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Biomedical subjects

J Hakim

Publications and source records attributed to J Hakim.

At least 37 records · Page 2Linked to original sources
PubMed1998

Interleukin-8 production by polymorphonuclear neutrophils in patients with rapidly progressive periodontitis: an amplifying loop of polymorphonuclear neutrophil activation.

Polymorphonuclear neutrophils (PMN) are the most abundant immune cells in inflammatory gingival sites of patients with early onset periodontitis, localized juvenile periodontitis, and rapidly progressive periodontitis (RPP). In the latter, the large number of PMN in connective tissue may explain the marked gingival destruction. Because interleukin-8 (IL-8) is a potent PMN chemoattractant, we evaluated circulating levels and gingival mRNA expression of IL-8. We found high IL-8 plasma levels as well as strong IL-8 mRNA expression in both epithelial and connective gingival cells from patients with RPP. Moreover, the gingival PMN themselves contained IL-8 mRNA, suggesting an autoamplification of PMN recruitment and activation in the gingiva. We also measured the expression of adhesion molecules at the PMN surface as well as the oxidative burst in whole blood from 14 patients with RPP, using flow cytometry to avoid irrelevant stimulations and to analyze single cells. In RPP patients, resting PMN showed reduced L-selectin, Lewis x, and sialyl Lewis x antigen expression as well as increased H2O2 production. These modifications of PMN adhesion molecule expression, together with their increased basal oxidative burst and excessive IL-8 production, may contribute to the noxious inflammatory reaction, which may in turn be autopotentiated by PMN production of IL-8. In addition, PMN showed a lack of increased response (H2O2 production) to formyl peptides after ex vivo priming with IL-8, possibly owing to IL-8 desensitization that may be involved in the increased susceptibility of RPP patients to infection. After appropriate treatment of RPP, the reduction in inflammation was associated with a return to control levels of both plasma IL-8 and PMN functions, suggesting that these features are linked.

Adult↗
PubMed1998

Absence of regulation of human polymorphonuclear oxidative burst by interleukin-10, interleukin-4, interleukin-13 and transforming growth factor-beta in whole blood.

Cytokines such as IL-10, IL-4, IL-13 and TGF-beta play a major role in the regulation of immune responses and are considered as anti-inflammatory agents mainly due to their actions on monocytes. These cytokines are also known to participate in the regulation of PMN activities. However, few and contradictory results have been reported on their direct and priming effects on the PMN oxidative burst, which is essential for killing bacteria. We used a flow cytometry method to study the effects of these cytokines on the PMN oxidative burst; we also used whole blood to avoid PMN activation related to isolation procedures and in order to simulate the in vivo situation more closely. None of the cytokines tested had direct or priming effects on PMN H2O2 production. We also show for the first time that these cytokines do not modulate TNF priming of the PMN oxidative burst in response to N-formyl peptides (fMLP). These results show that the anti-bacterial activity of PMN, in terms of the PMN respiratory burst, is not down regulated by these anti-inflammatory cytokines in whole blood.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1997

Intracellular pool of vascular endothelial growth factor in human neutrophils.

Vascular endothelial growth factor (VEGF ), an endothelial cell mitogen, is a potent angiogenic factor produced by several cell types. Whether human neutrophils are potential producers of VEGF has not yet been described. The present work shows that phorbol-12-myristate 13-acetate (PMA), fMet-Leu-Phe, and tumor necrosis factor-alpha (TNF-alpha) triggered a time-dependent secretion of VEGF by human neutrophils. Cells incubated with 50 ng/mL of PMA released significant amounts of VEGF after 15 minutes. Because the extracellular content of VEGF in human neutrophils supernatants remained constant over a period of 2 to 24 hours and because PMA is a potent inducer of human neutrophil degranulation, the PMA-induced secretion of VEGF may be due to a pre-existing intracellular pool of this molecule. This hypothesis was reinforced by the absence of cycloheximide effect on the PMA-induced secretion of VEGF. The existence of an intracellular pool of VEGF was confirmed by measuring the intracellular content of VEGF in resting neutrophils. A dosedependent inhibition of PMA-induced VEGF secretion was observed when the cells were incubated in the presence of pentoxifylline, a methylxanthine known to inhibit neutrophil degranulation. To confirm the implication of neutrophil degranulation in VEGF release, the effects of two inducers of physiologic degranulation, fMet-Leu-Phe and TNF-alpha, were determined. Both agonists induced a release of VEGF in the absence of cytochalasin B, confirming the involvement of neutrophil degranulation and suggesting the intracellular localization of VEGF in the specific granule fraction. In addition, the kinetics of fMet-Leu-Phe- and TNF-alpha-induced secretion of lactoferrin were similar to those of VEGF release induced by these two both agonists. The subcellular fractionation of human neutrophils showed a granule-specific distribution of the intracellular pool of VEGF in resting neutrophils. The finding that human neutrophils contain an intracellular pool of VEGF, secreted in the extracellular space under PMA-, fMet-Leu-Phe-, and TNF-alpha-induced degranulation, suggests a role for human neutrophils as cellular effectors of physiologic as well as pathologic angiogenesis.

