Effects of an aqueous extract from leaves of Ligustrum vulgare on mediators of inflammation in a human neutrophils model.

Leaves of Ligustrum vulgare (common privet) have been used for treatment of oropharyngeal inflammations or as antirheumatic, diuretic, and hypotensive agents in folk medicine in southern Europe. Taking into account that neutrophils are involved in the inflammation, the aim of the study was to determine the effect of an aqueous extract prepared from leaves of Ligustrum vulgare on neutrophil functions. The extract was characterized by the HPLC-DAD-MSn method. The inhibition of reactive oxygen species production by formyl-met-leu-phenylalanine- or phorbol 12-myristate 13-acetate-stimulated neutrophils was determined using luminol- or lucigenin-dependent chemiluminescence. The effect on myeloperoxidase, metalloproteinase 9, and interleukin 8 production by neutrophils was measured by an enzyme-linked immunosorbent assay. Neutrophil elastase release was established spectrophotometrically. The expression of adhesion molecules on neutrophils was analyzed with flow cytometry. The main compounds detected were flavonoids, phenylpropanoids, hydroxycinnamates, and secoiridoids. The inhibition of oxidative burst by the extract was comparable in both stimuli models (formyl-met-leu-phenylalanine: IC50 = 18.2 ± 4.0 µg/mL; phorbol 12-myristate 13-acetate: IC50 = 19.8 ± 3.0 µg/mL). The extract in the concentration range of 5-50 µg/mL inhibited neutrophil elastase release by 23.9-34.1 % and myeloperoxidase release by 24.2-37.4 %. The inhibitory effect on metalloproteinase 9 and interleukin 8 production was around 20 %. The extract in the highest concentration modulated the expression of L-selectin and ß2 integrin. Our results partly support the traditional use of common privet leaves as an anti-inflammatory agent.

Involvement of adhesion molecule in in vitro plaque-like formation of macrophages stimulated with Aggregatibacter actinomycetemcomitans lipopolysaccharide.

BACKGROUND AND OBJECTIVE: Inflammatory agents, such as lipopolysaccharide (LPS), in periodontal pockets may promote atherogenesis by activating leukocytes. In our previous study, we developed a microchannel chip to observe the cell adhesion process in a fluid system. The objective of this investigation was to examine the mechanism by which periodontopathic bacterial LPS enhances plaque-like formation on a microchannel chip. MATERIAL AND METHODS: To evaluate the effect of Aggregatibacter actinomycetemcomitans LPS on the expression of adhesion molecules, e.g. intercellular adhesion molecule 1 (ICAM-1), lymphocyte function-associated antigen 1 (LFA-1) and L-selectin, on the surface of murine macrophage RAW264.7 cells, the expression of each adhesion molecule was examined by flow cytometry and western blot analysis. Moreover, a flow test on the microchannel chip involving anti-adhesion molecule antibodies was conducted to clarify which adhesion molecule is related to plaque-like formation of RAW264.7 cells. RESULTS: The expressions of ICAM-1 and LFA-1 on the surface of RAW 264.7 cells increased following 12 h culture with LPS; L-selectin expression was unaffected. An increase in ICAM-1 expression was also confirmed by western blot analysis. The flow test revealed that anti-ICAM-1 antibody inhibited plaque-like formation of LPS-stimulated macrophages on the micropillars of the microchannel chip. CONCLUSION: These findings indicate that ICAM-1 plays an important role in plaque-like formation of LPS-stimulated macrophages. Our microchannel chip is a suitable tool for the investigation of etiological factors of atherosclerosis, including periodontitis, in vitro.

Inhibition of L-selectin binding by polyacrylamide-based conjugates under defined flow conditions.

