Viral Immunoregulatory Genes and Hepatocyte Transplants

病毒免疫调节基因与肝细胞移植

基本信息

批准号：
7347624
负责人：
JAYANTA ROY-CHOWDHURY
金额：
$ 34.14万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-02-01 至 2010-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7347624
关键词：
Adenoviruses Allogenic Allografting Animal Model Animals Apoptotic Autoimmune Process Beta Cell Bilirubin Biological Assay CCL2 gene CXCL10 gene Cell Transplants Cell surface Clonal Deletion Complex Condition Crigler-Najjar Syndrome Cytolysis Defect Dipeptidyl Peptidases Down-Regulation Engraftment Eukaryotic Initiation Factors Excision Gene Expression Genes Genetic Transcription Goals Gunn Rats Hepatocyte Homologous Transplantation Hyperbilirubinemia IL8 gene Immune system Immunologics Immunology Immunosuppression Inbred F344 Rats Inherited Insulin-Dependent Diabetes Mellitus Invasive Islets of Langerhans Lentivirus Infections Lentivirus Vector Liver MHC Class I Genes Marshal Metabolic Methods Modeling Mus Open Reading Frames Pancreas Pathway interactions Plasmids Prevention Procedures Process Proteins Rattus SV40 T Antigens Signal Transduction Pathway Simian virus 40 Site Splenocyte Subfamily lentivirinae Suppressor-Effector T-Lymphocytes System TNF gene TNFSF10 gene Technology Tetanus Helper Peptide Therapeutic immunosuppression Transcription Factor AP-1 Transfection Transgenes Transgenic Mice Transplantation Tumor Necrosis Factor Receptor Upper arm Viral base chemokine in vivo irradiation islet liver transplantation novel prevent promoter protein expression receptor recombinase reconstitution research study response

项目摘要

DESCRIPTION (provided by applicant): Our objective is to develop safe and efficient methods to genetically modify donor liver cells to abrogate allograft rejection after hepatocyte transplantation without the use of long-term immunosuppression. Adenovirus (Ad) early transcription region three (E3) encodes immunomodulatory proteins, which we have used previously to generate transgenic mice expressing the E3 genes in pancreatic islets. These islets did not undergo autoimmune destruction of beta-cells in model murine systems of type 1 diabetes and were not rejected when transplanted into allogeneic recipients. Our hypothesis is that insertion of some or all of the AdE3 genes into primary or immortalized hepatocytes will prevent their rejection upon transplantation into allogeneic recipient jaundiced Gunn rats, which are an animal model for Crigler-Najjar syndrome type 1 (CN1). CN1 is a potentially lethal inherited deficiency of bilirubin conjugation. Our goals are to determine which of the various potential mechanisms of AdE3 suppression of the immune system are most important in facilitating the survival and function of allogeneic hepatocyte transplants. Among the possible immunologic targets acting alone or together are the downregulation of class I MHC by Ad-gp19; inhibition of TNF cytolysis by Ad E3-14.7K; reduction of cell surface expression of pro-apoptotic receptors such as FAS, TRAIL and TNFR, by the complex of Ad E3- 10.4/14.5K; inhibition of signal transduction pathways such as NFkappaB and AP-1 by Ad E3-10.4/14.5K; and prevention of chemokine (MCP-1, IL-8, IP-10) synthesis in response to TNFalpha. In addition, we will attempt to dissect whether Ad E3 acts only on the hepatocyte by intrinsic mechanisms or also alters the effector arm of the immune system by tolerizing by clonal deletion or induction of suppressor cells. The experiments proposed will use both hepatocytes conditionally immortalized with SV40 T antigen (Tag) and primary hepatocytes. Hepatocytes present new challenges and opportunities, because unlike pancreatic islets, the host liver can be manipulated by controlled destruction of endogenous hepatocytes to permit extensive repopulation of the liver with the progeny of the transplanted cells. We will use a conditionally immortalized Fisher rat hepatocyte line to insert Ad E3 genes in new combinations expeditiously by transfection or lentivirus infection. The lentivirus technology also will allow us to use primary hepatocytes to assess their efficacy in reconstituting normal bilirubin processing in the liver.

描述（由申请人提供）：我们的目标是开发安全有效的方法，以基因修饰供体肝细胞，以消除肝细胞移植后消除同种异体移植，而无需使用长期免疫抑制。腺病毒（AD）早期转录区域三（E3）编码免疫调节蛋白，我们以前曾用过这种蛋白质来生成在胰岛中表达E3基因的转基因小鼠。这些小岛没有在1型糖尿病的模型鼠系统中遭受β细胞的自身免疫性破坏，并且在移植到同种异体受体中时不会被拒绝。我们的假设是，将某些或全部ADE3基因插入主要或不朽的肝细胞中，将阻止它们在将其移植到同种异体受体中的Jaund Cunn大鼠中后排斥反应，这是Crigler-Najjar综合征类型1（CN1）的动物模型。 CN1是胆红素结合的潜在致命遗传缺陷。我们的目标是确定免疫系统ADE3抑制的各种潜在机制中的哪种对于促进同种异体肝细胞移植物的生存和功能最重要。在可能的免疫目标中，单独或共同起作用的是AD-GP19对I类MHC的下调；通过AD E3-14.7K抑制TNF胞解；通过AD E3- 10.4/14.5K的复合物，促凋亡受体（例如FAS，TRAIL和TNFR）的细胞表面表达降低；通过AD E3-10.4/14.5K抑制信号转导途径，例如NFKAPPAB和AP-1；以及预防趋化因子（MCP-1，IL-8，IP-10）响应TNFALPHA的合成。此外，我们将尝试剖析AD E3是否仅通过固有机制在肝细胞上作用，或者通过通过克隆缺失或诱导抑制细胞耐受性来改变免疫系统的效应臂。提出的实验将同时使用与SV40 T抗原（TAG）和原代肝细胞有条件永生的肝细胞。肝细胞提出了新的挑战和机遇，因为与胰岛不同，可以通过控制内源性肝细胞的控制来操纵宿主肝脏，以允许使用移植细胞的后代对肝脏进行广泛的重生。我们将使用有条件永生的Fisher大鼠肝细胞系来通过转染或慢病毒感染迅速地将AD E3基因插入新组合。慢病毒技术还将使我们能够使用原发性肝细胞评估其在重建肝脏中正常胆红素加工方面的功效。