Fig 2.

Biochemical and histologic effects of EMSY knockdown in a skin organotypic model in vitro. A, qPCR showing knockdown of EMSY mRNA after 10 days in culture (7 days after lifting to the air-liquid interface) normalized to a nontargeting control–treated sample (n = 7, mean 36% reduction compared with the nontargeting control). B and C, Western blot showing knockdown of EMSY protein after 10 days in culture (Fig 2, B) and densitometry to quantify protein knockdown normalized to a nontargeting control (Fig 2, C; n = 7). D, Appearance of organotypic cultures showing an increased granular layer and thickened stratum corneum in response to EMSY siRNA knockdown; representative images were replicated in 10 independent donor experiments. Scale bar = 25 μm. E, Relative thickness of epidermal layers (n = 10). F, Transmission electron microscopy of stratum corneum showing thickness of the stratum corneum (white arrows) with an increase in the number of layers and number of corneodesmosomes (black arrowheads). G, Western blot showing filaggrin expression in skin organotypic cultures untreated (mock) and treated with nontargeting siRNA, FLG siRNA, and EMSY siRNA 7 days after lifting to the air-liquid interface. **P < .001 and ***P < .0001, paired t test. GAPDH, Glyceraldehyde-3-phosphate dehydrogenase; NT, nontargeting control siRNA. Bars show means and SEMs.