羧甲基壳聚糖锌多肽复合材料对人牙周膜成纤维细胞的毒性

Cytotoxicity of carboxymethyl chitosan zinc and peptide to human periodontal ligament cells

目的 :研究羧甲基壳聚糖锌多肽复合材料(carboxymethyl chitosan zinc and peptide,CMC-Zn+-P)对人牙周膜成纤维细胞(human periodontal ligament cells,HPDLCs)的细胞毒性,为材料的安全使用提供依据。方法:体外培养人牙周膜成纤维细胞,并进行形态学及免疫组织化学鉴定。使用含不同浓度材料浸提液的细胞培养液培养细胞,并进行分组,1~4组为实验组,材料浸提液浓度分别为100%、75%、50%和25%,第5组为对照组,不含材料浸提液。采用CCK-8法测定复合材料不同浓度浸提液对细胞增殖活性的影响,通过细胞相对增殖率进行细胞毒性评级。采用SPSS 17.0软件包对数据进行统计学分析。结果 :原代培养的细胞呈成纤维细胞形态,免疫组织化学染色结果显示细胞波形丝蛋白染色阳性,角蛋白染色阴性,属中胚层组织来源。不同浓度材料浸提液对细胞均有促进增殖的作用,相对增殖率均大于100%,细胞毒性分级为0级。结论:CMC-Zn+-P复合材料对体外培养的HPDLCs无细胞毒性,符合国家标准的要求。

PURPOSE： The purpose of this investigation was to study the cytotoxicity of carboxymethyl chitosan zinc and peptide to human periodontal ligament cells in vitro, in order to provide a reference for understanding materials＇ safety.METHODS： The primary cells were obtained from human periodontal tissues and cultured. The cultured cells were identified by observing the shape under microscope and by immunocytochemical method. HPDLCs were cultured in different concentrations of the tested materials extract and the activity of cells was evaluated using CCK-8 assay. The cytotoxicity grade was determined in term of the cell relative growth rates. The concentration of the tested materials extract was 100%, 75%, 50% and 25% in group 1-4, respectively. The fifth group had no materials. All statistical analysis was performed using SPSS 17.0 software package. RESULTS： The primary cells owned fibroblasts＇ shape. Immunocytochemical analysis showed the cells were stained positively to antibodies against vimentin, and negatively to antibodies against cytokeratin, which indicated that they were external embryo mesenchymal cell. The cell relative growth rates were more than100%, no matter the concentrations of materials extract were. The cytotoxicity grade of CMC-Zn＋-P was 0. CONCLUSIONS：CMC-Zn＋-P exhibits no cytotoxicity to HPDLCs in vitro, which meets the requirements of the national standard.