article : Google Scholar : PubMed/NCBI
10	Pérez-Garijo A and Steller H: Spreading the word: Non-autonomous effects of apoptosis during development, regeneration and disease. Development. 142:3253–3262. 2015. View Article : Google Scholar : PubMed/NCBI
11	Martin LJ: Neuronal cell death in nervous system development, disease, and injury (Review). Int J Mol Med. 7:455–478. 2001.PubMed/NCBI
12	Mosmann T: Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. J Immunol Methods. 65:55–63. 1983. View Article : Google Scholar : PubMed/NCBI
13	Ozeki N, Mogi M, Nakamura H and Togari A: Differential expression of the Fas-Fas ligand system on cytokine-induced apoptotic cell death in mouse osteoblastic cells. Arch Oral Biol. 47:511–517. 2002. View Article : Google Scholar : PubMed/NCBI
14	Tano T, Okamoto M, Kan S, Nakashiro K, Shimodaira S, Koido S, Homma S, Sato M, Fujita T, Kawakami Y and Hamakawa H: Prognostic impact of expression of Bcl-2 and Bax genes in circulating immune cells derived from patients with head and neck carcinoma. Neoplasia. 15:305–314. 2013. View Article : Google Scholar : PubMed/NCBI
15	Jiang W, Wang Q, Chen S, Gao S, Song L, Liu P and Huang W: Influenza A virus NS1 induces G0/G1 cell cycle arrest by inhibiting the expression and activity of RhoA protein. J Virol. 87:3039–3052. 2013. View Article : Google Scholar : PubMed/NCBI
16	Zhang W, Wang F, Xu P, Miao C, Zeng X, Cui X, Lu C, Xie H, Yin H, Chen F, et al: Perfluorooctanoic acid stimulates breast cancer cells invasion and up-regulates matrix metalloproteinase-2/-9 expression mediated by activating NF-κB. Toxicol Lett. 229:118–125. 2014. View Article : Google Scholar : PubMed/NCBI
17	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
18	Cerella C, Sobolewski C, Chateauvieux S, Henry E, Schnekenburger M, Ghelfi J, Dicato M and Diederich M: COX-2 inhibitors block chemotherapeutic agent-induced apoptosis prior to commitment in hematopoietic cancer cells. Biochem Pharmacol. 82:1277–1290. 2011. View Article : Google Scholar : PubMed/NCBI
19	Brunk UT, Dalen H, Roberg K and Hellquist HB: Photo-oxidative disruption of lysosomal membranes causes apoptosis of cultured human fibroblasts. Free Radic Biol Med. 23:616–626. 1997. View Article : Google Scholar : PubMed/NCBI
20	Zhang H, An X, Zhou Y, Zhang B, Zhang S, Li D, Chen Z, Li Y, Bai S, Lv J, et al: Effect of oxidative stress induced by Brevibacterium sp. BS01 on a HAB causing species-Alexandrium tamarense. PLoS One. 8:e630182013. View Article : Google Scholar : PubMed/NCBI
21	Izumi K, Nakajima T, Maeda T, Cheng J and Saku T: Adenosquamous carcinoma of the tongue: Report of a case with histochemical, immunohistochemical, and ultrastructural study and review of the literature. Oral Surg Oral Med Oral Pathol Oral Radiol Endod. 85:178–184. 1998. View Article : Google Scholar : PubMed/NCBI
22	Görögh T, Lippert BM, Gottschlich S, Folz B and Werner JA: Establishment and characterization of 2 cell lines of squamous epithelial carcinoma of the mouth floor and tongue. Laryngorhinootologie. 74:684–690. 1995. View Article : Google Scholar : PubMed/NCBI
23	Chen AK, Reuveny S and Oh SK: Application of human mesenchymal and pluripotent stem cell microcarrier cultures in cellular therapy: Achievements and future direction. Biotechnol Adv. 31:1032–1046. 2013. View Article : Google Scholar : PubMed/NCBI
24	Wilmarth PA, Riviere MA, Rustvold DL, Lauten JD, Madden TE and David LL: Two-dimensional liquid chromatography study of the human whole saliva proteome. J Proteome Res. 3:1017–1023. 2004. View Article : Google Scholar : PubMed/NCBI
25	Yu YS, Sun XS, Jiang HN, Han Y, Zhao CB and Tan JH: Studies of the cell cycle of in vitro cultured skin fibroblasts in goats: Work in progress. Theriogenology. 59:1277–1289. 2003. View Article : Google Scholar : PubMed/NCBI
26	Papa F, Scacco S, Vergari R, De Benedittis M, Petruzzi M, Lo Muzio L and Serpico R: Expression and subcellular distribution of Bcl-2 and BAX proteins in serum-starved human keratinocytes and mouth carcinoma epidermoid cultures. Life Sci. 73:2865–2872. 2003. View Article : Google Scholar : PubMed/NCBI