In vitro properties of chitosan nanoparticles induce apoptosis in human lymphoma SUDHL-4 cell line

DOI: 10.4236/abb.2013.412148   PDF   HTML     4,556 Downloads   6,781 Views   Citations


In this study, the possible mechanisms were investigated with chitosan nanoparticles using sodium tripoly-phosphate and effects on human lymphoma SUDHL-4 in vitro. It was characterized by XRD, FTIR, TGA, particle Size, zeta potential, SEM & TEM. Different techniques such as cell proliferation, ultra structure changes, DNA fragmentation, phase distribution of cell cycle, MTT assay, MMP, agarose gel electrophoresis of DNA, flow cytometry and electron microscopy were used with treatment of different concentrations of CH-NPs (25, 50, 75, 100 μg/ml) at different time periods. Electron microscopy study revealed that the chitosan nanoparticles showed 78 nm particle size which is a high surface charge as 52 mV. Inhibition of chitosan nanoparticles after 48h treatment was marked in cell proliferation of SUDHL-4 with an IC50 value of 5 μg/ml. Electron microscopy showed typical necrotic cell morphology after treatment of chitosan nanoparticles. The DNA degradation related with necrosis was determined using agarose electrophoresis and loss of MMP & occurrence of apoptosis was analyzed by flow cytometry. Chitosan nanoparticles with low molecular weight (LMW) were comparatively stable in medium containing aqueous and rate of dissolution was slow in acidic medium. Results of this present study clearly provided information that the chitosan nanoparticles effectively inhibit the proliferation of SUDHL-4 through multiple mechanisms in vitro and this novel formulation can open a new avenue against human Lymphoma.

Share and Cite:

Parida, U. , Rout, N. and Bindhani, B. (2013) In vitro properties of chitosan nanoparticles induce apoptosis in human lymphoma SUDHL-4 cell line. Advances in Bioscience and Biotechnology, 4, 1118-1127. doi: 10.4236/abb.2013.412148.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Parida, U.K., Nayak, A.K., Bindhani, B.K. and Nayak, P.L. (2011) Synthesis and characterization of chitosan-polyvinyl alcohol blended with cloisite 30B for controlled release of the anticancer drug curcumin. Journal of Biomaterials and Nanobiotechnology, 2, 414-425.
[2] Qian, C., Xu, X., Shen, Y., Li, Y. and Guo, S. (2013) Synthesis and preliminary cellular evaluation of phosphonium chitosan derivatives as novel non viral vector. Carbohydrate Polymers, 97, 676-683.
[3] Shukla, S.K., Mishra, A.K., Arotiba, O.A. and Mamba, B.B. (2013) Chitosan-based nanomaterials: A state-of-the-art review. International Journal of Biological Macromolecules, 59, 46-58.
[4] Di Martino, A., Sittinger, M. and Risbud, M.V. (2005) Chitosan: A versatile biopolymer for orthopaedic tissueengineering. Biomaterials, 26, 5983-5990.
[5] Senel, S. and Mc Clure, S.J. (2004) Potential applications of chitosan in veterinary medicine. Advanced Drug Delivery Reviews, 56, 1467-1480.
[6] Khor, E. and Lim, L.Y. (2003) Implantable applications of chitin and chitosan. Biomaterials, 24, 2339-2349.
[7] Bumgardner, J.D., Wiser, R., Gerard, P.D., Bergin, P., Chestnutt, B., Marini, M., Ramsey, V., Elder, S.H. and Gilbert, J.A. (2003) Chitosan: Potential use as a bioactive coating for othopaedic and craniofacial/dental implants. Journal of Biomaterials Science, Polymer Edition, 14, 429-438.
[8] Kumar, M.N.V.R. (2000) A review of chitin and chitosan applications. Reactive and Functional Polymers, 46, 1-27.
[9] Synowiecki, J. and Khateeb, N.A. (2003) Production, properties and some new applications of chitin and its derivatives. Critical Reviews in Food Science and Nutrition, 43, 145-171.
