Antitumor activity of bacterial exopolysaccharides from the endophyte Bacillus amyloliquefaciens sp. isolated from Ophiopogon japonicus

Chen,Yi-Tao; Yuan,Qiang; Shan,Le-Tian; Lin,Mei-Ai; Cheng,Dong-Qing; Li,Chang-Yu

doi:10.3892/ol.2013.1284

Antitumor activity of bacterial exopolysaccharides from the endophyte Bacillus amyloliquefaciens sp. isolated from Ophiopogon japonicus

Authors:
- Yi-Tao Chen
- Qiang Yuan
- Le-Tian Shan
- Mei-Ai Lin
- Dong-Qing Cheng
- Chang-Yu Li
View Affiliations

Affiliations: College of Pharmacy, Zhejiang Chinese Medical University, Hangzhou 310053, P.R. China, The Second Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou 310053, P.R. China, Institute of Orthopaedics and Traumatology, Zhejiang Chinese Medical University, Hangzhou 310053, P.R. China, College of Life Science, Zhejiang Chinese Medical University, Hangzhou 310053, P.R. China
Published online on: April 3, 2013 https://doi.org/10.3892/ol.2013.1284
Pages: 1787-1792

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

The endophytic bacterium, MD-b1, was isolated from the medicinal plant Ophiopogon japonicas and identified as the Bacillus amyloliquefaciens sp. with 99% similarity based on the partial sequence analysis of 16S rDNA. Exopolysaccharides were extracted from the endophyte for the evaluation of its antitumor activity against gastric carcinoma cell lines (MC-4 and SGC-7901). 3-(4,5-dimethylthiazol-2-yl)-2,5‑diphenyltetrazolium bromide (MTT) assays and microscopy were performed to estimate the cell viability and morphological changes of the MC-4 and SGC-7901 cells following treatment with the exopolysaccharides at 14, 22 and 30 µg/µl. The results revealed that the exopolysaccharides displayed concentration‑dependent inhibitory effects against the MC-4 and SGC-7901 cells, with an IC50 of 19.7 and 26.8 µg/µl, respectively. The exopolysaccharides also induced morphological abnormalities in the cells. These effects indicated the the exopolysaccharides had an antitumoral mechanism of action associated with the mitochondrial dysfunction of the treated cells. This is the first study to investigate the endophytic microorganism isolated from O. japonicas and also the first discovery of such antitumoral exopolysaccharides derived from the genus Bacillus. This provides a promising and reproducible natural product source with high therapeutic value for anticancer treatment, thereby facilitating the development of new anticancer agents.