Cells, Cultured↗
PubMed1997

Phosphorylation of the respiratory burst oxidase subunit p67(phox) during human neutrophil activation. Regulation by protein kinase C-dependent and independent pathways.

The respiratory burst oxidase of phagocytes and B lymphocytes catalyzes the reduction of oxygen to superoxide anion (O-2) at the expense of NADPH. This multicomponent enzyme is dormant in resting cells but is activated on exposure to an appropriate stimulus. The phosphorylation-dependent mechanisms regulating the activation of the respiratory burst oxidase are unclear, particularly the phosphorylation status of the cytosolic component p67(phox). In this study, we found that activation of human neutrophils with formyl-methionyl-leucyl-phenylalanine (fMLP), a chemotactic peptide, or phorbol myristate acetate (PMA), a stimulator of protein kinase C (PKC), resulted in the phosphorylation of p67(phox). Using an anti-p67(phox) antibody or an anti-p47(phox) antibody, we showed that phosphorylated p67(phox) and p47(phox) form a complex. Phosphoamino acid analysis of the phosphorylated p67(phox) revealed only 32P-labeled serine residues. Two-dimensional tryptic peptide mapping analysis showed that p67(phox) is phosphorylated at the same peptide whether fMLP or PMA is used as a stimulus. In addition, PKC induced the phosphorylation of recombinant GST-p67(phox) in vitro, at the same peptide as that phosphorylated in intact cells. PMA-induced phosphorylation of p67(phox) was strongly inhibited by the PKC inhibitor GF109203X. In contrast, fMLP-induced phosphorylation was minimally affected by this PKC inhibitor. Taken together, these results show that p67(phox) is phosphorylated in human neutrophils by different pathways, one of which involves protein kinase C.

Enzyme Activation↗
PubMed1997

Induction of interleukin-1 and subsequent tissue factor expression by anti-proteinase 3 antibodies in human umbilical vein endothelial cells.

OBJECTIVE: To assess the ability of anti-proteinase 3 (anti-PR3) classic antineutrophil cytoplasmic antibodies (cANCA) to stimulate endothelial expression of tissue factor (TF), which is the main initiator of the coagulation cascade that can lead to endothelial injury and thrombosis in patients with Wegener's granulomatosis. METHODS: Human umbilical vein endothelial cells (HUVEC) were grown to confluence and stimulated with affinity-purified anti-PR3 antibodies, Igs from healthy subjects, and endotoxin (lipopolysaccharide) as positive control. RESULTS: TF activity was generated in anti-PR3-stimulated cells, as shown by a chromogenic test. This activity was inhibited by specific anti-TF antibodies. TF messenger RNA (mRNA) was found in anti-PR3-stimulated cells, as detected by reverse transcriptase-polymerase chain reaction, but not in cells stimulated with irrelevant human Igs or Igs from normal control sera. TF expression reached maximum levels 12 hours after exposure to the anti-PR3 cANCA, and did not require complement. TF mRNA expression was inhibited by cycloheximide, suggesting a requirement for protein synthesis. When added to the incubation medium, interleukin-1 (IL-1) receptor antagonist inhibited the induced TF mRNA expression, suggesting that cANCA-stimulated cells initiate IL-1 synthesis. Moreover, cANCA induced IL-1alpha mRNA before TF mRNA. CONCLUSION: This study showed that anti-PR3 treatment of HUVEC induces sequential expression of IL-1alpha mRNA and TF mRNA, as well as their corresponding proteins. Both proteins could have pathogenic roles in the vasculitic process, since TF is the main initiator of the coagulation cascade.

Antibodies, Antineutrophil Cytoplasmic↗
PubMed1997

Antibodies to proteinase-3 mediate expression of intercellular adhesion molecule-1 (ICAM-1, CD 54).

