Soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1) Is Elevated in Bronchoalveolar Lavage Fluid of Patients with Acute Respiratory Distress Syndrome.

INTRODUCTION: Pulmonary vascular endothelial activation has been implicated in acute respiratory distress syndrome (ARDS), yet little is known about the presence and role of endothelial activation markers in the alveolar space in ARDS. We hypothesized that endothelial activation biomarkers would be differentially expressed in bronchoalveolar lavage fluid from patients with ARDS compared with healthy volunteers, and that biomarker concentrations would be associated with ARDS severity. METHODS: We performed a cross-sectional analysis of data from 26 intubated patients with ARDS undergoing evaluation for clinically suspected ventilator-associated pneumonia and five healthy volunteers. Patients underwent bronchoalveolar lavage a median of five days after intubation. Healthy volunteers also underwent bronchoalveolar lavage. Endothelial activation biomarkers (soluble vascular cell adhesion molecule-1 [sVCAM-1], soluble endothelial selectin [sESEL], angiopoietin-1 [Ang-1] and angiopoietin-2 [Ang-2]) were measured in bronchoalveolar lavage fluid. Clinically suspected ventilator-associated pneumonia was confirmed with microbiologic culture data. RESULTS: Patients with ARDS had significantly higher median sVCAM-1 concentrations in the bronchoalveolar lavage fluid compared with healthy volunteers (985 vs 119 pg/mL, p = 0.03). Additionally, there was a trend toward greater bronchoalveolar lavage fluid sVCAM-1 concentrations among patients with moderate/severe compared to mild ARDS (1395 vs 209 pg/mL, p = 0.06). We did not detect significant differences in bronchoalveolar lavage fluid levels of sESEL, Ang-1 or Ang-2 between patients with ARDS and healthy volunteers. Median bronchoalveolar lavage fluid biomarker levels did not differ between patients with and without microbiologically-confirmed ventilator-associated pneumonia. CONCLUSIONS: sVCAM-1 concentrations were significantly higher in the bronchoalveolar lavage fluid of patients with ARDS compared to healthy controls, and tended to be higher in moderate/severe ARDS compared to mild ARDS. Our findings add to the growing evidence supporting the concept that endothelial activation plays an important mechanistic role in the pathogenesis of ARDS. Further studies are necessary to characterize the role and/or clinical significance of sVCAM-1 and other endothelial activation markers present in the alveolar space in ARDS.

Markers of mechanical asphyxia: immunohistochemical study on autoptic lung tissues.

Forensic pathologists are often asked to provide evidence of asphyxia death in the trial and a histological marker of asphyxiation would be of great help. Data from the literature indicate that the reaction of lung tissue cells to asphyxia may be of more interest for forensic purposes than migrating cells. The lungs of 62 medico-legal autopsy cases, 34 acute mechanical asphyxia (AMA), and 28 control cases (CC), were immunostained with anti-P-selectin, anti-E-selectin, anti-SP-A, and anti-HIF1-α antibodies, in order to verify if some of them may be used as markers of asphyxia death. Results show that P- and E-selectins expression in lung vessels, being activated by several types of trigger stimuli not specific to hypoxia, cannot be used as indicator of asphyxia. Intra-alveolar granular deposits of SP-A seem to be related to an intense hypoxic stimulus, and when massively present, they can suggest, together with other elements, a severe hypoxia as the mechanism of death. HIF1-α was expressed in small-, medium-, and large-caliber lung vessels of the vast majority of mechanical asphyxia deaths and CO intoxications, with the number and intensity of positive-stained vessels increasing with the duration of the hypoxia. Although further confirmation studies are required, these preliminary data indicate an interesting potential utility of HIF1-α as a screening test for asphyxia deaths.

Serum inflammatory markers in obstructive sleep apnea: a meta-analysis.

