Leukocyte Adhesion Molecules Research Articles

Endothelium-derived markers and antioxidant status in the blood of obstructive sleep apnea males

ObjectiveThe relationship between obstructive sleep apnea (OSA) and cardiovascular disease is intensively discussed. Endothelial leukocyte adhesion molecule (E-selectin) is one of factors facilitating leukocyte migration to the subendothelial layer which could be considered proatherogenic. The aim of the study was to determine E-selectin levels and total plasma antioxidant status (TAS) in the blood of different stage OSA patients.MethodsNon-smoking, OSA-suspected males, aged 30-63, were selected for the study. An EMBLA polysomnographic system was used to establish the severity of apneic episodes. The results of apnea/hypopnea index (AHI) allowed dividing patients into the following groups: OSA-0 with AHI 0-4.9 (n = 14), OSA-1 with AHI 5-15 (n = 14), OSA-2 with AHI 16-30 (n = 13), OSA-3 with AHI ≥ 30 (n = 13). Complete blood count (CBC), glycemia during oral glucose tolerance test, fasting plasma lipid profile, uric acid, and high sensitivity C-reactive protein (hsCRP) were estimated among routine parameters. We determined plasma concentrations of E-selectin and total antioxidant status.ResultsWe found progressively decreasing concentrations of TAS (P = 0.03) and increased concentrations of E-selectin (P = 0.0001) from OSA-0 to OSA-3 subjects. No correlation between E-selectin and metabolic parameters was noted.ConclusionIn the studied OSA groups, E-selectin appeared an independent proatherogenic factor.

The role of leukocyte adhesion molecules in the development of severe sepsis

The role of leukocyte adhesion molecules in the development of severe sepsis

Heparan sulfate proteoglycans in extravasation: assisting leukocyte guidance

Heparan sulfate proteoglycans (HSPGs) are glycoconjugates that are implicated in various biological processes including development, inflammation and repair, which is based on their capacity to bind and present several proteins via their carbohydrate side chains (glycosaminoglycans; GAGs). Well-known HSPGs include the family of syndecans and glypicans, which are expressed on the plasma membrane and perlecan, agrin and collagen type XVIII, which are present in basement membranes. In this review, we provide an overview of the current knowledge on the role and regulation of HSPGs in leukocyte extravasation. In the non-inflamed endothelial glycocalyx HSPGs are anti-adhesive, and there are several indications that active regulation of HSPG core protein expression and/or GAG modification occurs upon inflammation. We address the current evidence for the role of HSPGs in leukocyte extravasation through interaction with the leukocyte adhesion molecule L-selectin, chemokines and other binding partners. Finally, a number of possibilities to use HSPGs as therapeutics or targets in anti-inflammatory strategies are discussed.

Comparative study of SPR and ELISA methods based on analysis of CD166/ALCAM levels in cancer and control human sera

Surface plasmon resonance (SPR), as a label free method for analysis of various analytes, has significantly advanced in recent years. However, assessment of the performance of SPR compared to label-based immunoassays such as the commonly used multiplexed enzyme-linked immunosorbent assay (ELISA) is limited, particularly for applications involving complex media. In this work, an optimized SPR assay was implemented and its performance compared with an ELISA assay for CD166/activated cell leukocyte adhesion molecule (ALCAM), as candidate pancreatic cancer marker, based on direct and amplified detection in buffer and in human serum samples from healthy individuals and subjects with cancer. ALCAM antibody was immobilized on the surface of a four-channel SPR sensor via physical adsorption onto charged amine-terminated alkanethiolates to mimic the ELISA plate surface. Excellent correlations between SPR and ELISA results were achieved in buffer and in human serum. SPR detected the target protein with a similar sensitivity to sandwich ELISA, with a detection limit below ng/mL. The detection time, sample consumption, throughput, signal referencing, and surface blocking and washing for detection in human serum were evaluated. It is demonstrated that SPR can distinguish between ALCAM levels in cancer and control sera using direct detection without the need for additional amplification steps.