View Figures

View References

1	GLOBOCAN 2008 (IARC): Section of Cancer Information WHO. http://www.who.int/mediacentre/factsheets/fs297/en/ Accessed July 20, 2012.
2	Gutierrez RM, Gonzalez AM and Ramirez AM: Compounds derived from endophytes: a review of phytochemistry and pharmacology. Curr Med Chem. 19:2992–3030. 2012. View Article : Google Scholar : PubMed/NCBI
3	Igarashi Y, Trujillo ME, Martínez-Molina E, Yanase S, Miyanaga S, Obata T, Sakurai H, Saiki I, Fujita T and Furumai T: Antitumor anthraquinones from an endophytic actinomycete Micromonospora lupine sp, nov. Bioorg Med Chem Lett. 17:3702–3705. 2007. View Article : Google Scholar : PubMed/NCBI
4	Taechowisan TC, Lu CH, Shen YM and Lumyong S: Antitumor activity of 4-Arylcoumarins from endophytic Streptomyces aureofaciens CMUAc130. J Cancer Res Ther. 3:86–91. 2007. View Article : Google Scholar
5	Blunt JW, Copp BR, Hu WP, Munro MH, Northcote PT and Prinsep MR: Marine natural products. Nat Prod Rep. 25:35–94. 2008. View Article : Google Scholar
6	Hallmann J, Quadt-Hallmann A, Mahaffee WF and Kloepper JW: Bacterial endophytes in agricultural crops. Can J Microbiol. 43:895–914. 1997. View Article : Google Scholar
7	Kobayashi DY and Palumbo JD: Bacterial endophytes and their effects on plants and uses in agriculture. Microbial Endophytes. Bacon CW and White JF Jr: Marcel Dekker; NY, New York: pp. 199–233. 2000
8	Zinniel DK, Lambrecht P, Harris NB, Feng Z, Kuczmarski D, Higley P, Ishmaru CA, Arunakumari A, Barletta RG and Vidaver AK: Isolation and characterization of endophytic colonizing bacteria from agronomic crops and prairie plants. Appl Environ Microbiol. 68:2198–2208. 2002. View Article : Google Scholar : PubMed/NCBI
9	Azevedo JL, Maccheroni W Jr, Pereira JO and Araújo WL: Endophytic microorganisms: a review on insect control and recent advances on tropical plants. Electron J Biotechnol. 3:e1–e4. 2000. View Article : Google Scholar
10	Sturz AV and Nowak J: Endophytic communities of Rhizobacteria and the strategies required to create yield enhancing associations with crops. Appl Soil Ecol. 15:183–190. 2000. View Article : Google Scholar
11	Lodewyck C, Vangronsveld J, Porteous F, Moore ERB, Taghavi S, Mezgeay M and van der Lelie D: Endophytic bacteria and their potential application. Crit Rev Plant Sci. 86:583–606. 2002. View Article : Google Scholar
12	Mundt JO and Hinkle NF: Bacteria within ovules and seeds. Appl Environ Microbiol. 32:694–698. 1976.PubMed/NCBI
13	McInroy JA and Kloepper JW: Population dynamics of endophytic bacteria in field-grown sweet corn and cotton. Can J Microbiol. 41:895–901. 1995. View Article : Google Scholar
14	Cho KM, Hong SY, Lee SM, Kim YH, Kahng GG, Lim YP, Kim H and Yun HD: Endophytic bacterial communities in ginseng and their antifungal activity against pathogens. Microbial Ecol. 54:341–351. 2007. View Article : Google Scholar : PubMed/NCBI
15	Tiwari R, Kalra A, Darokar MP, Chandra M, Aggarwal N, Singh AK and Khanuja SP: Endohytic bacteria from Ocimum sanctum and their yield enchancing capabilities. Curr Micobiol. 60:167–171. 2010.
16	Vendan RT, Yu YJ, Lee SH and Rhee YH: Diversity of endophytic bacteria in ginseng and their potential for plant growth promotion. J Microbiol. 48:559–565. 2010. View Article : Google Scholar : PubMed/NCBI
17	Yu BY and Xu GJ: Studies on resource utilization of Chinese drug Dwarf lilyturf (Ophiopogon japonicus). J Chin Herbs. 26:205–210. 1995.
18	Xiao PG: Modern Chinese Materia Medica. Chemical Indestry Press; Beijing: pp. 772002
19	Kou J, Yu B and Xu Q: Inhibitory effects of ethanol extract from Radix Ophiopogon japonicus on venous thrombosis linked with its endothelium-protective and anti-adhesive activities. Vascul Pharmacol. 43:157–163. 2005.PubMed/NCBI
20	Zhou YF, Qi J, Zhu DN and Yu BY: Two new steroidal glycosides from Ophiopogon japonicus. Chin Chem Lett. 19:1086–1088. 2008. View Article : Google Scholar
21	China Pharmacopoeia Committee: Pharmacopoeia of the People’s Republic of China. China Chemical Industry Press; I. Beijing, China: 64. 2010
22	Kou J, Sun Y, Lin Y, Cheng Z, Zheng W, Yu B and Xu Q: Anti-inflammatory activities of aqueous extract from Radix Ophiopogon japonicus and its two constituents. Biol Pharm Bull. 28:1234–1238. 2005. View Article : Google Scholar : PubMed/NCBI
23	Lin X, Xu DS, Feng Y, Li SM, Lu ZL and Shen L: Release-controlling absorption enhancement of enterally administered Ophiopogon japonicus polysaccharide by sodium caprate in rats. J Pharm Sci. 95:2534–2542. 2006. View Article : Google Scholar : PubMed/NCBI
24	Li N, Zhang JY, Zeng KW, Zhang L, Che YY and Tu PF: Anti-inflammatory homoisoflavonoids from the tuberous roots of Ophiopogon japonicus. Fitoterapia. 83:1042–1045. 2012. View Article : Google Scholar : PubMed/NCBI
25	Wang LY, Wang Y, Xu DS, Ruan KF and Wang S: MDG-1, a polysaccharide from Ophiopogon japonicas exerts hypoglycemic effects through the PI3K/Akt pathway in a diabetic KKAy mouse model. J Ethnopharmacol. 143:347–354. 2012.PubMed/NCBI