We investigated the role of intercellular adhesion molecule-1 (ICAM-1) in the adhesion of polymorphonuclear neutrophils (PMN) to classic antineutrophil cytoplasmic antibody (C-ANCA)-treated endothelial cells, independently of cytokines. Human umbilical vein endothelial cells (HUVEC) grown to confluence in cytokine-free conditions were stimulated with C-ANCA sera and affinity-purified anti-proteinase 3 antibodies (PR3) from Wegener's granulomatosis (WG) patients. Non-activated PMN were added to treated HUVEC and adhesion was measured. In parallel experiments, treated HUVEC were fixed and ICAM-1 and E-selectin were assayed by cyto-ELISA; in other experiments anti-ELAM-1 and anti-ICAM-1 antibodies were assessed. In this in vitro model, adhesion of non-activated PMN to anti-PR3-stimulated HUVEC was enhanced. Adhesion was greater with anti-PR3 antibodies than with control and normal immunoglobulins, and correlated with the level of anti-PR3 antibodies. Neutralization of anti-PR3 antibodies by neutrophil azurophilic granule proteins abolished adhesion. This adhesion increased at the fourth hour after simulation, peaked at the twelfth hour and then decreased. This phenomenon occurred mainly through endothelial expression of ICAM-1 (the main counter-receptor for integrins, involved in firm PMN adhesion and migration) and E-selectin on HUVEC membranes. Anti-adhesion molecule antibodies inhibited this adhesion. This work supports the hypothesis of a direct effect of C-ANCA in endothelial stimulation, namely, on endothelium-PMN adhesion, and strengthens the major role of ICAM-1, directly involved in firm sticking of PMN to HUVEC, besides E-selectin. C-ANCA upregulate endothelial adhesiveness and thus participate in inflammatory reactions by providing endothelial adhesive structures for neutrophils. This might be one of the first steps leading to clinical expression of the disease. These results provide new insights into the pathogenesis of C-ANCA-related diseases.

Antibodies↗
PubMed1997

Effect of advanced glycation end product-modified albumin on tissue factor expression by monocytes. Role of oxidant stress and protein tyrosine kinase activation.

Diabetes is associated with a hypercoagulable state that contributes to macrovascular complications, including cardiovascular events. The glycation reaction, a consequence of chronic hyperglycemia, has also been implicated in the pathogenesis of diabetic complications. Glycated proteins have receptors on monocytes and generate reactive oxygen species that can regulate the expression of a number of genes. As abnormal monocyte expression of tissue factor (TF), the main initiator of the coagulation cascade, is responsible for thrombosis in a number of clinical settings, we studied the effect of glycated albumin on monocyte TF expression. Mononuclear cells were incubated with glycated albumin for 24 hours, and monocyte TF activity was measured with a plasma recalcification time assay; TF antigen was measured by ELISA and TF mRNA by RT-PCR. Glycated albumin induced blood monocyte expression of the procoagulant protein TF at the mRNA level. Oxidative stress appeared to be involved in this effect, as the antioxidant N-acetylcysteine diminished TF mRNA accumulation in stimulated monocytes. Hydroxyl radicals, which may be generated inside cells from H2O2 via the Fenton reaction, also appeared to be involved in this effect, as hydroxyl radical scavengers downregulated TF activity and antigen levels (but not TF mRNA). Finally, the involvement of activated protein tyrosine kinase in the transmission of the signal from the membrane to the nucleus was suggested by the inhibitory effect of herbimycin A. These results point to a new mechanism for the hypercoagulability often described in diabetic patients and suggest that antioxidants or protein tyrosine kinase inhibitors might be of therapeutic value in this setting.

Acetylcysteine↗
PubMed1996

Inhibition of human neutrophil binding to hydrogen peroxide-treated endothelial cells by cAMP and hydroxyl radical scavengers.

