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At least 523 records · Page 29Linked to original sources
PubMed2001

RhoA is required for monocyte tail retraction during transendothelial migration.

Transendothelial migration of monocytes is the process by which monocytes leave the circulatory system and extravasate through the endothelial lining of the blood vessel wall and enter the underlying tissue. Transmigration requires coordination of alterations in cell shape and adhesive properties that are mediated by cytoskeletal dynamics. We have analyzed the function of RhoA in the cytoskeletal reorganizations that occur during transmigration. By loading monocytes with C3, an inhibitor of RhoA, we found that RhoA was required for transendothelial migration. We then examined individual steps of transmigration to explore the requirement for RhoA in extravasation. Our studies showed that RhoA was not required for monocyte attachment to the endothelium nor subsequent spreading of the monocyte on the endothelial surface. Time-lapse video microscopy analysis revealed that C3-loaded monocytes also had significant forward crawling movement on the endothelial monolayer and were able to invade between neighboring endothelial cells. However, RhoA was required to retract the tail of the migrating monocyte and complete diapedesis. We also demonstrate that p160ROCK, a serine/threonine kinase effector of RhoA, is both necessary and sufficient for RhoA-mediated tail retraction. Finally, we find that p160ROCK signaling negatively regulates integrin adhesions and that inhibition of RhoA results in an accumulation of beta2 integrin in the unretracted tails.

Amides↗
PubMed1999

Characterization of the 4C8 antigen involved in transendothelial migration of CD26(hi) T cells after tight adhesion to human umbilical vein endothelial cell monolayers.

In extravasation of T cells, little is known about the mechanisms of transendothelial migration subsequent to the T cells' tight adhesion to endothelium. To investigate these mechanisms, we developed a monoclonal antibody (mAb), termed anti-4C8, that blocks transmigration but not adhesion in a culture system in which high CD26-expressing (CD26(hi)) T cells preferentially migrate through human umbilical vein endothelial cell (HUVEC) monolayers cultured on collagen gels. Anti-4C8 reacted with all CD3(+) T cells and monocytes but not neutrophils or HUVECs. The structure defined by this antibody was an 80-kD molecule. The mAb at 1 mug/ml inhibited 80-90% of migration of CD3(+) T cells through unstimulated and interferon gamma-stimulated HUVEC monolayers without interfering with adhesion and cell motility. When added to the cultures after the adhesion, anti-4C8 completely blocked subsequent transmigration of adherent T cells. Phase-contrast and electron microscopy revealed that T cells are arrested at the intercellular junctions of HUVECs in the presence of anti-4C8. Anti-4C8 exhibited agonistic effects on resting T cells without other stimuli under culture conditions in which anti-4C8 can stimulate T cells. First, in the checkerboard assay using collagen gels, the antibody promoted chemokinetic migration of the cells in a dose-dependent manner from 0.1 to 10 mug/ml. The predominant population of T cells that migrated into collagen gels with impregnated anti-4C8 were CD26(hi). Second, solid-phase-immobilized anti-4C8 induced adhesion of T cells to the substrate, often with polarizations in cell shape and large pseudopods rich in filamentous (F-) actin. Third, soluble anti-4C8 augmented F-actin content preferentially in CD26(hi) T cells when added to T cells at a high dose of 10 mug/ml. Finally, both anti-4C8-induced chemokinetic migration and transendothelial migration were inhibited by pretreatment of T cells with pertussis toxin. These findings suggest that stimulation via the 4C8 antigen increases cell motility of CD26(hi) cells with profound cytoskeletal changes through signaling pathways including G proteins. The 4C8 antigen may be involved in preferential transmigration of CD26(hi) cells adherent to HUVECs.

Actins↗
PubMed2001

L-selectin shedding regulates leukocyte recruitment.

