An HLA-D region restriction fragment length polymorphism associated with celiac disease.

This study is the first to describe a molecular marker that distinguishes the celiac disease HLA-D region haplotype from a serologically identical haplotype in unaffected controls. Using a DQ beta chain cDNA probe and the restriction endonuclease Rsa I, we have detected a polymorphic 4.0 kb fragment which, in DQw2 individuals, is associated with a 40-fold increased relative risk of developing celiac disease. This finding should permit the identification of the celiac disease susceptibility gene(s) in the HLA-D region and facilitate a more precise dissection of the molecular and immunogenetic mechanisms involved in the pathogenesis of that disease.

The neutrophil-activating protein (HP-NAP) of Helicobacter pylori is a protective antigen and a major virulence factor.

Helicobacter pylori infection induces the appearance of inflammatory infiltrates, consisting mainly of neutrophils and monocytes, in the human gastric mucosa. A bacterial protein with neutrophil activating activity (HP-NAP) has been previously identified, but its role in infection and immune response is still largely unknown. Here, we show that vaccination of mice with HP-NAP induces protection against H. pylori challenge, and that the majority of infected patients produce antibodies specific for HP-NAP, suggesting an important role of this factor in immunity. We also show that HP-NAP is chemotactic for human leukocytes and that it activates their NADPH oxidase to produce reactive oxygen intermediates, as demonstrated by the translocation of its cytosolic subunits to the plasma membrane, and by the lack of activity on chronic granulomatous disease leukocytes. This stimulating effect is strongly potentiated by tumor necrosis factor alpha and interferon gamma and is mediated by a rapid increase of the cytosolic calcium concentration. The activation of leukocytes induced by HP-NAP is completely inhibited by pertussis toxin, wortmannin, and PP1. On the basis of these results, we conclude that HP-NAP is a virulence factor important for the H. pylori pathogenic effects at the site of infection and a candidate antigen for vaccine development.

Pilot translational study of dietary vitamin C supplementation in Barrett's esophagus.

The transcription factor Nuclear factor kappa B (NF-kappaB) is central to the regulation of genes encoding for mediators of inflammation and carcinogenesis. In the esophagus, NF-kappaB is progressively activated from inflammation to Barrett's metaplasia and adenocarcinoma. Vitamin C, an antioxidant, can inhibit NF-kappaB in in vitro models, and the aim of this study was to prospectively assess the effect of supplemental vitamin C on NF-kappaB and associated cytokines in patients with Barrett's esophagus. Twenty-five patients with long-segment Barrett's and specialized intestinal metaplasia received dietary vitamin C (1000 mg/day) orally for four weeks, and had pre- and post-vitamin C endoscopic biopsies. NF-kappaB activity (activated p50 and p65 subunits) of nuclear extracts was assessed using the Active Motif NF-kappaB assay, and cytokines and growth factors were measured using the Evidence Investigator biochip array. NF-kappaB and related pro-inflammatory cytokines and growth factors (IL-8, VEGF, IL-10) were activated in all Barrett's tissue pre-treatment. Down-regulation in activated NF-kappaB and cytokines was observed in 8/25 (35%) patients. Dietary vitamin C supplementation may down-regulate pro-inflammatory markers in a subset of Barrett's patients. Further studies with larger numbers of endpoints will be needed to further evaluate this effect.

Coeliac disease-associated risk variants in TNFAIP3 and REL implicate altered NF-kappaB signalling.

