Expression of intercellular adhesion molecule-1 on macrophages in vitro as a marker of activation

Macrophage activation is a major component of wound healing. It also determines the extent of inflammatory reactions and the response of the body to implanted materials. We have previously shown, using an in vitro model, that the extent of spreading of macrophages on different materials is a marker of activation, and that a soluble inducer has a dose-response effect on the secretion of cytokines in the culture medium. This work investigates the expression of three different cell surface markers [macrophages MAC-1, MAC-3 and intercellular adhesion molecule-1 (ICAM-1)] on macrophages in vitro using confocal microscopy and shows that ICAM-1 is also a marker of macrophage activation in this model. We observed increased amounts of ICAM-1 on activated macrophages compared to unactivated macrophages, whereas MAC-1 and MAC-3 were either expressed constitutively or demonstrated no quantitative change in expression after activation under the same experimental conditions. We also tested the expression of ICAM-1 with various concentrations of soluble inducers (lipopolysaccharide, 0.001, 0.01, 0.1, 1 and 10 micrograms ml-1. S-27609, 0.1, 0.25, 0.5, 1, 2 and 3 micrograms ml-1 and on a sheet of polylactic acid alone or in combination with soluble inducers. All doses of soluble inducers induced the expression of ICAM-1 on cells grown in glass chamber slides. The induction was not dose related but seemed to work rather in an on-off manner. There was no effect of material on ICAM-1 expression on the cell surface when no soluble inducer was added. This was similar to cytokine secretion, which was not induced by our material alone. When either lipopolysaccharide or S-27609 was used in combination with the material, there was an increase in the average measured intensity of ICAM-1. In this in vitro model, ICAM-1 staining as measured by confocal microscopy is a marker for macrophage activation. Our results suggest that the extent of macrophage activation as measured by ICAM-1 and by cytokine secretion is more sensitive to soluble inducers than to the action of the flat sheet of polylactic acid.     

Expression of intercellular adhesion molecule-1 in mice with Pseudomonas-induced pyelonephritis

PURPOSE: To investigate the role of intercellular adhesion molecule-1 (ICAM-1) in the renal inflammatory process, we studied the time-course fluctuation of ICAM-1 expression on inflammatory lesions in mice with experimentally induced bacterial pyelonephritis and the effect of in vivo administration of an anti-ICAM-1 monoclonal antibody (mAb) on leukocytic migration. MATERIALS AND METHODS: Ascending pyelonephritis was induced by transurethral instillation of Pseudomonas aeruginosa, and the expression of ICAM-1 in the pyelonephritic lesions was studied by immunohistochemical methods. RESULTS: The expression of ICAM-1 on the pyelonephritic lesions closely paralleled the degree of infiltration of neutrophils and macrophages until 3 days after infection. At 7 days after infection, though the degree of infiltration of these cells was quite high, expression of ICAM-1 was reduced. Treatment with the anti-ICAM-1 mAb in mice with bacterial pyelonephritis resulted in suppression of influx of neutrophils and macrophages in the infected sites until 3 days after infection. However, at 7 days after infection inhibition of the influx of these cells was not seen. CONCLUSIONS: These results suggest that ICAM-1 expression is transient and plays a key role in the influx of neutrophils and macrophages associated with the early-phase response, and that in the late phase ICAM-1 independent adhesion molecules may be more predominant.     

Expression of membrane and soluble intercellular adhesion molecule-1 in Graves'' disease

Understanding the epidemiology of equine colic is directly relevant to the management of individual horses with colic. In this article, the epidemiology of colic is reviewed with emphasis on epidemiologic studies that have identified specific factors associated with increased risk of colic and epidemiologic studies that are designed to predict the need for surgery and prognosis in horses with colic. Despite the magnitude of the problem of equine colic, much remains to be learned about the epidemiology of this disease.     