The physiologic role of L-selectin shedding is unknown. Here, we investigate the effect of L-selectin shedding on firm adhesion and transmigration. In a tumor necrosis factor alpha-induced model of inflammation, inhibition of L-selectin shedding significantly increased firm adhesion and transmigration by a lymphocyte function-associated antigen (LFA)-1 and intercellular adhesion molecule (ICAM)-1-dependent mechanism. We examined the quality of leukocyte rolling and L-selectin-mediated signaling. Blockade of L-selectin shedding significantly reduced the "jerkiness" of leukocyte rolling, defined as the variability of velocity over time. A low level of jerkiness was also observed in the rolling of microbeads conjugated with L-selectin, a model system lacking the mechanism for L-selectin shedding. Inhibition of L-selectin shedding potentiated activation of LFA-1 and Mac-1 induced by L-selectin cross-linking as shown by activation epitope expression and binding of ICAM-1-conjugated beads. We conclude that inhibition of L-selectin shedding increases leukocyte adhesion and transmigration by (a) increasing leukocyte exposure to the inflamed endothelium by decreasing jerkiness and (b) promoting leukocyte activation by outside-in signaling. These observations help to resolve the apparent discrepancy between the minor contribution of L-selectin to rolling and the significant leukocyte recruitment defect in L-selectin knockout mice.

Animals↗
PubMed2006

Venular basement membranes contain specific matrix protein low expression regions that act as exit points for emigrating neutrophils.

The mechanism of leukocyte migration through venular walls in vivo is largely unknown. By using immunofluorescence staining and confocal microscopy, the present study demonstrates the existence of regions within the walls of unstimulated murine cremasteric venules where expression of key vascular basement membrane (BM) constituents, laminin 10, collagen IV, and nidogen-2 (but not perlecan) are considerably lower (<60%) than the average expression detected in the same vessel. These sites were closely associated with gaps between pericytes and were preferentially used by migrating neutrophils during their passage through cytokine-stimulated venules. Although neutrophil transmigration did not alter the number/unit area of extracellular matrix protein low expression sites, the size of these regions was enlarged and their protein content was reduced in interleukin-1beta-stimulated venules. These effects were entirely dependent on the presence of neutrophils and appeared to involve neutrophil-derived serine proteases. Furthermore, evidence was obtained indicating that transmigrating neutrophils carry laminins on their cell surface in vivo. Collectively, through identification of regions of low extracellular matrix protein localization that define the preferred route for transmigrating neutrophils, we have identified a plausible mechanism by which neutrophils penetrate the vascular BM without causing a gross disruption to its intricate structure.

Animals↗
PubMed2005

Induction and detection of human mesenchymal stem cell migration in the 48-well reusable transwell assay.

The induction and detection of chemotactic migration in mesenchymal stem cells (MSC) are complicated by their adherent nature. We describe here the experimental details for inducing the transmigration of MSC seeded in microchemotaxis chambers and reliably identifying the translocated cells. A combination of 8-microm pore-sized membranes and a two-step staining procedure resulted in the detection of MSC migration against a gradient of human plasma which plateaued after 4 h. Microscopic enumeration of the transmigrated cells clearly distinguished contaminating nonmigrated cells from transmigrated cells. Chemokinetic and chemotactic effects could be separated and were influenced by precoated fibronectin or vitronectin. Thus, the technique described allows rapid and reliable induction and determination of migration in adherent MSC.

Bone Marrow Cells↗
PubMed2005

Neutrophil migration across tight junctions is mediated by adhesive interactions between epithelial coxsackie and adenovirus receptor and a junctional adhesion molecule-like protein on neutrophils.

Neutrophil (polymorphonuclear leukocytes [PMN]) transepithelial migration during inflammatory episodes involves a complex series of adhesive interactions and signaling events. Previous studies have shown that key adhesive interactions between leukocyte CD11b/CD18 and basally expressed fucosylated glycoproteins followed by binding to desmosomal-associated JAM-C are key elements of the transmigration response. Here we provide the first evidence that PMN-expressed junctional adhesion molecule-like protein (JAML) regulates transmigration via binding interactions with epithelial coxsackie and adenovirus receptor (CAR). Experiments with a JAML fusion protein revealed specific binding of JAML to epithelial CAR expressed at tight junctions in T84 cell monolayers and normal human colonic mucosa. Furthermore, JAML-CAR binding is mediated via the membrane distal immunoglobulin (Ig) loop of CAR and the membrane proximal Ig loop of JAML. PMN bound to immobilized CAR but not JAML in a divalent cation-independent manner. Lastly, in assays of PMN transepithelial migration, JAML/CAR fusion proteins and their antibodies significantly inhibited transmigration in a specific manner. Taken together, these results indicate that JAML and CAR are a novel pair of adhesion molecules that play an important role in modulating PMN migration cross epithelial tight junctions. These findings add a new element to a multistep model of PMN transepithelial migration and may provide new targets for anti-inflammatory therapies.