OBJECTIVE: Our previous coeliac disease genome-wide association study (GWAS) implicated risk variants in the human leucocyte antigen (HLA) region and eight novel risk regions. To identify more coeliac disease loci, we selected 458 single nucleotide polymorphisms (SNPs) that showed more modest association in the GWAS for genotyping and analysis in four independent cohorts. DESIGN: 458 SNPs were assayed in 1682 cases and 3258 controls from three populations (UK, Irish and Dutch). We combined the results with the original GWAS cohort (767 UK cases and 1422 controls); six SNPs showed association with p<1 x 10(-04) and were then genotyped in an independent Italian coeliac cohort (538 cases and 593 controls). RESULTS: We identified two novel coeliac disease risk regions: 6q23.3 (OLIG3-TNFAIP3) and 2p16.1 (REL), both of which reached genome-wide significance in the combined analysis of all 2987 cases and 5273 controls (rs2327832 p = 1.3 x 10(-08), and rs842647 p = 5.2 x 10(-07)). We investigated the expression of these genes in the RNA isolated from biopsies and from whole blood RNA. We did not observe any changes in gene expression, nor in the correlation of genotype with gene expression. CONCLUSIONS: Both TNFAIP3 (A20, at the protein level) and REL are key mediators in the nuclear factor kappa B (NF-kappaB) inflammatory signalling pathway. For the first time, a role for primary heritable variation in this important biological pathway predisposing to coeliac disease has been identified. Currently, the HLA risk factors and the 10 established non-HLA risk factors explain approximately 40% of the heritability of coeliac disease.

High sensitivity cytokine detection in acute coronary syndrome reveals up-regulation of interferon gamma and interleukin-10 post myocardial infarction.

Inflammation is an important element in the development and destabilization of atherosclerotic plaque. Using a high sensitivity multiplex assay, previously untested in the context of atherosclerotic disease, we determined serum concentrations of GM-CSF, IFNgamma, IL-1beta, IL-2, IL-10, IL-12p70, TNF alpha, IL-6, and IL-8 in 48 Myocardial Infarction (MI) patients, 14 Unstable Angina (UA) patients and 12 healthy controls. IFNgamma levels were significantly higher in MI compared to UA (p=0.0091) and Control groups (p=0.0014). IL-10 also showed higher expression levels between MI, UA groups and Controls (p=0.0299).This up-regulation may reflect the extent of plaque instability and/or rupture in MI patients.Our observations provide evidence that IFNgamma and IL-10 merit further investigation in atherosclerotic disease states as potential markers of disease and therapeutic targets.

Functional characterization of Ost3p. Loss of the 34-kD subunit of the Saccharomyces cerevisiae oligosaccharyltransferase results in biased underglycosylation of acceptor substrates.

Within the lumen of the rough endoplasmic reticulum, oligosaccharyltransferase catalyzes the en bloc transfer of a high mannose oligosaccharide moiety from the lipid-linked oligosaccharide donor to asparagine acceptor sites in nascent polypeptides. The Saccharomyces cerevisiae oligosaccharyltransferase was purified as a heteroligomeric complex consisting of six subunits (alpha-zeta) having apparent molecular masses of 64 kD (Ost1p), 45 kD (Wbp1p), 34 kD, 30 kD (Swp1p), 16 kD, and 9 kD. Here we report a structural and functional characterization of Ost3p which corresponds to the 34-kD gamma-subunit of the oligosaccharyltransferase. Unlike Ost1p, Wbp1p, and Swp1p, expression of Ost3p is not essential for viability of yeast. Instead, ost3 null mutant yeast grow at wild-type rates on solid or in liquid media irrespective of culture temperature. Nonetheless, detergent extracts prepared from ost3 null mutant membranes are twofold less active than extracts prepared from wild-type membranes in an in vitro oligosaccharyltransferase assay. Furthermore, loss of Ost3p is accompanied by significant underglycosylation of soluble and membrane-bound glycoproteins in vivo. Compared to the previously characterized ost1-1 mutant in the oligosaccharyltransferase, and the alg5 mutant in the oligosaccharide assembly pathway, ost3 null mutant yeast appear to be selectively impaired in the glycosylation of several membrane glycoproteins. The latter observation suggests that Ost3p may enhance oligosaccharide transfer in vivo to a subset of acceptor substrates.

The essential OST2 gene encodes the 16-kD subunit of the yeast oligosaccharyltransferase, a highly conserved protein expressed in diverse eukaryotic organisms.