STUDY OBJECTIVES: Obstructive sleep apnea (OSA) has been linked to and is associated with increased cardiovascular and cerebrovascular morbidity. Ongoing inflammatory responses play an important role in this association. Multiple small size studies addressing the profile of the inflammatory markers in OSA are available therefore we performed a meta-analysis. METHODS: Systematic review of medical literature was conducted using PubMed, Cochrane, and EMBASE databases from 1968 to 2011 by utilizing the key words obstructive sleep apnea, C-Reactive protein, tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), interleukin 8 (IL-8), intercellular adhesion molecule (ICAM), vascular cell adhesion molecule (VCAM) and Selectins. Inclusion criteria were: full text English articles; studies with adult population; reported values for at least one of the markers of interest; with at least two separate groups (subjects with OSA and control group); OSA was defined as AHI of ≥ 5/h. RESULTS: Five hundred and twelve studies were reviewed for inclusion with 51 studies pooled for analysis (30 studies for CRP, 19 studies for TNF-α, 8 studies for ICAM, 18 studies for IL-6, six studies for VCAM and 5 studies for Selectins). The levels of inflammatory markers were higher in patients with OSA compared to control group. Standardized pooled Mean differences were calculated to be 1.77 for CRP, 1.03 for TNF-α, 2.16 for IL-6, 4.22 for IL-8, 2.93 for ICAM, 1.45 for Selectins and 2.08 for VCAM. CONCLUSIONS: In this meta-analysis, the levels of systemic inflammatory markers were found to be higher in OSA patients compared to control subjects.

Target gene expression signatures in neutrophils and lymphocytes from cattle administered with dexamethasone at growth promoting purposes.

The glucocorticoid dexamethasone (DEX), when used as a growth promoter, cause morphological and functional alterations in cattle lymphoid organs and cells. In the present experiment, the transcriptional effects of an illicit DEX protocol upon six target genes were investigated in cattle neutrophils (NEU) and lymphocytes (LFC). Blood samples were taken before (T(0)) and 2, 3, 10, 19, 31 and 43 days from the beginning of DEX administration (T(1)-T(6)). Leukocytes were counted and cells isolated by gradient centrifugation; then, glutathione peroxidase 1 and 3 (GPX1 and GPX3), glucocorticoid receptor alpha (GRα), l-selectin, nuclear factor κB, subunit p65 (NFκB) and tumor necrosis factor alpha (TNFα) mRNA amounts were measured through a quantitative Real Time RT-PCR approach. A significant change vs controls in NEU/LFC ratio was noticed from T(3) forward. Compared to T(0), DEX significantly increased to a variable extent all candidate gene mRNAs abundances in NEU; in contrast, only l-selectin, GRα and GPX1 were significantly up-regulated in LFC. Present results suggest that illicit DEX affects transcription in cattle immune cells, that might be considered as a promising surrogate tissue for the screening of DEX abuse in cattle farming.

Alteraciones inmunológicas y de los valores de selectinas en pacientes hipertensos tratados según criterios actuales / Immunological alterations and selectin values in hypertensive patients treated according to current criteri

Fundamento y objetivo: Calcular la relación entre las selectinas P y E, y los cambios inmunológicos que se producen en la hipertensión esencial. Pacientes y método:Se incluyeron 261 pacientes con hipertensión esencial con y sin hipertrofia del ventrículo izquierdo (HVI), a los que se les determinó las concentraciones de selectinas y citocinas específicas. Resultados:Los valores de selectina-P y E estuvieron elevados respecto al grupo control (media [DE] de 75 [51] ng/mL frente a 45 [17] ng/mL, p<0,0001, y 47 [18] ng/mL frente a 38 [15] ng/mL, p=0,049), así como las concentraciones de citocinas (p<0,0001). Además, las selectinas se relacionaron significativamente con los cambios en marcadores inmunológicos. El análisis multivariado mostró que la citocina interleucina (IL) 1ra (p<0,0001) y selectina-P (p=0,013) fueron factores independientes de los valores de selectina-E. Para selectina-P, lo fueron IL-1ra (p=0,005), IL-8 (p<0,0001), y selectina-E (p=0,013). Por último, las concentraciones de selectinas no fueron diferentes entre pacientes con y sin hipertrofia ventricular. Conclusiones: La activación inmunológica influye en los valores de selectinas en pacientes asintomáticos con hipertensión esencial. La HVI no indujo cambios significativos (AU)

Background and objective: To calculate the relationsship of E and P selectins with immunological changes in essential hypertension.Patients and method: We included 261 patients with essential hypertension with or without left ventricular hypertrophy (LVH), in whom selectins and specific cytokines were quantified. Results:E-and P-selectins were increased in hypertension compared to controls [75(51) ng/mL versus 45(17) ng/mL, (p<0.0001) and 47(18) ng/mL versus 38(15) ng/mL, (p=0.049), respectively], as well as cytokine concentrations (p<0.0001). Furthermore, selectin values were significantly correlated with changes in immunological markers. The multivariate analysis showed that IL-1ra (p<0.0001) and P-selectin (p=0.013) were independent factors of E-selectin values. For P-selectin, IL-1ra (p=0.005), IL-8 (p<0.0001) and selectin-E (p=0.013) influenced on its concentration. Finally, selectin values were similar between patients with or without left ventricular hypertrophy. Conclusions: Immunological activation influenced on selectin concentrations in cardiologic asymptomatic patients with essential hypertension. LVH did not induce significant changes (AU)

Soluble P selectin in synovial fluid level is correlated with the radiographic severity of knee osteoarthritis.