Selectins mediate tethering and rolling of leukocytes along the endothelium in a shear force-dependent manner. This key step in the cellular immune response is a target for experimental anti-inflammatory therapies. In the present paper we have examined the inhibitory activity of the minimal selectin ligand sialyl Lewis x (SiaLe(x)), its isomer sialyl Lewis a (SiaLe(a)) and sulfated tyrosine (sTyr) residues under dynamic flow reflecting the rheological conditions in the blood stream. The monomeric ligands were compared to multivalent polyacrylamide (PAA)-based conjugates under defined flow conditions on the molecular level, using surface plasmon resonance (SPR) technology, and on the cellular level, using a parallel-plate flow chamber. SPR measurements showed that a spatial arrangement of binding epitopes mimicking the selectin binding motif of the natural ligand PSGL-1 inhibits L-selectin binding successfully with IC(50) values in the nanomolar range. Using a flow chamber adhesion assay it could be shown that the multivalent inhibitors efficiently blocked rolling and tethering of NALM-6 pre-B cells transfected with human L-selectin to activated endothelium and that the inhibitory activity increased with rising shear stress. While PAA-conjugates were almost not inhibitory at low shear stress, NALM-6 cell rolling was nearly completely inhibited at high shear stress. The results indicate that multimeric conjugates of SiaLe(x), SiaLe(a) and sTyr are highly effective inhibitors of L-selectin-mediated cell adhesion particularly under flow conditions. Consequently, SiaLe(x), SiaLe(a) and/or sTyr on macromolecular carriers may be promising candidates for anti-inflammatory therapy.

Leukocyte and endothelial adhesion molecules in patients with hypercholesterolemia: the effect of atorvastatin treatment.

Atherogenesis involves the migration of leukocytes into vascular subendothelial space, a process mediated by endothelial and leukocyte cell adhesion molecules. Endothelial molecules are assessed indirectly via serum levels, but leukocyte molecules can be assessed directly. We have therefore hypothesized that leukocyte adhesion molecules are altered to a greater degree in hypercholesterolemia than serum endothelial adhesion molecules. We examined 29 subjects with hypercholesterolemia and 27 controls at baseline and after 12 weeks of atorvastatin treatment (20 mg/day). Expression of leukocyte integrins CD11a, CD11b, CD18, and CD49d and of L-selectin was measured by flow cytometry. Serum ICAM-1, E-selectin and von Willebrand factor were measured by ELISA. Expression of leukocyte adhesion molecules was significantly higher in patients at baseline than in the controls, except for CD11a. Expression significantly decreased after atorvastatin in most adhesion molecules except for CD11b. In contrast, there was no effect of hypercholesterolemia and/or atorvastatin on the serum endothelial molecules. Leukocyte but not endothelial adhesion molecules were influenced by hypercholesterolemia and by lipid lowering treatment. Leukocyte molecules may therefore be a more sensitive marker of atherogenesis than endothelial molecules. Our results support the role of increased leukocyte adhesiveness in atherogenesis.

Early Targeting of L-Selectin on Leukocytes Promotes Recovery after Spinal Cord Injury, Implicating Novel Mechanisms of Pathogenesis.

L-selectin, a lectin-like receptor on all leukocyte classes, functions in adhesive and signaling roles in the recruitment of myeloid cells from the blood to sites of inflammation. Here, we consider L-selectin as a determinant of neurological recovery in a murine model of spinal cord injury (SCI). Spinal cord-injured, L-selectin knock-out (KO) mice (male) showed improved long-term recovery with greater white matter sparing relative to wild-type (WT) mice and reduced oxidative stress in the injured cord at 72 h post-SCI. There was a partial and transient reduction in accumulation of neutrophils in the injured spinal cords of KOs at 24 h post-injury. To complement these findings with KO mice, we sought a pharmacologic means for lowering L-selectin levels. We found that diclofenac, a nonsteroidal anti-inflammatory drug (NSAID), induced the shedding of L-selectin from the cell surface of myeloid subsets, specifically neutrophils and non-classical monocytes, in the blood and the injured spinal cord. Diclofenac administration to injured WT mice enhanced neurological recovery to a level comparable to that of KOs but did not improve recovery in KOs. While diclofenac treatment had no effect on myeloid cell accumulation, there was a reduction in oxidative stress at 72 h post-SCI. These findings implicate L-selectin in secondary pathogenesis beyond a role in leukocyte recruitment and raise the possibility of repurposing diclofenac for the treatment of SCI.

