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Effects of Carboxymethyl Chitosan on the Growth of Tumor Cells and Tumor Tissue

Author: ZhengMeiLing
Tutor: LiuWanShun
School: Ocean University of China
Course: Biochemistry and Molecular Biology
Keywords: Carboxymethyl chitosan Tumor Biomaterial Cytokine
CLC: R96
Type: Master's thesis
Year: 2011
Downloads: 39
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Abstract

Purpose: Carboxymethyl chitosan (CM-CTS) is a water soluble chitosan derivative by introducing–COOH groups to chitosan molecule and endows it some outstanding biological properties. CM-CTS has been a research focus in the field of pharmaceutical and biomedical industry for its unique physicochemical and biological properties such as biocompatibility, biodegradability, degradation controllability and antimicrobial activity. There have also been some reports on the therapeutic utilities of the CM-CTS in the fields of antimicrobes, hemostatic, wound healing, reducing the formation of post surgical adhesions, drug carrying, enzyme inhibitor and so on. As versatile biomedical material, CM-CTS always conjugates or entraps or self-assembles with agent and extensively applies in tumor therapy research. However, safety evaluation of CM-CTS has been focused on the compounds and in vitro model, little reported work of CM-CTS perse safety both in vitro and vivo has been discussed, especially in tumor therapy. Moreover, the actual metabolic pathway and degradation products of CM-CTS in the body have been as yet unknown. Therefore, additional biological safety research of CM-CTS in application is necessary.In this study, the biological safety of CM-CTS with different molecular weights (MW), different substituted degrees (DS) and different degree of deacetylation (DD) on clinical application is investigated both in vitro and in vivo. CM-CTS is purified, characterized and its cytotoxicity is investigated by using normal cell and three tumor cells in vitro, and the influence of CM-CTS on the growth of sarcoma 180 and Heps tumor and immune response of tumor-bearing mice are also investigated, providing experimental basis for CM-CTS utilization in tumor application as biomedical material. Methods:1. Four CM-CTS samples with different MW, different DS and different DD are purified and their mainly physicochemical properties are measured. The MW of CM-CTS are measured with gel permeation chromatography using HPLC; DS and DD of CM-CTS are determined by potentiometric; The moisture and ash properties of CM-CTS are also determined.2. Human normal liver cell L02, human hepatoma cell Bel-7402, human gastric cancer cell SGC-7901 and human cervical carcinoma cell Hela are used to evaluate the cytotoxicity effects of CM-CTS by MTT assay in vitro and the morphology of cell is observed by inverted microscope.3. The levels of human transforming growth factor-α(TGF-α) and human vascular endothelial growth factor (VEGF) secreted by L02 and Bel-7402 cells are measured by ELISA.4. In vivo, the transplant tumor models of Heps and sarcoma 180 are established in mice and 340kDa CM-CTS is administered through intraperitoneal injection to investigate tumor inhibitory effect. The tumors are weighed to calculate the tumor inhibition rate. Immune organs including spleen and thymus of the mice are also removed and weighed to obtain the index of the spleen and thymus. Serum IL-2 and TNF-αlevel of sarcoma-180-bearing mice are also investigated.Results:1. Four CM-CTS samples with different MW, different DS and different DD are purified and their main chemical and physical parameters are measured.2. Four CM-CTS samples with different different MW, different DS and different DD could promote the proliferation of L02 cells within the concentration of 50~800μg/ml, but not promote even inhibit the proliferation of Bel-7402, SGC-7901 and Hela cells in certain concentration ranges and incubation time. The RGR value are 114%~126% of L02 cells but 80%~95% for that of three tumor cell lines: Bel-7402, SGC-7901 and Hela cells.3. Four CM-CTS samples with different MW, different DS and different DD also could improve the TGF-αsecretion of L02 cells, whereas decrease that of Bel-7402 cells. The level of VEGF secreted by Bel-7402 cells is also inhibited by CM-CTS to some extent.4. CM-CTS exhibits anticancer effect in a dose-dependent manner. The tumor inhibition rates of low, medium and high doses of CM-CTS are 12.9 %, 18.2 % and 22.1 % to the growth of Heps tumor and 29.4 %, 31.6 % and 39.9 % to the growth of sarcoma 180 tumors, respectively. CM-CTS also could enhance body immune function in the certain dose via elevation of serum IL-2 and TNF-αlevel and the spleen index and thymus index in treated mice.Conclusion: CM-CTS with different MW, different DS and different DD are safe to cell and exhibit twofold bioactivities in the culture of human normal liver cell L02 and three tumor cells: Bel-7402, SGC-7901 and Hela. Results at cytokine levels are compatible with the observations at liver cell levels. In vivo, CM-CTS are nontoxic to body and could enhance body immune function and exhibited anticancer effect to some extent and these results strongly suggest that CM-CTS is safe in tumor application as biomedical material.

Effects of Carboxymethyl Chitosan on the Growth of Tumor Cells and Tumor Tissue

Abstract

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