Xu Qingxiang, Sun Xitai, Qiu Yudong, Zhang Heyuan, Ding Yitao
Hepatobiliary Surgery Department, Affiliated Drum Tower Hospital of Medical College, Nanjing University, Nanjing, Peoples Republic of China.
Ann Clin Lab Sci. 2004 Winter;34(1):87-93.
A bioartificial liver (BAL) based on viable porcine hepatocytes can serve as a bridge to liver transplantation in patients with acute liver failure (ALF). To support liver functions, an adequate mass of hepatocytes is needed, which depends upon the cell density in the BAL device. This study evaluated the optimal density of hepatocytes within BAL devices that were constructed by perfusing porcine hepatocyte suspensions mixed with cytodex-3 into polysulfon hollow-fibers. The BAL devices were prepared with 6 different cell densities. The mass of hepatocytes in each device was evaluated for (a) cell viability, (b) ability to degrade diazepam, (c) ability to synthesize urea, (d) incorporation of [3H]-leucine into protein, (e) glucose-6-phosphatase activity, (f) total RNA content, and (g) p53 gene expression. Hepatocyte viability was about 90% in each device. With increasing hepatocyte density, the diazepam concentration in the medium decreased from 9.26 +/- 0.96 mg/L at 1 x 10(5) cells/ml to a minimum of 5.25 +/- 1.02 mg/L at 5 x 10(6) cells/ml and thereafter remained at low levels. Urea production and [3H]-leucine incorporation into protein increased progressively until the cell density reached 5 x 10(6)/ml and thereafter remained at high levels. Glucose-6-phosphatase activity and total RNA content stayed at high levels until the cell density reached 5 x 10(6)/ml and then progressively decreased. p53 gene expression differed from the other parameters, since it increased only when the cell density reached 5 x 10(7)/ml. In conclusion, the density of 5 x 10(6) cells/ml is a critical inflection point for most of the functional parameters, although p53 gene expression is not elevated at this cell density. These findings suggest that 5 x 10(6) cells/ml is the optimal hepatocyte density in the hollow-fiber BAL device.
基于活猪肝细胞的生物人工肝（BAL）可作为急性肝衰竭（ALF）患者肝移植的桥梁。为支持肝功能，需要足够数量的肝细胞，这取决于BAL装置中的细胞密度。本研究评估了通过将与cytodex-3混合的猪肝细胞悬液灌注到聚砜中空纤维中构建的BAL装置内肝细胞的最佳密度。制备了具有6种不同细胞密度的BAL装置。对每个装置中的肝细胞数量进行了以下评估：（a）细胞活力，（b）地西泮降解能力，（c）尿素合成能力，（d）[3H] - 亮氨酸掺入蛋白质的能力，（e）葡萄糖-6-磷酸酶活性，（f）总RNA含量，以及（g）p53基因表达。每个装置中的肝细胞活力约为90%。随着肝细胞密度的增加，培养基中的地西泮浓度从1×10（5）个细胞/毫升时的9.26±0.96毫克/升降至5×10（6）个细胞/毫升时的最低值5.25±1.02毫克/升，此后保持在低水平。尿素生成和[3H] - 亮氨酸掺入蛋白质的量逐渐增加，直到细胞密度达到5×10（6）/毫升，此后保持在高水平。葡萄糖-6-磷酸酶活性和总RNA含量在细胞密度达到5×10（6）/毫升之前保持在高水平，然后逐渐下降。p53基因表达与其他参数不同，因为它仅在细胞密度达到5×10（7）/毫升时增加。总之，5×10（6）个细胞/毫升的密度是大多数功能参数的关键拐点，尽管在此细胞密度下p53基因表达未升高。这些发现表明，5×10（6）个细胞/毫升是中空纤维BAL装置中最佳的肝细胞密度。