Antibodies, Monoclonal↗
PubMed2000

Transendothelial migration of 27E10+ human monocytes.

The myeloid-related proteins MRP8 (S100A8) and MRP14 (S100A9), two members of the S100 family of calcium-binding proteins, are co-expressed and form a cell-surface and cytoskeleton-associated heterodimer upon calcium mobilization which is recognized by the mAb 27E10. The heterodimer is abundantly expressed in the cytoplasm of granulocytes and a subpopulation of blood monocytes. Previously, we and others demonstrated endothelium-associated MRP8/14 in inflamed tissues in the vicinity of transmigrating leukocytes, suggesting a function of the proteins in this process. Here, we demonstrate that 27E10(+) cells represent a fast-migrating monocyte subpopulation which preferentially utilizes an ICAM-1-dependent mechanism. The following observations imply a function of MRP8/14 in the transmigration process: (i) higher secretion of MRP8/14 from 27E10(+) monocytes compared to 27E10(-) monocytes after interaction with activated endothelium, (ii) higher expression of CD11b on 27E10(+) compared to 27E10(-) monocytes, (iii) up-regulation of CD11b on 27E10(-) monocytes in the presence of MRP14 or MRP8/14 heterodimers but not MRP8 and (iv) active participation of MRP14 but not of MRP8 in transmigration as shown by blocking with respective antibodies. We show that the interaction of 27E10(+) monocytes with activated endothelium leads to MRP8/14 release which may account for the high MRP8/14 concentrations in body fluids of patients with acute or chronic inflammatory diseases. Released MRP8/14 may serve a function by enhancing CD11b expression and/or affinity in human monocytes and by participating in the transendothelial migration mechanism. Thus, MRP8/14 substantially contributes to the recruitment of monocytes to an inflammatory site.

Antibodies, Monoclonal↗
PubMed1997

Immunosurveillance modelled in vitro: naive and memory T cells spontaneously migrate across unstimulated microvascular endothelium.

As a model for T cell immigration into non-lymphoid tissue we set up an in vitro assay that would allow us to investigate the phenotype of T lymphocytes from peripheral lymph nodes (PLN), mesenteric lymph nodes (MLN) or peripheral blood (PBL) of mice, which were able to spontaneously migrate across unstimulated microvascular endothelium. The transendothelial migrating T cell population was enriched for T lymphocytes expressing a "recently activated/memory' phenotype: LFA-1/CD44/ICAM-1high, but also contained CD45RBhigh and LFA-1low T cells, which in the case of MLN T cells were phenotyped as CD4+ and thus characterized as naive T cells. Transmigrated T cells could be further distinguished from their original populations and from each other by their distinct but heterogeneous expression patterns for L-selectin, alpha 4 beta 7-integrin and PECAM-1. This observation suggests the presence of phenotypically different migratory T cells among MLN, PLN and PBL. Additional studies provided evidence that the capacity to migrate across unstimulated microvascular endothelium was a characteristic of a T cell population that could phenotypically be differentiated from activated T cells. The endothelial cells were found to play an active role in selecting the traversing T cell population, as they controlled the number and phenotype of spontaneously transmigrating T cells. Our studies suggest that the capacity to transmigrate across unstimulated microvascular endothelium and hence to immigrate into non-lymphoid tissue is owned by a phenotypically heterogeneous T cell population, which is enriched for memory T cells but not devoid of naive T cells.

Animals↗
PubMed2006

Trans-basement membrane migration of human basophils: role of matrix metalloproteinase-9.