BACKGROUND: Recent studies provide evidence that inflammation is a feature of the disease process in Osteoarthritis (OA). The clinical significance of P selectin (Ps) in OA has not been adequately studied and the association between Ps level and OA severity remains unknown. METHODS: We enrolled 120 knee OA subjects and 45 controls. All patients were scored for Kellgren-Lawrence grade (0-4). The Ps in serum and synovial fluid (SF) as well as serum C-reactive protein (CRP) levels were detected. RESULTS: The mean Ps level in OA subjects was markedly increased than that in controls. In OA patients, the SF Ps levels increased with the severity of KL scores and significantly correlated with severity of disease (r=0.546, P<0.001) and serum CRP level (r=0.488, P<0.001). However, the serum Ps level did not show a significant correlation with the severity of OA. CONCLUSION: The Ps levels in SF were significantly correlated with the severity of OA, suggesting that it may be used as a biomarker to evaluate the progression of OA.

CD19 regulates the development of bleomycin-induced pulmonary fibrosis in a mouse model.

OBJECTIVE: The contribution of CD19 and B lymphocytes to pulmonary fibrosis is controversial. The aim of this study was to address the role of CD19 during the development of pulmonary fibrosis. METHODS: Mice lacking or overexpressing the B cell surface molecule CD19, which is known as a positive regulator of B cell activation, were used in a model of bleomycin-induced pulmonary fibrosis. Ten or sixteen days after intratracheal injection of bleomycin, lung sections from mice were evaluated by histologic analysis. Seven days after instillation, the total leukocyte count and the number of B cells in bronchoalveolar lavage fluid (BALF) were determined, using a hemocytometer and flow cytometry. Bleomycin was also administered into selectin-deficient or intercellular adhesion molecule 1-deficient mouse strains. The level of CXCR3 expression on B cells was determined by flow cytometry. RESULTS: CD19 deficiency significantly reduced susceptibility to intratracheal bleomycin challenge on day 16, while CD19 overexpression augmented fibrosis even on day 10. Furthermore, the survival rate and number of B cells in BALF also correlated with CD19 expression levels. The accumulation of B cells in BALF was dependent on CD19 levels, whereas there was no association with the levels of selectins or intercellular adhesion molecule 1. Additionally, CXCR3 was up-regulated in BALF B cells, while it was rarely expressed on circulating B cells. Furthermore, CD19 signaling facilitated B cell CXCR3 up-regulation in response to stimulation in vitro. CONCLUSION: These results suggest that CD19 signaling is associated with the development of pulmonary fibrosis by controlling B cell infiltration into lung tissue, which may be associated with CXCR3 up-regulation.

Cell adhesion molecules: potential therapeutic & diagnostic implications.

The role of cell adhesion molecules (CAM) and extracellular matrix proteins (ECM) in various pathological processes including angiogenesis, thrombosis, apoptosis, cell migration & proliferation are well documented. These processes can lead to both acute and chronic disease states such as ocular diseases, metastasis, unstable angina, myocardial infarction, stroke, osteoporosis, a wide range of inflammatory diseases, vascular remodeling, and neurodegenerative disorders. A key success in this field is evident from the potential role of the platelet GPIIb/IIIa integrin in the prevention and diagnosis of various thromboembolic disorders. Additionally, the use of soluble adhesion molecules as potential diagnostic markers for acute and chronic leukocyte, platelet, and endothelial cellular insult are increasingly utilized. The development of various therapeutic and diagnostic candidates based on the key role of CAM, with special emphasis on integrins in various diseases as well as the structure-function aspects of cell adhesion and signaling of the different CAM and ECM are highlighted.

Association of classical and related inflammatory markers with high-sensitivity C-reactive protein in healthy individuals: results from the Stanislas cohort.

