NIH Research Festival
The human gamma-herpesvirus, Kaposi's sarcoma-associated herpesvirus (KSHV) is known to be the causative agent of Kaposi's sarcoma (KS), and two B-cell lymphoproliferative disorders: primary effusion lymphoma (PEL) and Multicentric Castleman’s disease(MCD). Like all herpesviridae, KSHV can replicate through both latent and lytic cycles. KS is the most common tumor in AIDS patients and tumor formation involves a large amount of angiogenesis and inflammation. Tumor cells found within KS lesions are spindle-like cells of endothelial origin harboring KSHV in its latent state. In cell culture, low levels of virus are produced from a myriad of cell types infected with KSHV. This stymies the advancement of KSHV studies and no chronically infected stable endothelial cell-lines are available. In an attempt to resolve these confounding problems, we engineered novel inducible producer telomerase immortalized endothelial (TIME) cells that harbor (r)KSHV.219 virus and the replication and transcription activator (RTA) under the control of a doxycycline-inducible Tet-On trans-activator system. The resulting cells (iTIME.219) fluoresced green (EGFP), but not red (DS-RED) indicating that the virus was latently maintained. Upon characterization, exposing the cells to either Sodium Butyrate (SB) or doxycycline resulted in increased RFP detection in addition to EGFP. This indicated that cells had entered lytic replication in response to SB and a more natural (Dox/RTA) method of induction. Our data shows that inducing the iTIME.219 cells produces large quantities of virus and high levels of infection when titered on Vero endothelial cells. These iTIME.219 cells can be used for KSHV experiments testing pathogenesis and viral spread.
Scientific Focus Area: Virology
This page was last updated on Friday, March 26, 2021