A new continuous monitoring system for simultaneous measurements of intracellular calcium and either endothelial permeability or leukocyte extravasation

Objective and Design: We have developed a continuous monitoring system that will quantify lymphocyte extravasation or intercellular permeability with intracellular calcium measurements in a single preparation. Materials: Human microvascular endothelial cells, human lymphocytes and his tamine. Treatment: Endothelial barrier function and intracellular calcium were exam ined upon application of either human lymphocytes (1 x 10(5) cells/ml) or h istamine (1 mu M). Methods: Endothelial cells labeled with FURA-2 were examined for both chang es in endothelial barrier function and mobilization of intracellular calciu m using fluorescence spectroscopy and ratio imaging, respectively. Results: Increases in endothelial intracellular calcium occur in the first minute and are followed by increases in endothelial intercellular permeabil ity to albumin (5-7 min) or lymphocytes (15-20 min). Approximately 60 min f ollowing calcium mobilization, there was a second and larger extravasation. Conclusion: The continuous monitoring system expands our understanding of t he time course of the permeability and extravasation events to mobilization of endothelial intracellular calcium.