Neutrophil Chemoattractant CXCL1 Research Articles

WFDC1/ps20: A host factor that influences the neutrophil response to murine hepatitis virus (MHV) 1 infection

The whey acidic protein family member, WFDC1/ps20 is a permissivity factor in HIV infection. Herein we describe a contrasting role for ps20 in limiting MHV-1 infection. Intranasal MHV-1 infection produces a respiratory infection in mice. Using ps20 knockout mice we provide evidence that intranasal MHV-1 infection results in increased lung viral titers in ps20−/− compared to ps20+/+ mice. Accompanying MHV-1 infection we observe an increase in the number of neutrophils infiltrating the BAL and an increase in the percentage of neutrophils in the lung draining lymph nodes of ps20−/− compared with ps20+/+ mice. Gene expression levels for the neutrophil chemoattractants CXCL1 and CXCL2 are elevated in the lungs of ps20−/− mice post-MHV-1 infection. Characterization of the immune cell profile in naïve ps20−/− mice revealed an increase in circulating neutrophils compared to ps20+/+ mice. No notable differences in other immune cell profiles were observed between the ps20+/+ and ps20−/− mice. Accordingly, we examined MHV-1 infection of neutrophils and provide evidence that neutrophils isolated from ps20−/− mice are more susceptible to MHV-1 infection than neutrophils isolated from ps20+/+ mice. These data suggest roles for ps20 in regulating expression of neutrophil-specific chemotactic factors, thereby potentially modulating neutrophil migration, and in modulating neutrophil susceptibility to MHV-1 infection.

Human cardiac fibroblasts express ICAM-1, E-selectin and CXC chemokines in response to proinflammatory cytokine stimulation

Neutrophil attraction and adhesion to endothelial cells occurs via well defined mechanisms, yet the ability of other cell types to express neutrophil-binding adhesion molecules is not well studied. Cardiac fibroblasts (CF) are a key cell type involved in repair of the infarcted myocardium, a scenario in which neutrophil recruitment is perceived to be detrimental. Here we determined the effects of proinflammatory cytokines on expression of neutrophil-binding adhesion molecules and neutrophil-attracting chemokines in CF cultured from multiple patients, and explored the underlying regulatory mechanisms. An adhesion molecule focused RT-PCR array identified 5 transcripts that were increased markedly in human CF treated with the proinflammatory cytokine interleukin-1 (IL-1, 10 ng/ml, 6 h); including intercellular cell adhesion molecule (ICAM-1) and E-selectin. Real-time RT-PCR verified the array data and immunoblotting confirmed cytokine-induced ICAM-1 and E-selectin protein expression. Treatment with a panel of pharmacological inhibitors identified the NF-κB pathway as mediating IL-1-induced ICAM-1 and E-selectin mRNA and protein expression. Additionally, E-selectin expression in human CF was markedly potentiated by the JNK inhibitor SP600125, but this was not observed when a more selective inhibitor ((L)-JNKI-1) was used, or in human vascular endothelial cells. IL-1 also stimulated CF to secrete the neutrophil chemoattractant CXCL8 via a p38- and NF-κB-dependent mechanism, as well as inducing CXCL1, CXCL2 and CXCL5 mRNA expression. In conclusion, human CF express neutrophil-binding adhesion molecules and neutrophil chemoattractants in response to proinflammatory cytokines suggesting that, in addition to EC, CF may play an important role in regulating neutrophil recruitment into the infarcted myocardium.

Human Neutrophil Clearance of Bacterial Pathogens Triggers Anti-Microbial γδ T Cell Responses in Early Infection

