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|Title:||Characterization of HCMV Clinical Strains in Epithelial Cells|
|Abstract:||Human cytomegalovirus (HCMV) is a DNA virus infecting over 80% of the world population. Though HCMV rarely harms healthy individuals, it can cause serious and potentially fatal complications in fetuses and immunocompromised adults, and is the most common cause of congenital brain infection in humans. HCMV establishes latency and persists, occasionally resurfacing to replicate even in the presence of robust host immunity. Laboratory and clinical strains propagate effectively in fibroblast cells, but have had limited success in infecting and replicating within epithelial cells. Epithelial cells are a relevant cell type in which to study HCMV; the virus is spread through bodily fluids like saliva and urine, often first infecting the epithelial cells lining body cavities. Since little is known about the way HCMV navigates epithelial cell infection in terms of regulation of cellular processes like transcription, translation, and modulation of immune defenses, we propose that there is potential for novel findings within this area of study. We have characterized two clinical strains with high tropism for epithelial cells, specif- ically within the primary cell type ARPE-19. These strains were grown in epithelial cells and have a functional UL128 locus. Performing a systematic analysis of HCMV infection in ARPE-19 allowed us to understand these viruses and their patterns of replication and gene expression within epithelial cells, an area of research that has been largely abandoned due to limited success in infecting epithelial cells. Merlin RL13-UL128+ (Merlin-Epi) and TB40 RL13-UL128+ (TB40-Epi), both grown from epithelial cells, were able to infect ARPE-19 cells at the high rate of 70% while also demonstrating higher than anticipated levels of viral DNA replication. RT-qPCR indicated miRNA and mRNA expression at various time points, completing our characterization of Merlin-Epi and TB40-Epi.|
|Type of Material:||Princeton University Senior Theses|
|Appears in Collections:||Molecular Biology, 1954-2020|
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