Enosawa Shin, Yuan Wenji, Douzen Masaharu, Nakazawa Atsuko, Omasa Takeshi, Fukuda Akinari, Sakamoto Seisuke, Shigeta Takanobu, Kasahara Mureo
Clinical Research Center, National Center for Child Health and Development , Tokyo , Japan.
† Department of Pathology, National Center for Child Health and Development , Tokyo , Japan.
Cell Med. 2012 May 14;3(1-3):13-18. doi: 10.3727/215517912X639469. eCollection 2012 Jan.
Hepatocyte transplantation is hoped to be an alternative treatment to certain cases of liver transplantation. The most promising indication is for congenital metabolic diseases, especially in infants. To establish a safe protocol for hepatocyte transplantation into infants, we examined physiological changes during treatment using infantile pigs. Recipient pigs (domestic, crossbred with Large-Yorkshire, Landrace, and Duroc; 2.5 kg; 7-14 days old) were anesthetized with isoflurane; a midline incision of minimum length provided access to the superior mesenteric vein. A double lumen catheter was inserted through the vein to within 1 cm of the hepatic portal region. Physiological parameters such as heart rate, systemic blood pressure, and portal pressure were monitored. Cryopreserved porcine hepatocytes isolated from the same strain were suspended in physiological saline and transfused through the catheter. In experiments requiring tracing, cells were stained with fluorescent dye prior to transfusion. Recipient pigs were kept for 1 day and sacrificed for histological liver examination. After preliminary experiments, the optimized number and concentration for hepatocyte transplantation were determined to be 1 × 10 cells/kg and 1 × 10 cells/ml. The cell suspension was transfused at a rate of 0.67 ml/min. No marked anomaly of physiological parameters was observed, whereas light tachycardia occurred in preliminary trials when the transfusion rate was faster than our standard protocol. All five pigs transfused with the established method recovered from anesthesia and survived with good vital signs. Histology revealed that the transfused hepatocytes were integrated in the hepatic tissue.
肝细胞移植有望成为某些肝移植病例的替代治疗方法。最有前景的适应症是先天性代谢疾病，尤其是在婴儿中。为了建立一种安全的将肝细胞移植到婴儿体内的方案，我们使用幼猪研究了治疗过程中的生理变化。受体猪（国内品种，与大约克夏、长白和杜洛克杂交；体重2.5千克；7 - 14日龄）用异氟醚麻醉；做最小长度的中线切口以暴露肠系膜上静脉。将双腔导管通过静脉插入至肝门区域内1厘米处。监测心率、体循环血压和门静脉压力等生理参数。从同一品系分离的冷冻保存的猪肝细胞悬浮于生理盐水中，并通过导管输注。在需要追踪的实验中，细胞在输注前用荧光染料染色。受体猪饲养1天，然后处死进行肝脏组织学检查。经过初步实验，确定肝细胞移植的最佳数量和浓度分别为1×10⁹个细胞/千克和1×10⁸个细胞/毫升。细胞悬液以0.67毫升/分钟的速度输注。未观察到生理参数有明显异常，而在初步试验中，当输注速度快于我们的标准方案时出现了轻度心动过速。用既定方法输注的所有5头猪均从麻醉中恢复，生命体征良好，存活下来。组织学显示输注的肝细胞整合到了肝组织中。