Human blood Vγ9/Vδ2 T cells, monocytes and neutrophils share a responsiveness toward inflammatory chemokines and are rapidly recruited to sites of infection. Studying their interaction in vitro and relating these findings to in vivo observations in patients may therefore provide crucial insight into inflammatory events. Our present data demonstrate that Vγ9/Vδ2 T cells provide potent survival signals resulting in neutrophil activation and the release of the neutrophil chemoattractant CXCL8 (IL-8). In turn, Vγ9/Vδ2 T cells readily respond to neutrophils harboring phagocytosed bacteria, as evidenced by expression of CD69, interferon (IFN)-γ and tumor necrosis factor (TNF)-α. This response is dependent on the ability of these bacteria to produce the microbial metabolite (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), requires cell-cell contact of Vγ9/Vδ2 T cells with accessory monocytes through lymphocyte function-associated antigen-1 (LFA-1), and results in a TNF-α dependent proliferation of Vγ9/Vδ2 T cells. The antibiotic fosmidomycin, which targets the HMB-PP biosynthesis pathway, not only has a direct antibacterial effect on most HMB-PP producing bacteria but also possesses rapid anti-inflammatory properties by inhibiting γδ T cell responses in vitro. Patients with acute peritoneal-dialysis (PD)-associated bacterial peritonitis – characterized by an excessive influx of neutrophils and monocytes into the peritoneal cavity – show a selective activation of local Vγ9/Vδ2 T cells by HMB-PP producing but not by HMB-PP deficient bacterial pathogens. The γδ T cell-driven perpetuation of inflammatory responses during acute peritonitis is associated with elevated peritoneal levels of γδ T cells and TNF-α and detrimental clinical outcomes in infections caused by HMB-PP positive microorganisms. Taken together, our findings indicate a direct link between invading pathogens, neutrophils, monocytes and microbe-responsive γδ T cells in early infection and suggest novel diagnostic and therapeutic approaches.

Steroid-Insensitive ERK1/2 Activity Drives CXCL8 Synthesis and Neutrophilia by Airway Smooth Muscle

Severe or refractory asthma affects 5 to 15% of all patients with asthma, but is responsible for more than half of the health burden associated with the disease. Severe asthma is characterized by a dramatic increase in smooth muscle and airway inflammation. Although glucocorticoids are the mainstay of treatment in asthma, they are unable to fully control the disease in individuals with severe asthma. We found that airway smooth muscle cells (ASMCs) from individuals with severe asthma showed elevated activities of the ERK1/ERK2 and p38 MAPK pathways despite treatment with oral and inhaled glucocorticoids, which increased the expression of DUSP1, a phosphatase shown to limit p38 MAPK activity. In ex vivo ASMCs, TNF-α but not IL-17A induced expression of the neutrophil chemoattractant CXCL8. Moreover, TNF-α led to up-regulation of the ERK1/ERK2 and p38 MAPKs pathways, with only the latter being sensitive to pretreatment with the glucocorticoid dexamethasone. In contrast to epithelial and endothelial cells, TNF-α-stimulated CXCL8 synthesis was dependent on ERK1/ERK2 but not on p38 MAPK. Moreover, suppressing ERK1/ERK2 activation prevented neutrophil recruitment by ASMCs, whereas suppressing p38 MAPK activity had no impact. Taken together, these results highlight the ERK1/ERK2 MAPK cascade as a novel and attractive target in severe asthma because the activation of this pathway is insensitive to the action of glucocorticoids and is involved in neutrophil recruitment, contributing the to inflammation seen in the disease.

CXCL1 and its receptor, CXCR2, mediate murine sickle cell vaso‐occlusion during hemolytic transfusion reactions

Hemolytic transfusion reactions (HTR) can produce acute vaso-occlusive crisis (VOC), a major cause of morbidity and mortality in sickle cell disease (SCD) patients; however, the mechanisms remain unknown. We established a model of alloimmune IgG-mediated HTR using a well-characterized humanized murine model of SCD. In this model, HTR induced acute VOC, resulting in shortened survival of the SCD mice (by 3 h, P<0.05, log-rank test). Acute VOC was associated with elevated circulating inflammatory chemokine levels, including the neutrophil chemoattractant CXCL1, which was strikingly elevated (139 ± 35 vs 43 ± 10 pg/ml, P<0.05). Recombinant CXCL1 administration corresponding to the HTR-elicited endogenous CXCL1 levels induced acute VOC in non-transfused SCD mice, characterized by leukocyte recruitment in venules (2571 ± 193 vs 1464 ± 194 adherent leukocytes per mm2, P<0.01), marked interactions with circulating red blood cells, reduced venular flow rates (430 ± 34 vs 310 ± 36 nL/s, P<0.05), and shortened survival (by 4 h, P<0.001, log-rank test). In contrast, blocking its receptor, CXCR2, prevented HTR-elicited acute VOC and prolonged the survival of SCD mice (by 6 h, P<0.01, log-rank test). Thus, CXCL1 is a key inflammatory mediator of acute VOC in SCD mice and targeted inhibition of CXCL1 and/or CXCR2 may provide new approaches for treating acute VOC in SCD patients.

