RIPK3 Expression in Fibroblasts in an in vivo and in vitro Skin Wound Model: A Controversial Result.

Abstract

One of the major problems of regenerative medicine is the development of hypertrophic scars and keloids. The protein kinase RIPK3 is involved in necroptosis; however, recent evidence indicates that it also has non-canonical functions, including its involvement in the development of renal fibrosis. The aim of our work was to study the expression of RIPK3 in mouse and human skin models of fibrotic processes. A subpopulation of RIPK3+Vim+ cells was found in both human keloid and a mouse wound, with the cell number being significantly greater in the mouse wound bed compared to healthy skin. Real-time polymerase chain reaction (RT-PCR) detected expression of the Ripk3 and fibroblast biomarkers Acta2, Fap, Col1a1, and Fn1 in the cells isolated from the wound bed, indicating that RIPK3 can be expressed by wound bed fibroblasts. An analysis of the human fibroblasts stained with anti-RIPK3 antibodies demonstrated an increase in the fluorescence intensity in the presence of lipopolysaccharide (LPS) at concentrations of 5, 10, 25, 50, and 100 ng/ml and TGF-β at concentrations of 0.1, 1, 2, and 5 ng/ml compared to the control. At the same time, the expression levels of RIPK3 and fibroblast activation markers in the presence of TGF-β and LPS did not differ significantly from the control. It is possible that RIPK3 expression in wound fibroblasts is not directly associated with fibrotic processes, and that kinase plays a different, yet unknown role in wound healing. KEYWORDS scarring, keloid, skin, fibroblasts, cell culture, RIPK3.