Characterization of Dengue Virus Serotype 1 and 2 Infection in Human Dermal Fibroblasts

Abstract

Dengue virus (DENV) is transmitted to human cells in the skin after the bite of a mosquito. As an abundant cell type in the skin, fibroblasts are hypothesized as one of the cell types in the skin susceptible to DENV infection and replication. Their innate immune response to DENV might be critical for regulating DENV early during infection, although this has yet to be fully characterized. Thus, the aim of this work was to characterize DENV infection of fibroblasts, and gain insight surrounding the innate immune response to infection. This was evaluated by comparing their characteristics in vitro against BHK-21 cells, a fibroblast cell line known to grow rapidly in culture and that is also highly susceptible to DENV. Results show that regardless of seeding density, fibroblasts demonstrate slower growth kinetics compared to BHK-21 cells in vitro. Infection with DENV-1 16007 and DENV-2 16681 at 0.01 and 0.1 MOI does not induce cytopathic effects in cultures of human fibroblasts. Infectious DENV titers in viral supernatants collected from fibroblasts are also lacking at low MOIs, especially for DENV-1. Conversely, BHK-21 cells proved to be permissible to both DENV-1 and DENV-2 infection at 0.01 and 0.1 MOI as early as 1 day post infection and infectious DENV titers were measured for both serotypes and MOIs. When the seeding density of human fibroblasts and the MOI of infection were increased, differences in the amount of floating cells were observed for infection at 0.1 and 1 MOI. At 5 and 10 MOI though, cytopathic effects were observed by days 10 and 6 for DENV-1 and DENV-2 infected fibroblasts respectively. Infection from DENV-2 also appeared to be produce stronger CPE, similarly to BHK-21 cells. Infectious DENV titers in viral supernatants collected from DENV-1 infected fibroblast did not appear to increase post infection for any MOI while titers for DENV-2 slightly increased after day 0. Lastly, an 18-plex analyte Luminex kit identified several cytokines and chemokines (IL-6, IL-8, CCL2, CXCL10, CCL5, CCL7, and CXCL1) that were strongly expressed by fibroblasts infected with DENV-2 at 10 MOI days 0-9 post infection. Overall, these results establish an infection model for investigating DENV-1 and DENV-2 infection in human fibroblast. Further insight surrounding the role human fibroblasts may play during the innate immune response to DENV-1 and DENV-2 is also uncovered.