Evolvulus alsinoides methanolic extract triggers apoptosis in HepG2 cells

Document Type: Original Research Article

Authors

1 Department of Pharmacology, Saveetha Dental College, Saveetha Institute of Medical and Technical Sciences, Chennai, Tamil Nadu, India

2 Saveetha Institute of Medical and Technical Sciences

3 Biomedical Research Unit and Laboratory Animal Centre, Saveetha Dental College, Chennai, Tamil Nadu, India – 600 077

Abstract

Objective: The objective of the present study was to evaluate the cytotoxic potentials of Evolvulus alsinoides in human hepatoma HepG2 cells.
Materials and Methods: HepG2 cells were treated with methanolic extract of E. alsinoides at 20, 40 and 80 µg/ml for 24 hr and cytotoxic effect was analyzed by MTT assay. The apoptosis rate was investigated by Hoechst 33342 and annexin V/propidium iodide staining. Mitochondrial membrane potential was evaluated by rhodamine staining. Also, the expression of catenin – β 1 protein was analyzed by western blotting.
Results: E. alsinoides methanolic extract treatment caused significant cytotoxicity in HepG2 cells in a concentration-dependent manner.Dual staining assayconfirmed the presence of early and late apoptotic cells only in extract-treated groups. Plant extract treatment also caused nuclear fragmentation and chromatin condensation in HepG2 cells. Mitochondrial membrane potential also reduced upon E. alsinoides treatments.This treatment also modulated the catenin – β 1 protein expression.
Conclusion:In this study, we demonstrated theproapoptotic potential E. alsinoides in HepG2 cells; thus, this plant may be beneficial in the treatment of liver cancer.

Keywords

Main Subjects


Agrawal S. 2016. Looking Beyond Sorafenib to Treat Advanced Hepatocellular Carcinoma. J Clin Exp Hepatol, 6:339-342.

Arora A, Kumar A. 2014. Treatment Response Evaluation and Follow-up in Hepatocellular Carcinoma. J Clin Exp Hepatol, 4:S126-129.

Baecker A, Liu X, La Vecchia C, Zhang ZF. 2018. Worldwide incidence of hepatocellular carcinoma cases attributable to major risk factors. Eur J Cancer Prev, 27:205-212.

Baskić D, Popović S, Ristić P, Arsenijević NN. 2006. Analysis of cycloheximide-induced apoptosis in human leukocytes: fluorescence microscopy using annexin V/propidium iodide versus acridin orange/ethidium bromide. Cell Biol Int, 30:924-932.

Crowley LC, Marfell BJ, Waterhouse NJ. 2016. Analyzing Cell Death by Nuclear Staining with Hoechst 33342. Cold Spring Harb Protoc, 2016:9.

Do MT, Kim HG, Choi JH, Khanal T, Park BH, Tran TP, Jeong TC, Jeong HG. 2013.Antitumor efficacy of piperine in the treatment of human HER2-overexpressing breast cancer cells. Food Chem, 141:2591-2599.

Efferth T, Saeed MEM, Mirghani E, Alim A, Yassin Z, Saeed E, Khalid HE, Daak S. 2017.Integration of phytochemicals and phytotherapy into cancer precision medicine. Oncotarget, 8:50284-50304.

Ezhilarasan D. 2018. Herbal Therapy for Cancer. In: Prakash Srinivasan Timiri Shanmugam, editor. Understanding Cancer Therapies, CRC Press, pp.129-166.

Ezhilarasan D, Evraerts J, Sid B, Calderon PB, Karthikeyan S, Sokal E, Najimi M. 2017. Silibinin induces hepatic stellate cell cycle arrest via enhancing p53/p27 and inhibiting Akt downstream signaling protein expression. Hepatobiliary Pancreat Dis Int, 16:80-87.

Feng XL, Fei HZ, Hu L. 2018. Dexamethasone induced apoptosis of A549 cells via the TGF-β1/Smad2 pathway. Oncol Lett, 15:2801-2806.

Ganju L, Karan D, Chanda S, Srivastava KK, Sawhney RC, Selvamurthy W. 2003. Immunomodulatory effects of agents of plant origin. Biomed Pharmacother, 57:296-300.

Gomathi D, Kalaiselvi M, Ravikumar G, Devaki K, Uma C. 2015. GC-MS analysis of bioactive compounds from the whole plant ethanolic extract of Evolvulus alsinoides (L.) L. J Food Sci Technol, 52:1212-1217.

Gomathi D, Ravikumar G, Kalaiselvi M, Devaki K, Uma C. 2013. Efficacy of Evolvulus alsinoides (L.) L. on insulin and antioxidants activity in pancreas of streptozotocin induced diabetic rats. J Diabetes Metab Disord, 12:39.

Gomathi D, Ravikumar G, Kalaiselvi M, Devaki K, Uma C. 2014. Antioxidant activity and functional group analysis of Evolvulus alsinoides. Chin J Nat Med, 12:827-832.

Hartke J, Johnson M, Ghabril M. 2017. The diagnosis and treatment of hepatocellular carcinoma. Semin Diagn Pathol, 34:153-159.

