Filaggrin (FLG) loss-of-function mutations and the T helper 2 (Th2) dominated cytokine milieu are assumed to cause an impaired skin barrier function in atopic dermatitis (AD), but this presumed mechanism is still largely hypothetical. Previous studies have used in vitro skin equivalents to provide experimental evidence for the role of FLG deficiency but different experimental setups and incomplete knockdown approaches make these data difficult to compare and interpret. Using 3D epidermal equivalents, we here addressed the question if FLG deficiency alters skin barrier function. We excluded interplay of FLG mutations with other AD-typical concomitant factors like inflammation or altered microbiome. We therefore used keratinocytes of ichthyosis vulgaris patients that innately carry homozygous FLG null mutations. This approach uses genetically defined cells without detectable filaggrin protein and avoids potential off-target effects of knockdown approaches. FLG did not alter constitutive or Th2-cytokine dependent expression of differentiation-associated proteins. We observed a decrease of tight junction protein expression, which, however, did not lead to alteration of the outside-in nor did it affect inside-out stratum corneum barrier as measured by permeability for low molecular weight tracers (Lucifer Yellow or biotin-SH). Although these findings do not completely rule out alterations in epidermal permeability for molecules with other biophysical properties, our study contradicts previous work that suggests an increased permeability for low molecular weight polar solutes in FLG deficient epidermis.
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