In allergic disorders, basophils migrate from the blood stream to inflamed tissue sites. Since trans-basement membrane migration is an important step for local basophil accumulation, we performed a human basophil transmigration assay using a model basement membrane, Matrigel. IL-3 in the upper chamber was critical for basophil trans-basement membrane migration over baseline levels, since none of the chemoattractants placed in the lower chambers induced migration. RANTES, IL-8, 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE) and platelet-activating factor (PAF) significantly up-regulated the transmigration of IL-3-treated basophils. Neutralizing experiments indicated the involvement of beta2 integrin and matrix metalloproteinase (MMP)-2/9 in basophil transmigration. Real-time quantitative PCR revealed that basophils constitutively expressed transcripts for MMP-9, and at lower levels, MMP-2, but cell-surface expression was only detected for MMP-9. MMP-9 was also detected in the cytoplasm and culture supernatant of the basophils. Treatment with IL-3 up-regulated the surface level of MMP-9 on the basophils. Our results suggest that basophils possess a unique regulatory mechanism for trans-basement membrane migration which is affected by cytokines, chemoattractants, beta2 integrin and MMPs, especially MMP-9. MMP-9 may be critically involved in the pathogenesis of local basophil influx in allergic diseases.

Basement Membrane↗
PubMed1998

Effect of tissue-plasminogen activator on leukocyte-endothelial interactions at the microcirculatory level.

In free tissue transfer and replantation surgery, there is a debate over whether any pharmacologic agents should be used to improve vessel patency and tissue survival. Because tissue-plasminogen activator (t-PA) is a highly effective and safe fibrinolytic, it may be useful in obtaining and maintaining vessel patency. The direct effects of t-PA on skeletal muscle hemodynamics and leukocyte activation at the microcirculatory level were investigated. Male Sprague-Dawley rats (n = 20) were divided into three experimental groups: control (n = 8), vehicle (n = 6), and t-PA (n = 6). Using the cremaster muscle flap model and intravital microscopy, red blood cell velocity, vessel diameter, capillary perfusion, endothelial edema index, and leukocyte-endothelial interactions (rolling, adhering, and transmigrating leukocytes) in postcapillary venules were measured. In the vehicle and t-PA groups, vehicle or t-PA was infused by means of a catheter inserted into the lower abdominal aorta for local infusion. Except for a significant reduction in the diameter of the first order arterioles from 117 microm to 82 microm (medians; p = 0.026), t-PA did not significantly affect red blood cell velocity, vessel diameter, or capillary perfusion compared with vehicle. However, leukocyte-endothelial interactions did differ significantly in postcapillary venules. Adhering leukocytes counted per visual field decreased from 4.67 in the vehicle group and 3.50 in the control group to 1.67 in the t-PA group (medians; p = 0.015 and p = 0.005, respectively); transmigrating leukocytes in the t-PA group decreased from 4.75 in the vehicle group and 3.50 in the control group to 1.67 in the t-PA group (medians; p = 0.002 and p = 0.043, respectively). t-PA treatment significantly decreased the number of both adhering and transmigrating leukocytes. These novel findings on leukocyte-endothelial interactions suggest that t-PA has anti-inflammatory effect.

Animals↗
PubMed1999

Blockade of platelet endothelial cell adhesion molecule-1 (PECAM-1) protects against ischemia-reperfusion injury in muscle flaps at microcirculatory level.

