Authors:

Arushi Virmani

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About Paper:

Atopic Dermatitis (AD; also called eczema) is a common inflammatory skin condition that can reduce quality of life, causing itching, treatment burden, and sleep disruption. Central to disease pathogenesis is a reduction in the expression of filaggrin (FLG), whether a primary genetic deficiency or secondary to the pathologic increases in type 2 cytokines, and disruption of the epidermal ceramide barrier. The essential sphingoid backbones of these ceramides are synthesized by the serine palmitoyltransferase (SPT) complex, and RNAseq data demonstrates that AD lesional skin shows significant reductions in expression of skin-specific SPT subunits, SPTLC3 and SPTSSB. However, the function of the unique sphingoid bases synthesized by these SPTs is poorly understood. We hypothesized that altering the SPT complex composition by knocking down (KD) SPTLC3 and SPTssb in keratinocytes would compromise the barrier function. SPTLC3 and SPTSSB were KD in keratinocytes using shRNA-expressing-lentivirus, followed by the generation of 3D skin organoids. After 12 days of differentiation, keratinocytes were harvested for mRNA analysis and immunofluorescent staining of differentiation markers and SPT complex subunits. KD of either SPTLC3 or SPTssb reduced expression of the FLG gene, which is known to be deficient in AD. FLG protein was also reduced based on immunofluorescence staining, as were early (DSG1, K10) and other late (LOR) differentiation markers. SPTLC3 KD reduced expression of other SPT complex subunits, suggesting its role in controlling SPT complex activity overall, whereas SPTSSB KD selectively lowered expression of SPTLC2, perhaps as a compensatory response to limit short chain LCBs. These findings suggest that the recently observed reductions in SPTLC3 and SPTSSB in AD may contribute to both the decreased FLG expression and the characteristic dry skin of AD, in addition to altering ceramide composition via changes in the SPT complex.

Source:

Northwestern University

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Co-authors:

Arushi Virmani