Leukocyte recruitment and control of vascular permeability in acute inflammation

Abstract

The inflammatory process is fundamental in host defense against tissue injury or infection. However, the inflammatory reaction may itself cause harm to the host and contribute to tissue damage and organ dysfunction. Leukocyte recruitment and edema formation are key components of the inflammatory response. This thesis reports experiments that were undertaken to further elucidate the mechanisms controlling leukocyte extravasation and concurrent alteration of vascular permeability in acute inflammation.

In order for leukocytes to penetrate the vessel wall they need to sequentially interact with the endothelial lining and the perivascular basement membrane (BM) of which laminin-411 is a major constituent. The role of BM laminin-411 in leukocyte recruitment to inflammatory loci was addressed using α4 chain deficient (Lam4-/-) and wild-type (WT) mice. Recruitment of all major leukocyte subsets (neutrophils, monocytes, and lymphocytes) was reduced in Lam4-/- mice compared to WT. With the use of intravital microscopy it was concluded that this decrease was due to impaired diapedesis through the vessel wall.

Concurrent with neutrophil recruitment to extravascular tissue, there is an increase in vascular permeability. However, the mechanism behind this alteration is unknown. It was shown that stimulation of neutrophils with the potent chemoattractant leukotriene B4 (LTB4) leads to degranulation and release of, amongst others, heparin binding protein (HBP). Further, postsecretory supernatants from LTB4-stimulated neutrophils induced intracellular calcium mobilization in endothelial cells in vitro and increase in vascular permeability in vivo. Selective removal of HBP from the supernatant significantly reduced these activities indicating a role for HBP in LTB4-induced plasma extravasation. The mechanism behind neutrophil-induced alteration of endothelial barrier function was further investigated and revealed a pivotal role of the kallikrein-kinin system. Neutrophil activation was shown to enable proteolytic processing of high molecular weight kininogen bound to endothelial cells. Accordingly, plasma exudation in vivo in response to challenge with leukocyte chemoattractants was largely annulled by antagonists of the kallikrein-kinin system. Collectively, the data provide novel insight into the regulation of neutrophil-induced plasma extravasation and may help to identify better therapeutic strategies for interventions in inflammatory disease.

To investigate the role of neutrophil-induced alterations in vascular permeability in a clinically relevant setting, experiments were performed using controlled cortical impact (CCI) as a model for traumatic brain injury (TBI) in normal mice and in mice that were depleted of neutrophils. Neutrophil depletion did not significantly affect plasma leakage across the bloodbrain barrier after CCI. Yet, neutrophils were found to play a role in edema formation in brain tissue after injury. At a later phase, neutropenic mice displayed a decreased number of activated microglia, and an attenuation of tissue loss after injury. These results suggest that neutrophils contribute to the secondary injury following TBI. Altogether, this thesis provides insight into the role of the BM in leukocyte recruitment and clarifies the mechanism behind neutrophil-induced edema formation in acute inflammation.