Several lines of evidence show that platelet endothelial cell adhesion molecule-1 (PECAM-1), a component of endothelial cell junctions, is required for leukocyte transmigration through endothelial cell monolayers. Polymorphonuclear leukocytes play an important role in ischemia-reperfusion injury. We sought to determine whether administering an anti-PECAM-1 antibody would prevent or attenuate ischemia-reperfusion injury in a rat cremaster muscle flap injury model. Eighteen male Sprague-Dawley rats were divided into three groups. Group I (control): Cremaster muscle island flaps were dissected for baseline measurements of eight indicators: numbers of rolling, sticking, and transmigrating neutrophils, numbers of rolling and sticking lymphocytes, number of perfused capillaries, endothelial edema, and vessel permeability. Group II: The prepared cremaster flap was subjected to 4 hours of ischemia and 24 hours of reperfusion. Group III: The muscle flap was subjected to ischemia and reperfusion as in group II, and anti-PECAM-1 antibodies (1 mg/kg) were injected subcutaneously 15 minutes before reperfusion. Blood vessels were observed in vivo under an intravital microscopy system. Microvascular permeability was made visible with injected fluorescein isothiocyanate-labeled albumin and evaluated with Kontron Elektronik computer software. The ischemia-reperfusion-alone group (group II) presented a 225-percent increase in the activation of sticking leukocytes (2.4 +/- 0.4 to 7.8 +/- 0.8, p < 0.05) (p < 0.01). This leukocyte activation was reduced by 83 percent following anti-PECAM-1 monoclonal antibody treatment (1.3 +/- 0.5 per 100 microm) (p < 0.01). At 24 hours, endothelial injury in group II was confirmed by a 4-fold increase in the number of transmigrating leukocytes into the interstitial space (7.6 +/- 1.2 per field versus 1.9 +/- 0.4 per field in controls). This phenomenon was reduced by 85 percent following anti-PECAM-1 monoclonal antibody treatment (1.1 +/- 0.2 per field) (p < 0.01). Analysis showed that the number of flowing capillaries was 67 percent lower in group II (6.8 +/- 0.3 to 2.2 +/- 0.7, p < 0.01). Anti-PECAM-1 antibody treatment caused a 2.5-fold increase in this number (5.6 +/- 0.5, p < 0.01). Microcirculatory permeability index showed a 180-percent increase in group II (p < 0.05) when compared with baseline values. This increased albumin leakage was effectively attenuated by antibody treatment (p < 0.05). Blocking the action of PECAM-1 in vivo by administering monoclonal antibodies significantly attenuated ischemia-reperfusion injury, presumably by inhibiting transendothelial migration of neutrophils and by increasing capillary perfusion at a muscle flap microcirculatory level.

Animals↗
PubMed2001

Proinflammatory effects of bacterial lipoprotein on human neutrophil activation status, function and cytotoxic potential in vitro.

Bacterial lipoprotein (BLP) is the most abundant protein in gram-negative bacterial cell walls, heavily outweighing lipopolysaccharide (LPS). Herein we present findings demonstrating the potent in vitro effects of BLP on neutrophil (PMN) activation status, function, and capacity to transmigrate an endothelial monolayer. PMNs are the principal effectors of the initial host response to injury or infection and constitute a significant threat to invading bacterial pathogens. The systemic inflammatory response syndrome (SIRS) is characterised by significant host tissue injury mediated, in part, by uncontrolled regulation of PMN cytotoxic activity. We found that BLP-activated human PMN as evidenced by increased CD11b/CD18 (Mac-1) expression. Up-regulation of PMN Mac-1 in response to BLP occurred independently of membrane-bound CD14 (mCD14). A similar up-regulation of intercellular adhesion molecule-1 (ICAM-1) on endothelial cells was observed whilst E-Selectin expression was unaffected. PMN transmigration across a human umbilical vein endothelial cell (HUVEC) monolayer was markedly increased after treating either PMN's or HUVEC independently with BLP. This increased transmigration did not occur as a result of any direct effect of BLP on HUVEC monolayer permeability, assessed objectively using the passage of FITC-labeled Dextran-70. BLP primed PMN for enhanced respiratory burst and superoxide anion production in response to PMA, but did not influence phagocytosis of opsonized Escherichia coli. BLP far exceeds LPS as a gram-negative bacterial wall component, these findings therefore implicate BLP as an additional putative mediator of SIRS arising from gram-negative infection.

Antigens, CD↗
PubMed2004

Neutrophils secrete MIP-1 beta after adhesion to laminin contained in basement membrane of blood vessels.

