微胶囊固定化培养重组盘基网柄菌提高人类可溶性Fas配体的表达

Improving the Expression Level of Soluble Human Fas Ligand by Cultivating Recombinant Dictyostelium discoideum in CMC-ALG Microcapsules

首先研究了应用于液芯羧甲基纤维素钠-海藻酸钙(CMC-ALG)微胶囊制备中的各种化学组分对重组盘基网柄菌生长的影响,然后考察不同组分浓度制备而成的各种CMC-ALG微胶囊内重组盘基网柄菌的生长情况,从而得到较适的微胶囊制备的组分配比(CMC12g·L-1,SA8g·L-1,CaCl2100g·L-1)。结果表明,在以上较适条件下制备的微胶囊内重组盘基网柄菌的生长得到了极大的改善,最大的细胞密度比游离培养时提高了4倍,达到了8.0×107mL-1;相应的人类可溶性Fas配体(FasL)的表达水平也提高了1.5倍,达到了315μg·L-1。最后,开展了微胶囊化重组盘基网柄菌的二次重复发酵FasL的研究,结果表明,经过二次重复批次培养,最大细胞密度可达到1.24×108mL-1,为游离培养的8～10倍,而且FasL的表达水平还能维持高水平(280μg·L-1),为游离培养时的2倍。

The biocompatibility between the growth of Dictyostelium discoideum AX3-pLu8 and chemical components used for the preparation of carboxy methyl cellulose-alginate （CMC-ALG） microcapsules was evaluated at first, and then the effects of the concentrations of each component forming the capsule on the growth behaviors of D. discoideum cultivated in it were studied. For microencapsulated cultivating D. discoideum, it was found that the most suitable capsule components are 12 g.L^-1 of CMC, 8 g.L^-1of sodium alginate （SA） and 100 g.L^-1 of CaCl2. Using the microcapsules with mentioned composition, the cell density in it can reach 8.0×10^7 mL^-1, this value is about 5 times that could be observed in suspension culture, and correspondently, in the capsules a high concentration （315 μg.L^-1） of soluble human Fas ligand （shFasL） was detected, which is about 2.5 times higher than that obtained in suspension culture under the same culture condition. In addition, the immobilized cells could be used effectively for repeated batch cultivation and in the second repeated batch cultivation, a very high cell density （up to 1.24× 10^8 mL^-1） and still a high expression of FasL （280μg.L^-1） could be obtained.