Establishment and Characterization of Human Cell Lines Stably Infected With Bovine Leukemia Virus In Vitro Reveals Apoptosis After Latency and Silencing.

Bovine leukemia virus (BLV) is the etiological agent of B-cell leukemia/lymphoma in cattle worldwide. Although previous studies have linked BLV to breast cancer in women, its zoonotic potential remains controversial. Moreover, no previous studies have clearly investigated the BLV latency in human cells. In this study, we aimed to establish and characterize human cell lines stably infected with BLV as in vitro models of natural BLV infection. Human epithelial 293T, human cervical cancer HeLa cells (HeLa), and human breast cancer MCF7 cells were infected with cell-free virus from persistently infected fetal lamb kidney (FLK-BLV) cultures. Single clones of the three infected cell lines were tested every 2 weeks for 18 months. The three infected clones maintained the full-length BLV provirus. None of the three infected cell lines showed cellular expression of viral gp51 and p24 proteins, cell-to-cell infectivity, or detectable virion p24 and RNA throughout the study, indicating BLV silencing in human cells. A significant delay in the doubling time of BLV-infected 293T and MCF7 cells with cytopathic effects on cellular morphology was observed after BLV latency. The findings revealed that latent BLV-induced apoptosis in 293T and MCF7 cells but not in HeLa. Additionally, analysis of apoptosis-related pathways showed that BLV infection did not increase expression of antiapoptotic markers in all infected cell lines. In conclusion, we successfully established three stably infected human cell lines by BLV silencing. Of critical importance to public health, human cells latently infected with BLV undergo apoptosis but not malignant transformation. Further studies to analyze the mechanisms of BLV-human interaction in established models are in progress.