CXCL1 and its receptor, CXCR2, mediate murine sickle cell vaso-occlusion during hemolytic transfusion reactions

Hemolytic transfusion reactions (HTRs) can produce serious and potentially life-threatening complications in sickle cell disease (SCD) patients; however, the mechanisms underlying these complications remain undetermined. We established a model of alloimmune, IgG-mediated HTRs in a well-characterized humanized murine model of SCD. HTRs induced acute vaso-occlusive crisis (VOC), resulting in shortened survival of SCD mice. Acute VOC was associated with elevated circulating inflammatory chemokine levels, including striking elevation of the levels of the neutrophil chemoattractant CXCL1. Recombinant CXCL1 administration was sufficient to induce acute VOC in SCD mice, characterized by leukocyte recruitment in venules, capture of circulating red blood cells, reduction of venular flow, and shortened survival. In contrast, blockade of the CXCL1 receptor, CXCR2, prevented HTR-elicited acute VOC and prolonged survival in SCD mice. These results indicate that CXCL1 is a key inflammatory mediator of acute VOC in SCD mice. Targeted inhibition of CXCL1 and/or CXCR2 may therefore represent a new therapeutic approach for acute VOC in SCD patients.

Novel role of regulatory T cells in limiting early neutrophil responses in skin

It is clear that CD4(+) CD25(+) Foxp3(+) regulatory T (Treg) cells inhibit chronic inflammatory responses as well as adaptive immune responses. Among the CD4(+) T-cell population in the skin, at least one-fifth express Foxp3. As the skin is constantly exposed to antigenic challenge and is a common site of vaccination, understanding the role of these skin-resident Treg cells is important. Although the suppressive effect of Treg cells on T cells is well documented, less is known about the types of innate immune cells influenced by Treg cells and whether the Treg cells suppress acute innate immune responses in vivo. To address this we used a mouse melanoma cell line expressing Fas ligand (B16FasL), which induces an inflammatory response following subcutaneous injection of mice. We demonstrate that Treg cells limit this response by inhibiting neutrophil accumulation and survival within hours of tumour cell inoculation. This effect, which was associated with decreased expression of the neutrophil chemoattractants CXCL1 and CXCL2, promoted survival of the inoculated tumour cells. Overall, these data imply that Treg cells in the skin are rapidly mobilized and that this activity serves to limit the amplification of inflammatory responses at this site.

T-cell regulation of neutrophil infiltrate at the early stages of a murine colitis model

T-cells are a main target for antiinflammatory drugs in inflammatory bowel disease. As the innate immune system is also implicated in the pathogenesis of these diseases, T-cell suppressors may not only inhibit T-cell-dependent production of proinflammatory mediators but also affect innate immune cell function. Specifically, these drugs may impair innate immune cell recruitment and activation through inhibition of T-cells or act independent of T-cell modulation. We explored the extent of immune modulation by the T-cell inhibitor tacrolimus in a murine colitis model. We assessed the effects of tacrolimus on trinitro-benzene sulphonic acid (TNBS) colitis in wildtype and Rag2-deficient mice. The severity of colitis was assessed by means of histological scores and weight loss. We further characterized the inflammation using immunohistochemistry and by analysis of isolated intestinal leukocytes at various stages of disease. Tacrolimus-treated wildtype mice were less sensitive to colitis and had fewer activated T-cells. Inhibition of T-cell function was associated with strongly diminished recruitment of infiltrating neutrophils in the colon at the early stages of this model. In agreement, immunohistochemistry demonstrated that tacrolimus inhibited production of the neutrophil chemoattractants CXCL1 and CXCL2. Rag2-deficient mice displayed an enhanced baseline level of lamina propria neutrophils that was moderately increased in TNBS colitis and remained unaffected by tacrolimus. Both the innate and the adaptive mucosal immune system contribute to TNBS colitis. Tacrolimus suppresses colitis directly through inhibition of T-cell activation and by suppression of T-cell-mediated recruitment of neutrophils.