Monocyte chemoattractant protein-1 in the choroid plexus: A potential link between vascular pro-inflammatory mediators and the CNS during peripheral tissue inflammation

During peripheral tissue inflammation, inflammatory processes in the CNS can be initiated by blood-borne pro-inflammatory mediators. The choroid plexus, the site of cerebrospinal fluid (CSF) production, is a highly specialized interface between the vascular system and CNS, and thus, this structure may be an important element in communication between the vascular compartment and the CNS during peripheral tissue inflammation. We investigated the potential participation of the choroid plexus in this process during peripheral tissue inflammation by examining expression of the small inducible cytokine A2 (SCYA2) gene which codes for monocyte chemoattractant protein-1 (MCP-1). MCP-1 protein was previously reported to be induced in a variety of cells during peripheral tissue inflammation. In the basal state, SCYA2 is highly expressed in the choroid plexus as compared with other rat CNS tissues. During hind paw inflammation, SCYA2 expression was significantly elevated in choroid plexus, whereas it remained unchanged in a variety of brain regions. The SCYA2-expressing cells were strongly associated with the choroid plexus as vascular depletion of blood cells by whole-body saline flush did not significantly alter SCYA2 expression in the choroid plexus. In situ hybridization suggested that the SCYA2-expressing cells were localized to the choroid plexus stroma. To elucidate potential molecular mechanisms of SCYA2 increase, we examined genes in the nuclear factor-kappa B (NF-κB) signaling cascade including tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β) and inhibitor of kappa B alpha (IκBα) in choroid tissue. Given that we also detected increased levels of MCP-1 protein by ELISA, we sought to identify potential downstream targets of MCP-1 and observed altered expression levels of mRNAs encoding tight junction proteins TJP2 and claudin 5. Finally, we detected a substantial up-regulation of the transcript encoding endothelial leukocyte adhesion molecule 1 (E-selectin), a molecule which could participate in leukocyte recruitment to the choroid plexus along with MCP-1. Together, these results suggest that profound changes occur in the choroid plexus during peripheral tissue inflammation, likely initiated by blood-borne inflammatory mediators, which may modify events in CNS.

Inflammatory Cytokines in Acute Ischemic Stroke

Three major cytokines, namely, tumor necrosis factor (TNF-alpha), interleukin (IL)-1, and IL-6 are produced by cultured brain cells after various stimuli such as ischemia. Neurones, astrocytes, microglia and oligodendrocytes can produce inflammatory mediators, and cytokine receptors are expressed constitutionally throughout the Central Nervous System (CNS), albeit at low levels. Cytokines are involved in virtually every facet of stroke and they have numerous pro-inflammatory and pro-coagulant effects on endothelium. TNF-alpha expression after stroke stimulates expression of tissue factor and adhesion molecules for leukocytes, release of interleukin-1 (IL-1), nitric oxide, factor VIII/von Willebrand factor, platelet-activating factor and endothelin, suppression of the thrombomodulin-protein C-protein S system, reduction of tissue-plasminogen activator and release of plasminogen activator inhibitor-1. Research into the actions of IL-1beta in the brain initially focused on its role in host defence responses to systemic disease. IL-1beta can also elicit an array of responses which could either inhibit, exacerbate or induce neuronal damage and death. IL-6 can be induced by a variety of molecules including IL-1, TNF-alpha, transforming growth factor-beta and prostaglandins (PGs), and many other mediators such as b-amyloid, interferon-g (IFNg) and IL-4 can potentiate these primary inducers, highlighting the complex nature of IL-6 modulation. Several studies reported that plasma levels of TNF-alpha and IL-6 are associated with prognosis after ischemic stroke and our group showed that plasma levels of cytokines such as TNF-alpha, IL-1beta are different in every diagnostic subtype of ischemic stroke, and how plasma levels of some immunoinflammatory markers and thrombotic-phybrinolitic markers are predictive of acute ischemic stroke diagnosis in the acute setting.

Multimodality Cardiovascular Molecular Imaging, Part I

In Part I of this consensus article, the imaging methodology, evolving imaging technology, and development of novel targeted molecular probes relevant to the developing field of cardiovascular molecular imaging were reviewed. Novel reporter gene and reporter probe imaging approaches for tracking of cardiac transgene expression were also discussed and have important future implications for evaluation of gene- and cell-based therapies for the failing heart. The current role of metabolic and receptor imaging was also briefly reviewed, as these represent the beginning of our clinical application of molecular imaging within the cardiovascular system. Part II will summarize the available targeted imaging probes as well as specific future applications of molecular imaging for identification and evaluation of critical pathophysiological processes of the cardiovascular system.

