Development of In Vitro Co-Culture Model to Mimic the Cell to Cell Communication in Response to Urban PM 2.5
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Background: Airborne particulate matter (PM), a widespread air contaminant, is a complex mixture of solids and aerosols composed of particles suspended in the air. PM is associated with inflammatory responses and may worsen inflammatory skin diseases. However, the mechanisms through which PM affects atopic dermatitis (AD) remain unclear. Objective: To establish an in vitromodel that more accurately mimics AD using human ke- ratinocyte (HaCaT), dermal fibroblast (HDF), and mast cell (HMC-1) and using this model to investigate the mechanism through which PMs affect AD. Methods: An AD-like in vitro model was established by seeding HaCaT, HDF, and HMC- 1 cells with recombinant human interleukin (IL)-1α and polyinosinic:polycytidylic acid. We confirmed the effect of PM on the inflammatory cytokine expression of a triple-cell culture model. SRM 1649b Urban Dust, which is mainly composed of polycyclic aromatic hydrocarbons, was used as the reference PM. The effects of PM on the expression levels of proinflammatory cytokines and skin barrier markers were assessed using quantitative real-time polymerase chain reaction and western blotting. Inflammatory cytokine levels were measured using an enzyme-linked immunosorbent assay. Results: Interactions between various skin cell types were evaluated using a co-culture system. PM treatment increased mRNA and protein levels of the inflammatory cytokines IL-6, IL-1α, tumor necrosis factor-α, IL-4, and IL-1β and decreased the expression of the skin barrier markers filaggrin and loricrin. Conclusion: Our results suggest that an in vitro triple-cell culture model using HaCaT, HDF, and HMC-1 cells may be reliable for obtaining more physiological, functional, and reproducible data on AD and skin barriers.
Abstract
Background: Airborne particulate matter (PM), a widespread air contaminant, is a complex mixture of solids and aerosols composed of particles suspended in the air. PM is associated with inflammatory responses and may worsen inflammatory skin diseases. However, the mechanisms through which PM affects atopic dermatitis (AD) remain unclear. Objective: To establish an in vitromodel that more accurately mimics AD using human ke- ratinocyte (HaCaT), dermal fibroblast (HDF), and mast cell (HMC-1) and using this model to investigate the mechanism through which PMs affect AD. Methods: An AD-like in vitro model was established by seeding HaCaT, HDF, and HMC- 1 cells with recombinant human interleukin (IL)-1α and polyinosinic:polycytidylic acid. We confirmed the effect of PM on the inflammatory cytokine expression of a triple-cell culture model. SRM 1649b Urban Dust, which is mainly composed of polycyclic aromatic hydrocarbons, was used as the reference PM. The effects of PM on the expression levels of proinflammatory cytokines and skin barrier markers were assessed using quantitative real-time polymerase chain reaction and western blotting. Inflammatory cytokine levels were measured using an enzyme-linked immunosorbent assay. Results: Interactions between various skin cell types were evaluated using a co-culture system. PM treatment increased mRNA and protein levels of the inflammatory cytokines IL-6, IL-1α, tumor necrosis factor-α, IL-4, and IL-1β and decreased the expression of the skin barrier markers filaggrin and loricrin. Conclusion: Our results suggest that an in vitro triple-cell culture model using HaCaT, HDF, and HMC-1 cells may be reliable for obtaining more physiological, functional, and reproducible data on AD and skin barriers.
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