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目的:高浓度肝实质细胞与非实质细胞原代共培养,研究其功能活性。方法:应用原位胶原酶灌流法分离大鼠肝细胞,获得有活性的肝细胞,并应用高浓度实质和非实质肝细胞共培养的方法原代培养（共培养组）,并以微囊肝细胞培养为对照组（微囊组）进行了比较。结果:两组均维持白蛋白分泌、尿素合成功能7天;共培养组的白蛋白分泌与微囊组一样为下降趋势,共培养组从（0.870±0.102）降至（0.492±0.040）g·L-1·10-6cells·24h-1,微囊组从（1.147±0.099）降至（0.375±0.012）g·L-1·10-6cells·24h-1;共培养组的肝细胞第4天后维持在一个较稳定的水平,而微囊组肝细胞前3天明显高于共培养组,而后两天显著低于共培养组（P<0.05）。共培养组的尿素合成功能,由（4.50±0.56）降至（4.37±0.19）μmol·L-1·10-6cells·90min-1,微囊组由（5.42±0.81）降至（3.60±0.33）μmol·L-1·10-6cells·90min-1,前3天微囊组高于共培养组,后2天共培养组明显高于微囊组（P<0.05）。共培养2～7天肝细胞的对氨基苯甲酸（PABA）浓度稳定在 7.2～9.9mg/L·10-6cells·24h-1。结论:高浓度肝实质细胞与非实质细胞共培养,可使肝细胞的特异性功能维持7d,而且较微囊肝细胞更适合应用于中空纤维舱型生物人工肝。
OBJECTIVE: To study the functional activity of high-concentration hepatic parenchymal cells co-cultured with non-parenchymal cells. Methods: Rat liver cells were isolated by in situ collagenase perfusion and activated hepatocytes were obtained. Primary cultured cells (co-culture group) were co-cultured with high concentration of parenchymal and non-parenchymal hepatocytes. Cell culture was compared to the control group (microcapsule group). Results: Albumin secretion and urea synthesis function were maintained in both groups for 7 days. The albumin secretion in co-culture group was the same as that in the microcapsule group. The co-culture group decreased from (0.870 ± 0.102) to (0.492 ± 0.040) g · L-1 · 10-6cells · 24h-1, and the number of hepatocytes in co-culture group decreased from (1.147 ± 0.099) to (0.375 ± 0.012) g · L-1 · 10-6cells · 24h- Day after the maintenance of a more stable level, while the microcapsule group of hepatocytes in the first 3 days was significantly higher than the co-culture group, and then two days was significantly lower than the co-culture group (P <0.05). In the co-culture group, the urea synthesis function decreased from (4.50 ± 0.56) to (4.37 ± 0.19) μmol·L-1 · 10-6cells · 90min-1 and from 5.42 ± 0.81 to 3.60 ± 0.33 ) Micromol·L-1 · 10-6cells · 90min-1. The microcapsule group was higher than the co-cultured group in the first 3 days and the co-cultured group was significantly higher than the microcapsule group on the 2nd day (P <0.05). The concentration of p-aminobenzoic acid (PABA) in hepatocytes co-cultured for 2-7 days was stable at 7.2-9.9 mg / L · 10-6 cells · 24h-1. CONCLUSION: Co-culture of high-concentration hepatic parenchyma cells with non-parenchymal cells can maintain the specific function of hepatocytes for 7 days, and is more suitable for the hollow-fiber capsule bioartificial liver than the microcapsule hepatocytes.