Effects of parenteral structured lipid emulsion on modulating the inflammatory response in rats undergoing a total gastrectomy

Objectives Structured lipid emulsion improves the nitrogen balance and is rapidly cleared from the blood of moderately catabolic patients. However, the effects of structured lipids on inflammatory reactions during major surgery are not clear. This study investigated the effect of a parenteral structured triacylglycerol emulsion on leukocyte adhesion molecule expression and inflammatory mediator production in rats undergoing a total gastrectomy. Methods Normal rats with internal jugular catheters were assigned to three experimental groups and received total parenteral nutrition. At the same time, a total gastrectomy was performed on the experimental groups. The total parenteral nutrition solutions were isonitrogenous and identical in nutrient compositions except for differences in the composition of the fat emulsion. Group 1 received a conventional fat emulsion with long-chain triacylglycerols (LCTs), group 2 received a physical mixture of medium-chain triacylglycerols (MCTs) and LCTs (MCT/LCT), and group 3 received structured lipids composed of MCTs and LCTs (STG). Half of the rats in each respective group were sacrificed 1 d and the other half 3 d after surgery to examine the analytical parameters. Results Plasma cholesterol and free fatty acid levels in the STG group were lower than those in the other groups after surgery. The STG group had lower leukocyte CD11a/CD18 expressions than the MCT/LCT group 3 d after surgery, and CD11b/CD18 expressions in the STG group were lower than those in the LCT group on postoperative days. The STG group had higher monocyte chemotactic protein-1 and macrophage inflammatory protein-2 levels in peritoneal lavage fluid than did the other two groups. Conclusion These results suggest that, compared with the LCT and MCT/LCT groups, rats administered STG had lower plasma lipid concentrations and leukocyte integrin expressions. In addition, STG administration may cause increased recruiting of neutrophils and monocytes at the site of injury and enhance antipathogenicity in rats undergoing a total gastrectomy.

Immunopathogenesis of Folliculitis Decalvans

Folliculitis decalvans (FD) is a rare variant of primary cicatricial alopecia, for which the etiopathogenesis remains unclear. Our purpose was to evaluate whether certain immunologic mechanisms might have a significant role in the pathogenesis of FD. Lesional scalp biopsy specimens from 7 patients with FD, 7 with lichen planopilaris, and 4 with alopecia areata were studied immunohistochemically by using monoclonal antibodies to CD1a, CD3, CD4, CD8, CD20, CD25, HLA-DR, interleukin (IL)-1beta, IL-4, IL-8, interferon gamma, tumor necrosis factor alpha, basic fibroblast growth factor (b-FGF), transforming growth factor (TGF)-beta, endothelial leukocyte adhesion molecule 1, intercellular adhesion molecule (ICAM)-1, and vascular cell adhesion molecule. We showed that early FD lesions are characterized by an infiltration of activated T-helper cells, featuring mixed TH1/TH2 polarization. IL-8 and ICAM-1 may contribute to the infiltration of neutrophils, whereas b-FGF and TGF-beta may represent important mediators of the fibrosis that characterizes late-phase FD.

Review Paper: Pathophysiology of Myocardial Reperfusion Injury: The Role of Genetically Engineered Mouse Models

Coronary heart disease is the leading cause of death worldwide, affecting millions of men and women each year. Following an acute myocardial infarction, early and successful reperfusion therapy with thrombolytic therapy or primary percutaneous coronary intervention plays an important role in minimizing tissue injury associated with cessation of blood flow. The process of restoring blood flow to the ischemic myocardium, however, can induce additional injury. This phenomenon, termed myocardial ischemia-reperfusion (MI-R) injury, can paradoxically reduce the beneficial effects of myocardial reperfusion. MI-R injury is characterized by the formation of oxygen radicals upon reintroduction of molecular oxygen to the ischemic tissue, resulting in widespread lipid and protein oxidative modifications, mitochondrial injury, and cell death. In addition, studies have shown that MI-R is characterized by an inappropriate immune response in the microcirculation, resulting in leukocyte-endothelial cell interactions mediated by the upregulation of both leukocyte and endothelial cell adhesion molecules. Furthermore, MI-R ameliorates the production of certain cardioprotective factors such as nitric oxide. Advances in the generation of genetically modified mouse models enable researchers to identify the functional importance of genes involved in these processes.

