Investigating Fibroblast Involvement in Vascular Inflammation Using Co-Culture Models

Vascular tissue-engineered in vitro models are increasing in popularity but often lack the complexity required to examine the underlying inflammatory response of diseases such as atherosclerosis. Most models also focus on the interactions between the endothelial cells and smooth muscle cells. However, fibroblasts found in the tunica adventitia are known to express pro-inflammatory cytokines such as Tumour Necrosis Factor Alpha (TNF-α) and Interleukin-1 (IL-1) which significantly affect the expression of adhesion proteins, such as Intercellular Adhesion Molecule 1 (ICAM-1) directing leukocyte infiltration and the recurrence of acute coronary syndromes. Consequently, the aim was to develop an in vitro co-culture model to examine whether the TNF-α induced expression of ICAM-1 could be up-regulated in endothelial and smooth muscle cells when co-cultured with fibroblasts.
Human Umbilical Vein Endothelial Cells (HUVEC), Human Umbilical Artery Smooth Muscle Cells (HUASMC) and Normal Human Dermal Fibroblasts (NHDF) were cultured either alone or in co-culture and were stimulated with 25 U/mL TNF-α for up to 12 hours to assess the surface expression of ICAM-1.
NHDF increased HUVEC TNF-α-induced ICAM-1 expression after 12 hours, in a contact-dependent manner, when compared to monoculture (p<0.001, n=3). The contrary was observed in NHDF, where the fold increase in TNF-α-induced ICAM-1 expression was significantly reduced in co-culture with HUVEC (p<0.001, n=3) and completely inhibited in co-culture with HUASMC.
These results identify the existence of a 2-way signalling pathway between the endothelial cell and the fibroblast, which may be used to control the vascular pro-inflammatory response, fibroblast recruitment and infiltration of leukocytes into the vessel wall during the initial stages of atherosclerotic plaque formation, suggesting that fibroblasts should be included more routinely in vascular co-culture model designs.