Regulation of NF-κB-mediated inflammatory responses by miR-378a-3p in 3D epidermal model

Abstract

miR-378a-3p has previously been shown to be upregulated under the inflammatory stimulus and in psoriatic skin lesions, suggesting a possible role in psoriasis-associated inflammatory signalling. Previous studies have used two-dimensional (2D) keratinocyte culture model and reported that miR-378a-3p activates NF-κB signalling, resulting in increased CXCL8 (IL-8) expression. However, the effects of miR-378a-3p in differentiated keratinocyte culture models have not previously been investigated. Therefore, this thesis focuses on analysing the role of miR-378a-3p in three-dimensional (3D) keratinocyte culture models by comparing its effects on inflammatory gene expression in conventional 2D and differentiated 3D keratinocyte cultures. The study examined the effect of miR-378a-3p transfection on inflammatory gene expression, with particular focus on NF-κB pathway. In the course of the study, expression levels of CXCL8 expression and selected NF-κB-associated genes (TRAF6, NFKBIA, and EGFR) were evaluated under both unstimulated and IL-17A-stimulated conditions. In addition, the effect of miR-378a-3p transfection on keratinocyte differentiation- and proliferation-related genes (KRT5, KRT14, KRT10, and INV) were investigated. The obtained results demonstrated distinct effect of miR-378a-3p between 2D and 3D culture system. In 2D cultures, miR-378a-3p increased CXCL8 expression, consistent with previously reported pro-inflammatory effects. In contrast, differentiated 3D cultures showed reduced CXCL8 expression together with downregulation of TRAF6 and EGFR following miR-378a-3p transfection. Overall, the findings suggest that the effect of miR-378a-3p may depend on differentiation state and culture conditions.