Acute Pulmonary Lipopolysaccharide Tolerance Decreases TNF-α without Reducing Neutrophil Recruitment

Pulmonary LPS exposure plays a key role in exacerbation of lung diseases such as chronic obstructive pulmonary disease and asthma. However, little is known about the effects of repeated LPS exposure in the lung microenvironment. We have developed a novel murine model of pulmonary LPS tolerance induced by intratracheal (i.t.) administration of LPS. First, we show that pulmonary LPS exposure does not induce whole-body refractoriness to systemic LPS, because i.t. administration followed by i.p. administration did not decrease plasma TNF-alpha. However, a local refractory state can be induced with two i.t. LPS exposures. Pulmonary LPS tolerance was induced by i.t. administration of 100 ng LPS at time 0 and 48 h. Nontolerant mice received PBS at time 0 and LPS at 48 h. Bronchoalveolar lavage levels of TNF-alpha were significantly attenuated in tolerant mice vs nontolerant mice (1597 pg/ml vs 7261 pg/ml). TNF-alpha mRNA was significantly reduced in bronchoalveolar lavage cells (5-fold) and lung tissue (10-fold). No reduction was seen in neutrophil numbers in the bronchoalveolar lavage fluid, myeloperoxidase activity, or expression of neutrophil chemoattractants CXCL1 and CXCL2, reflecting the specificity of the response. The reduction in TNF-alpha was accompanied by a significant increase in soluble receptors, TNF-SRI (159 pg/ml vs 206 pg/ml) and TNF-SRII (1366 pg/m vs 2695 pg/ml). In conclusion, pulmonary LPS tolerance results in a specific reduction in TNF-alpha expression, while the neutrophilic response is unaffected. This response may be a mechanism to limit tissue damage by reducing TNF-alpha levels, while still maintaining the antimicrobial capacity of the lung.

Toll-Like Receptor Prestimulation Increases Phagocytosis ofEscherichia coliDH5α andEscherichia coliK1 Strains by Murine Microglial Cells

Meningitis and meningoencephalitis caused by Escherichia coli are associated with high rates of mortality. When an infection occurs, Toll-like receptors (TLRs) expressed by microglial cells can recognize pathogen-associated molecular patterns and activate multiple steps in the inflammatory response that coordinate the brain's local defense, such as phagocytosis of invading pathogens. An upregulation of the phagocytic ability of reactive microglia could improve the host defense in immunocompromised patients against pathogens such as E. coli. Here, murine microglial cultures were stimulated with the TLR agonists Pam(3)CSK(4) (TLR1/TLR2), lipopolysaccharide (TLR4), and CpG oligodeoxynucleotide (TLR9) for 24 h. Upon stimulation, levels of tumor necrosis factor alpha and the neutrophil chemoattractant CXCL1 were increased, indicating microglial activation. Phagocytic activity was studied after adding either E. coli DH5alpha or E. coli K1 strains. After 60 and 90 min of bacterial exposure, the number of ingested bacteria was significantly higher in cells prestimulated with TLR agonists than in unstimulated controls (P < 0.01). Addition of cytochalasin D, an inhibitor of actin polymerization, blocked >90% of phagocytosis. We also analyzed the ability of microglia to kill the ingested E. coli strains. Intracellularly surviving bacteria were quantified at different time points (90, 150, 240, and 360 min) after 90 min of phagocytosis. The number of bacteria killed intracellularly after 6 h was higher in cells primed with the different TLR agonists than in unstimulated microglia. Our data suggest that microglial stimulation by the TLR system can increase bacterial phagocytosis and killing. This approach could improve central nervous system resistance to infections in immunocompromised patients.

Anthrax Toxins Inhibit Neutrophil Signaling Pathways in Brain Endothelium and Contribute to the Pathogenesis of Meningitis

BackgroundAnthrax meningitis is the main neurological complication of systemic infection with Bacillus anthracis approaching 100% mortality. The presence of bacilli in brain autopsies indicates that vegetative bacteria are able to breach the blood-brain barrier (BBB). The BBB represents not only a physical barrier but has been shown to play an active role in initiating a specific innate immune response that recruits neutrophils to the site of infection. Currently, the basic pathogenic mechanisms by which B. anthracis penetrates the BBB and causes anthrax meningitis are poorly understood.Methodology/Principal FindingsUsing an in vitro BBB model, we show for the first time that B. anthracis efficiently invades human brain microvascular endothelial cells (hBMEC), the single cell layer that comprises the BBB. Furthermore, transcriptional profiling of hBMEC during infection with B. anthracis revealed downregulation of 270 (87%) genes, specifically key neutrophil chemoattractants IL-8, CXCL1 (Groα) and CXCL2 (Groβ), thereby strongly contrasting hBMEC responses observed with other meningeal pathogens. Further studies using specific anthrax toxin-mutants, quantitative RT-PCR, ELISA and in vivo assays indicated that anthrax toxins actively suppress chemokine production and neutrophil recruitment during infection, allowing unrestricted proliferation and dissemination of the bacteria. Finally, mice challenged with B. anthracis Sterne, but not the toxin-deficient strain, developed meningitis.Conclusions/SignificanceThese results suggest a significant role for anthrax toxins in thwarting the BBB innate defense response promoting penetration of bacteria into the central nervous system. Furthermore, establishment of a mouse model for anthrax meningitis will aid in our understanding of disease pathogenesis and development of more effective treatment strategies.