Oligosaccharyltransferase catalyzes the transfer of a preassembled high mannose oligosaccharide from a dolichol-oligosaccharide donor to consensus glycosylation acceptor sites in newly synthesized proteins in the lumen of the rough endoplasmic reticulum. The Saccharomyces cerevisiae oligosaccharyltransferase is an oligomeric complex composed of six non-identical subunits (alpha-zeta). The alpha, beta, gamma, and delta subunits of the oligosaccharyltransferase are encoded by the OST1, WBP1, OST3, and SWP1 genes, respectively. Here we describe the functional characterization of the OST2 gene that encodes the epsilon-subunit of the oligosaccharyltransferase. Genomic disruption of the OST2 locus was lethal in haploid yeast showing that expression of the Ost2 protein is essential for viability. Overexpression of the Ost2 protein suppresses the temperature-sensitive phenotype of the wbp1-2 allele and increases in vivo and in vitro oligosaccharyltransferase activity in a wbp1-2 strain. An analysis of a series of conditional ost2 mutants demonstrated that defects in the Ost2 protein cause pleiotropic underglycosylation of soluble and membrane-bound glycoproteins. Microsomal membranes isolated from ost2 mutant yeast show marked reductions in the in vitro transfer of high mannose oligosaccharide from exogenous lipid-linked oligosaccharide to a glycosylation site acceptor tripeptide. Surprisingly, the Ost2 protein was found to be 40% identical to the DAD1 protein (defender against apoptotic cell death), a highly conserved protein initially identified in vertebrate organisms. The protein sequence of ost2 mutant alleles revealed mutations at highly conserved residues in the Ost2p/DAD1 protein sequence.

The alpha subunit of the Saccharomyces cerevisiae oligosaccharyltransferase complex is essential for vegetative growth of yeast and is homologous to mammalian ribophorin I.

Oligosaccharyltransferase mediates the transfer of a preassembled high mannose oligosaccharide from a lipid-linked oligosaccharide donor to consensus glycosylation acceptor sites in newly synthesized proteins in the lumen of the rough endoplasmic reticulum. The Saccharomyces cerevisiae oligosaccharyltransferase is an oligomeric complex composed of six nonidentical subunits (alpha-zeta), two of which are glycoproteins (alpha and beta). The beta and delta subunits of the oligosaccharyltransferase are encoded by the WBP1 and SWP1 genes. Here we describe the functional characterization of the OST1 gene that encodes the alpha subunit of the oligosaccharyltransferase. Protein sequence analysis revealed a significant sequence identity between the Saccharomyces cerevisiae Ost1 protein and ribophorin I, a previously identified subunit of the mammalian oligosaccharyltransferase. A disruption of the OST1 locus was not tolerated in haploid yeast showing that expression of the Ost1 protein is essential for vegetative growth of yeast. An analysis of a series of conditional ost1 mutants demonstrated that defects in the Ost1 protein cause pleiotropic underglycosylation of soluble and membrane-bound glycoproteins at both the permissive and restrictive growth temperatures. Microsomal membranes isolated from ost1 mutant yeast showed marked reductions in the in vitro transfer of high mannose oligosaccharide from exogenous lipid-linked oligosaccharide to a glycosylation site acceptor tripeptide. Microsomal membranes isolated from the ost1 mutants contained elevated amounts of the Kar2 stress-response protein.

Comparison of three cell fixation methods for high content analysis assays utilizing quantum dots.

