Neutrophil Chemotaxis In Response Research Articles

Neutrophil chemotaxis induced by corneal epithelial cells after herpes simplex virus type 1 infection

Purpose. Neutrophil invasion is a primary event in the development of herpetic keratitis. It has been reported that HSV-1 infection of keratocytes induces the synthesis of IL-8, a potent neutrophil chemoattractant, while corneal epithelium does not. Nevertheless, little is known about the correlation between neutrophil migration and the production of chemotactic factors by HSV-1-infected corneal cells, especially in epithelial cells which form an initial barrier of the ocular surface. We examined whether human corneal epithelial cells as well as keratocytes could induce neutrophil chemotaxis in response to HSV-1 infection.Methods. Human corneal epithelial cells immortalized with SV40 (HCE) and human keratocytes were infected with HSV-1. The culture fluids collected at 4, 12, 24 h after infection were assayed for human neutrophil chemotaxis using a modified Boyden chamber method. IL-8 levels in these supernatants were measured using enzyme-linked immunosorbent assay (ELISA).Results. The chemotactic activity induced by HCE and keratocytes after MP strain of HSV-1 infection peaked as early as 4 h postinfection, then declined. Chemotactic activity induced by HSV-1-infected HCE and IL-8 levels on these supernatants paralleled with the infectious virus titer. It was inhibited by monoclonal anti-IL-8 antibody. UV-inactivation of MP strain abrogated neither the induction of chemotactic activity nor IL-8 secretion of infected HCE.Conclusions. At the early phase of HSV-1 infection, corneal epithelial cells play an important role in inducing neutrophil chemotaxis, which was mediated by IL-8. Curr. Eye Res. 17:687–693, 1998.This work has been conducted in accordance with the Declaration of Helsinki.

Human neutrophil chemotaxis in response to diepoxides of linolenic acid.

Diepoxides of linolenic acid were found to be chemoattractants. The concentration that produces 50% of maximal chemotaxis was 4.5 10(-7) mol/l for human neutrophils when investigated in a chemotaxis test based on the method of spectrophotometric determination of myeloperoxidase activity with buffers containing bovine serum albumin. Leukotriene B4 was used as positive control. The concentration of leukotriene B4 that produces 50% of maximal chemotaxis was 1.8 nmol/l. No chemotactic activity was observed when monoepoxides of linoleic or linolenic acid, diepoxides of linoleic acid or triepoxides of linolenic acid were used. Mono-, di- and triepoxides of polyunsaturated fatty acids were synthesized with meta-chloroperbenzoic acid, separated by TLC and HPLC and identified by GC/MS.

Does cytosolic free Ca2+ signal neutrophil chemotaxis in response to formylated chemotactic peptide?

Cytosolic free Ca2+ concentration was measured and imaged in human neutrophils moving towards a source of formylated peptide in a micropipette held close to the cells. Under these conditions, neutrophils changed shape and displayed chemotaxis without significant or persistent global or localised elevations in cytosolic free Ca2+. A rear-to-front persistent Ca2+ gradient of less than 0.5 nM/micron was present in the migrating neutrophils, until they reached the zone of higher peptide concentration, when an abrupt rise in cytosolic free Ca2+ concentration was triggered and chemotaxis stopped. Small localised rises in cytosolic free Ca2+, which were occasionally observed during neutrophil manoeuvring, were attributed to the effect of local deformation of the neutrophil membrane, since deformation of the membrane with a blunt micropipette caused similar Ca2+ changes. These data suggest that neutrophil chemotaxis towards a source of formylated peptide occurs without significant changes in Ca2+ signalling.

Interaction of Fibronectin (FN) Cell Binding Fragments and Interleukin-8 (IL-8) in Regulating Neutrophil Chemotaxis

This study investigated the possible interaction of FN fragments in regulating IL-8-mediated neutrophil chemotaxis in vitro using Neuroprobe microchambers. Human neutrophil suspensions were incubated with purified FN fragments or an RGD-containing peptide and allowed to migrate in response to chemotactically active concentrations of human recombinant IL-8. The 120-kD fragment of FN containing the RGD sequence or an RGD peptide (GRGDSP) inhibited IL-8-mediated neutrophil chemotaxis; however, these RGD peptides did not inhibit neutrophil chemotaxis in response to other chemotactic agents. Furthermore, FN fragments not containing the RGD sequence had no effect on IL-8-mediated chemotaxis. These data suggest that directed migration of neutrophils in response to IL-8 is inhibited in the presence of cell-binding fragments of FN and may represent a local mechanism for terminating neutrophil migration at areas of tissue injury.

Neutrophil chemotaxis in response to TGF-beta isoforms (TGF-beta 1, TGF-beta 2, TGF-beta 3) is mediated by fibronectin.