BACKGROUND: Although high-sensitivity C-reactive protein (hs-CRP) has emerged as a cardiovascular marker, questions arise regarding the relative information provided by other inflammatory molecules. Therefore, as a first step, we examined interrelationships between serum hs-CRP concentrations and inflammatory, adhesion and growth factors in healthy adults. METHODS: Circulating concentrations of hs-CRP, haptoglobin, orosomucoid, interleukin-6 (IL-6), IL-8, IL-18, tumor necrosis factor-alpha (TNF-alpha), TNF-receptor II (TNF-RII), E-, P-, and L-selectins, intercellular adhesion molecule-1 (ICAM-1), monocyte chemoattractant protein-1, endothelial growth factor (EGF), vascular EGF (VEGF), insulin-like growth factor-1 (IGF-1) and IGF-binding protein (IGFBP-3) were measured in 154 men and 161 women of the Stanislas cohort. Leukocyte and platelet counts were also determined. RESULTS: Correlations were significant between hs-CRP concentrations and leukocyte and platelet counts, as well as haptoglobin, orosomucoid, IL-6, and ICAM-1 concentrations (p< or =0.001). Correlation coefficients for ICAM-1 were higher in men than in women (p< or =0.05). When stratifying subjects according to hs-CRP levels, the group with high hs-CRP levels had significantly higher haptoglobin and orosomucoid concentrations than the others, in addition to higher leukocyte counts and IL-6 concentrations in women, and platelet counts and ICAM-1 concentrations in men. CONCLUSIONS: Further studies are warranted to explain the association pattern for hs-CRP. Partition of these factors according to their association with hs-CRP concentration opens a new perspective for choice of the best factors in terms of cardiovascular risk in relation to hs-CRP, while non-associated markers could be used to give additional information.

Mathematical modeling of cell adhesion in shear flow: application to targeted drug delivery in inflammation and cancer metastasis.

Cell adhesion plays a pivotal role in diverse biological processes that occur in the dynamic setting of the vasculature, including inflammation and cancer metastasis. Although complex, the naturally occurring processes that have evolved to allow for cell adhesion in the vasculature can be exploited to direct drug carriers to targeted cells and tissues. Fluid (blood) flow influences cell adhesion at the mesoscale by affecting the mechanical response of cell membrane, the intercellular contact area and collisional frequency, and at the nanoscale level by modulating the kinetics and mechanics of receptor-ligand interactions. Consequently, elucidating the molecular and biophysical nature of cell adhesion requires a multidisciplinary approach involving the synthesis of fundamentals from hydrodynamic flow, molecular kinetics and cell mechanics with biochemistry/molecular cell biology. To date, significant advances have been made in the identification and characterization of the critical cell adhesion molecules involved in inflammatory disorders, and, to a lesser degree, in cancer metastasis. Experimental work at the nanoscale level to determine the lifetime, interaction distance and strain responses of adhesion receptor-ligand bonds has been spurred by the advent of atomic force microscopy and biomolecular force probes, although our current knowledge in this area is far from complete. Micropipette aspiration assays along with theoretical frameworks have provided vital information on cell mechanics. Progress in each of the aforementioned research areas is key to the development of mathematical models of cell adhesion that incorporate the appropriate biological, kinetic and mechanical parameters that would lead to reliable qualitative and quantitative predictions. These multiscale mathematical models can be employed to predict optimal drug carrier-cell binding through isolated parameter studies and engineering optimization schemes, which will be essential for developing effective drug carriers for delivery of therapeutic agents to afflicted sites of the host.

Quantitative measurement of selectin-ligand interactions: assays to identify a sweet pill in a library of carbohydrates.

Soluble oligosaccharides and glycoproteins can inhibit leukocyte adhesion during a range of vascular ailments including inflammation, thrombosis, and cancer metastasis. The design of such molecules in many cases is based on the structure of naturally occurring selectin ligands. In this case, synthetic selectin-ligand mimetics act as competitive inhibitors of cell adhesion. In an alternate approach, cell-permeable, small-molecule oligosaccharides have been shown to alter the metabolic pathways that lead to the biosynthesis of functional selectin-ligands. The addition of such molecules results in glycoproteins that are defective in their ability to bind selectins. Quantitative in vitro testing of the efficacy of the above inhibition strategies ideally requires the application of assays that mimic the in vivo physiological milieu in terms of the valency of selectin and selectin-ligands, the physiological fluid-flow conditions, and the use of blood cells. Assays that are performed in small volumes are preferable when the quantity of available inhibitor is scarce. Finally, the measurements must account for the rapid on- and off-rates of selectin-mediated binding interactions. This chapter addresses these issues by presenting methods to measure selectin function in enzyme-linked immunosorbent assay and flow cytometry-based static assays, cell-adhesion assays performed under shear flow in cone-plate viscometers, and Biacore surface plasmon resonance measurements of molecular-binding kinetics. Examples are presented where such methods are applied to measure the ability of simple oligosaccharides based on sialyl Lewis-X and complex molecules with the core-2 structure to block selectin function. Such methods may be extended to identify potent selectin antagonists in a library of carbohydrates.