     Kumar M, Ahmad A, Rawat P, Khan MF, Rasheed N, Gupta P, Sathiamoorthy B, Bhatia G, Palit G, Maurya R. 2010. Antioxidant flavonoid glycosides from Evolvulus alsinoides. Fitoterapia, 81:234-242.

Kumar R, Bashyam MD. 2017. Multiple oncogenic roles of nuclear beta-catenin. J Biosci, 42:695-707.

Lin Y, Xu J, Liao H, Li L, Pan L. 2014. Piperine induces apoptosis of lung cancer A549 cells via p53-dependent mitochondrial signaling pathway. Tumour Biol, 35:3305-3310.

Morin PJ. 1999. beta-catenin signaling and cancer. Bioessays, 21:1021-1030.

Naikawadi VB, Ahire ML, Lahiri A, Nikam TD. 2016. In vitro propagation and cell cultures of memory tonic herb Evolvulus alsinoides: a best source for elicited production of scopoletin. Appl Microbiol Biotechnol, 100:3463-3476.

Rajan V, Tripathi J, Variyar P, Pandey BN. 2014. Mechanism of cytotoxicity by Psoralea corylifolia extract in human breast carcinoma cells. J Environ Pathol Toxicol Oncol, 33:265-277.

Rieger AM, Nelson KL, Konowalchuk JD, Barreda DR. 2011. Modified annexin V/propidium iodide apoptosis assay for accurate assessment of cell death. J Vis Exp, 50: pii: 2597.

Safadi FF, Xu J, Smock SL, Kanaan RA, Selim AH, Odgren PR, Marks SC Jr, Owen TA, Popoff SN. 2003.Expression of connective tissue growth factor in bone: its role in osteoblast proliferation and differentiation in vitro and bone formation in vivo. J Cell Physiol, 196:51-62.

Sakamuru S, Attene-Ramos MS, Xia M. 2016. Mitochondrial Membrane Potential Assay. Methods Mol Biol, 1473:17-22.

Salhab M, Canelo R. 2011. An overview of evidence-based management of hepatocellular carcinoma: a meta-analysis. J Cancer Res Ther, 7:463-475.

Sharma S, Shrivastava N. 2016. Internal transcribed spacer guided multiplex PCR for species identification of Convolvulus prostratus and Evolvulus alsinoides. Acta Pharm Sin B, 6:253-258.

Singh T, Sharma SD, Katiyar SK. 2011. Grape proanthocyanidins induce apoptosis by loss of mitochondrial membrane potential of human non-small cell lung cancer cells in vitro and in vivo. PLoS One, 6:e27444.

Thakur R, Mishra DP. 2013. Pharmacological modulation of beta-catenin and its applications in cancer therapy. J Cell Mol Med, 17:449-456.

Thandassery RB, Goenka U, Goenka MK. 2014.  Role of local ablative therapy for hepatocellular carcinoma. J Clin Exp Hepatol, 4:S104-111.

Tsai CH, Tzeng SF, Hsieh SC, Yang YC, Hsiao YW, Tsai MH, Hsiao P. 2017. A standardized herbal extract mitigates tumor inflammation and augments chemotherapy effect of docetaxel in prostate cancer. Sci Rep, 7:15624.

Wang CZ, Calway TD, Wen XD, Smith J, Yu C, Wang Y, Mehendale SR, Yuan CS. 2013.Hydrophobic flavonoids from Scutellaria baicalensis induce colorectal cancer cell apoptosis through a mitochondrial-mediated pathway. Int J Oncol, 42:1018-1026.

Wang Z, Zhang H, Hou J, Niu J, Ma Z, Zhao H, Liu C. 2015. Clinical implications of β-catenin protein expression in breast cancer. Int J Clin Exp Pathol, 8:14989-14994.

Woo SM, Lee KM, Lee GR, Park JY, Lee HJ, Bahn HJ, Yoon HS, Kim JY, Shin YC, Cho SG, Ko SG. 2018.Novel herbal medicine LA16001 ameliorates cisplatin-induced anorexia. Mol Med Rep, 17:2665-2672.

Yaffe PB, Doucette CD, Walsh M, Hoskin DW. 2013. Piperine impairs cell cycle progression and causes reactive oxygen species-dependent apoptosis in rectal cancer cells. Exp Mol Pathol, 94:109-114.

Yang J, Liu X, Bhalla K, Kim CN, Ibrado AM, Cai J, Peng TI, Jones DP, Wang X. 1997.

Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science, 275:1129-1132.

Yin SY, Wei WC, Jian FY, Yang NS. 2013. Therapeutic applications of herbal medicines for cancer patients. Evid Based Complement Alternat Med, 2013:302426.

Yue Q, Gao G, Zou G, Yu H, Zheng X. 2017. Natural Products as Adjunctive Treatment for Pancreatic Cancer: Recent Trends and Advancements. Biomed Res Int, 2017:8412508.

Zhang Y, Qi Y, Zhao Y, Sun H, Ge J, Liu Z. 2018. Activin A induces apoptosis of mouse myeloma cells via the mitochondrial pathway. Oncol Lett, 15:2590-2594.