Hydrogen peroxide (H2O2) increases adherence of human polymorphonuclear neutrophils (PMN) to cultured human umbilical vein endothelial cells (HUVEC). Catalase and HO. scavengers did not affect the increased PMN adherence to HUVEC stimulated by other compounds such as phorbol myristate acetate (PMA) and thrombin, showing that the observed effect was H2O2- and HO.-specific. This effect was inhibited by hydroxyl radicals (HO.) scavengers and not by iron-chelators that do not penetrate the cells, suggesting the involvement of intracellular HO. in the increased adherence mechanism. An increase in cAMP inhibited H2O2-induced adherence, as observed with isoproterenol, isobutylmethylxanthine, and dibutyryl-cAMP. Similarly, pentoxifylline (Ptx), an HO. scavenger that also increases cAMP, inhibited H2O2-mediated adherence but had no effect on that induced by PMA or thrombin. PKA inhibitors cancelled the Ptx-induced inhibition of H2O2-mediated adherence. However, PKA inhibitors or atrial natriuretic peptide that decreases cAMP did not increase adherence, showing that decrease in cAMP is not responsible for increased adherence. HO. scavengers did not alter the H2O2-induced reduction in cAMP levels, but did inhibit the effect of H2O2 on adherence. We conclude that HO. mediates the H2O2-induced increased in PMN adherence to HUVEC, and that the increase in cAMP that mediates PKA activation downregulates this effect.

1-Methyl-3-isobutylxanthine↗
PubMed1995

Redistribution of protein kinase C isoforms in human neutrophils stimulated by formyl peptides and phorbol myristate acetate.

The redistribution of protein kinase C (PKC) isoforms between the cytosolic and plasma membrane fractions of stimulated human polymorphonuclear leukocytes (PMN) was analysed by means of western blotting with antibodies against PKC beta I, beta II and Zeta. Treatment of PMN with 1 microM formyl-methionyl-leucyl-phenylalanine (fMLP) induced a rapid (5-10 sec) and sustained (at least 10 min) increase in the membrane association of PKC beta I, beta II, and the two immunoreactive proteins (76-81 kDa) recognized by the antibody directed against PKC zeta. Optimal translocation of PKC isoforms to the plasma membrane occurred in the presence of 10(-6) M fMLP and was not associated with a detectable fall in cytosolic PKC. In the absence of external calcium, the translocation of all PKC isoforms induced by fMLP was rapid (5 sec) but the membrane association of PKC was lost within one minute. Unlike fMLP, phorbol myristate acetate (PMA) induced a concentration-dependent translocation of the PKC isoforms, which persisted in the membrane in the absence of external calcium. These data provide the first evidence of redistribution of PKC isoforms by a chemoattractant. They further indicate that external calcium plays a crucial role in the persistence of the membrane association of PKC beta I, beta II and zeta induced by formyl peptides.

Blotting, Western↗
PubMed1995

Contrasting effects of calyculin A and okadaic acid on the respiratory burst of human neutrophils.

The involvement of serine/threonine protein-phosphatases in the production of superoxide (respiratory burst) by human neutrophils was investigated using calyculin A, a potent inhibitor of both protein phosphatases type 1 and 2A, and okadaic acid, which preferentially inhibits protein phosphatase type 2A. Treatment of neutrophils with calyculin A (25-75 nM) or okadaic acid (1-4 microM) had no stimulatory effect but potently enhanced total superoxide production induced by an optimal fMLP (N-formyl-methionyl-leucyl-phenylalanine) concentration (0.1 microM). The maximum increase plateaued with 50-75 nM calyculin A and 2-4 microM okadaic acid, reaching approximately 120 and 200% of control values, respectively. Unlike calyculin A, okadaic acid also primed the initial rate of superoxide production, suggesting that protein phosphatases may down-regulate both initiation and termination of respiratory burst. Optimal stimulation of the respiratory burst by PMA (160 nM) was inhibited by calyculin A and okadaic acid, with an IC50 of 60 nM and 2 microM, respectively, although both drugs caused protein hyperphosphorylation. The inhibition was partially prevented by a nonstimulatory concentration of A23187, indicating a role of calcium in the inhibitory effects of the drugs. Unlike the optimal respiratory burst, suboptimal respiratory burst induced by PMA (1-7 nM) was enhanced by calyculin A and okadaic acid. Unprimed and primed respiratory bursts were depressed by a selective antagonist of protein kinase C (GF 109203X), indicating positive regulation of these responses by protein kinase C. Thus, the use of calyculin A and okadaic acid distinguishes two regulatory processes of superoxide production.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Impairment of polymorphonuclear neutrophil function in HIV-infected patients.

