@article{mbs:/content/journal/jgv/10.1099/0022-1317-80-8-2149, author = "Hukkanen, Veijo and Mikola, Hannamari and Nykänen, Marja and Syrjänen, Stina", title = "Herpes simplex virus type 1 infection has two separate modes of spread in three-dimensional keratinocyte culture", journal= "Journal of General Virology", year = "1999", volume = "80", number = "8", pages = "2149-2155", doi = "https://doi.org/10.1099/0022-1317-80-8-2149", url = "https://www.microbiologyresearch.org/content/journal/jgv/10.1099/0022-1317-80-8-2149", publisher = "Microbiology Society", issn = "1465-2099", type = "Journal Article", abstract = "This study describes the outcome of herpes simplex virus type 1 (HSV-1) infection in an organotypic raft culture of spontaneously immortalized HaCat keratinocytes and human fibroblasts, as related to the virus load and epithelial stratification and differentiation. In this model, a confluent monolayer of HaCat keratinocytes was formed 60 h after seeding. Inoculation of HSV-1 before induction of differentiation by lifting of the culture to the air–liquid interface always resulted in a productive infection, but the virus yield was highest when the inoculation took place 72 h after seeding. Even at 0·1 p.f.u. per culture, the HaCat cultures became HSV positive. Infection of the full-thickness epithelium at 5 p.f.u. per culture resulted in a productive infection of the whole epithelium. The HaCat cells were about 10 times more sensitive to HSV-1 infection than the Vero cells in which the virus stocks were titrated. The raft cultures infected 30 min after lifting were negative by HSV-1 culture, and no HSV-1 antigen was detected by immunocytochemistry. PCR showed the presence of HSV-1 DNA and in situ hybridization showed reactivity with a latency-associated RNA probe, indicating the presence of a non-productive infection. Two different patterns of virus spread in epithelia were found: (i) lateral spread through the superficial layers of the epithelium and (ii) a demarcated infection throughout the whole thickness of the epithelium at the margins of the culture.", }