Staphylococcal enterotoxin-A-induced in-vitro adhesion of HL-60 cells to endothelial cells involves both selectin and integrin families of cell adhesion molecules.
In-vivo exposure to the bacterial superantigen Staphylococcal enterotoxin-A (SEA) induces an inflammatory response characterized by rapid extravasation of leucocytes and release of excessive amounts of cytokines. We have utilized an in-vitro adhesion assay to understand the molecular mechanisms responsible for SEA-induced extravasation of leucocytes. Stimulation of human umbilical cord endothelial cells (HUVEC) with increasing concentrations of recombinant SEA (rSEA) did not influence the in-vitro adhesion of HL-60 cells to HUVEC, whereas stimulation of HUVEC by interleukin (IL)-1beta supported adhesion of HL-60 cells. Increased adhesion of HL-60 cells to HUVEC was noted upon stimulation of endothelium with culture medium obtained from human peripheral blood mononuclear cells (PBM) stimulated with recombinant SEA for 24 (CM-SEA 24 h), 72 (CM-SEA 72 h) and 120 h (CM-SEA 120 h), but not after stimulation with culture medium obtained from control human peripheral blood mononuclear cells (CM), suggesting that soluble factors present in the supernatants play a major role in SEA-induced cell adhesion. While CM-SEA 24 and 72 h induced both a rapid (4 h) and delayed type of adhesion, CM-SEA 120 h only induced a delayed type of adhesion. Stimulation of PBM by SEA resulted in increased levels of IL-1beta, IL-2 and interferon (IFN)-gamma after 24h. Further stimulation for 72-120h resulted in a significant increase in the levels of IL-1beta, IFN-gamma and tumour necrosis factor (TNF). Stimulation of PBM with SEA also resulted in increased levels of soluble and L-selectin in the cell supernatants. Increased cell-surface expression of E-selectin, ICAM-1, HLA-DR and VCAM-1 was detected on HUVEC stimulated with CM-SEA media. While E-selectin and VCAM were induced on HUVEC within a few hours, induction of ICAM and HLA-DR required a longer induction period. Adhesion of HL-60 cells to HUVEC treated with CM-SEA was inhibited by monoclonal antibodies (MoAbs) against both the selectin and integrin families of cell adhesion molecules, suggesting that multiple pathways contribute to SEA-induced leucocyte extravasation. The results suggested that selectin-dependent adhesion was more prominent during the early phase while integrin-induced adhesion occurred at a later stage.
Srinivas
U
U
Department of Clinical Chemistry, Faculty of Health Sciences, Hospital Pharmacy, University Hospital, Linköping, Sweden.
Dohlsten
M
M
Kalland
T
T
Lundblad
A
A
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface / Q000032:analysis; D002448:Cell Adhesion; D017077:Culture Media, Conditioned; D004727:Endothelium / Q000166:cytology*; D004768:Enterotoxins / Q000494:pharmacology*; D018922:HL-60 Cells / Q000166:cytology*; D006801:Humans; D016023:Integrins / Q000378:metabolism / Q000502:physiology*; D007375:Interleukin-1 / Q000378:metabolism; D019041:L-Selectin / Q000378:metabolism / Q000502:physiology*; D014470:Umbilical Cord / Q000166:cytology / Q000378:metabolism; D015854:Up-Regulation
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D017077:Culture Media, Conditioned; D004768:Enterotoxins; D016023:Integrins; D007375:Interleukin-1; D019041:L-Selectin; C026066:enterotoxin A, Staphylococcal