{ label needed for ref[@id='b26-ol-05-06-1787'] }	[26] Koyama R, Matsumoto A, Inahashi Y, Ōmura S and Takahashi Y: Isolation of actinomycetes from the root of the plant, Ophiopogon japonicus, and proposal of two new species, Actinoallomurus liliacearum sp nov and Actinoallomurus vinaceus sp nov. J Antibiot (Tokyo). 65:335–340. 2012.PubMed/NCBI
27	Rosenblueth M and Martinez-Romero E: Rhizobium etli maize populations and their competitiveness for root colonization. Arch Microbiol. 181:337–344. 2004. View Article : Google Scholar
28	Chen YT, Ding LX, Cheng DQ, Ding ZS, Lin MA and Pan PL: Isolation and identification of endofungi from Liriope spicata. J Laiyang Agri Col (Nat Sci). 23:13–16. 2006.(In Chinese).
29	Miller KI, Qing C, Sze DM and Neilan BA: Investigation of the biosynthetic potential of endophytes in traditional Chinese anticancer herbs. PLoS One. 7:e359532012. View Article : Google Scholar : PubMed/NCBI
30	Rosenblueth M and Martínez-Romero E: Bacterial endophytes and their interactions with hosts. Mol Plant Microbe Interact. 19:827–837. 2006. View Article : Google Scholar : PubMed/NCBI
31	Compant S, Mitter B, Colli-Mull JG, Gangl H and Sessitsch A: Endophytes of grapevine flowers, berries, and seeds: identification of cultivable bacteria, comparison with other plant parts, and visualization of niches of colonization. Microbial Ecol. 62:188–197. 2011. View Article : Google Scholar
32	Bonmatin J, Laprévote O and Peypoux F: Diversity among microbial cyclic lipopeptides: iturins and surfactins. Activity-structure relationships to design new bioactive agents. Comb Chem High Throughput Screen. 6:541–556. 2003. View Article : Google Scholar
33	Snook ME, Mitchell T, Hinton DM and Bacon CW: Isolation and characterization of leu7-surfactin from the endophytic bacterium Bacillus mojavensis RRC 101, a biocontrol agent for Fusarium verticillioides. J Agric Food Chem. 57:4287–4292. 2009. View Article : Google Scholar : PubMed/NCBI
34	Krishnan P, Bhat R, Kush A and Ravikumar P: Isolation and functional characterization of bacterial endophytes from Carica papaya fruits. J Appl Microbiol. 113:308–317. 2012. View Article : Google Scholar : PubMed/NCBI
35	Newman DJ and Cragg GM: Natural products as sources of new drugs over the last 25 years. J Nat Prod. 70:461–477. 2007.PubMed/NCBI
36	Strobel G, Daisy B, Castillo U and Harper J: Natural products from endophytic microorganisms. J Nat Prod. 67:257–268. 2004. View Article : Google Scholar : PubMed/NCBI
37	Yang MP, Wu H, Yin L, Zhang X and Duan JA: Advances in research of saponins and polysaccharides in Ophiopogonis japonicus. Chin Arch Trad Chin Med. 10:2169–2171. 2008.
38	Chen YT, Lu QY, Lin MA, Cheng DQ, Ding ZS and Shan LT: A PVP-extract fungal protein of Omphalia lapideacens and its antitumor activity on human gastric tumors and normal cells. Oncol Rep. 26:1519–1526. 2011.PubMed/NCBI
39	Xia M, Huang R, Witt KL, Southall N, Fostel J, Cho MH, Jadhav A, Smith CS, Inglese J, Portier CJ, Tice RR and Austin CP: Compound cytotoxicity profiling using quantitative high-throughput screening. Environ Health Perspect. 116:284–291. 2008. View Article : Google Scholar : PubMed/NCBI
40	Chiu TH, Lai WW, Hsia TC, Yang JS, Lai TY, Wu PP, Ma CY, Yeh CC, Ho CC, Lu HF, Wood WG and Chung JG: Aloe-emodin induces cell death through S-phase arrest and caspase-dependent pathways in human tongue squamous cancer SCC-4 cells. Anticancer Res. 29:4503–4511. 2009.PubMed/NCBI
41	Zheng JY, Tao LY, Liang YJ, Chen LM, Mi YJ, Zheng LS, Wang F, She ZG, Lin YC, To KK and Fu LW: Anthracenedione derivatives as anticancer agents isolated from secondary metabolites of the mangrove endophytic fungi. Mar Drugs. 8:1469–1481. 2010. View Article : Google Scholar : PubMed/NCBI
42	Tan RX and Zou WX: Endophytes: a rich source of functional metabolites. Nat Prod Rep. 18:448–459. 2001.PubMed/NCBI
43	Thompson JD, Higgins DG and Gibson TJ: CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22:4673–4680. 1994. View Article : Google Scholar
44	Thompson JD, Gibson TJ, PleWniak F, Jeanmougin F and Higgins DG: The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25:4876–4882. 1997. View Article : Google Scholar : PubMed/NCBI
45	Tamura K, Dudley J, Nei M and Kumar S: MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol. 24:1596–1599. 2007. View Article : Google Scholar : PubMed/NCBI
46	Saitou N and Nei M: The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol. 4:406–425. 1987.PubMed/NCBI
47	Felsenstein J: Evolutionary trees from DNA sequences: a maximum likelihood approach. J Mol Evol. 17:368–376. 1981. View Article : Google Scholar : PubMed/NCBI
48	Fitch WM: Toward defining the course of evolution: minimum change for a specific tree topology. Syst Zool. 20:406–416. 1971. View Article : Google Scholar
49	Masuko T, Minami A, Iwasaki N, Majima T, Nishimura S and Lee YC: Carbohydrate analysis by a phenol-sulfuric acid method in microplate format. Anal Biochem. 339:69–72. 2005. View Article : Google Scholar : PubMed/NCBI
50	Tang QY and Feng MG: DPS Data Processing System: Experimental Design, Statistical Analysis, and Data Mining. Science Press; Beijing: pp. 146–164. 2007