Role of the TRAF binding site and NF-kappaB activation in Epstein-Barr virus latent membrane protein 1-induced cell gene expression

In this study, we investigated the induction of cellular gene expression by the Epstein-Barr Virus (EBV) latent membrane protein 1 (LMP1). Previously, LMP1 was shown to induce the expression of ICAM-1, LFA-3, CD40, and EBI3 in EBV-negative Burkitt lymphoma (BL) cells and of the epidermal growth factor receptor (EGF-R) in epithelial cells. We now show that LMP1 expression also increased Fas and tumor necrosis factor receptor-associated factor 1 (TRAF1) in BL cells. LMP1 mediates NF-kappaB activation via two independent domains located in its C-terminal cytoplasmic tail, a TRAF-interacting site that associates with TRAF1, -2, -3, and -5 through a PXQXT/S core motif and a TRADD-interacting site. In EBV-transformed B cells or transiently transfected BL cells, significant amounts of TRAF1, -2, -3, and -5 are associated with LMP1. In epithelial cells, very little TRAF1 is expressed, and only TRAF2, -3, and -5, are significantly complexed with LMP1. The importance of TRAF binding to the PXQXT/S motif in LMP1-mediated gene induction was studied by using an LMP1 mutant that contains alanine point mutations in this motif and fails to associate with TRAFs. This mutant, LMP1(P204A/Q206A), induced 60% of wild-type LMP1 NF-kappaB activation and had approximately 60% of wild-type LMP1 effect on Fas, ICAM-1, CD40, and LFA-3 induction. In contrast, LMP1(P204A/Q206A) was substantially more impaired in TRAF1, EBI3, and EGF-R induction. Thus, TRAF binding to the PXQXT/S motif has a nonessential role in up-regulating Fas, ICAM-1, CD40, and LFA-3 expression and a critical role in up-regulating TRAF1, EBI3, and EGF-R expression. Further, D1 LMP1, an LMP1 mutant that does not aggregate failed to induce TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 expression confirming the essential role for aggregation in LMP1 signaling. Overexpression of a dominant form of IkappaBalpha blocked LMP1-mediated TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 up-regulation, indicating that NF-kappaB is an important component of LMP1-mediated gene induction from both the TRAF- and TRADD-interacting sites.     

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.     

新生儿败血症血清降钙素原和可溶性细胞间黏附分子-1的变化

目的 观察新生儿败血症患儿血清降钙素原(PCT)和可溶性细胞间黏附分子-1(sICAM-1)的变化.方法 新生儿败血症患儿56例,其中30例患儿血培养阳性(病原确诊组),26例血培养阴性(临床诊断组),取同期出生的新生儿25名作为对照组,分别应用化学发光法和酶联免疫吸附法测定患儿血清PCT和sICAM-1水平,同时进行新生儿危重评分.结果 败血症组血清sICAM-1和PCT明显高于对照组,且病原确诊组的血清PCT和sICAM-1水平又明显高于临床诊断组,差异均有统计学意义(P＜0.01).血清sICAM-1和PCT水平与新生儿危重评分呈负相关(γ=-0.778,P＜0.01;γ=-0.851,P＜0.01).结论 PCT和sICAM-1可作为新生儿败血症早期诊断的敏感指标.     

Rickettsia conorii infection enhances vascular cell adhesion molecule-1- and intercellular adhesion molecule-1-dependent mononuclear cell adherence to endothelial cells

Leukocyte adherence to the endothelium is an essential component of the inflammatory response during rickettsial infection. In vitro, Rickettsia conorii infection of endothelial cells enhances the expression of adhesive molecules E-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) in a time- and dose-dependent manner. Rickettsial lipopolysaccharide does not seem to be involved, because polymyxin B does not reduce their expression. The intracellular presence of the organism and de novo host protein synthesis are required for expression of cell adhesive molecules, since rickettsial inactivation by formol and pretreatment of cells with cycloheximide inhibits an increase in expression. The contribution of interleukin-1alpha (IL-1alpha) to this endothelial adhesive phenotype was shown by inhibitory experiments 8 and 24 h after infection with IL-1 receptor antagonist and IL-1alpha blocking antibodies. Enhanced adherence of mononuclear cells to infected endothelial cells involved VCAM-1- and ICAM-1-dependent mechanisms at the late phase of the inflammatory response. This endothelial adhesive phenotype may constitute a key pathophysiologic mechanism in R. conorii-induced vascular injury.     