[10] Shon, D.H. (2001) Chitosan oligosaccharides for functional foods and microbial enrichment of chitosan oligosaccharides in soy-paste. Proceedings of the International Workshop on Bioactive Natural Products, Tokyo, 56-66.
[11] Tharanathan, R.N. and Kittur, F.S. (2003) Chitin—The undisputed biomolecule of great potential, Critical Reviews in Food Science and Nutrition, 43, 61-87.
[12] Sekiguchi, S., Miura,Y., Kancko, H., Nishimura, S.L., Nishi, N., Iwase, M. and Tokura, S. (1994) Molecular weight dependency of antimicrobial activity by chitosan oligomers. In: Nishimuri E. and Doi, E., Eds., Food Hydrocolloids: Structure, Properties and Function, Plenum Press, New York, 71-76.
[13] Jeon, Y.J., Park, P.J. and Kim, S.K. (2001) Antimicrobial effect of chitooligosaccharides produced by bioreactor. Carbohydrate Polymers, 44, 71-76.
[14] Kondo, Y., Nakatani, A., Hayashi, K. and Ito, M. (2000) Low molecular weight chitosan prevents the progression of low dose streptozotocin induced slowly progressive diabetes mellitus in mice. Biological & Pharmaceutical Bulletin, 23, 1458-1464.
[15] Tsai, G.J., Wu, Z.Y. and Su, W.H. (2000) Antibacterial activity of a chitooligosaccharide mixture prepared by cellulase digestion of shrimp chitosan and its application in milk preservation. Journal of Food Protection, 63, 747-752.
[16] Tsai, G.J., Wu, Z.Y. and Su, W.H. (2000) Antibacterial activity of a chitooligosaccharide mixture prepared by cellulase digestion of shrimp chitosan and its application in milk preservation. Journal of Food Protection, 63, 747-752.
[17] Richardson, S.C.W., Kolbe, H.V.J. and Duncan, R. (1999) Potentials of low molecular mass chitosan as a DNA delivery system: Biocompatibility, body distribution and ability to complex and protect DNA. International Journal of Pharmaceutics, 178, 231-243.
[18] Vimal, S., Abdul Majeed, S., Taju, G., Nambi, K.S., Sundar Raj, N., Madan, N., Farook, M.A., Rajkumar, T., Gopinath, D. and Sahul Hameed, A.S. (2013) Chitosan tripolyphosphate (CS/TPP) nanoparticles: Preparation, characterization and application for gene delivery in shrimp. Acta Tropica, 128, 486-493.
[19] Oliveira, A.V., Silva, A.P., Bitoque, D.B., Silva, G.A. and Rosa da Costa, A.M. (2013) Transfection efficiency of chitosan and thiolated chitosan in retinal pigment epithelium cells: A comparative study. Journal of Pharmacy and Bioallied Sciences, 5, 111-118.
[20] Qi, L.F., Xu, Z.R., Li, Y., Jiang, X. and Han, X.Y. (2005) In vitro effects of chitosan nanoparticles on proliferation of human gastric carcinoma cell line MGC803 cells. World Journal of Gastroenterology, 11, 5136-5141.
[21] Singh, D.J., Lohade, A.A., Parmar, J.J., Hegde, D.D., Soni, P., Samad, A. and Menon, M.D. (2012) Development of Chitosan-based Dry Powder inhalation System of Cisplatin for Lung Cancer. Indian Journal of Pharmaceutical Sciences, 74, 521-526.
[22] Del Turco, S., Ciofani, G., Cappello. V., Gemmi, M., Cervelli, T., Saponaro, C., Nitti, S., Mazzolai, B., Basta, G. and Mattoli, V. (2013) Cytocompatibility evaluation of glycol-chitosan coated boron nitride nanotubes in human endothelial cells. Colloids Surf B: Biointerfaces, 111C, 142-149.