Impaired polymorphonuclear neutrophil (PMN) function may contribute to the onset of certain bacterial and fungal infections and to tissue damage in human immunodeficiency virus (HIV)-infected patients. Published data on PMN function in HIV infection are controversial, possibly because most studies have involved PMNs isolated from the normal blood environment by various procedures that may modify PMN responses. We therefore used flow cytometry to study the expression of adhesion molecules at the PMN surface, actin polymerization, and the oxidative burst of whole-blood PMNs in 42 HIV-infected patients at different stages of the disease. These PMNs were activated in vivo, as shown by increased expression of the adhesion molecule CD11b/CD18, reduced L-selectin antigen expression, increased actin polymerization, and increased H2O2 production. The alterations were present in asymptomatic patients with CD4+ cell counts above 500/microliters and did not increase with progression of the disease. This PMN activation could contribute to the oxidative stress described in HIV infection. Stimulation by bacterial N-formyl peptides showed dysregulation of L-selectin shedding and decreased H2O2 production after cx vivo priming with tumor necrosis factor-alpha or interleukin-8. These latter impairments, which correlated with the decrease in CD4+ lymphocyte numbers, could contribute to the increased susceptibility of HIV-infected patients to bacterial infections.

Actins↗
PubMed1995

Effects of pentoxifylline on the adherence of polymorphonuclear neutrophils to oxidant-stimulated human endothelial cells: involvement of cyclic AMP.

Cultured human umbilical vein endothelial cells (HUVECs) treated with reactive oxygen species (ROS) show increased adherence of polymorphonuclear leukocytes (PMNs). Because pentoxifylline (PTX) is known to inhibit cell interactions, we studied PMN adherence to ROS-stimulated HUVECs pretreated with PTX. ROS were generated by the oxidation of hypoxanthine by xanthine oxidase, giving rise to superoxide anion and hydrogen peroxide. Human PMNs were then added to HUVEC monolayers. After various times, the cultures were washed and the number of adherent PMNs was estimated by measuring myeloperoxidase in the total cell homogenate. PTX inhibited adherence in a concentration-dependent manner. Moreover, the increase in intracellular cAMP content varied with the PTX concentration. Isobutylmethylxanthine (IBMX) and isoproterenol (ISO) which increase intracellular cAMP content, also inhibited the adherence of PMNs to ROS-stimulated HUVECs. We conclude that cAMP is probably involved in the intracellular regulation of ROS-mediated PMN adherence to endothelial cells.

1-Methyl-3-isobutylxanthine↗
PubMed1995

Pharmacologic control of intracellular signaling pathways: from research to therapy.

Intracellular signaling pathways mediate the main events in a cell's life cycle, such as growth, proliferation, differentiation, performance of specialized functions, and apoptosis. Intracellular signaling is initiated either by extracellular signals or by intracellular gene products, e.g., those resulting from appropriately or inappropriately activated oncogenes. Extracellular signals either are generated by the general environment or are secretory products of other cells (e.g., growth factors, cytokines, hormones). Both act on specific receptors and, with some similarities to endogenous products, activate intracellular signaling and cell functional responses. Extracellular and intracellular signals coordinate the activities of the various cells in multicellular organisms. Dysregulation of signaling pathways by extracellular stimuli such as pathogens, or by gene abnormalities such as oncogene activation or loss of tumor suppressor gene activity, induces disease. The usual therapeutic targets of drugs are the stimuli and, in cancer, the oncogene or its direct product, the mRNA. Molecular components of cell signaling pathways are candidate targets for therapy. This approach may itself be sufficient to modify the positive or negative effects of a molecule in the signaling pathway. Drugs targeting cell signaling molecules can be combined with drugs acting on oncogenes, extracellular stimuli, or receptors. The main molecules involved in intracellular signaling are those of the transduction pathways and the second messengers and transcription factors. These molecules and their interactions are first recalled and are then analyzed to determine how, by the use of "specific" inhibitors, antisense methods, and directed mutagenesis, their functionality has been established. Finally, we will consider whether or not these methods can be adapted to clinical use.

Animals↗
PubMed1995

Protein tyrosine kinase activation is required for LPS and PMA induction of tissue factor mRNA in human blood monocytes.