We have recently demonstrated that granulocyte colony-stimulating factor (G-CSF) stimulated the production of epithelial-cell-derived-neutrophil attractant-78 (ENA-78) by neutrophils and that ENA-78 might promote the accumulation of neutrophils that had migrated from the intravascular space into inflammatory tissues. In this study, we examined whether other chemokines could be secreted by neutrophils that had accumulated after migrating from the intravascular space into the inflammatory tissues. We demonstrated that adhesion to laminin contained in the basement membrane and Matrigel, which is an artificial basement membrane model, induced macrophage inflammatory protein-1 beta (MIP-1 beta) in neutrophils and that MIP-1 beta secreted by neutrophils induced the chemotaxis of dendritic cells. These findings suggest that neutrophils transmigrating through the basement membrane are stimulated to secrete MIP-1 beta by the basement membrane, inducing the transmigration of dendritic cells from the intravascular space into inflammatory tissues. We propose that neutrophils intervene between innate immune response and specific immune response by secreting MIP-1 beta during the transmigration through the basement membrane.

Basement Membrane↗
PubMed2000

Neutrophil migration across intestinal epithelium.

Transmigration of neutrophils across epithelial surfaces is the hallmark of inflammatory mucosal diseases of diverse organs. In disorders such as Crohn's disease, ulcerative colitis, pyelonephritis, and bronchitis, for example, neutrophil transmigration correlates with clinical disease activity, is associated morphologically with injury to the epithelium, and is central to disease pathophysiology. The mechanisms by which neutrophils transmigrate across epithelia are, therefore, of considerable significance for numerous pathologic states. In this paper, we discuss current evidence that defines these mechanisms in intestinal epithelium, emphasizing the structural constituents determining adhesive interactions and a subset of the complex regulatory signals between neutrophils and epithelium.

Animals↗
PubMed1997

Porphyromonas gingivalis infection of oral epithelium inhibits neutrophil transepithelial migration.

Periodontal diseases are inflammatory disorders caused by microorganisms of dental plaque that colonize the gingival sulcus and, subsequently, the periodontal pocket. As in other mucosal infections, the host response to plaque bacteria is characterized by an influx of polymorphonuclear leukocytes (PMNs) to the gingival crevice. Neutrophil migration through the epithelial lining of the gingival pocket is thought to be the first line of defense against plaque bacteria. In order to model this phenomenon in vitro, we used the oral epithelial cell line KB and human PMNs in the Transwell system and examined the impact of Porphyromonas gingivalis-epithelial cell interactions on subsequent PMN transepithelial migration. We demonstrate here that P. gingivalis infection of oral epithelial cells failed to trigger transmigration of PMNs. Furthermore, it significantly inhibited neutrophil transmigration actively induced by stimuli such as N-formylmethionyl leucyl phenylalanine, interleukin-8 (IL-8), and the intestinal pathogen enterotoxigenic Escherichia coli. The ability of P. gingivalis to block PMN transmigration was strongly positively correlated with the ability to adhere to and invade epithelial cells. In addition, P. gingivalis attenuated the production of IL-8 and the expression of intercellular adhesion molecule 1 by epithelial cells. The ability of P. gingivalis to block neutrophil migration across an intact epithelial barrier may critically impair the potential of the host to confront the bacterial challenge and thus may play an important role in the pathogenesis of periodontal disease.

Bacterial Adhesion↗
PubMed2003

Divergent interactions of Ehrlichia chaffeensis- and Anaplasma phagocytophilum-infected leukocytes with endothelial cell barriers.

Human anaplasmosis (formerly human granulocytic ehrlichiosis) and human monocytic ehrlichiosis (HME) are emerging tick-borne infections caused by obligate intracellular bacteria in the family Anaplasmataceae. Clinical findings include fever, headache, myalgia, leukopenia, thrombocytopenia, and hepatic inflammatory injury. Whereas Ehrlichia chaffeensis (HME) often causes meningoencephalitis, this is rare with Anaplasma phagocytophilum infection. The abilities of infected primary host monocytes and neutrophils and of infected HL-60 cells to cross human umbilical vein endothelial cell-derived EA.hy926 cell barriers and human brain microvascular cells (BMEC), a human blood-brain barrier model, were studied. Uninfected monocyte/macrophages crossed endothelial cell barriers six times more efficiently than neutrophils. More E. chaffeensis-infected monocytes transmigrated than uninfected monocytes, whereas A. phagocytophilum suppressed neutrophil transmigration. Differences were not due to barrier dysfunction, as transendothelial cell resistivities were the same for uninfected cell controls. Similar results were obtained for HL-60 cells used as hosts for E. chaffeensis and A. phagocytophilum. Differential transmigration of E. chaffeensis- and A. phagocytophilum-infected leukocytes and HL-60 cells confirmed a role for the pathogen in modifying cell migratory capacity. These results support the hypothesis that Anaplasmataceae intracellular infections lead to unique pathogen-specific host cell functional alterations that are likely important for pathogen survival, pathogenesis, and disease induction.

