Interferon‐γ Enhancement of E‐Selectin Expression on Endothelial Cells is Inhibited by Monensin

Abstract

Strindhall J, Lundblad A, Påhlsson P. Interferon-g Enhancement of E-Selectin Expression on Endothelial Cells is Inhibited by Monensin. Scand J Immunol 1997;46:338-343 The expression of E-selectin reaches a maximum 4-6 h after stimulation of human umbilical vein endothelial cells (HUVEC) in vitro with tumour necrosis factor-a (TNF-a) and then declines to basal level within 24 h. If interferon-g (IFN-g) is added to the cell culture medium together with TNF-a the surface expression of Eselectin is augmented and prolonged in a synergistic way. The aim of the present study was to investigate if altered protein glycosylation could explain the IFN-g induced persistent surface expression of E-selectin. SDS-PAGE analysis of HUVEC glycoproteins, metabolically radiolabelled in the carbohydrate portion, indicated that addition of IFN-g produced an altered protein glycosylation. Lectin blot analysis using the Sambucus nigra agglutinin lectin also indicated differences in protein glycosylation when HUVEC were incubated with IFN-g/TNF-a compared to TNF-a alone. The kinetics of surface expression of E-selectin were measured using a cell ELISA assay. When HUVEC were incubated with monensin, a potent inhibitor of late Golgi function, together with both TNF-a and IFN-g, the additive effect of IFN-g on E-selectin expression was almost abolished. Since monensin is known to affect glycosylation processing, this experiment suggested that the IFN-g induced change in protein glycosylation might induce the prolonged surface expression of Eselectin. However, when HUVEC were cultured with IFN-g/TNF-a in the presence of several different inhibitors of N-glycosylation processing, no significant effect of E-selectin expression was observed. Regulation of adhesion molecule expression after activation of endothelial cells is likely to play a pivotal role for the inflammatory response. Further studies are needed to understand the mechanisms underlying this regulation.

Insulin and IGF1 receptors in human cardiac microvascular endothelial cells: metabolic, mitogenic and anti-inflammatory effects

“…We found that TNF-a had a pronounced pro-inflammatory effect in HMVEC-Cs and also in HUVECs, whereas high glucose, insulin, IGF1 or IL6 did not affect E-selectin expression. Prominent effects of TNF-a on E-selectin expression have previously been shown in HUVECs and in human aortic endothelial cells (Strindhall et al 1997, Ewart et al 2008, Mako et al 2010. The lack of insulin or IGF1 effects is in agreement with observations made with HUVECs (Balaram et al 1997, Montagnani et al 2002, Madonna et al 2004.…”

“…Expression of E-selectin was detected using an ELISA as described by Strindhall et al (1997). Briefly, HUVECs or HMVEC-Cs, 10 000 cells/well, were seeded in a 96-well microplate and incubated for 2 days.…”

Interferon-Mediated Block in Cell Cycle and Altered Integrin Expression in a Differentiated Salivary Gland Cell Line (HSG) Cultured on Matrigel

“…First, IFN-g treatment affects processing of specific oligosaccharide side chains for some, but not all, transmembrane proteins. (32)(33)(34) Second, inhibitors of specific glycosylation processing steps alter the electrophoretic mobility for a3 integrin in a manner strikingly similar to those shown here. (35,36) Based on these two lines of evidence and lack of altered splicing in the carboxyl-terminal domain of the a3 subunit, our working hypothesis is that IFN-g may inhibit glycosylation of one or more of the integrin subunits.…”

Expression of MUC1 mucin in human umbilical vein endothelial cells (HUVEC).

“…Human umbilical vein endothelial cells (HUVEC) ELISA was performed according to the method described by Strindhall et al [17], with some modifications. Briefly, cells were grown in M199 medium in 96-well microtitre plates to a confluent cobblestone-like monolayer, and then incubated with TNF-α (40 ng/ml) for different periods of time (0, 2, 6 and 18 h).…”