灌注式培养复方壳多糖组织工程皮肤的营养条件

NUTRITIONAL CONDITION FOR COMPOSITE CHITOSAN TISSUE ENGINEERED SKIN IN PERFUSION CULTURE

目的探索灌注培养条件下不同营养浓度对复方壳多糖组织工程皮肤形态结构及细胞凋亡的影响,为进一步优化灌注营养条件提供依据。方法常规构建组织工程皮肤后,将复方壳多糖组织工程皮肤分别置于浓度为40%和80%的培养基中进行灌注式培养,分别于第5天及第10天取材进行HE染色观察细胞形态,利用TUNEL染色检测细胞凋亡情况。结果复方壳多糖组织工程皮肤呈典型双层结构,表真皮界限清晰,表皮细胞各层次界限不清楚,且均可见明显的角化过度;真皮区成纤维细胞成梭形,排列有序。40%的培养基培养第5天时真皮和表皮中凋亡细胞数显著高于同时间80%培养基的凋亡细胞数(P<0.05)。80%的培养基培养第10天表皮中凋亡细胞数显著高于培养第5天的凋亡细胞数(P<0.05)。结论灌注式培养可用于组织工程皮肤的培养,80%培养基浓度较40%培养基浓度培养效果更佳。

Objective To explore the influence of different nutritional concentrations on morphological structure and cell apoptosis of composite chitosan tissue engineered skin under perfusion culture, in order to provide the basis for further optimized perfusion nutritional condition and application of tissue engineered skin. Methods The composite chitosan tissue engineered skin was placed on substrates with concentrations of 40% , and 80% re- spectively for perfusion culture after the regular tissue engineered skin was constructed. HE staining was used for observing cellular morphology 5 d, and 10 d later respectively . TUNEL staining was applied to test the number of the apoptotic cells. Results The composite chitosan tissue engineered skin showed typical bilayer structure. The de- marcation was clear between the epidermis and dermis, but was not clear among various layers of the epidermal cells; and obvious hyperkeratinization was found in all of them. Fibroblasts in the＂ dermis were fusiform in shape, and orderly arranged. The number of apoptotic cells in the substrate with a concentration of 40% was significantly higher than that in the substrate with a concentration of 80% after 5 d-perfusion culture （ P〈0.05 ）. Moreover, for tissue engineered skin cultivated in the substrate with a concentration of 80% , the number of apoptotic cells of the epidermis after 10 d-culture was higher sue engineered skin; furthermore, the concentration of 40%. than that after 5 d-culture. Conclusion Perfusion culture can be used for tis- substrate with a concentration of 80% yields better effect than that with a