Anaplasma phagocytophilum↗
PubMed2004

Increased rate of apoptosis and diminished phagocytic ability of human neutrophils infected with Afa/Dr diffusely adhering Escherichia coli strains.

The proinflammatory effect of Afa/Dr diffusely adhering Escherichia coli (Afa/Dr DAEC) strains have been recently demonstrated in vitro by showing that polymorphonuclear leukocyte (PMN) transepithelial migration is induced after bacterial colonization of apical intestinal monolayers. The effect of Afa/Dr DAEC-PMN interaction on PMN behavior has been not investigated. Because of the putative virulence mechanism of PMN apoptosis during infectious diseases and taking into account the high level of expression of the decay-accelerating factor (DAF, or CD55), the receptor of Afa/Dr DAEC on PMNs, we sought to determine whether infection of PMNs by Afa/Dr DAEC strains could promote cell apoptosis. We looked at the behavior of PMNs incubated with Afa/Dr DAEC strains once they had transmigrated across polarized monolayers of intestinal (T84) cells. Infection of PMNs by Afa/Dr DAEC strains induced PMN apoptosis characterized by morphological nuclear changes, DNA fragmentation, caspase activation, and a high level of annexin V expression. However, transmigrated and nontransmigrated PMNs incubated with Afa/Dr DAEC strains showed similar elevated global caspase activities. PMN apoptosis depended on their agglutination, induced by Afa/Dr DAEC, and was still observed after preincubation of PMNs with anti-CD55 and/or anti-CD66 antibodies. Low levels of phagocytosis of Afa/Dr DAEC strains were observed both in nontransmigrated and in transmigrated PMNs compared to that observed with the control E. coli DH5alpha strain. Taken together, these data strongly suggest that interaction of Afa/Dr DAEC with PMNs may increase the bacterial virulence both by inducing PMN apoptosis through an agglutination process and by diminishing their phagocytic capacity.

Annexin A5↗
PubMed2004

Contribution of proteoglycans to human immunodeficiency virus type 1 brain invasion.

As a neurotropic virus, human immunodeficiency virus type 1 (HIV-1) invades the brain and causes severe neuronal, astrocyte, and myelin damage in AIDS patients. To gain access to the brain, HIV-1 must migrate through brain microvascular endothelial cells (BMECs), which compose the blood-brain barrier (BBB). Given that BMECs lack the entry receptor CD4, HIV-1 must use receptors distinct from CD4 to enter these cells. We previously reported that cell surface proteoglycans serve as major HIV-1 receptors on primary human endothelial cells. In this study, we examined whether proteoglycans also impact cell-free HIV-1 invasion of the brain. Using an artificial BBB transmigration assay, we found that both heparan and chondroitin sulfate proteoglycans (HSPGs and CSPGs, respectively) are abundantly expressed on primary BMECs and promote HIV-1 attachment and entry. In contrast, the classical entry receptors, CXCR4 and CCR5, only moderately enhanced these processes. HSPGs and CSPGs captured HIV-1 in a gp120-dependent manner. However, no correlation between coreceptor usage and transmigration was identified. Furthermore, brain-derived viruses did not transmigrate more efficiently than lymphoid-derived viruses, suggesting that the ability of HIV-1 to replicate in the brain does not correlate with its capacity to migrate through the BBB as cell-free virus. Given that HIV-1-proteoglycan interactions are based on electrostatic contacts between basic residues in gp120 and sulfate groups in proteoglycans, HIV-1 may exploit these interactions to rapidly enter and migrate through the BBB to invade the brain.

Blood-Brain Barrier↗