Basic mechanisms behind the effects of n -3 fatty acids on cardiovascular disease

The epidemiological association between high intakes of n -3 fatty acids (FA) and decreased morbidity and mortality from cardiovascular disease (CVD) can be explained by two main basic mechanisms: (a) an effect on atherothrombosis, and (b) an effect on cardiac arrhythmias. These mechanisms probably reflect different beneficial influences of n -3 FA on cardiovascular biology. Effects on atherothrombosis include the modulation of the expression of pro-atherogenic genes (e.g., endothelial leukocyte adhesion molecules, inflammatory cytokines and cyclooxygenase (COX)-2) and the hepatic synthesis of very low density lipoproteins (VLDL), and are slow in onset, requiring incorporation into cell membrane phospholipids, and usually doses in humans in the order of 3 g/day or higher. Effects on cardiac arrhythmias include complex interactions with ion channels (sodium, potassium and calcium channels), typically requiring the presence of free FA in extracellular fluids and usually occurring with lower doses (around 1 g/day) of nutritional or pharmacological intake. We have focused most of our research effort in unraveling the pathophysiological background of protection by n -3 FA from atherothrombosis. As the result of incorporation of n -3 FA in the sn -2 position predominantly of the phosphatidyl ethanolamine pool in the inner leaflet of the plasma membrane, n -3 FA appear on the one hand to increase the production of bioactive lipid mediators (protectins and resolvins) affecting cytokine-induced signal transduction; and on the other hand to directly interfere with the generation of reactive oxygen species (mostly hydrogen peroxide), directly responsible for the activation of the transcription factor nuclear factor (NF)- κB, which controls the expression of a variety of pro-inflammatory and pro-atherogenic genes, including those encoding for interleukin (IL)-1, IL-6, IL-8, tumor necrosis factor (TNF) α, vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and COX-2. The upstream-direct or indirect-inhibition of cytokine- and other atherogenic trigger-induced signaling pathway may involve interference with the activation of protein kinase (PK) C isoforms and NADP(H) oxidase. Such interference may also explain the blunt anti-inflammatory effect of n -3 FA in many experimental models and clinical conditions of inflammation. All together, these mechanisms may provide an integrated view of how n -3 FA may affect CVD.

Blood serum levels of vascular cell adhesion molecule (sVCAM-1), intercellular adhesion molecule (sICAM-1) and endothelial leucocyte adhesion molecule-1 (ELAM-1) in diabetic retinopathy

Interaction between cells via intimate cell-cell contact is facilitated by a cell surface molecules, termed adhesion molecules. The aim of the study was to evaluate the blood serum concentration of soluble forms of vascular cell adhesion molecule (VCAM-1), intercellular adhesion molecule (ICAM-1) and endothelial leukocyte adhesion molecule-1 (ELAM-1) in patients with type 1 diabetes mellitus without and with diabetic retinopathy. The study was performed in 75 patients with type 1 diabetes mellitus, 35 without retinopathy (group 1) and 40 with retinopathy (group 2). Soluble forms of VCAM-1, ICAM-1 and ELAM-1 were determined by enzyme-linked immunosorbent assay (ELISA). The serum concentration of sICAM-1 and sELAM-1 were significantly elevated and the concentration sVCAM-1 was elevated but not significantly in diabetic patients when compared with control subjects. There was a significant difference in VCAM-1 concentrations between the control group and group 2 (965.9 +/- 229.0 vs. 1283.7 +/- 387.6 ng/ml, p < 0.05) and between group 1 and group 2 (1115.0 +/- 285.5 vs. 1283.7 +/- 387.6 ng/ml, p < 0.05). There were significant differences in sICAM-1 concentrations between the control group and group 1 (p < 0.05) and between the control group and group 2 (p < 0.05). Where was no significant difference in sICAM-1 concentration between group 1 and 2 (405.2 +/- 135.9 vs. 443.1 +/- 112.7 ng/ml, p = 0.08). ELAM-1 concentration was significantly elevated in group 2 (120.5 +/- 49.3 ng/ml) when compared with the control group (51.7 +/- 18.1 ng/ml, p < 0.005) and with group 1 (81.2 +/- 27.7 ng/ml, p < 0.05). The correlations found between sVCAM-1, sICAM-1 and sELAM-1 and the presence of retinopathy suggest that cellular adhesion and neovascularization may be linked processes.