Tissue factor (TF) is a transmembrane glycoprotein which assembles with factor VIIa on cell surfaces to form a proteolytically active cofactor-enzyme complex; the TF/VIIa complex initiates the coagulation protease cascade. In response to bacterial lipopolysaccharide (LPS) and phorbol-12 myristate 13-acetate (PMA), monocytes synthesize and express TF on their surface. However, the mechanisms by which LPS and PMA activate TF synthesis by human blood monocytes are not fully understood. As it has been established that LPS and PMA activate protein tyrosine kinase (PTK) in monocytes, we studied the role of PTK in LPS and PMA induction of TF by human blood monocytes. Both LPS- and PMA-induced TF activity was inhibited in a concentration-dependent manner by the protein tyrosine kinase-specific inhibitors herbimycin A and genistein. TF antigen determination confirmed that LPS- and PMA-induced cell surface TF protein levels decreased in parallel to TF functional activity under herbimycin A and genistein treatment. Northern blot analysis of total RNA from LPS- and PMA-stimulated monocytes showed a concentration-dependent decrease in TF mRNA levels in response to herbimycin A and genistein. The rate of decay of LPS-induced TF mRNA, evaluated after the arrest of transcription by actinomycin D was not affected by genistein and herbimycin A, suggesting that the inhibitory effects occur at least partly at the transcriptional level. We conclude that LPS- and PMA-induced TF production by human monocytes is dependent on tyrosine kinase activation.

Antibodies, Monoclonal↗
PubMed1994

Okadaic acid, an inhibitor of type 1 and type 2A phosphatases, modulates the activation of phospholipase D in formyl peptide- and mastoparan-stimulated human neutrophils.

The activation of phospholipase D (PLD) induced by formyl peptides (fMLP), as evaluated by production of tritiated phosphatidylethanol (PEt) and phosphatidic acid (PA) in polymorphonuclear leukocytes (PMN), was markedly enhanced (50-125%) by low okadaic acid concentrations (0.25-0.5 microM) but inhibited by higher concentrations (2-3 microM), although the drug caused protein hyperphosphorylation. Both effects of okadaic acid were amplified when PLD activation was primed with cytochalasin B. Stimulation of PMN with mastoparan, a wasp venom toxin that activates Pertussis toxin(PTX)-sensitive G proteins, resulted in a weak calcium-dependent production of PEt which was respectively enhanced and inhibited by okadaic acid (1-2 microM) in unprimed and cytochalasin-primed PMN. The results show that low okadaic acid concentrations primed fMLP-mediated activation of PLD, in keeping with a down-regulatory role of protein phosphatases. The contrasting effects of okadaic acid in mastoparan-stimulated PMN further suggest that protein phosphatases may regulate the generation of second messengers through alteration of major signaling events at/or downstream of PTX-sensitive G proteins (Gi).

Dose-Response Relationship, Drug↗
PubMed1994

Interleukin-10 and pentoxifylline inhibit C-reactive protein-induced tissue factor gene expression in peripheral human blood monocytes.

Fibrin deposition is an integral feature of the inflammatory response. In response to C-reactive protein (CRP), an acute-phase reactant, blood monocytes synthesize and express tissue factor (TF), the main initiator of blood coagulation. We report the inhibitory effect of interleukin 10 (IL-10) and that of pentoxifylline, a methyl xanthine derivative, on monocyte expression of TF activity, TF protein and TF mRNA in response to CRP. These agents may be of use in diseases where a TF-induced prothrombotic state is detrimental.

C-Reactive Protein↗
PubMed1994

Polymorphonuclear neutrophils from human immunodeficiency virus-infected patients show enhanced activation, diminished fMLP-induced L-selectin shedding, and an impaired oxidative burst after cytokine priming.

Impaired polymorphonuclear neutrophil (PMN) function may contribute to the onset of certain life-threatening bacterial and fungal infections in human immunodeficiency virus (HIV)-infected patients. Published data on PMN functional activity in HIV infection are controversial, possibly because most studies have involved PMNs isolated from their blood environment by means of various procedures that may differently affect surface receptor expression and thereby alter cellular responses. We therefore used flow cytometry to study the expression of adhesion molecules at the PMN surface, actin polymerization, and the oxidative burst of whole-blood polymorphonuclear neutrophils in 42 HIV-infected patients at different stages of the disease. These PMNs were activated in vivo, as demonstrated by increased expression of the adhesion molecule CD11b/CD18, reduced L-selectin antigen expression, increased actin polymerization, and increased H2O2 production. The alterations were present in asymptomatic patients with CD4+ cell counts greater than 500/microL and did not increase with the progression of the disease. Stimulation by bacterial N-formyl peptides showed dysregulation of L-selectin shedding and decreased H2O2 production after ex vivo priming with tumor necrosis factor alpha or interleukin-8 (IL-8). These latter impairments, which correlated with the decrease in CD4+ lymphocyte numbers and with IL-8 and IL-6 plasma levels, could contribute to the increased susceptibility of HIV-infected patients to bacterial infections.

Acquired Immunodeficiency Syndrome↗