Expression of Wnt5a and its effect on oxLDL uptake in THP‐1 cells (LB534)

Abstract

Objective: To study Wnt5a expression and determine its effect on oxLDL uptake and foam cell development.Introduction: Atherosclerosis is a chronic inflammatory disease involving plaque buildup in the arteries. This disease can lead to a heart attack and stroke, which are two of the biggest causes of death in this country. A key step in the pathogenesis of atherosclerosis is oxLDL uptake by macrophages and foam cell formation in the intima of the artery. Our group has been investigating the role of Wnt5a signaling in atherosclerosis. Previous results in the lab have shown that oxLDL up‐regulated Wnt5a mRNA expression in human monocyte‐derived macrophages and THP‐1 cells (an acute monocytic leukemia cell line). It was also found that Frizzled 5 (Fz5), a Wnt5a receptor, is highly expressed in foam cells present in human atherosclerotic plaques. Our hypothesis is that Wnt5a stimulates oxLDL uptake in macrophages.Methods: THP‐1 cells were cultured in RPMI 1640 with 10% FBS. The cells were differentiated with Phorbol‐12‐Myristate‐13‐Acetate (PMA) for 24 hours. Cells were treated with oxLDL, nLDL, and rWnt5a in combinations to determine their effects on oxLDL uptake. Oil Red staining was used to assess lipid uptake in‐vitro. Expression of CD68, CD36, scavenger receptor, ROR2, Fz5, and Wnt5a were evaluated using immunofluorescence and western blot. RT‐PCR was done to evaluate mRNA expression.Results: Oil Red staining showed a significant increase in the lipid uptake in macrophages treated with oxLDL. Immunofluorescence showed that oxLDL increased CD68 and Wnt5a expression in THP‐1 cells. Wnt5a also increased Fz5 and scavenger receptor expression in THP‐1 cells. Western blot showed that oxLDL significantly increased CD36 expression in THP‐1 cells. RT‐PCR showed that oxLDL induced Wnt5a expression in THP‐1cells.Conclusions: The results show that oxLDL increases the expression of Wnt5a in THP‐1cells. Furthermore, Wnt5a increases the expression of receptors involved in the uptake of oxLDL. While the mechanism of this uptake is still unknown, our results suggest that Wnt5a plays a role in atherogenesis.