The ITGB2 immunomodulatory gene (CD18), enterocolitis, and Hirschsprung's disease

Hirschsprung's disease (HSCR)–associated enterocolitis (HAEC) remains a major contributor to morbidity and mortality associated with HSCR, being sometimes difficult to diagnose in its subclinical form. Its pathogenesis appears to include impaired local defense mechanisms as well as dysfunctional immune response and leukocyte function. In this context, the ITGB2 (CD18) immunomodulation-related gene is a possible candidate in HAEC pathogenesis as it codes for the β-subunit of leukocyte adhesion molecule lymphocyte function-associated antigen 1, which has an established role in T-cell development and function. ITGB2/CD18 has also been linked to chronic colitis in both human and animal models involving defense mechanisms within colonic mucosa. There is therefore a fairly compelling case for the potential involvement of the ITGB2 (CD18) in HAEC pathogenesis. Aim The aim of this study was to investigate the ITGB2 immunomodulatory gene (CD18) in a cohort of patients with HSCR and explore its correlation with enterocolitis. Patients and methods Screening for mutations of the ITGB2 (CD18) gene was performed on DNA extracted from colonic tissue samples and whole blood of 33 HSCR patients controlled by analysis of 60 unaffected individuals from the diverse South African population. Polymerase chain reaction amplification was performed, followed by heteroduplex single-strand conformation polymorphism analysis and bidirectional semiautomated DNA sequencing analysis. Results Heteroduplex single-strand conformation polymorphism banding patterns of the ITGB2 gene showed variations in 22 HSCR patients (66%), 13 of whom had severe episodes of HAEC, and 6 others had milder symptoms. Of the 13, 6 (46%) had Down's syndrome–associated HSCR. Genetic variations included 1 mutation (D77N), 2 known (V367, V441), and 4 novel polymorphisms (−111T/C, 24G/T, 295G/A, 892A/G). Significant associations were identified in the exon 5′ untranslated promotor region ( P < .0001), exon 10 ( P < .0007), and the 3′ untranslated promotor region at 122G/A ( P < .0001) and 370 G/T positions ( P = .04). Those regions of the gene most frequently associated with HAEC and severe symptoms were those with more than 1 variant identified in the gene. Conclusions This study shows that impaired CD18 leukocyte and T regulatory cell regulation can probably be linked to a genetic ( ITGB2) predisposition to HAEC. It furthermore provides a possible genetic link to HAEC patient selection, identifying a potential molecular target.

Ultrasound molecular imaging of cardiovascular disease

Myocardial contrast echocardiography utilizes intravenously injected gas-filled microspheres as acoustically active red blood cell tracers. During ultrasound imaging, unimpeded microsphere transit through the intramyocardial microcirculation causes transient myocardial opacification, which can be mapped and quantified as myocardial perfusion. Ultrasound molecular imaging utilizes similar acoustically active microspheres, which are modified to bear a receptor-specific ligand on the surface, conferring microsphere binding to a disease-specific endothelial epitope. Because the microspheres adhere to the endothelium, ultrasound imaging reveals a persistent, rather than transient, contrast effect, indicating the presence and location of the molecule of interest in real time. Molecular contrast echocardiography has been developed to detect upregulated leukocyte adhesion molecules during microvascular inflammation, such as occurs in cardiac transplant rejection and ischemia-reperfusion. Principles of microsphere targeting and ultrasound imaging of microvascular epitopes have been extended to larger vessels to image molecular markers of atherosclerosis. This Article summarizes the current status of cardiovascular ultrasound molecular imaging. Experimental proofs of concept will be outlined and the clinical extension of these concepts to the molecular imaging of cardiovascular disease using clinical ultrasound technology will be discussed.

