Stress activated p38 MAPK regulates cell cycle via AP-1 factors in areca extract exposed human lung epithelial cells
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Areca nut chewing habits are associated with several oral manifestations like leukoplakia, submucous fibrosis and oral squamous cell carcinoma. Although numerous evidence on areca toxicity is known but the mechanistic pathway of disease causation is to be studied. Aqueous areca nut extract treated A549 cells showed reduced cell viability by 48 h with IC50 value of 0.50%. The toxic nature of areca nut induced the production of reactive oxygen species with decreased anti-oxidant glutathione S transferase levels lead to altered redox homeostasis. PCR studies showed decreased mRNA levels of Jun and Fos AP-1 subunits on extract treatment by 48 h. The protein levels of PCNA, CDK4, RB, p53, c-Jun and c-Fos were found to be downregulated with upregulated CDK inhibitor p21 on extract treatment as compared to control. Results of FACS analysis further confirm G1/S phase cell cycle arrest on areca nut extract exposure. The regulation of downstream AP-1 subunits by MAPKs was studied by using specific inhibitors of ERK, JNK and p38 along with areca nut extract. Results showed the redox activation of MAP kinases down regulated the mRNA levels of AP-1 subunits in aqueous areca nut extract treated cells. Hence the present study aids in elucidating the role of MAP kinases in regulating the AP-1 subunits and their implications on target genes that are involved regulation of various cellular processes. Further, it would help in understanding the mechanistic aspects of the diseased state which may facilitate in designing of new therapeutic modalities that could help in cancer management.
KeywordsAP-1 factors Areca nut extract A549 cells Cell cycle regulators MAPK
Areca nut extract
Cyclin dependent kinase
Proliferative cell nuclear antigen
Semi quantitative reverse transcriptase-Polymerase Chain Reaction
The authors wish to express their gratitude to the University Grant Commission-Centre with Potential for Excellence Area (UGC-CPEPA) [8-2/2008(NS/PE)] and Department of Science and Technology-Promotion of University Research and Scientific Excellence (DST-PURSE) [SR/59/Z-23/2010/38(c)], New Delhi for providing financial support. Author RN is grateful to UGC-CPEPA for providing fellowships. Authors also wish to express their gratitude to the Department of Microbiology and Biotechnology, Bangalore University, Bengaluru, for providing the DST-FIST, UGC-SAP and department instrumentation facility.
Compliance with ethical standards
Conflict of interest
Authors declare that there are no conflicts of interest.
- Angel P, Karin M (1991) The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation. Biochem Biophys Acta 1072:129–157Google Scholar
- Blagosklonny MV, Pardee AB (2002) The restriction point of the cell cycle. Cell Cycle 1:103–110Google Scholar
- Chang MC et al (2004) The induction of prostaglandin E2 production, interleukin-6 production, cell cycle arrest, and cytotoxicity in primary oral keratinocytes and KB cancer cells by areca nut ingredients is differentially regulated by MEK/ERK activation. J Biol Chem 279:50676–50683. https://doi.org/10.1074/jbc.M404465200 CrossRefGoogle Scholar
- Hsu JC, Cressman DE, Taub R (1993) Promoter-specific trans-activation and inhibition mediated by JunB. Cancer Res 53:3789–3794Google Scholar
- IARC (1985) Betel-quid and areca-nut chewing. IARC Monogr Eval Carcinog Risk Chem Hum 37:137–202Google Scholar
- Ji WT, Yang SR, Chen JY, Cheng YP, Lee YR, Chiang MK, Chen HR (2012) Arecoline downregulates levels of p21 and p27 through the reactive oxygen species/mTOR complex 1 pathway and may contribute to oral squamous cell carcinoma. Cancer Sci 103:1221–1229. https://doi.org/10.1111/j.1349-7006.2012.02294.x CrossRefGoogle Scholar
- Mannervik B (1985) The isoenzymes of glutathione transferase. Adv Enzymol Relat Areas Mol Biol 57:357–417Google Scholar
- Shackelford RE, Kaufmann WK, Paules RS (1999) Cell cycle control, checkpoint mechanisms, and genotoxic stress. Environ Health Perspect 107:5–24Google Scholar
- Sundqvist K, Liu Y, Nair J, Bartsch H, Arvidson K, Grafstrom RC (1989) Cytotoxic and genotoxic effects of areca nut-related compounds in cultured human buccal epithelial cells. Cancer Res 49:5294–5298Google Scholar
- Tseng YH, Chang CS, Liu TY, Kao SY, Chang KW, Lin SC (2007) Areca nut extract treatment down-regulates involucrin in normal human oral keratinocyte through P13K/AKT activation. Oral Oncol 43:670–679. https://doi.org/10.1016/j.oraloncology.2006.08.003 CrossRefGoogle Scholar
- Vahrmeijer AL, van Dierendonck JH, Schutrups J, van de Velde CJ, Mulder GJ (1999) Effect of glutathione depletion on inhibition of cell cycle progression and induction of apoptosis by melphalan (l-phenylalanine mustard) in human colorectal cancer cells. Biochem Pharmacol 58:655–664CrossRefGoogle Scholar
- World Health Organization ROftWP (2012) Review of areca (betel) nut and tobacco use in the Pacific: a technical report WHO Regional Office for the Western Pacific. Philippine, ManilaGoogle Scholar