The effects of ethanol on beta2-integrin and l-selectin on the surface of leukocytes in human whole blood.

BACKGROUND: Acute alcohol intoxication is associated with increased susceptibility to infection. In host defense, the expression of adhesion molecules such as beta2-integrin and l-selectin on leukocytes is involved in leukocyte migration to inflamed organ tissue. To elucidate the mechanisms underlying the immunosuppressive effects of ethanol, we investigated whether ethanol pretreatment may influence the changes in adhesion molecule expression induced by lipopolysaccharide (LPS) or interleukin (IL)-8 in human whole blood. METHODS: Ethanol was added to samples of human whole blood (final concentration: 0%, 0.2%, 0.4%, and 0.8%). Samples were assigned to an unstimulated group and an LPS-stimulated group. In another set of experiments, stimulation was induced by IL-8. After fluorescence labeling of alphaM-subunit of beta2-integrin (CD11b) and l-selectin (CD62L), the expression of CD11b and CD62L were measured using flow cytometry. RESULTS: Stimulation with LPS significantly upregulated CD11b expression (5.9 +/- 0.9 to 16.3 +/- 1.8, p < 0.05). Ethanol inhibited this LPS-induced upregulation of CD11b (p < 0.001). Stimulation with IL-8 significantly upregulated CD11b expression (5.3 +/- 1.7 to 7.5 +/- 2.7, p < 0.01) and this IL-8-induced upregulation of CD11b was also inhibited by ethanol pretreatment (p < 0.001). In contrast, ethanol did not modify CD62L expression in either unstimulated or stimulated groups. CONCLUSION: The impairment of CD11b expression on leukocytes suggests that alcohol intake interferes with the migration of leukocytes to sites of inflammation, which may explain, in part, why alcohol intoxication increases susceptibility to infection.

Down-Regulation of CD62L Shedding in T Cells by CD39+ Regulatory T Cells Leads to Defective Sensitization in Contact Hypersensitivity Reactions.

Injection of regulatory T cells (Tregs) followed by sensitization with 2,4,6-trinitrochlorobenzene induced a transient increase in size and cellularity of skin-draining lymph nodes (LNs) in mice. This led us to hypothesize that Tregs may affect the trafficking of T cells from and to peripheral LNs. Two to three hours after sensitization, we found fewer CD8+ T cells expressing CD62L in LNs compared with untreated controls. Injection of wild-type Tregs prevented this down-regulation of CD62L. In contrast, Tregs devoid of the adenosine triphosphate (ATP)-degrading ecto-enzyme CD39 were unable to do so. As for the mechanism of CD62L regulation, we found that ATP, which is released in skin upon hapten-exposure, is inducing the protease ADAM17 in LN-residing T cells via engagement of P2X7 ATP receptors. ADAM17 cleaves CD62L from the surface of CD8+ T cells, which in turn provide a signal for T cells to leave the LNs. This regulation of CD62L is disturbed by the presence of Tregs, because Tregs remove extracellular ATP from the tissue by activity of CD39 and, therefore, abrogate the shedding of CD62L. Thus, these data indicate that the regulation of ATP turnover by Tregs in skin and LNs is an important modulator for immune responses.

Effects of dexamethasone treatment and respiratory vaccination on rectal temperature, complete blood count, and functional capacities of neutrophils in beef steers.