LFA-1 decreases the antigen dose for T cell activation in vivo

Leukocyte adhesion molecule leukocyte function-associated antigen (LFA)-1 not only mediates intercellular binding but also delivers co-stimulatory signals in T cells. LFA-1 has been shown to decrease the threshold of TCR signal and an antigen dose required for T cell activation and proliferation in vitro. However, physiological significance of the role of LFA-1 in TCR signal has remained unclear. We examined whether LFA-1 decreased the antigen dose for T cell activation in vivo. We showed here that, although collagen-induced arthritis (CIA) could not be induced by immunization and challenge with a standard amount of type-II collagen in LFA-1-deficient mice, a higher dose of the antigen did induce CIA in the absence of LFA-1. We also showed that CD4+ T cells could be primed by immunization with a high, but not low, dose of ovalbumin antigen in LFA-1-deficient mice. These results suggest that LFA-1 decreases the threshold of TCR signal for T cell activation in vivo as well as in vitro. Further studies using TCR-transgenic LFA-1-deficient mice showed that LFA-1 cooperated with TCR in sustained Erk1/2 phosphorylation. Moreover, TCR could induce sustained Erk1/2 phosphorylation in the absence of LFA-1 when T cells were stimulated with a high, but not low, dose of antigen, suggesting that LFA-1 may cooperate with TCR in sustaining Erk1/2 phosphorylation.

Molecular imaging of cardiovascular disease using ultrasound

Molecular imaging using probes that specifically home to function- or disease-specific targets is a promising tool for both basic research investigations as well as clinical diagnostics. Ultrasound-based molecular imaging utilizes acoustically active particles (contrast agents) bearing targeting ligands that specifically bind to a molecule of interest. In the presence of an ultrasound field, the bound particles are detectable as a persistent contrast effect during ultrasound imaging. Different types of targeted contrast agents have been reported, most of which share in common the presence of a gas encapsulated by a shell of varying chemical formulation. These agents, or "microbubbles," are typically 2 to 4 mum in diameter, and have a natural resonance frequency that corresponds to the frequencies used in diagnostic echocardiography. This attribute makes it possible to induce microbubble resonance and non-linear oscillation at diagnostic ultrasound frequencies, leading to acoustic emissions from the microbubbles that can be detected as specific signals during two dimensional ultrasound imaging. Targeting ligands that have been attached to microbubbles include monoclonal antibodies, peptides, and the naturally occurring ligands for the receptor of interest, such as vascular endothelial growth factor. Because the contrast agents stay within the intravascular space, they are ideally suited for detection of endothelial epitopes, such as leukocyte adhesion molecules or angiogenesis receptors. Ultrasound molecular imaging with targeted contrast agents has been used to detect inflammation association with ischemia/reperfusion (ischemic memory), cardiac transplant rejection, early atherosclerosis, and angiogenesis. Application to tumor angiogenesis has also been reported using peptides that specifically bind to angiogenic tumor endothelium. Translation of ultrasound molecular imaging to the clinical arena will require optimization of contrast agent design to maximize specific binding, and customization of imaging systems to sensitively detect the binding events.

Impaired granulocyte oxidative burst and decreased expression of leucocyte adhesion molecule-1 (LAM-1) in patients with Wegener's granulomatosis.

Neutrophils are the target of autoantibodies in Wegener's granulomatosis (WG). In this study, granulocyte function and surface marker expression were investigated in patients with WG. The oxidative burst in response to phorbol myristate acetate (PMA) was tested with granulocytes of 25 patients with histologically proven WG. A significantly diminished percentage of oxygen radical-producing cells was found in patients with active disease. Surface antigen expression of CD11b and LAM-1 was analysed on granulocytes of 20 patients with WG. Whereas the expression of CD11b was normal, surface expression of LAM-1 was decreased in nine cases with WG. The decrease of LAM-1 correlated with disease activity. Phagocytosis of Escherichia coli was tested in 10 patients with WG, and normal values were found in all cases. We conclude that down-regulation of LAM-1 may be a marker of disease activity in WG. The altered response to PMA may indicate functional changes in granulocyte reactivity due to autoantibody-induced damage of the granulocyte membrane.