Activation of OX40 signal transduction pathways leads to tumor necrosis factor receptor-associated factor (TRAF) 2- and TRAF5-mediated NF-kappaB activation

We investigated the intracellular signaling of OX40, a member of the tumor necrosis factor receptor family. Activation of NF-kappaB in OX40-transfected HSB-2 cells was detected by electrophoretic mobility shift assay within 30 min after the binding of the ligand gp34. In vitro binding experiments showed that tumor necrosis factor receptor-associated factor (TRAF) 1, TRAF2, TRAF3, and TRAF5 but not TRAF4 associated with glutathione S-transferase-OX40 fusion protein. The cotransfection experiments using human embryo kidney cell derived HEK 293T cells showed that TRAF2, TRAF3, and TRAF5 associated with OX40 in vivo. Studies with OX40 deletion mutants demonstrated that the cytoplasmic portion consisting of amino acid sequence 256-263 (GGSFRTPI) was required for the association with TRAFs and NF-kappaB activation. The introduction of the dominant negative mutants of TRAF2 and TRAF5 into HSB-2-OX40 cells suppressed NF-kappaB activation in a dose-dependent manner. In addition, the introduction of TRAF3 together with the dominant negative mutants of TRAF2 or TRAF5 further reduced NF-kappaB activation. These results indicate that the NF-kappaB activation resulting from OX40 stimulation is mediated by both TRAF2 and TRAF5, and is likely to be negatively modulated by TRAF3.     

Reduced susceptibility to collagen-induced arthritis in mice deficient in intercellular adhesion molecule-1

Intercellular adhesion molecule-1 (ICAM-1) plays an important role in the firm adhesion of leukocytes to venular endothelium and facilitates leukocyte extravasation from the vasculature into inflamed tissue. In addition, ICAM-1 is an important costimulatory molecule during Ag presentation to lymphocytes. Using mice deficient in ICAM-1, we have investigated the role of this molecule in the development of collagen-induced arthritis. After immunization with type II collagen, 71% of wild-type mice developed arthritis compared with 50% of ICAM-1 heterozygote mutants and 18% of ICAM-1 homozygous mutants. In those ICAM-1 mutants that developed arthritis, the mean day of onset, the mean number of involved paws, and the severity of paw inflammation were not significantly different from those in wild-type mice. The reduced incidence of arthritis in the ICAM-1 homozygous mutant mice was not due to lack of immunity to type II collagen, since these mice developed similar levels of anti-type II collagen IgG compared with wild-type mice and had a positive delayed-type hypersensitivity reaction to type II collagen. The reduction of arthritis in heterozygous as well as homozygous deficient mice indicates that expression of ICAM-1 can be a pivotal variable in the pathogenesis of collagen-induced arthritis in mice. The results suggest that naturally occurring genetic variation in the expression of ICAM-1 or related inflammatory cell adhesion molecules might influence susceptibility to the complex disease of rheumatoid arthritis in humans and support the concept that pharmacologic approaches to chronic reduction in the expression or the function of ICAM-1 may be of therapeutic value.     

Improved survival and amelioration of nephrotoxic nephritis in intercellular adhesion molecule-1 knockout mice

Intercellular adhesion molecule-1 (ICAM-1) expression is upregulated in nephrotoxic nephritis, a model of human rapidly progressive glomerulonephritis. To evaluate the pathogenetic relevance of ICAM-1 in this model, nephrotoxic nephritis was induced in ICAM-1 knockout mice and genetic controls. Mice were preimmunized with rabbit IgG in complete Freund's adjuvant. Seven days later they received rabbit anti-mouse glomerular basement membrane IgG. The early humoral immune responses (levels of circulating mouse anti-rabbit IgG, glomerular deposition of rabbit and mouse IgG and mouse C3c) were not altered in ICAM-1 knockout mice. During 28 d of follow-up, 3 of 19 control nephritic mice and 0 of 16 ICAM-1 knockout mice died. Proteinuria was high in nephritic control mice (means 10 to 12 mg/24 h at all time points investigated) and significantly reduced in nephritic ICAM-1 knockout mice (means <4.4 mg). Mean serum creatinine rose from 29 micromol/L at day -7 to 48 micromol/L (day 28) in nephritic control mice. This increase in serum creatinine was significantly lower in ICAM-1 knockout mice: 27 (day -7) and 36 micromol/L (day 28). Histologic analysis at day 28 revealed that ICAM-1 deficiency in nephrotoxic nephritis mice led to significantly reduced glomerular crescent formation (2+/-3% in ICAM-1 knockout mice versus 13+/-8% in nephritic controls) and tubulointerstitial injury (score 0.4+/-0.4 versus 2.0+/-1.1). By immunohistochemistry, ICAM-1 deficiency in nephritic mice led to significantly reduced (peri-)glomerular and/or interstitial macrophage influx, alpha-smooth muscle actin expression, and type IV collagen accumulation. These data indicate that ICAM-1 is a central mediator of glomerular and tubulointerstitial injury in murine nephrotoxic nephritis.     