[23] Paiva, D., Ivanova, G., do Carmo Pereira, M. and Rocha, S. (2013) Chitosan conjugates for DNA delivery. Physical Chemistry Chemical Physics, 15, 11893-11899.
[24] Liang, X., Li, X., Chang, J., Duan, Y. and Li, Z. (2013) Properties and evaluation of quaternized chitosan/lipid cation polymeric liposomes for cancer-targeted gene delivery. Langmuir, 29, 8683-8693.
[25] Sahu, A., Goswami, P. and Bora U. (2009) Microwave mediated rapid synthesis of chitosan. Journal of Materials Science: Materials in Medicine, 20, 171-175.
[26] Cheng, C.Y. and Li, Y.K. (2000) An Aspergillus chitinase with potential for largescale preparation of chitosan oligosaccharides, Biotechnol. Biotechnology and Applied Biochemistry, 32, 197-203.
[27] Baxter, A., Dillon, M. and Anthony Taylor, K.D. (1992) Improved method for i.r. determination of the degree of N-acetylation of chitosan. International Journal of Biological Macromolecules, 14, 66-69.
[28] Lavertu, M., Xia, Z., Serrequ, A.N., Berrada, M., Rodrigues, A., Wang, D., Buschmann, M.D. and Gupta, A. (2003) A validated 1H-NMR method for the determination of the degree of deacetylation of chitosan. Journal of Pharmaceutical and Biomedical Analysis, 32, 1149-1158.
[29] Venkatesan, P., Puvvada, N., Dash, R., Prashanth Kumar, B.N., Sarkar, D., Azab, B., Pathak, A., Kundu, S.C., Fisher, P.B. and Mandal, M. (2011) The potential of celecoxib-loaded hydroxyapatite-chitosan nanocomposite for the treatment of colon cancer. Biomaterials, 32, 3794-3806.
[30] Huang, Y. and Lapitsky, Y. (2011) Monovalent salt enhances colloidal stability during the formation of chotosan/tripolyphosphate microgels. Langmuir, 27, 10392-10399.
[31] Jeevitha, D. and Amarnath, K. (2013) Chitosan/PLA nanoparticles as a novel carrier for the delivery of anthraquinone: Synthesis, characterization and in vitro cytotoxicity evaluation. Colloids Surfaces B: Biointerfaces, 101, 126-134.
[32] Kim, H., You, S., Kong, B.W., Foster, L.K., Farris, J. and Foster, D.N. (2001) Necrotic cell death by hydrogen peroxide in immortal DF-1 chicken embryo fibroblast cells expressing deregulated MnSOD and catalase. Biochimica et Biophysica Acta, 1540, 137-146.
[33] Behl, G., Sharma, M., Dahiya, S., Chhikara, A. and Chopra, M. (2011) Synthesis, characterization, and evaluation of radical scavenging ability of ellagic acid-loaded nanogels. Journal of Nanomaterials, 1-9.
[34] Zhang, Z.P. and Feng, S.-S. (2006) The drug encapsulation efficiency, in vitro drug release, cellular uptake and cytotoxicity of paclitaxel-loaded poly (lactide)-tocopheryl polyethylene glycol succinate nanoparticles. Biomaterials, 27, 4025-4033.
[35] Stacey, N.B., Samantha, M.Y., Karl, R.F., Areti, T., Omid, K. and Ipsita, A.B. (2011) Ellagic acid promoted biomimetic synthesis of shape-controlled silver nanochains. Nanotechnology, 22, 1-10.
[36] Sonaje, K., Italia, J.L., Sharma, G., Bhardwaj, V., Tikoo, K. and Ravi Kumar, M.N.V. (2007) Development of biodegradable nanoparticles for oral delivery of ellagic acid and evaluation of their antioxidant efficacy against cyclosporine A-induced nephrotoxicity in rats. Pharmaceutical Research, 24, 899-908.
[37] Gautam, B., Monal, S., Saurabh, D., Aruna, C. and Madhu, C. (2011) Synthesis, characterization, and evaluation of radical scavenging ability of ellagic acid-loaded nanogels. Journal of Nanomaterials, 2011, Article ID: 695138.
