Establishment of the novel method for induction of repopulation after hepatocyte transplantation

肝细胞移植后诱导增殖新方法的建立

• 批准号: 18591181
• 负责人: OGAWA Atsushi
• 金额: $ 2.53万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18591181/
• 关键词: Hepatocyte transplantation; Apoptosis; CrmA

Hepatocyte transplantation is expected to be a next generation of treatment for the patient with inborn error of metabolism. Organ transplantation is now performed to the patient who is difficult to be controlled with diet or drug therapy. However organ transplantation is very invasive to the patients and other problems such as a shortage of donor organ or life long immunosuppresive therapy are serious. Hepatocyte transplantation was performed to 18 patients with inborn error of metabolism so far now, but the clinical improvement was not enough, some of the patients were received organ transplantation later. Several methods were investigated in animal model to get a higher rate of repopulation in recipient liver. However the reported methods or drugs used were too invasive or toxic to apply to clinical settings.The purpose of this study was to establishment of a novel method for induction of repopulation after hepatocyte transplantation It is well known that chenodeoxycholic acid induc … More es apoptosis to hepatocytes via Fas signal pathway. We plan to use CDCA to induce apoptosis to hepatocytes of recipient, To prevent the donor hepatocytes from apoptosis with CDCA, CrmAgene, that inhibits caspase 1 and 8, was planed to be transduced with Lentivirus vector CrmA gene was inserted downstream of intestinal bile acid binding protein (I-BABP) promoter donor hepatocytes were expected to be resistant for apoptosis only under the high dose administration of CDCA After hepatocyte transplantation, in which donor hepatocytes were transduced with I-BABP-CrmA gene fragment with Lentivirus vector, CDCA is planed to be administered to the recipient. It is expected that apoptosis is induced to hepatocytes of recipient, on the other hand, donor cell become resistant to apoptosis because of expression of CrmA protein and get repopulated in the recipient liver.The results are as follows. 1) I-BABP promoter was cloned from the genome of C57bl/6 mouse. 2) I-BABP promoter was activated with CDCA. 3) Lentivirus vector was produced containing of gene fragment, I-BABP promoter, EGFP and CrmA gene 4) this gene fragment was transduced to HepG2 cells 5) Expression of EGFP-CrmA protein was observed under the fluorecent microscopy and checked by western blot as well. 6) HepG2 cells transduced the gene fragment were susceptible for apoptosis without CDCA, however, the cells become resistant to apoptosis under the presence of CDCA. Less

预计肝细胞移植将是天生代谢错误的患者的下一代治疗。现在，对难以通过饮食或药物治疗控制的患者进行了器官移植。然而，器官移植对患者和其他问题（例如供体器官或寿命短的免疫培养疗法）非常侵入性。到目前为止，对18例具有代谢的天生误差的患者进行了肝细胞移植，但是临床改善还不够，一些患者后来接受了器官移植。在动物模型中研究了几种方法，以获得受体肝脏中较高的重生率。然而，所报道的方法或药物过于侵入性或有毒，无法应用于临床环境。该研究的目的是建立一种新的方法来诱导肝细胞移植后诱导重生的方法，众所周知，Chenexyoxycholic Acid诱导的……更多的ES ES凋亡通过FAS信号途径引起肝细胞。我们计划使用CDCA诱导受体的肝细胞凋亡，以防止供体肝细胞与CDCA（Crmagene，crmagene）凋亡，抑制caspase 1和8的供体肝细胞，该供体抑制了caspase 1和8仅在肝细胞移植后CDCA的高剂量施用下才能抗凋亡，其中将供体肝细胞用I-BABP-CRMA基因片段与慢病毒载体转移，CDCA计划被施用给接收者。预计凋亡被诱导到受体的肝细胞，另一方面，供体细胞由于CRMA蛋白的表达而对凋亡具有抗性，并在受体肝脏中转移。结果如下。 1）从C57BL/6小鼠的基因组克隆了I-BABP启动子。 2）用CDCA激活I-BABP启动子。 3）慢病毒载体的产生含有基因片段，I-BABP启动子EGFP和CRMA基因4）在荧光显微镜下观察到EGFP-CRMA蛋白的表达转化为HEPG2细胞5）表达EGFP-CRMA蛋白，并通过Western Blot很好地检查。 6）转移基因片段的HEPG2细胞在没有CDCA的情况下易感凋亡，但是，在CDCA存在下，细胞对凋亡具有抗性。较少的

项目成果

先天代謝異常症に対する肝細胞移植治療

肝细胞移植治疗先天性代谢缺陷

• 发表时间: 2007
• 作者: Yoneshige;A.;Kubo;N.;Suzuki;K.;Matsuda;J;小川 真司
• 通讯作者: 小川 真司