Delay of migrating leukocytes by the basement membrane deposited by endothelial cells in long-term culture

Abstract

We investigated the migration of human leukocytes through endothelial cells (EC), and particularly their underlying basement membrane (BM). EC were cultured for 20 days on 3 μm-pore filters or collagen gels to form a distinct BM, and then treated with tumour necrosis factor-α, interleukin-1β or interferon-γ. Neutrophil migration through the cytokine-treated EC and BM was delayed for 20-day compared to 4-day cultures. The BM alone obstructed chemotaxis of neutrophils, and if fresh EC were briefly cultured on stripped BM, there was again a hold-up in migration. In studies with lymphocytes and monocytes, we could detect little hold-up of migration for 20-day versus 4-day cultures, in either the filter- or gel-based models. Direct microscopic observations showed that BM also held-up neutrophil migration under conditions of flow. Treatment of upper and/or lower compartments of filters with antibodies against integrins, showed that neutrophil migration through the endothelial monolayer was dependent on β2-integrins, but not β1- or β3-integrins. Migration from the subendothelial compartment was supported by β1- and β2-integrins for all cultures, but blockade of β3-integrin only inhibited migration effectively for 20-day cultures. Flow cytometry indicated that there was no net increase in expression of β1- or β3-integrins during neutrophil migration, and that their specific subendothelial function was likely dependent on turnover of integrins during migration. These studies show that BM is a distinct barrier to migration of human neutrophils, and that β3-integrins are particularly important in crossing this barrier. The lesser effect of BM on lymphocytes and monocytes supports the concept that crossing the BM is a separate, leukocyte-specific, regulated step in migration.

TIMP-2 Targets Tumor-associated Myeloid Suppressor Cells With Effects in Cancer Immune Dysfunction and Angiogenesis

“…These studies showed that activation of MMPs in the tumor increased angiogenesis mainly by increasing bioavailability of VEGF bound to the extracellular matrix at the initial steps of tumor angiogenesis and subsequent signaling for mobilization of BM inflammatory cells including MDSCs 26,49 . However, more recent studies have shown that MDSCs produce MMPs and directly promote angiogenesis and tumor immune suppression 15,504951 . In addition, these studies have shown the pivotal role that VEGF-R1 play in many MDSC functions, including their recruitment by tumors and formation of the pre-metastatic “niche” 21,23 .…”

Tissue stroma as a regulator of leukocyte recruitment in inflammation

“…It is also possible to construct models, e.g., where ECs are cultured for prolonged periods on porous filters or on collagen gels with stromal cells in close proximity, in which leukocyte recruitment can be followed (e.g., refs. [33,42,53]). Such constructs may be more representative of the in vivo situation, where the proximity of the stroma to the endothelium allows bidirectional cell-cell messages.…”

On, Around, and Through: Neutrophil-Endothelial Interactions in Innate Immunity

“…Although the focus of this review is on the endothelium and its glycocalyx, it is important to acknowledge that other cellular and noncellular elements such as pericytes and the basement membrane are involved in regulation of transmigration of neutrophils (16). For instance, regions of the basement membrane that are low in laminins appear to be preferentially used by migrating neutrophils (120, 124).…”