The objective of this research was to examine the effects of dexamethasone (DEX) treatment on various aspects of immunity following administration of a multivalent respiratory vaccine, using a model intended to mimic acute versus chronic stress. Angus × Hereford steers ( = 32; 209 ± 8 kg) were stratified by BW and randomly assigned to 1 of 3 treatments: 1) acute stress (ACU), in which 0.5 mg/kg BW DEX was intravenously administered at 1000 h only on d 0; 2) chronic stress (CHR), in which 0.5 mg/kg BW DEX was intravenously administered at 1000 h on d -3 to 0; or 3) control (CON), in which no DEX was administered. Steers were fitted with indwelling jugular catheters and rectal temperature (RT) recording devices on d -4 relative to vaccination and placed in individual stanchions in an environmentally controlled facility. Blood samples were collected and serum was isolated at -74, -50, and -26 h; at 0.5-h intervals from -4 to 6 h; and at 12, 24, 36, 48, and 72 h relative to multivalent respiratory vaccination at 1200 h on d 0. Additional blood samples were used to analyze complete blood cell count (CBC) and functional capacities of neutrophils. There was a treatment × time interaction ( < 0.01) for RT such that DEX treatment in CHR and ACU steers decreased RT on d -3 and 0, respectively. A treatment × time interaction ( < 0.01) was observed for total white blood cells (WBC), neutrophils, lymphocytes, and monocytes. Specifically, DEX increased WBC and neutrophils in CHR and ACU steers ( < 0.001) yet decreased lymphocytes in CHR steers ( = 0.02) compared with CON steers. Neutrophil concentration increased rapidly, within 2 h of the DEX infusion, in ACU steers. Monocytes transiently increased ( < 0.001) in response to DEX treatment in CHR and ACU steers. In contrast, eosinophils were greater ( < 0.01) in CON steers than in ACU and CHR steers. A treatment × time interaction ( = 0.004) was observed for interferon-γ, with CON cattle exhibiting greater concentrations than the ACU and CHR cattle at 5 h after vaccination, through d 3. Treatment also influenced ( ≤ 0.001) the expression of L-selectin on the surface of neutrophils. The percentage of neutrophils engaging in phagocytosis and the oxidative burst were suppressed ( ≤ 0.001) among only the CHR steers, whereas the intensity of the oxidative burst was suppressed ( ≤ 0.001) for both ACU and CHR steers. These data suggest that our model induced acute and chronic immunosuppression and defined the acute response to a multivalent vaccine in CON steers.

Effects of tibolone on selectins in postmenopausal women.

OBJECTIVE: The first step in atherosclerosis is characterized by the adherence of lymphocytes and monocytes to cell adhesion molecules expressed by endothelial cells. The precise mechanism by which steroid hormones may be exerting a protective action against atherogenesis remains unclear. Therefore, we wanted to investigate the effect of tibolone on the circulating levels of various selectins in postmenopausal women. METHODS: Thirty healthy postmenopausal women were enrolled in a prospective, randomized, double blind, placebo-controlled outpatient trial. RESULTS: Patients treated with tibolone revealed a significant decrease for the variables sE-selectin, sL-selectin, and sPECAM-1 after 8 weeks of treatment. CONCLUSIONS: By reducing leukocyte adhesion molecule expression on human endothelial cells, tibolone may have the intrinsic potential to exert additional, lipid-independent, cardiovascular protective effects that may explain the clinical benefits of cardiovascular diseases in postmenopausal women.

Synthetic glycoprotein mimics inhibit L-selectin-mediated rolling and promote L-selectin shedding.

