Jaagsiekte sheep retrovirus is necessary and sufficient to induce a contagious lung cancer in sheep
Jaagsiekte sheep retrovirus is necessary and sufficient to induce a contagious lung cancer in sheep
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DOI：
10.1128/jvi.73.8.6964-6972.1999
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发表时间：
1999-08-01
影响因子：
5.4
通讯作者：
Fan, H
中科院分区：
文献类型：
--
作者：
Palmarini, M;Sharp, JM;Fan, H
绵羊肺腺瘤病（SPA）是一种传染性和实验性的绵羊肺癌，类似于人类细支气管肺泡癌。D型逆转录病毒，称为绵羊肺腺病毒（jaagsiekte sheep retrovirus，JSRV），与SPA的病因学有关，但由于缺乏（i）用于JSRV繁殖的组织培养系统和（ii）感染性JSRV分子克隆，其在肿瘤诱导中的确切作用尚不清楚。为了研究JSRV在SPA病因学中的作用，我们从源自SPA的天然病例的肿瘤基因组DNA文库中分离全长JSRV前病毒克隆pJSRV（21）。对pJSRV（21）进行了完全测序，并显示与JSRV的原始南非株推断的开放阅读框一致的开放阅读框。用阳离子脂质体复合的pJSRV（21）DNA，经体内感染3只新生羔羊，结果表明pJSRV（21）是一个感染性分子克隆。通过高度敏感的JSRV-U3半巢式PCR在转染后2周至6个月的不同时间点在转染动物的外周血单核细胞（PBMC）中检测到病毒DNA。此外，在转染后9个月处死的两只羔羊的PBMC、肺和纵隔淋巴结中检测到前病毒DNA，但未诱导肉眼或组织学SPA损伤。我们通过用JSRV构建体（pCMV 2 JS（21））瞬时转染293 T细胞来制备JSRV颗粒，其中上游U3被巨细胞病毒早期启动子替换。用这种方法生产的JSRV（21）颗粒接种4只新生羔羊，其中2只在感染后4个月出现典型的SPA症状。结果表明，转基因株的JSRVDNA和蛋白质均呈阳性。因此，JSRV（21）是一种感染性和致病性的分子克隆，是诱导绵羊肺腺瘤病的必要和充分条件。
Sheep pulmonary adenomatosis (SPA) is a contagious and experimentally transmissible lung cancer of sheep resembling human bronchiolo-alveolar carcinoma. A type D retrovirus, known as jaagsiekte sheep retrovirus (JSRV), has been associated with the etiology of SPA, but its exact role in the induction of the tumor has not been clear due to the lack of (i) a tissue culture system for the propagation of JSRV and (ii) an infectious JSRV molecular clone. To investigate the role of JSRV in the etiology of SPA, we isolated a full-length JSRV proviral clone, pJSRV(21), from a tumor genomic DNA library derived from a natural case of SPA. pJSRV(21) was completely sequenced and showed open reading frames in agreement with those deduced for the original South African strain of JSRV. In vivo transfection of three newborn lambs by intratracheal inoculation with pJSRV(21) DNA complexed with cationic lipids showed that pJSRV(21) is an infectious molecular clone. Viral DNA was detected in the peripheral blood mononuclear cells (PBMCs) of the transfected animals by a highly sensitive JSRV-U3 heminested PCR at various time points ranging from 2 weeks to 6 months posttransfection. In addition, proviral DNA was detected in the PBMCs, lungs, and mediastinal lymph nodes of two lambs sacrificed 9 months posttransfection, but no macroscopic or histological SPA lesion was induced. We prepared JSRV particles by transient transfection of 293T cells with a JSRV construct (pCMV2JS(21)) in which the upstream U3 was replaced with the cytomegalovirus early promoter. Four newborn lambs were inoculated with JSRV(21) particles produced in this manner, and two of them showed the classical signs of SPA 4 months postinfection. The resulting turners were positive for JSRV DNA and protein. Thus, JSRV(21) is an infectious and pathogenic molecular clone and is necessary and sufficient to induce sheep pulmonary adenomatosis.