Imaging molecular expression on vascular endothelial cells by in vivo immunofluorescence microscopy.

Molecular expression on the vascular endothelium is critical in regulating the interaction of circulating cells with the blood vessel wall. Leukocytes as well as circulating cancer cells gain entry into tissue by interacting with adhesion molecules on the endothelial cells (EC). Molecular targets on the EC are increasingly being explored for tissue-specific delivery of therapeutic and imaging agents. Here we use in vivo immunofluorescence microscopy to visualize the endothelial molecular expression in the vasculature of live animals. High-resolution images are obtained by optical sectioning through the intact skin using in vivo confocal and multiphoton microscopy after in situ labeling of EC surface markers with fluorescent antibodies. Other vascular beds such as the bone marrow and ocular blood vessels can be imaged with little or no tissue manipulation. Live imaging is particularly useful for following the dynamic expression of inducible molecules such as E-selectin during an inflammatory response.

Expression of cell adhesion molecules in oesophageal carcinoma and its prognostic value.

Oesophageal carcinoma remains a disease of poor prognosis. Surgical cure rates are compromised by the fact that most patients are diagnosed at a late stage of disease because of the delayed onset of symptoms, by which time metastases and organ infiltration may have already occurred. Thus, invasion and metastases play a key role in influencing patient survival, and the search for novel treatments may therefore hinge on gaining insight into the mechanisms controlling these processes. It has been established that the initial step in the metastatic cascade is the detachment of tumour cells from the primary tumour via dysregulation of normal cell-cell and cell-matrix interactions. Distinct proteins known as cell adhesion molecules (CAMs) mediate these interactions. In recent years, a plethora of information has contributed to the in depth understanding of these molecules. This review provides a brief description of five families of CAMs (cadherins, integrins, CD44, immunoglobulin superfamily, and selectins) and highlights their altered expression in relation both to prognosis and tumour behaviour in squamous cell carcinoma and adenocarcinoma of the oesophagus.

Markers of low-grade inflammation and soluble cell adhesion molecules in Chinese patients with coronary artery disease.

BACKGROUND: Inflammation plays an important role in atherosclerosis. Markers of low-grade chronic inflammation, such as C-reactive protein (CRP) and soluble cell adhesion molecules (sCAMs), have been associated with coronary artery disease (CAD). OBJECTIVE: To evaluate the significance of inflammatory markers as novel risk factors for CAD in the Chinese population. METHODS: High-sensitivity CRP (hs-CRP); sCAMs, including vascular cell adhesion molecule-1 (sVCAM-1), intercellular cell adhesion molecule-1 (sICAM-1), P-selectin (sP-selectin) and E-selectin (sE-selectin); and white blood cell (WBC) count were measured in 170 angiographically defined CAD patients (70% or greater stenosis affecting at least one vessel) and 177 healthy control subjects in the Chinese population in Singapore. RESULTS: The levels of hs-CRP, sVCAM-1 and sP-selectin, and the WBC count were higher in CAD patients than in control subjects (P<0.001, P<0.05, P<0.05 and P<0.001, respectively). There were no significant differences in the levels of sICAM-1 and sE-selectin between the two groups. Patients with unstable angina or myocardial infarction had higher levels of hs-CRP, and higher WBC and monocyte counts than those with stable angina or atypical chest pain (all P<0.05). The level of sP-selectin in patients with multivessel disease was higher than in those with single-vessel disease (P<0.05). Overall, the levels of hs-CRP and sCAMs showed a significant correlation with the lipid profile and the WBC count. CONCLUSIONS: The present study suggests that inflammatory markers, including hs-CRP and WBC count, together with sP-selectin and sVCAM-1, could serve as markers of atherogenesis in Chinese patients with CAD, with potential diagnostic and therapeutic implications.