L-selectin is a leukocyte cell-surface protein that facilitates the rolling of leukocytes along the endothelium, a process that leads to leukocyte migration to a site of infection. Preventing L-selectin-mediated rolling minimizes leukocyte adhesion and extravasation; therefore, compounds that inhibit rolling may act as anti-inflammatory agents. To investigate the potential role of multivalent ligands as rolling inhibitors, compounds termed neoglycopolymers were synthesized that possess key structural features of physiological L-selectin ligands. Sulfated neoglycopolymers substituted with sialyl Lewis x derivatives (3',6-disulfo Lewis x or 6-sulfo sialyl Lewis x) or a sulfatide analog (3,6-disulfo galactose) inhibited L-selectin-mediated rolling of lymphoid cells. Functional analysis of the inhibitory ligands indicates that they also induce proteolytic release of L-selectin. Thus, their inhibitory potency may arise from their ability to induce shedding. Our data indicate that screening for compounds that promote L-selectin release can identify ligands that inhibit rolling.

Effects of hypertonic saline on expression of human polymorphonuclear leukocyte adhesion molecules.

Hypertonic saline prevents vascular adherence of neutrophils and ameliorates ischemic tissue injury. We hypothesized that hypertonic saline attenuates N-formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated expression of adhesion molecules on human polymorphonuclear leukocytes (PMNLs). fMLP-stimulated up-regulation of beta2-integrins was diminished by hypertonic saline but not by hypertonic choline chloride-, mannitol-, or sucrose-modified Hanks' buffered salt solution. Shedding of L-selectin was decreased by hypertonic saline and choline chloride but not by hypertonic mannitol or sucrose. When the effects of hypertonic sodium chloride- and choline chloride-modified media were compared, neither solution affected fMLP-receptor binding but both equally inhibited fMLP-stimulated increase in intracellular calcium, ionophore A23187, and phorbol myristate acetate (PMA)-stimulated numerical up-regulation of beta2-integrins. Analysis of mitogen-activated protein (MAP) kinases p38 and p44/42 for phosphorylation revealed that hypertonic solutions did not differ in preventing fMLP-stimulated increases in phospho-p38 and phospho-p44/42. Resting PMNLs shrunk by hypertonic saline increased their volume during incubation and further during chemotactic stimulation. Addition of amiloride further enhanced inhibition of up-regulation of beta2-integrins. No fMLP-stimulated volume changes occurred in PMNLs exposed to hypertonic choline chloride, resulting in significant cell shrinkage. Results suggest a sodium-specific inhibitory effect on up-regulation of beta2-integrins of fMLP-stimulated PMNLs, which is unlikely to be caused by alterations of fMLP receptor binding, decrease in cytosolic calcium, attenuation of calcium or protein kinase C-dependent pathways, suppression of p38- or p44/42 MAP kinase-dependent pathways, or cellular ability to increase or decrease volumes.

Mechanism of the immunosuppressive effect in vivo of novel immunosuppressive drug beta-SQAG9, which inhibits the response of the CD62L+ T-cell subset.

INTRODUCTION: We synthesized sulfo-glycolipid, beta-SQAG9 (designate square beta-SQAG9 liposome, because it efficiently forms a liposome structure) that possessed immunosuppressive effects such as inhibition of T-cell responses in human allogeneic MLR and skin allograft survival in rats, and bound to CD62L (L-selectin) in vitro. In this study, we further investigated the immunosuppressive mechanism in vivo by beta-SQAG9 liposome in a skin-allografted rat model. METHODS: ACI rats (RT1(a)) were grafted skin of LEW rats (RT1(1)) treated with PBS or beta-SQAG9 liposome IV once a day for 7 days. Subsequently, we investigated the population of T cells and CD62L(+) T-cell subset in the spleen, axillary lymph nodes (ALNs), and peripheral blood of skin-allografted rats by two-color flow cytometry. RESULTS: Five of 11 (45.5%) rats that were treated with 50 mg/kg beta-SQAG9 liposome showed graft survival and another showed moderate rejection in graft. The CD62L(+) T-cell subset population in ALNs of beta-SQAG9 liposome-treated rats decreased in a dose-dependent manner. No significant difference in the T-cell population was observed between the beta-SQAG9 and control groups. These data suggest that beta-SQAG9 could bind to the CD62L(+) T-cell subset in vivo as well as in vitro and affect T-cell migration, which might lead to T-cell tolerance in vivo.

