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 alpha (TNF-alpha) 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-alpha (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-alpha. [CAM-1 is constitutively expressed in unstimulated mouse cremaster venules and slightly upregulated after 2 hours of TNF-alpha treatment. Under baseline conditions, neither E-selectin, P-selectin, nor ICAM-1 is detectable in arterioles or capillaries. After TNF-alpha 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.