ABB  Vol.3 No.8 , December 2012
Fermentation product of RS3 inhibited proliferation and induced apoptosis in colon cancer cell HCT-116
Abstract: Cell free supernatant containing short chain fatty acid (SCFA) resulted from fermentation of resistant starch type three (RS3) by Clostridium butyricum BCC B2571 or Eubacterium rectale DSM 17629 were investigated for their ability to inhibit proliferation and induce apoptosis of human colon cancer cell line HCT-116. HCT-116 was cultured in complete medium and after 50% confluent, incubation was continued for another 48 hours in the absence or presence cell free supernatant containing SCFA mixture at butyrate levels up to 10 mM. The study revealed that the proliferation inhibition effect was higher (>80%) on HCT-116 treated with supernatant of C. butyricum BCC B2571 than that (<70%) of HCT-116 treated with supernatant of E. rectale DSM 17629. The cells were induced to undergo apoptosis by both supernatant. The apoptosis occured through mitochondrial pathway by changing the expression of gene Bcl-2 and Bax, thus incresed the Bax/Bcl-2 ratio by more than 3.5 fold. The protein caspase-3 was increased by more than 250% in the presence of the cell free supernatant.
Cite this paper: Purwani, E. , Iskandriati, D. and Suhartono, M. (2012) Fermentation product of RS3 inhibited proliferation and induced apoptosis in colon cancer cell HCT-116. Advances in Bioscience and Biotechnology, 3, 1189-1198. doi: 10.4236/abb.2012.38145.

[1]   World Health Organization (2008) The globocan project.

[2]   World Cancer Research Fund/American Institute for Cancer Research (2011) Update project report summary, food, nutrition, physical activity, and the prevention of colorectal cancer. World Cancer Research Fund, England.

[3]   Englyst, H.N., Kingman, S.M. and Cummings, J.H. (1992) Classification and measurement of nutritionally important starch fractions. European Journal of Clinical Nutrition, 46, 2S33-2S50.

[4]   Sajilata, M.G., Singhal, R.S. and Kulkarni, P.R. (2006) Resistant starch—A review. Comprehensive Reviews in Food Science and Food Safety, 5, 1-17. doi:10.1111/j.1541-4337.2006.tb00076.x

[5]   Bird, A.R., Brown, I.L. and Topping, D.L., (2000) Starches, resistant starches, the gut microflora and human health. Current Issues in Intestinal Microbiology, 1, 2-37.

[6]   Louis, P. and Flint, H.J. (2009) Diversity, metabolism and microbial ecology of butyrate-producing bacteria from the human large intestine. FEMS Microbiol Letters, 294, 1-8. doi:10.1111/j.1574-6968.2009.01514.x

[7]   Ramsay, A.G., Scott, K.P., Martin, J.C., Rincon, M.T. and Flint, H.J. (2006) Cell-associated-amylases of butyrate-producing firmicute bacteria from the human colon. Journal of Microbiology, 152, 3281-3290. doi:10.1099/mic.0.29233-0

[8]   Augenlicht, L.H., Mariadason, J.M., Wilson, A., Arango, D., Yang, W.C., Heerdt, B.G. and Velcich, A. (2002) Short chain fatty acids and colon cancer. The American Society for Nutritional Sciences, 132, 3804S-3808S.

[9]   Bauer-Marinovic, M., Florian, S., Muller-Schmehl, K., Glatt, H., and Jacobasch, G. (2006) Dietary resistant starch type 3 prevents tumor induction by 1,2-dimethylhydrazine and alters proliferation, apoptosis and dedifferentiation in rat colon. Carcinogenesis, 27, 1849-1859. doi:10.1093/carcin/bgl025

[10]   Elmore, S. (2007) Apoptosis: A review of programmed cell death. Toxicologic Pathology, 35, 495-516. doi:10.1080/01926230701320337

[11]   Purwani, E.Y., Purwadaria, T. and Suhartono, M.T. (2012) Fermentation RS3 derived from sago and rice starch with Clostridium butyricum BCC B2571 or Eubacterium rectale DSM 17629. Anaerobe, 18, 55-61. doi:10.1016/j.anaerobe.2011.09.007

[12]   Goni, I., Garcia-Diz, L., Manas, E. and Saura-Calixto, F. (1996) Analysis of resistant starch: A method for foods and food products. Food Chemistry, 56, 445-449. doi:10.1016/0308-8146(95)00222-7

[13]   Nohara, K., Yokoyama, Y. and Kano, K. (2007) The important role of caspase-10 in sodium butyrate-induced apoptosis. Kobe Journal of Medical Sciences, 53, 265-273.

[14]   Pfaffl, M.W. (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Research, 29, 2002-2007. doi:10.1093/nar/29.9.e45

[15]   Emmenaker, N.J., Calaf, G.M., Cox, D., Basson, M.D. and Qureshi, N. (2001) Short-chain fatty acids inhibit invasive human colon cancer by modulating uPA, TIMP-1, TIMP-2, Mutant p53, Bcl-2, Bax, p21 and PCNA protein expresion in an in vitro cell culture model. American Society for Nutrition, 131, 3041S-3046S.