Facile supermolecular aptamer inhibitors of L-selectin.

Multivalent interactions occur frequently in nature, where they mediate high-affinity interactions between cells, proteins, or molecules. Here, we report on a method to generate multivalent aptamers (Multi-Aptamers) that target L-selectin function using rolling circle amplification (RCA). We find that the L-selectin Multi-Aptamers have increased affinity compared to the monovalent aptamer, are specific to L-selectin, and are capable of inhibiting interactions with endogenous ligands. In addition, the Multi-Aptamers efficiently inhibit L-selectin mediated dynamic adhesion in vitro and homing to secondary lymphoid tissues in vivo. Importantly, our method of generating multivalent materials using RCA avoids many of the challenges associated with current multivalent materials in that the Multi-Aptamers are high affinity, easily produced and modified, and biocompatible. We anticipate that the Multi-Aptamers can serve as a platform technology to modulate diverse cellular processes.

Dexamethasone down-regulates the expression of L-selectin on the surface of neutrophils and lymphocytes in humans.

OBJECTIVE: On the basis of previous animal studies, we hypothesized that dexamethasone may reduce the expression of L-selectin on neutrophils and lymphocytes in healthy men. METHODS: A double-blind, randomized, placebo-controlled, and three-way crossover trial was conducted in nine healthy men. Every subject received four identical infusions of saline solution, 0.04 mg/kg dexamethasone, or 1.0 mg/kg dexamethasone during three observation periods of 48 hours each. RESULTS: Dexamethasone time and dose dependently decreased the L-selectin expression on neutrophils and lymphocytes as measured by flowcytometry. This effect occurred with a time lag of 8 hours after start of treatment: the L-selectin binding index of neutrophils decreased by a maximum of -50% (confidence interval [CI], -37% to -63%) and that of lymphocytes by -26% (CI, -8% to -45%) at 32 hours after the start of treatment with high-dose dexamethasone (p < 0.016). Low-dose dexamethasone had only a transient effect on L-selectin expression of lymphocytes and a less pronounced effect on L-selectin expression of neutrophils. CONCLUSION: Dexamethasone time and dose dependently decreases L-selectin expression on neutrophils and lymphocytes in health men, an effect that is less pronounced than that previously reported for animals.

Improved enzymatic synthesis of a highly potent oligosaccharide antagonist of L-selectin.

The polylactosamine sLex beta1-3'(sLex beta1-6')LacNAc beta1-3'(sLex beta1-6')LacNAc beta1-3'(sLex beta1-6')LacNAc (7) (where sLex is Neu5Ac alpha2-3Gal beta1-4(Fuc alpha1-3)GlcNAc and LacNAc is Gal beta1-4GlcNAc) is a nanomolar L-selectin antagonist and therefore a potential anti-inflammatory agent (Renkonen et al. (1997) Glycobiology, 7, 453). Here we describe an improved synthesis of 7. The octasaccharide LacNAc beta1-3'LacNAc beta1-3'LacNAc beta1-3'LacNAc (4) was converted into the triply branched undecasaccharide LacNAc beta1-3'(GlcNAc beta1-6')LacNAc beta1-3'(GlcNAc beta1-6')LacNAc beta1-3'(GlcNAc beta1-6')LacNAc (5) by incubation with UDP-GlcNAc and the midchain beta1,6-GlcNAc transferase activity of rat serum. Glycan 5 was enzymatically beta1,4-galactosylated to LacNAc beta1-3'(LacNAc beta1-6')LacNAc beta1-3'(LacNAc beta1-6')LacNAc beta1-3'(LacNAc beta1-6')LacNAc (6). Combined with the enzymatic conversion of 6 to 7 (Renkonen et al., loc. cit.) and the available chemical synthesis of 4, our data improve the availability of 7 for full assessment of its anti-inflammatory properties.

