Regulation of E-selectin, P-selectin, and intercellular adhesion molecule 1 expression in mouse cremaster muscle vasculature

Objective: To investigate the expression of P- and E-selectin and intercellular adhesion molecule 1 (ICAM-1) in the vasculature of mouse cremaster muscles both with and without tumor necrosis factor α (TNF-α) treatment. Methods: Mice received injections of monoclonal antibody to P-selectin, E-selectin, or ICAM-1 before fixation to restrict detection, to antigen expressed on the endothelial surface. Whole-mount preparations of mouse cremaster muscles were fixed in acetone and stained using biotinylated secondary antibody and peroxidase-conjugated streptavidin. Results: P-selectin is expressed on the endothelial surface of cremaster muscle venules within 10 minutes after exteriorization. Expression increases upon treatment with TNF-α (2 hours), reflecting transcriptional regulation of P-selectin expression in situ. The baseline E-selectin expression is patchy and barely detectable but shows a significant upregulation after treatment with TNF-α. ICAM-1 is constitutively expressed in unstimnlated mouse cremaster ventiles and slightly upregulated after 2 hours of TNF-α treatment. Under baseline conditions, neither E-selectin, P-selectin, nor ICAM-1 is detectable in arterioles or capillaries. After TNF-α treatment, arterioles stain faintly for P-selectin, but not E-selectin or ICAM-1. Conclusion: The temporal and spatial pattern of expression of P- and E-selectin and ICAM-1 is consistent with the functional role of these molecules in mediating preferential leukocyte rolling and adhesion in mouse cremaster muscle venules.