Targeted Inflammation During Oncolytic Virus Therapy Severely Compromises Tumor Blood Flow

Oncolytic viruses (OVs) are selected or designed to eliminate malignancies by direct infection and lysis of cancer cells. In contrast to this concept of direct tumor lysis by viral infection, we observed that a significant portion of the in vivo tumor killing activity of two OVs, vesicular stomatitis virus (VSV) and vaccinia virus is caused by indirect killing of uninfected tumor cells. Shortly after administering the oncolytic virus we observed limited virus infection, coincident with a loss of blood flow to the interior of the tumor that correlated with induction of apoptosis in tumor cells. Transcript profiling of tumors showed that virus infection resulted in a dramatic transcriptional activation of pro-inflammatory genes including the neutrophil chemoattractants CXCL1 and CXCL5. Immunohistochemical examination of infected tumors revealed infiltration by neutrophils correlating with chemokine induction. Depletion of neutrophils in animals prior to VSV administration eliminated uninfected tumor cell apoptosis and permitted more extensive replication and spreading of the virus throughout the tumor. Taken all together, these results indicate that targeted recruitment of neutrophils to infected tumor beds enhances the killing of malignant cells. We propose that activation of inflammatory cells can be used for enhancing the effectiveness of oncolytic virus therapeutics, and that this approach should influence the planning of therapeutic doses.

Chemokine Signatures in the Skin Disorders of Lyme Borreliosis in Europe: Predominance of CXCL9 and CXCL10 in Erythema Migrans and Acrodermatitis and CXCL13 in Lymphocytoma

The three skin disorders of Lyme borreliosis in Europe include erythema migrans, an acute, self-limited lesion; borrelial lymphocytoma, a subacute lesion; and acrodermatitis chronica atrophicans, a chronic lesion. Using quantitative reverse transcription-PCR, we determined mRNA expression of selected chemokines, cytokines, and leukocyte markers in skin samples from 100 patients with erythema migrans, borrelial lymphocytoma, or acrodermatitis chronica atrophicans and from 25 control subjects. Chemokine patterns in lesional skin in each of the three skin disorders included low but significant mRNA levels of the neutrophil chemoattractant CXCL1 and the dendritic cell chemoattractant CCL20 and intermediate levels of the macrophage chemoattractant CCL2. Erythema migrans and particularly acrodermatitis lesions had high mRNA expression of the T-cell-active chemokines CXCL9 and CXCL10 and low levels of the B-cell-active chemokine CXCL13, whereas lymphocytoma lesions had high levels of CXCL13 and lower levels of CXCL9 and CXCL10. This pattern of chemokine expression was consistent with leukocyte marker mRNA in lesional skin. Moreover, using immunohistologic methods, CD3(+) T cells and CXCL9 were visualized in erythema migrans and acrodermatitis lesions, and CD20(+) B cells and CXCL13 were seen in lymphocytoma lesions. Thus, erythema migrans and acrodermatitis chronica atrophicans have high levels of the T-cell-active chemokines CXCL9 and CXCL10, whereas borrelial lymphocytoma has high levels of the B-cell-active chemokine CXCL13.

Genetic profiling of dendritic cells exposed to live‐ or ultraviolet‐irradiated Chlamydia muridarum reveals marked differences in CXC chemokine profiles

