The long term objective of our research program is to elucidate the basic mechanisms underlying accumulation of excess fluid in the extravascular spaces. In our current proposal, we focus on 1) the basic mirovascular, interstitial and lymphatic processes contributing to intrinsic regulation of transmicrovascular fluid balance and 2) the alterations in these processes elicited by neutrophil-dependent inflammatory reactions.
The specific aims of the proposed research are: 1) to quantify transport of water and proteins across venular endothelium under control conditions and following activation of neutrophils; 2) to delineate the cellular and pericellular routes of enhanced protein extravasation induced by neutrophil-dependent inflammatory reactions at the venular wall 3) to evaluate the impact of neutrophil-derived products on the physical properties of the extracellular matrix and 4) to examine the role of lymphatic contractility in governing lymph propulsion during intravenous volume loading and following exposure to neutrophil-derived chemicals. A variety of techniques are used to address the specific aims. At the tissue level, venular function is probed by intravital video microscopy applied to the intact microcirculation of hamster cheek pouch; similar techniques are utilized to study the pumping action of muscular lymphatics in the rat mesentery. Transport of water and solutes across single cheek pouch venules is examined with the isolated perfused microvessel approach. In vitro analysis of transport functions of venular endothelium is made possible by culture of endothelial cells derived from venules dissected from the cheek pouch. Electron microscopy and the immunogold antibody technique are used to probe cellular and interendothelial pathways of albumin and neutrophil extravasation across the venular membrane. Finally, corneal stroma and umbilical cord provide model matrices for probing the impact of neutrophil- derived substances on exclusion, diffusion and convection in basement membranes and interstitial spaces, respectively; alterations in matrix stiffness also are examined. To integrate the findings in these and other studies, computer simulations of microvascular, interstitial and lymphatic interactions under normal and stress conditions are utilized. The proposed research should yield new insights into the causes and treatment of inflammatory reactions in the microvasculature, interstitium and lymphatics.
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