Isolation and characterisation of potential respiratory syncytial virus receptor(s) on epithelial cells.

Respiratory syncytial virus (RSV) infection causes severe lower respiratory diseases in infancy, early childhood and the elderly. RSV infections respond poorly to current therapies. Therefore, we initiated a search for novel drug targets by investigating the characteristics and identity of RSV adhesion receptors on mammalian cells. Soluble human lectins, complex polysaccharides and a low molecular selectin antagonist, TBC1269, were used to characterise and isolate the RSV receptor on a human epithelial cell line (Hep2 cells). The binding characteristics of the RSV receptor on Hep2 cells were similar to those reported for L-selectin. The carbohydrate-based selectin antagonists, fucoidan and TBC 1269, inhibit RSV infection both in vitro and in a mouse model of infection. Furthermore, we have isolated annexin II as a potential RSV receptor on Hep2 cells. The expression of annexin II was increased after RSV infection. Recombinant annexin II binds to RSV G-protein, heparin and plasminogen and the binding is inhibited by a selectin antagonist, TBC1269. These findings indicate that inhibitors of annexin II could have potential in treating RSV infection.

IL-1beta stimulation induces paracrine regulation of PMN function and apoptosis.

Polymorphonuclear leukocytes (PMN) play a crucial role in the primary immunological defense against infectious agents. PMN activation and function is influenced in a paracrine manner by cytokines and bacterial products. While cell-cell communication has been demonstrated between PMN and other cell types, little data is available addressing PMN-PMN communication. Therefore, the aim of this study was to determine whether PMN were able to affect PMN function in vitro in a cell-contact independent manner, and whether IL-1beta influenced this effect. Conditioned medias (CM) were prepared by incubating PMN in HBSS +/- IL-1beta for 1-4 h. Incubation of fresh PMN in these conditioned medias had little or no effect on the expression of cell surface FcgammaR expression or oxidative metabolism. However, incubation of PMN in CM-IL1beta, but not control CM, increased phagocytotic activity and suppressed apoptosis. Additionally, CM-IL1beta, but not control CM, slowed the changes in Mac-1 and CR1 cell surface expression that occurred in HBSS within 2 h of incubation. Finally, control CM down-regulated the cell surface expression of PSGL-1; an effect that was not observed with CM-IL1beta. In conclusion, we demonstrate that PMN are able to communicate with and influence the immunological function of other PMN independent of cell-cell contact, and that this influence is regulated by cytokines such as IL-1beta. The major impact of this paracrine regulation is to down-regulate PMN apoptosis with the potential for an upregulated inflammatory response.

Elevated levels of soluble adhesion molecules in serum of patients with diffuse panbronchiolitis.

STUDY OBJECTIVE: Adhesion molecules have been implicated in the pathogenesis of inflammatory diseases. This study was designed to determine whether soluble adhesion molecules in serum reflect the disease activity in diffuse panbronchiolitis (DPB). PATIENTS AND METHODS: Using an enzyme-linked immunosorbent assay, we measured the serum levels of soluble L-, E-, and P-selectin (sL-, sE-, and sP-selectin), intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 in 27 patients with DPB, 13 with bronchiectasis, and 15 normal adults. BAL was also performed, and the levels of interleukin (IL)-8 and IL-1 beta in BAL fluid (BALF) were measured. RESULTS: The serum levels of these molecules were significantly elevated in DPB patients compared with the control subjects. DPB patients also had significant high levels of circulating sE- and sP-selectin compared with patients with bronchiectasis. There was a significant correlation between serum sE-selectin and the percentage of neutrophils in BALF in all patients. There was a significant inverse correlation between serum sE-selectin and percent vital capacity in DPB patients. In the same patients, the relationships between serum sE-selectin and BALF concentrations of IL-1 beta as well as between serum sL-selectin and BALF IL-8 were also significant. Treatment of DPB patients with macrolides significantly reduced the serum levels of these soluble adhesion molecules and BALF concentrations of IL-1 beta and IL-8. CONCLUSIONS: Our results suggest that these soluble adhesion molecules, particularly selectins, may reflect the disease activity of DPB, and that their levels may be regulated by cytokines produced in the lungs.