Soluble intercellular adhesion molecule-1 in patients with lung cancer and benign lung diseases

Intercellular adhesion molecule-1 (ICAM-1) expression correlates with tumour progression in patients with malignant melanoma or renal cell carcinoma. To assess the value of soluble ICAM-1 (sICAM-1) for lung cancer patients, sICAM-1 was determined by means of an enzyme-linked immunosorbent assay. Sera from 147 patients with lung cancer, from 75 patients with benign lung diseases and from 108 healthy adults were investigated for sICAM-1 expression. Significant differences in sICAM-1 levels were detected in lung cancer patients (387 +/- 176 ng/ml) and patients with benign lung diseases (365 +/- 110 ng/ml) compared to the group of healthy adults (310 +/- 90 ng/ml). There was no difference in sICAM-1 level among the subtypes of lung cancer. Advanced tumour stages and patients with progressive disease tended to be associated with higher sICAM-1 levels, the site of metastasis being relevant for the level attained. Patients with liver metastasis had the highest sICAM-1 levels (547 +/- 295 ng/ml) compared to patients with cerebral metastasis (317.8 +/- 92.2 ng/ml). An increase of sICAM-1 expression during the progression of the disease coincided with a poorer survival prognosis for the patients compared to patients with stable or falling sICAM-1 levels.     

Talc-induced expression of C-C and C-X-C chemokines and intercellular adhesion molecule-1 in mesothelial cells

Treatment of symptomatic carcinomatous pleural effusions is primarily directed at local palliation with a wide variety of sclerosing agents, of which talc is considered to be the most successful. The mechanism whereby talc achieves this effect is unknown. The objective of this study was to investigate whether talc stimulates pleural mesothelial cells (PMC) to release C-X-C and/or C-C chemokines and express adhesion molecules that initiate and amplify the inflammatory process in the pleural space. When PMC were challenged with talc in vitro, interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) levels were increased (p < 0.001) both at the protein and the mRNA level as compared with unstimulated cultures. Talc-stimulated PMC culture supernatant showed chemotactic activity for neutrophils and monocytes. The chemotactic activity of PMC culture supernatant was blocked by 44.2% with IL-8-specific antibody and by 55.7% with MCP-1-specific antibody, demonstrating that PMC-derived chemokines are bioactive. Talc also enhanced intercellular adhesion molecule-1 (ICAM-1) expression in PMC. The data demonstrate that talc stimulates PMC to release chemokines and express adhesion molecules that may play a critical role in pleurodesis.     

Circulating soluble adhesion molecules E-cadherin, E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in patients with gastric cancer

The concentrations of the soluble adhesion molecules E-cadherin, E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were investigated in 45 patients with gastric cancer before treatment and their correlation with clinical, histological and routine laboratory parameters was examined. Data were collected on tumour stage at presentation, presence and sites of metastatic disease, tumour pathology, survival and results of routine laboratory tests. Serum concentrations of ICAM-1 and VCAM-1 were significantly elevated in the patients with gastric cancer in comparison with the group of healthy subjects (P < 0.00001 and P < 0.0001 respectively). Increased serum concentrations of VCAM-1 were associated with locally advanced and metastatic disease whereas ICAM-1 was significantly elevated both in local and in advanced/metastatic disease. Soluble E-cadherin and E-selectin concentrations did not show any significant elevation in gastric cancer patients. Concentrations of soluble adhesion molecules showed significant correlation with each other (except E-selectin and VCAM-1) and with alkaline phosphatase. Soluble ICAM-1 and VCAM-1 were significantly associated with an elevated total white cell count. Patients with elevated VCAM-1 had significantly poorer survival in comparison with patients with normal serum levels (P = 0.0361).