RETROVIRAL ETIOLOGY OF PRIMARY BILIARY CIRRHOSIS

原发性胆汁性肝硬化的逆转录病毒病因

• 批准号: 6341532
• 负责人: Andrew L Mason
• 金额: $ 11.88万
• 依托单位: OCHSNER CLINIC FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-30 至 2002-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6341532
• 关键词: Retroviridae; complementary DNA; disease /disorder etiology; electron microscopy; gene expression; human tissue; in situ hybridization; liver; molecular cloning; nucleic acid sequence; polymerase chain reaction; primary biliary cirrhosis; representational difference analysis; tissue /cell culture; virus DNA; virus protein; western blottings

Primary biliary cirrhosis (PBC) is an idiopathic multisystem autoimmune disorder that primarily effects women. Patients with PBC have a pluriglandular syndrome resulting in cirrhosis and sicca syndrome, and a demonstrable autoimmune response to specific mitochondrial oxo-acid dehydrogenase E2 proteins. PBC patients are also prone to develop other autoimmune diseases such as Sjogren's syndrome, thyroiditis, and systemic lupus erythematosus. These autoimmune disorders have all been linked to rettroviral infection as by Western blot studies 30 to 35 percent of patients have indeterminate serum reactivity to HIV proteins and 85 to 05 percent have serum reactivity to human intracisternal A type particle (HIAP-I) which was isolated from salivary glands of patients with Sjogren's syndrome and also visualized by electron microscopy. Likewise, 15 percent of PBC patients have been reported to have false positive HIV ELISA reactivity and in preliminary studies, retrovirus particles have been observed by electron microscopy in the biliary epithelial cells of PBC patients but not controls. Using representational difference analysis, we have isolated and cloned novel retroviral nucleic acid sequences from the liver of a PBC patients. These clones have been used to screen a cDNA library made from bilary epithelium cells isolated from three transplant recipients with PBC. We have now identified more than ten qunique cDNA clones with sequence homology to HIV, SIV, HTLV-1, and IAP as well as the E2 mitochondrial autoantigens. To date, our RTPCR studies have revealed that all the RDA and 3 of 3 novel clones tested to data are not unique to PBC patients, suggesting that they may be derived from endogenous retroviruses. Also, we have conducted Western blot studies which reveal that approximately 74 percent of PBC patients sera have indeterminate reactivity to HIAP proteins and 35 percent of patients react to HIV p24 gag, compared to less than 5 percent of liver disease controls. This suggests that the putative PBC retrovirus shares antigenic determinants with HIV-I and HIAP-I but is a separate virus. In order to further characterize the agent associated with PBC, we plan to identify specific retroviral proteins and nucleic sequences with the ultimate goal of investigating the role this virus plays in the eitology of PBC. Our specific aims are to (1) Clone full length PBC retrovirus from hepatic tissue and PBC cDNA libraries; (2) Isolate the PBC retrovirus in lymphoblastoid and hepatic cell lines by co-culture with infected hepatic tissue; (3) Assess the prevalence of retroviral infection in PBC patients and controls by nucleic acid hybridization techniques and western blot experiments of recombinant or purified viral proteins.

原发性胆道肝硬化（PBC）是特发性多系统 主要影响女性的自身免疫性疾病。 患者 PBC患有多个综合征，导致肝硬化和 西卡综合征，以及对 特定的线粒体氧酸脱氢酶E2蛋白。 PBC 患者也容易发生其他自身免疫性疾病此类 作为Sjogren综合征，甲状腺炎和全身性狼疮 红斑。 这些自身免疫性疾病均已链接 通过Western印迹研究30至35 患者的百分比对艾滋病毒的血清反应不确定 蛋白质和85％至05％的蛋白质对人的血清反应性 肠内A型粒子（HIAP-I），该粒子是从中分离出来的 患有Sjogren综合征患者的唾液腺以及 通过电子显微镜可视化。 同样，PBC的15％ 据报道患者患有假阳性HIV ELISA 反应性和初步研究，逆转录病毒颗粒具有 通过电子显微镜在胆道上皮观察到 PBC患者的细胞，但没有对照。 使用代表性差异分析，我们已经孤立了， 克隆的新型逆转录病毒核酸序列来自肝脏的肝脏 PBC患者。 这些克隆已被用来筛选cDNA 由从三个分离的双拉上皮细胞制成的库 具有PBC的移植受者。 我们现在已经确定的超过 十个Qunique cDNA克隆具有与HIV，SIV的序列同源性的序列同源性 HTLV-1和IAP以及E2线粒体自身抗原。 迄今为止，我们的RTPCR研究表明所有RDA和3 在数据测试的3个新型克隆中，PBC患者不是独有的 表明它们可能来自内源性逆转录病毒。 另外，我们进行了Western印迹研究，揭示了 大约74％的PBC患者血清不确定 对HIAP蛋白的反应性和35％的患者对 艾滋病毒P24插科打，而肝病不到5％ 控件。 这表明推定的PBC逆转录病毒分享 HIV-I和HIAP-I的抗原决定因素，但是独立的 病毒。 为了进一步表征与代理相关的 使用PBC，我们计划确定特定的逆转录病毒蛋白和 核序列的最终目标是研究 该病毒在PBC的生物学中发挥作用。 我们的具体目标 是（1）从肝组织克隆全长PBC逆转录病毒 和PBC cDNA库； （2）隔离PBC逆转录病毒 淋巴母细胞和肝细胞系通过感染 肝组织； （3）评估逆转录病毒感染的患病率 在PBC患者和核酸杂交对照中 重组或重组的技术和蛋白质印迹实验 纯化的病毒蛋白。