Cardiopulmonary bypass induces significant platelet activation in children undergoing open-heart surgery.

BACKGROUND AND OBJECTIVES: To evaluate the effects of cardiopulmonary bypass (CPB) on platelet function in children undergoing open-heart surgery. METHODS: Data from 16 consecutive children undergoing cardiac surgery with CPB were prospectively collected. Blood samples of 10 mL were collected via the central venous line immediately before and after CPB for CD62 measurements by flow cytometry. RESULTS: Ten children had acyanotic heart disease (median age 3 yr, range 1.8-14) and six had a cyanotic defect (median age 4yr, range 2-14). The platelet count decreased significantly with CPB in both groups: from 163.5 (130-201) to 93.5 (57-186) x 10(3) microL(-1) in acyanotic children and from 139.5 (77-212) to 75 (43-99) x 10(3) microL(-1) in cyanotic children (P < 0.0001). The percentage of activated platelets was significantly lower in acyanotic children at baseline: 1% (0-23%) vs. 5% (3-8%) (P = 0.07). CPB increased the percentage of activated platelets significantly in both groups: post-bypass the values were 10% (range 1-17%) in acyanotic children and 7% (range 1-30%) in cyanotic children (P = 0.03). The increase in the percentage of activated platelets did not differ between the two study groups (P = 0.11). CONCLUSION: CPB induces significant platelet activation in children undergoing open-heart surgery.

Disruption of L-histidine decarboxylase reduces airway eosinophilia but not hyperresponsiveness.

Histamine has a variety of airway actions and is considered to be an important mediator in asthma. This study examined the role of endogenous histamine in allergic airway eosinophil recruitment and hyperresponsiveness using L-histidine decarboxylase gene knockout mice. Histamine levels of the airways in L-histidine decarboxylase knockout mice were largely diminished compared with wild-type mice. Inhalation challenge with ovalbumin (OVA) in OVA-sensitized wild-type mice caused eosinophil accumulation in the lung as well as airway hyperresponsiveness to methacholine 3 days after the challenge. The eosinophil recruitment was significantly reduced in the knockout mice. In the bone marrow, the proliferation of eosinophils was enhanced after OVA challenge in the wild-type mice; however, the proliferation was significantly reduced in the knockout mice. The induction of P-selectin in the lung after OVA challenge was also inhibited in the knockout mice. In contrast, airway hyperresponsiveness was not suppressed in the knockout mice. These results suggest that endogenous histamine is involved in the accumulation of eosinophils into the airways after allergic challenge, possibly acting in the bone marrow and producing P-selectin in the airways. Furthermore, allergen-induced airway hyperresponsiveness appeared to occur independently of airway eosinophilia in our present model.

Leukocyte count and vascular function in Type 2 diabetic subjects with treated hypertension.

BACKGROUND: Traditional cardiovascular risk factors may only partially explain abnormal vascular function in Type 2 diabetic patients. This study examined the associations between vascular function and markers of inflammation in Type 2 diabetic subjects with treated hypertension. METHODS: Flow-mediated dilatation (FMD) and glyceryl-trinitrate mediated dilatation (GTNMD) of the brachial artery were used to assess endothelium-dependent and -independent function, respectively, in 29 hypertensive Type 2 diabetic subjects (HbA1c <9%), and 17 healthy control subjects. Plasma C-reactive protein (CRP), fibrinogen, interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and leukocyte count were used as markers of inflammation. Soluble L-selectin, P-selectin, and von Willebrand factor (vWf) were measured to assess leukocyte, platelet and endothelial cell activation, respectively. RESULTS: Compared with controls, diabetic subjects had impaired FMD (3.9+/-3.0 vs. 5.5+/-2.4%, P=0.07) and GTNMD (11.4+/-4.8% vs. 15.4+/-7.1%, P=0.04). They also had higher levels of CRP (2.7+/-2.6 vs. 1.4+/-1.1 mg/l, P=0.03), fibrinogen (3.4+/-0.7 vs. 2.7+/-0.3 g/l, P<0.001) and TNF-alpha (20.9+/-13.4 vs. 2.5+/-1.7 pg/l, P<0.001). In diabetic subjects, after adjustment for age and gender, leukocyte count was an independent predictor of FMD (P=0.02), accounting for 17% of total variance. Similarly, leukocyte count (P<0.001) accounted for 23% and IL-6 (P=0.03) for 12% of the variance in GTNMD. vWf was correlated with leukocyte count (r=0.38, P=0.04), FMD (r=-0.35, P=0.06) and GTNMD (r=-0.47, P=0.009), whilst P-selectin correlated with fibrinogen (r=0.58, P=0.001). CONCLUSION: These cross-sectional observations are consistent with the hypothesis that reduced FMD and GTNMD in Type 2 diabetes is at least in part secondary to increased inflammation, with associated endothelial and platelet activation.