Induction of adhesion molecules on human schwann cells by proinflammatory cytokines, an immunofluorescence study.

The presence of cytokines in the peripheral nerve was positively correlated to the induction and progression of inflammation during experimental allergic neuritis (EAN) and Guillain Barré syndrome (GBS). We investigated the induction of adhesion molecules such as L-selectin, E-selectin, ICAM-1, VCAM-1 and Mac-1 on Schwann cells by proinflammatory cytokines. Cultured human Schwann cells from normal adult, fetal and diabetic nerves were studied by immunofluorescence at basal condition and after stimulation with cytokines for 6, 24, 48 and 96 h. Incubation of human Schwann cells with TNFalpha, IFNgamma and IL-1beta induces the expression of ICAM-1 starting at 6 h and reaching a peak at 24 h on more than 90% of cells. VCAM-1 expression was induced after 6 h of treatment with TNFalpha and IL-1beta on almost 100% of Schwann cells. Surprisingly, stimulation with TNFalpha, IFNgamma and IL-1beta also induced the expression of L-selectin on fetal and diabetic Schwann cells, but not on normal adult cells. E-selectin, an adhesion molecule classically upregulated during inflammation, as well as Mac-1, a ligand for ICAM-1, were not expressed on human Schwann cells at basal condition or after treatment with cytokines. No ICAM-1, VCAM-1 and L-selectin expression was found on unstimulated Schwann cells. Our results suggest that upregulation of adhesion molecules on Schwann cells may have a role in the pathogenesis of inflammation in the peripheral nerve.

The effects of dipyridamole stress test on plasma levels of soluble adhesion molecules intracellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin and L-selectin in patients with ischemic heart disease and patients with syndrome X.

BACKGROUND: Adhesion of activated leukocytes to the endothelium as a result of myocardial ischemia/reperfusion has been shown to be involved in the development of tissue injury. Leukocyte adhesion to the endothelium occurs via adhesion molecules expressed on the surface of both cell types. Upon cell activation these proteins may be released into the circulation and measured in a soluble form. AIM: To verify whether the dipyridamole stress test, performed in patients with ischemic heart disease (IHD) and in patients with syndrome X, modifies plasma levels of the soluble adhesion molecules vascular cell adhesion molecule-1 (VCAM-1), intracellular adhesion molecule-1 (ICAM-1), E-selectin and L-selectin. METHODS: Plasma levels of the soluble endothelial adhesion molecules ICAM-1, VCAM-1 and E-selectin, as well as of the soluble leukocyte adhesion molecule L-selectin, were measured in venous blood samples taken before and 7 min after administration of dipyridamole in patients with IHD, patients with syndrome X and healthy individuals. Myocardial perfusion was evaluated using single photon emission tomography. The plasma levels of soluble VCAM-1, ICAM-1, E-selectin and L-selectin were all measured using enzyme-linked immunosorbent assays. RESULTS: After infusion of dipyridamole, plasma levels of ICAM-1 increased significantly in patients with IHD, whereas they remained unchanged in patients with syndrome X and in the control group. In patients with IHD, the initial plasma levels of VCAM-1, E-selectin and L-selectin, before administration of dipyridamole, were higher than those observed in patients with syndrome X and than those in the control group. Plasma levels of soluble VCAM-1, E-selectin and L-selectin decreased significantly in patients with IHD following the dipyridamole stress test, whereas they remained unchanged in patients with syndrome X, and in the control group. CONCLUSION: In patients with IHD, administration of dipyridamole induces myocardial ischemia resulting in modification of plasma levels of the soluble adhesion molecules.