[38] Mostafa, A.D., Adel Zaki, E., Mohamed, M.A. and Dina, M.D.B. (2012) Thermal stability and degradation of chitosan modified by cinnamic acid. Open Journal of Polymer Chemistry, 2, 14-20.
[39] Zobir, M.H., Samer, H.A.A., Zulkarnain, Z. and Muhammad, N.H. (2011) Development of antiproliferative nanohybrid compound with controlled release property using ellagic acid as the active agent. International Journal of Nanomedicine, 6, 1373-1383.
[40] Yokoyama, M., Satoh, A., Sakurai, Y., Okano, T., Matsumura, Y., Kakizoe, T. and Kataoka, K. (1998) Incorporation of water-insoluble anticancer drug into polymeric micelles and control of their particle size. Journal of Controlled Release, 55, 219-229.
[41] Zhang, L.Y., Hu, Y., Jiang, X.Q., Yang, C.Z., Lu, W. and Yang, Y.H. (2004) Camptothecin derivative-loaded poly (caprolactone-co-lactide)-b-PEG-b-poly(caprolactone-colactide) nanoparticles and their biodistribution in mice. Journal of Controlled Release, 96, 135-148.
[42] Jinno, H., Ikeda, T., Matsui, A., Kitagawa, Y., Kitajima, M., Fujii, H., Nakamura, K. and Kubo, A. (2002) Section 5. Breast Sentinel lymph node biopsy in breast cancer using technetium-99m tin colloids of different sizes. Biomedicine and Pharmacotherapy, 56, 213-216.
[43] Takenaga, M. (1996) Application of lipid microspheres for the treatment of cancer. Advanced Drug Delivery Reviews, 20, 209-219.
[44] Yuan, X., Yang, X., Cai, D., Mao, D., Wu, J., Zong, L. and Liu, J. (2008) Intranasal immunization with chitosan/pCETP nanoparticles inhibits atherosclerosis in a rabbit model of atherosclerosis. Vaccine, 26, 3727-3734.
[45] Morrison, M.R. and Smith, M. (1963) Preparation of fatty acid methyl esters and dimethyl acetyls from lipids with boron fluoride methanol. Journal of Lipid Research, 5, 600-608.
[46] Focher, B., Naggi, A., Torri, G., Cosani, A. and Terbojevich, M. (1992) Chitosans from Euphausia superba. 2: Characterization of solid-state structure. Carbohydrate Polymers, 18, 43-49.
[47] Monti, M.G., Ghiaroni, S., Marverti, G., Montanari, M. and Moruzzi, M.S. (2004) Polyamine depletion switches the form of 2-deoxy-d-ribose induced cell death from apoptosis to necrosis in HL-60 cells. The International Journal of Biochemistry & Cell Biology, 36, 1238-1248.
[48] Vishu Kumar, A.B., Varadaraj, M.C., Lalitha, R.G. and Tharanathan, R.N. (2005) Characterization of chito-oligosaccharides prepared by chitosanolysis with the aid of papain and pronase, and their bactericidal action against Bacillus cereus and Escherichia coli. Biochemical Journal, 391, 167-175.
[49] Kanjanathaworn, N., Polpanich, D., Jangpatarapongsa, K. and Tangboriboonrat, P. (2013) Reduction of cytotoxicity of natural rubber latex film by coating with PMMA-chitosan nanoparticles. Carbohydrate Polymers, 97, 52-58.
[50] Yang, S.J., Chang, S.M., Tsai, K.C., Tsai, H.M., Chen, W.S. and Shieh, M.J. (2012) Enhancement of chitosan nanoparticle-facilitated gene transfection by ultrasound both in Vitro and in Vivo. Journal of Biomedical Materials Research. Part B, Applied Biomaterials, 100, 1746-1754.

comments powered by Disqus

Copyright © 2020 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.