TGF-beta isoforms regulate numerous cellular functions including cell growth and differentiation, the cellular synthesis and secretion of extracellular matrix proteins, such as fibronectin (Fn), and the immune response. We have previously shown that TGF-beta 1 is the most potent chemoattractant described for human peripheral blood neutrophils (PMNs), suggesting that TGF-beta s may play a role in the recruitment of PMNs during the initial phase of the inflammatory response. In our current studies, we demonstrate that the maximal chemotactic response was attained near 40 fM for all mammalian TGF-beta isoforms. However, there was a statistically significant difference in migratory distance of the PMNs: TGF-beta 2 (556 microM) > TGF-beta 3 (463 microM) > TGF-beta 1 (380 microM) (beta 2: beta 3, p < or = 0.010; beta 3: beta 1, p < or = 0.04; beta 2: beta 1, p < or = 0.0012). A mAb to the cell binding domain (CBD) of Fn inhibited the chemotactic response to TGF-beta 1 and TGF-beta 3 by 63% and to TGF-beta 2 by 70%, whereas the response to FMLP, a classic chemoattractant, was only inhibited by 18%. In contrast, a mAb to a C-terminal epitope of Fn did not retard migration (< 1.5%). The Arg-gly-Asp-ser tetrapeptide inhibited chemotaxis by approximately the same extent as the anti-CBD (52 to 83%). Furthermore, a mAb against the VLA-5 integrin (VLA-5; Fn receptor) also inhibited TGF-beta-induced chemotaxis. These results indicate that chemotaxis of PMNs in response to TGF-beta isoforms is mediated by the interaction of the Arg-gly-Asp-ser sequence in the CBD of Fn with an integrin on the PMN cell surface, primarily the VLA-5 integrin. TGF-beta isoforms also elicited the release of cellular Fn from PMNs; we observed a 2.3-fold increase in Fn (389 to 401 ng/ml) in the supernatants of TGF-beta-stimulated PMNs compared with unstimulated cells (173.6 ng/ml). The concentration of TGF-beta required to cause maximal release of Fn from PMNs (4000 fM) is a concentration at which TGF-beta is no longer chemotactic, suggesting that PMNs only use Fn that is constitutively expressed for migration. At higher concentrations of TGF-beta, the Fn released may accumulate basal to the cell, ultimately retarding cellular migration and modulating the chemotactic response.

Decreased Adherence, Chemotaxis and Phagocytic Activities of Neutrophils from Preterm Neonates

Using microanalytic assays various phagocytic functions of separated neutrophils from preterm neonates (mean birthweight 1,506 g, n = 13) were simultaneously studied. Adherence of neutrophils to nylon fibre was decreased in cells from preterm infants (77.1 +/- 3.1%) when compared with adult controls (86.9 +/- 2.1%, mean +/- 1 SD, p less than 0.05). In addition neutrophil chemotaxis in response to zymosan activated serum was reduced in preterm neonates (131.9 +/- 19.7, adults 166.6 +/- 11.1, p less than 0.001); directed migration towards Formyl-Methionyl-Leucyl-Phenylalanine was also decreased (preterm neonates 93.4 +/- 15, adults 111.1 +/- 16.8, p less than 0.05). Preterm infants had a higher percentage of slow moving neutrophils when compared with adults (p less than 0.001). Phagocytosis of Candida albicans was reduced in neutrophils from preterm neonates (phagocytic index: preterm neonates 41.4 +/- 12.7, adults 83 +/- 7.2). Adult neutrophils ingested more Candida per cell (p less than 0.001). Chemiluminescence, exocytosis of elastase and lactoferrin during uptake of opsonized zymosan was also reduced in neutrophils from preterm neonates. However, random migration, phagocytosis of Staphylococcus aureus and production of O2- in response to Phorbol myristate acetate or opsonized zymosan were identical in cells from either source. We conclude, that these abnormalities of neutrophils could predispose the preterm infant to serious and often overwhelming bacterial and fungal infections.

Inhibition of formylmethionyl-leucyl-phenylalanine-stimulated neutrophil chemiluminescence by human immunoglobulin A paraproteins.

Immunoglobulin A (IgA) paraproteins from patients with myeloma have been shown to inhibit human neutrophil chemotaxis to C5a, casein, and chemotactic factors produced by Escherichia coli. This study demonstrates that these paraproteins also inhibit neutrophil chemotaxis in response to the synthetic peptide formylmethionyl-leucyl-phenylalanine (f-MLP). Furthermore, the neutrophil chemiluminescence response stimulated by f-MLP was markedly suppressed by the presence of IgA paraprotein. Maximal inhibition of chemiluminescence was observed when the paraprotein was present during the chemiluminescence response. The inhibitory activity was substantially reduced by removal of the Fc region of IgA or by conversion of polymeric IgA to monomeric IgA by limited reduction and alkylation. Additional experiments showed that these IgA paraproteins inhibited C5a but not phorbol myristate acetate-stimulated chemiluminescence. These observations are constant with the hypothesis that polymeric forms of IgA bind to human neutrophils and interfere with the binding of chemotactic factor to its receptor or the consequent receptor-mediated oxidative burst or both.

Neutrophil chemotaxis in response to surface-bound C5a

Many biological functions of neutrophils can be initiated by surface-bound stimuli. This report describes experiments examining the role of surface-bound C5a on neutrophil chemotaxis. Cellulose-nitrate filters were preincubated with various concentrations of highly purified human C5a. After washing the filters, incubating for an additional hour and washing again, the filters were used in a standard two-filter chemotaxis assay with human neutrophils and no additional stimulus. The bound C5a induced an effective directional migration of the neutrophils. Establishment of a chemotactic gradient within the filters was suggested by the absence of directional migration when the pretreated C5a filters were inverted before the chemotactic assay. The presence of the chemotactic gradient was further shown by use of autoradiography and 125I-C5a. These studies suggest that interaction of neutrophils with surface-bound C5a can promote the movement of the neutrophil in the direction of increased concentrations of the bound ligand.