Examination of several potential mechanisms for the negative outcome in a clinical stroke trial of enlimomab, a murine anti-human intercellular adhesion molecule-1 antibody: a bedside-to-bench study.

BACKGROUND AND PURPOSE: Enlimomab, a murine monoclonal anti-human intercellular adhesion molecule (ICAM)-1 antibody, had a negative outcome in a multicenter acute-stroke trial. We did a bedside-to-bench study in standardized rat stroke models to explore mechanisms for these untoward results. METHODS: After focal brain ischemia in Wistar rats and spontaneously hypertensive rats (SHR), we administered murine anti-rat ICAM-1 antibody (1A29), subclass-matched murine immunoglobulin (IgG1), or vehicle intravenously. To examine whether rat anti-mouse antibodies were generated against the mouse protein and whether these were deleterious, we sensitized Wistar rats with 1A29 or vehicle 7 days before surgery. Infarct volume, tissue myeloperoxidase activity, neutrophil CD11b expression, and microvascular E-selectin, P-selectin, and ICAM-1 expression were examined 48 hours after surgery. Complement activation was serially assessed for 2 hours after a single injection of either 1A29 or vehicle. RESULTS: 1A29 treatment did not significantly reduce infarct size in either strain. 1A29 sensitization augmented infarct size and generated rat anti-mouse antibodies. Although 1A29 inhibited neutrophil trafficking shown by reduction in brain myeloperoxidase activity, circulating neutrophils were activated and displayed CD11b upregulation. Complement was activated in 1A29-sensitized Wistar rats and 1A29-treated SHR. E-selectin (SHR), endothelial P-selectin (Wistar and SHR), and ICAM-1 (SHR) were upregulated in animals treated with 1A29. CONCLUSIONS: Administration to rats of a murine antibody preparation against ICAM-1, 1A29, elicits the production of host antibodies against the protein, activation of circulating neutrophils, complement activation, and sustained microvascular activation. These observations provide several possible mechanisms for central nervous system-related clinical deterioration that occurred when Enlimomab was given in acute ischemic stroke.

Cyclosporine is associated with endothelial dysfunction but not with platelet activation in renal transplantation.

BACKGROUND: Cyclosporine (CsA) is associated with thrombotic micro-angiopathy and endothelial dysfunction. Markers of endothelial dysfunction may serve to identify patients at risk for development of vascular injury. In this study we measured von Willebrand Factor (vWF) and sP-selectin as possible markers for endothelial dysfunction in renal transplant recipients at different concentrations of CsA. Because sP-selectin can also be derived from platelets an additional in vitro study was performed to study the potential effect of CsA on the expression of P-selectin on platelet surface, while the effects of CsA on the interaction of platelets with Endothelial Cell Matrix (ECM) were studied under flow conditions in a perfusion chamber model. METHODS: CsA was stepwisely replaced by mycophenolate mofetil (MMF) in 15 renal transplant recipients (more than 6 months after transplantation). VWF and sP-selectin were measured at normal CsA (median trough level 130 microg/l), low CsA (trough level 45 microg/l) and after stopping CsA. MMF 2 g daily was added while lowering and stopping CsA. Platelet activation was investigated by measurement of P-selectin on platelet surface by flow-cytometry (FACS), after incubation with CsA (0, 2, 20 and 200 mg/l) in vitro and after perfusion of whole blood over ECM with CsA (0 or 2 mg/l, peak levels). RESULTS: Stepwise withdrawal of CsA gave a dose-related decrease of both vWF and sP-selectin, suggesting reversible endothelial dysfunction. FACS showed no expression of P-selectin on platelets by CsA. Also perfusion studies over ECM demonstrated no platelet activation by CsA but even inhibition of platelet adhesion and aggregation. CONCLUSIONS: The use of CsA is not accompanied by platelet activation. However endothelial dysfunction induced by CsA does occur as reflected by increased vWF and sP-selectin. (See Editorial p. 1).