T cell heterogeneity in patients with common variable immunodeficiency as assessed by abnormalities of T cell subpopulations and T cell receptor gene analysis.

T lymphocyte regulation of immunoglobulin production may be abnormal in some patients with common variable immunodeficiency (CVI). Phenotypic analysis of peripheral blood T lymphocytes from nine patients with CVI was conducted to examine whether an abnormal distribution could be detected in a functionally distinct T lymphocyte subpopulation. The percentage of CD8+ lymphocytes proved to be increased in some patients and decreased in others. In comparison with normal controls, many patients with CVI had reduced percentages of lymphocytes expressing both CD4 and CD45RA, a phenotype associate with naive CD4+ cells. There was no significant difference in CD4+ populations bearing CD29 or leucocyte adhesion molecule-1 (LAM-1) antigens. The pattern of gene rearrangement of the T cell antigen receptor (TCR) was studied using peripheral blood lymphocytes from these patients with CVI. Genomic DNA from freshly isolated lymphocytes as well as from selectively propagated CD4+ or CD8+ populations were examined using Southern blot analysis and a probe for the beta chain of the TCR. A polyclonal pattern of TCR gene rearrangement, without the appearance of dominant non-germline bands, was demonstrated in all patient samples. These data suggest that the T lymphocytes in patients with CVI have a polyclonal pattern of TCR rearrangement despite an abnormal distribution of T cell subpopulations in some patients.

A 19-year-old man with leucocyte adhesion deficiency. In vitro and in vivo studies of leucocyte function

We describe a male patient with leucocyte adhesion molecule deficiency (LAD) of moderate phenotype. Although diagnosis was made only 2 years before his death, the patient survived until 19 years of age. This enabled us to perform a number of novel investigation, both in vivo and in vitro, relating to his leucocyte biology. Monocytes cultured in vitro matured into morphologically normal, phagocytically capable macrophages, which were able to recognize aged 'apoptotic' neutrophils. By injection of radiolabelled autologous neutrophils we demonstrated a prolonged neutrophil half-life, but normal margination, de-margination on exercise, and splenic pooling. Neutrophil adherence in vitro to vascular endothelium was normal. Histological examination of the patient's lungs at post-mortem showed intravascular aggregation of polymorphonuclear leucocytes but a paucity of cells in the interstitium and alveolar spaces. These findings indicate that the peripheral blood leucocytosis commonly observed in these patients may be due to prolonged intravascular neutrophil survival, and suggest that CD11/18 molecules have an important role in facilitating neutrophil emigration from blood vessels at sites of inflammation.

Elevated tumour necrosis factor-alpha (TNF-alpha) biological activity in psoriatic skin lesions.

Lesions of the common inflammatory skin disease psoriasis are characterized by epidermal hyperproliferation, leucocyte adhesion molecule expression and leucocyte infiltration. The local release of proinflammatory cytokines, such as TNF-alpha, may play an important role in the induction of these events. We have, therefore, analysed aqueous extracts of lesional and uninvolved (clinically normal) stratum corneum for the presence of TNF-alpha immunoreactivity and biological activity. TNF-alpha immunoreactivity and bioactivity were consistently higher in lesional compared with uninvolved samples. By using an anti-TNF-alpha neutralizing antibody it was demonstrated that the biological activity measured was due to the presence of TNF-alpha alone. Concentrations of soluble TNF receptors (p55 and p75) were also higher in lesional stratum corneum extracts, with the p55 form predominating. The plasma of psoriatic patients was also found to contain elevated concentrations of soluble p55 compared with normal controls. These results confirm the presence of immunoreactive TNF-alpha and, for the first time, conclusively demonstrate TNF-alpha biological activity and quantifiable concentrations of soluble TNF receptors (p55 and p75) in lesional psoriatic samples. TNF-alpha recovery from stratum corneum probably reflects synthesis in deeper, viable layers, where it is likely to exert its biological effects. Local and systemic release of soluble TNF receptors, in particular p55, may serve to regulate the effects of TNF-alpha in psoriasis.