[16]   Le-Leu, R.K., Hu, Y. and Young, G.P. (2002) Effects of resistant starch and nonstarch polysaccharides on colonic luminal environment and genotoxin-induced apoptosis in the rat. Carcinogenesis, 23, 713-719. doi:10.1093/carcin/23.5.713

[17]   Li, C.J. and Elsasser, T.H. (2005) Butyrate-induced apoptosis and cell cycle arrest in bovine kidney epithelial cells: Involvement of caspase and proteasome pathways. Journal of Animal Sciences, 83, 89-97.

[18]   Ruemmele, F.M., Dionne, S., Qureshi, I., Sarma, D.S.R., Levy, E. and Seidman, E.G. (1999) Butyrate mediates Caco-2 cell apoptosis via up-regulation of pro-apoptotic BAK and inducing caspase-3 mediated cleavage of poly- (ADP-ribose) polymerase (PARP). Cell Death Different, 6, 729-735. doi:10.1038/sj.cdd.4400545

[19]   Sharp, R. and Macfarlane, G.T. (2000) Chemostat enrichments of human feces with resistant starch are selective for adherent butyrate-producing clostridia at high dilution rate. Applied and Environmental Microbiology, 66, 4212-4221. doi:10.1128/AEM.66.10.4212-4221.2000

[20]   Topping, D.L. and Clifton, P.M. (2001) Short-chain fatty acids and human colonic function: Roles of resistant starch and nonstarch polysaccharides. Physiological Reviews, 81, 1031-1064.

[21]   He, G.-Q., Kong, Q., Chen, Q.-H. and Ruan, H. (2005) Batch and fed-batch production of butyric acid by Clostridium butyricum ZJUCB. Journal of Zhejiang University Science, 6B, 1076-1080. doi:10.1631/jzus.2005.B1076

[22]   Reid, C.A., Hillman, K. and Henderson, C. (1998) Effect of retrogradation, pancreatin digestion and amylose/ amylopectin ratio on the fermentation of starch by Clostridium butyricum (NCIMB 7423). Journal of the Science of Food and Agriculture, 76, 221-225. doi:10.1002/(SICI)1097-0010(199802)76:2<221::AID-JSFA935>3.0.CO;2-O

[23]   Lesmes, U., Beards, E.J., Gibson, G.R., Tuohy, K.K. and Shimoni, E. (2008) Effects of resistant starch tipe III polymorphs on human colon microbiota and short chain fatty acids in human gut models. Journal of Agricultural and Food Chemistry, 56, 5415-5421. doi:10.1021/jf800284d

[24]   Hadjiagapiou, C., Schmidt, L., Dudeja, P.K., Layden, T.J. and Ramaswamy, K. (2000) Mechanism(s) of butyrate transport in Caco-2 cells: Role of monocarboxylate transporter 1. American Journal of Physiology-Gastrointestinal and Liver Physiology, 279, G775-G780.

[25]   Lecona, E., Olmo, N., Turnay, J., Santiago-Gomez, A., L′opez De Silanes, I., Gorospe, M. and Lizarbe, M.A. (2008) Kinetic analysis of butyrate transport in human colon adenocarcinoma cells reveals two different carrier-mediated mechanisms. Biochemical Journal, 409, 311-320. doi:10.1042/BJ20070374

[26]   Hatayama, H., Iwashita, J., Kuwajima, A. and Abe, T. (2007) The short-chain fatty acid, butyrate, stimulates MUC2 mucin production in the human colon cancer cell line, LS174T. Biochemical and Biophysical Research Communications, 356, 599-603. doi:10.1016/j.bbrc.2007.03.025

[27]   Avivi-Green, C., Polak-Charcon, S., Madar, Z. and Schwartz, B. (2002) Different molecular events account for butyrate-induced apoptosis in two human colon cancer cell lines. Journal of Nutrition, 132, 1812-1818.

[28]   Le-Leu, R.K., Brown, I.L., Hu, Y., Morita, T., Esterman, A. and Young, G.P. (2007) Effect of dietary resistant starch and protein on colonic fermentation and intestinal tumourigenesis in rat. Carcinogenesis, 28, 240-245. doi:10.1093/carcin/bgl245

[29]   Kim, E.J., Park, S.Y., Shin, H., Kwon, D.Y., Surh, Y. and Park, J.H.Y. (2007) Activation of caspase-8 contributes to 3,3-diindolylmethane-induced apoptosis in colon cancer cells. Journal of Nutrition, 137, 31-36.

[30]   Jan, G., Belzacq, A., Haouzi, D., Rouault A., Meativier, A., Kroemer, G. and Brenner, C. (2002) Propionibacteria induce apoptosis of colorectal carcinoma cells via shortchain fatty acids acting on mitochondria. Cell Death and Different, 9, 179-188. doi:10.1038/sj.cdd.4400935

[31]   Hsu, C., Lin, Y., Chou, C., Zhou, S., Hsu, Y., Liu, C., Ku, F. and Chung, Y. (2009) Mechanisms of grape seed procyanidin-induced apoptosis in colorectal carcinoma cells. Anticancer Research, 29, 283-290.