培养猪肝细胞中空纤维型生物反应器的制备

Preparation of hollow fiber bioreactor for culturing pig hepatocytes

目的 探讨培养猪肝细胞中空纤维型生物反应器的制备方法。方法 将两步法分离的新生实验小型猪肝细胞接种于中空纤维器内,用培养液循环式人工毛细管培养系统进行培养,观察生物反应器的蛋白分泌、利多卡因转换能力;检测培养猪肝细胞的乳酸脱氢酶(LDH)漏出量及细胞形态和活力。结果在反应器制备后第2、4、6天可检测出猪肝细胞分泌的白蛋白,生物反应器对利多卡因的转换率为89.6％～96.1％。培养结束时LDH漏出量由(23.7±4.6)U/L升高至(127.8±17.4)U/L(F=39.582,P<0.01),细胞活力由95.8％±0.3%降低至83.8％±4.7％(t=5.135,P<0.01)。结论 可用人工毛细管培养系统制备出1周内性能较好的培养猪肝细胞中空纤维型生物反应器。

Objective To study the method of preparing the hollow fiber bioreactor for culturing pig hepatocytes. Methods Hepatocytes were isolated from experimental suckling minipigs by two-step perfusion with collagenase, and seeded onto hollow fiber bioreactor, then cultured with an artificial capillary cell culture system. The albumin-excretion, lidocaine-transforming rate, lactate dehydrogenase (LDH) release and the cell viability in bioreactors were examined. Results The porcine albumin could be detected by SDS/PAGE on the 2nd, 4th, 6th day. The rates of lidocaine-transforming ranged from 89.6% to 96.1%. The release of LDH into the culture medium increased from (23.7 ± 4.6) U/L to (127.8 ± 17.4) U/L (F=39.582, P < 0.01) during the experiments, and the viability of pig hepatocytes in hollow fiber bioreactor reduced from 95.8% ± 0.3% to 83.8% + 4.7% (t = 5.135, P < 0.01). Conclusion The hollow fiber bioreactor for culturing pig hepatocytes can. be prepared by artificial capillary cell culture system, which provides a certain liver-specific function in 1 week.