Chlamydia trachomatis is a major cause of sexually transmitted disease worldwide for which an effective vaccine is being actively pursued. Current vaccine efforts will be aided by elucidating the interaction between Chlamydia and dendritic cells (DCs). Protective immunity appears to develop slowly following natural infection in humans, and early vaccine trials using inactivated C. trachomatis resulted in partial, short-lived protection with possible enhanced inflammatory pathology during re-infection. Thus, immunity following natural infection with live chlamydia may differ fundamentally from immune responses induced by immunization with inactivated chlamydia. We explored this conjecture by studying the response of DCs exposed to either viable or inactivated [ultraviolet (UV) -irradiated] chlamydia elementary bodies (EBs; designated as Live-EB and UV-EB, respectively) using Affymetrix GeneChip microarrays. Thirty-one immunologically characterized genes were differentially expressed by DCs following exposure to Live-EB or UV-EB, including two glutamic acid-leucine-arginine cysteine-X-cysteine (ELR CXC) neutrophil chemoattractant chemokines, Cxcl1 (KC), and Cxcl2 (MIP-2). Up-regulation of these genes by Live-EB as compared to UV-EB was verified by quantitative reverse transcription-polymerase chain reaction and increased chemokine secretion was confirmed by enzyme-linked immunosorbent assay both in vitro and in vivo. Immunofluorescence and fluorescence-activated cell sorter analysis of chlamydia-infected lung tissue confirmed that Live-EB but not UV-EB induced significant DC and neutrophil infiltration during infection. These observations demonstrate that the development of an antichlamydial immune response is dramatically influenced by chlamydial viability. This has implications as to why early inactivated chlamydial vaccines were ineffective and suggests that new vaccine design efforts may benefit from in vitro DC screening for ELR chemokine expression profiles.

Coexpression and interaction of CXCL10 and CD26 in mesenchymal cells by synergising inflammatory cytokines: CXCL8 and CXCL10 are discriminative markers for autoimmune arthropathies

Leukocyte infiltration during acute and chronic inflammation is regulated by exogenous and endogenous factors, including cytokines, chemokines and proteases. Stimulation of fibroblasts and human microvascular endothelial cells with the inflammatory cytokines interleukin-1β (IL-1β) or tumour necrosis factor alpha (TNF-α) combined with either interferon-α (IFN-α), IFN-β or IFN-γ resulted in a synergistic induction of the CXC chemokine CXCL10, but not of the neutrophil chemoattractant CXCL8. In contrast, simultaneous stimulation with different IFN types did not result in a synergistic CXCL10 protein induction. Purification of natural CXCL10 from the conditioned medium of fibroblasts led to the isolation of CD26/dipeptidyl peptidase IV-processed CXCL10 missing two NH2-terminal residues. In contrast to intact CXCL10, NH2-terminally truncated CXCL10(3–77) did not induce extracellular signal-regulated kinase 1/2 or Akt/protein kinase B phosphorylation in CXC chemokine receptor 3-transfected cells. Together with the expression of CXCL10, the expression of membrane-bound CD26/dipeptidyl peptidase IV was also upregulated in fibroblasts by IFN-γ, by IFN-γ plus IL-1β or by IFN-γ plus TNF-α. This provides a negative feedback for CXCL10-dependent chemotaxis of activated T cells and natural killer cells. Since TNF-α and IL-1β are implicated in arthritis, synovial concentrations of CXCL8 and CXCL10 were compared in patients suffering from crystal arthritis, ankylosing spondylitis, psoriatic arthritis and rheumatoid arthritis. All three groups of autoimmune arthritis patients (ankylosing spondylitis, psoriatic arthritis and rheumatoid arthritis) had significantly increased synovial CXCL10 levels compared with crystal arthritis patients. In contrast, compared with crystal arthritis, only rheumatoid arthritis patients, and not ankylosing spondylitis or psoriatic arthritis patients, had significantly higher synovial CXCL8 concentrations. Synovial concentrations of the neutrophil chemoattractant CXCL8 may therefore be useful to discriminate between autoimmune arthritis types.

Valproic acid induces expression of neutrophil chemoattractants of the CXC chemokine family in endothelial cells

The branched-chain fatty acid valproate (valproic acid; VPA) displays antitumoral properties by blocking tumor growth, progression and invasion. Recent data have shown that VPA reduces the angiogenic activity of endothelial cells. The object of this study was to investigate whether endothelial modulation might also influence the level of chemotactic mediators. Endothelial cells were isolated from human umbilical cord veins (HUVEC) and treated with VPA-concentrations ranging from 0.125 mM to 1 mM. The mRNA level of CXC-chemokines was investigated by reverse transcriptase-polymerase chain reaction. The proliferative activity of HUVEC was measured as well. VPA evoked a striking increase in the neutrophil chemoattractants CXCL1, CXCL3, CXCL4, CXCL5 and a moderate increase in CXCL6 with maximal effects after a 3-day incubation period. Other CXC-chemokines and CXC-receptors remained unaffected. HUVEC growth was diminished time- and dose-dependently by VPA. We conclude that VPA treatment leads to alterations in the chemokine expression profile of endothelial cells. This might allow more neutrophils to reach the tumor area and trigger cytolysis.