Semiconductor nanoparticles or quantum dots are being increasingly utilized as fluorescent probes in cell biology both in live and fixed cell assays. Quantum dots possess an immense potential for use in multiplexing assays that can be run on high content screening analysers. Depending on the nature of the biological target under investigation, experiments are frequently required on cells retaining an intact cell membrane or also on those that have been fixed and permeabilized to expose intracellular antigens. Fixation of cell lines before or after the addition of quantum dots may affect their localization, emission properties and stability. Using a high content analysis platform we perform a quantitative comparative analysis of three common fixation techniques in two different cell lines exposed to carboxylic acid stabilized CdTe quantum dots. Our study demonstrates that in prefixed and permeabilized cells, quantum dots are readily internalized regardless of cell type, and their intracellular location is primarily determined by the properties of the quantum dots themselves. However, if the fixation procedures are preformed on live cells previously incubated with quantum dots, other important factors have to be considered. The choice of the fixative significantly influences the fluorescent characteristics of the quantum dots. Fixatives, regardless of their chemical nature, negatively affected quantum dots fluorescence intensity. Comparative analysis of gluteraldehyde, methanol and paraformaldehyde demonstrated that 2% paraformaldehyde was the fixative of choice. The presence of protein in the media did not significantly alter the quantum dot fluorescence. This study indicates that multiplexing assays utilizing quantum dots, despite being a cutting edge tool for high content cell imaging, still require careful consideration of the basic steps in biological sample processing.

Characterization of tumour necrosis factor-alpha genetic variants and mRNA expression in patients with severe sepsis.

Tumour necrosis factor-alpha (TNFalpha) has been implicated in the pathogenicity of severe sepsis by both genetic association studies and animal models. Conflicting functional data have emerged in relation to genetic variants and TNFalpha protein production. Therefore, we assessed the functionality of TNFalpha genetic variants in terms of mRNA production and their potential influence on outcome in the setting of severe sepsis. Sixty-two Irish Caucasian patients presenting with severe sepsis were recruited and TNFalpha mRNA and protein levels were quantified. Patient DNA was analysed for the presence of common promoter polymorphisms and haplotypes were inferred. An A allele at position -863 was associated with more TNFalpha mRNA on day 1 compared to C homozygotes (P = 0.037). There was a trend for G homozygotes at position -308 to produce more TNFalpha mRNA on day 1 than those carrying an A allele (P = 0.059). The presence of an A allele at -863 was associated with greater levels of TNFalpha mRNA in comparison with patients carrying the A allele at -308 on day 1 (P = 0.02). Patients homozygous for the A allele at position -308 had a higher mortality than those carrying the G allele (P = 0.01). Our data are consistent with recent reports suggesting that a deficient proinflammatory response may be harmful in human sepsis. This deficient inflammatory response may be mediated in part by polymorphisms in the TNFalpha promoter.

A calcified lung tumour and microcytic anaemia in a young woman: partial expression of the Carney triad.

The Carney triad is the non-familial addociation of gastric stromal tumours (GISTs), pulmonar chondromas and extra-adrenal paragangliomas. Fewer than 100 cases of the disorder have been reported since its description in 1977. The condition has a predeliction for young women. Most patients exhibit only two of the three components. The tumours tend to be multifocal in the affected organ or system. Herein, we describe the case of a 27-year-old woman with multiple gastric GISTs and a pulmonary chondroma, partial expression of the Carney triad. It is important to be aware of the Carney triad when one of its constituent tumours is found, particularly if the patient is a young woman, so that a search can be made for and surveillance instituted for the other components. Treatment for the gastric tumours (sarcomas) and the paragangliomas (potentially malignant) is surgical. The lung chondromas are benign neoplasms and ordinarily not symptomatic. If a diagnosis of the tumour can be established by biopsy, surgical resection may not be necessary.

Blood flow, oxygenation, and bioenergetic status of tumors after erythropoietin treatment in normal and anemic rats.

Growth, blood flow, oxygenation, and bioenergetic status of experimental tumors were investigated in normal (control) and anemic animals after administration of recombinant human erythropoietin (rhEPO). DS sarcomas were implanted s.c. onto the hind foot dorsum of Sprague-Dawley rats. Tumor-associated anemia was induced by the development of an i.p. hemorrhagic ascites. rhEPO (1000 IU/kg) was administered s.c. three times per week over 14 days, after which it was found to have significantly increased hematocrit values in both normal and anemic animals. Tumor growth in anemic animals was slower than in normal animals, and rhEPO administration did not influence tumor growth in either group. Tumor blood flow in anemic animals was lower than in control animals and was only increased in larger tumors in animals in which anemia was prevented by prophylactic rhEPO application. Tumor oxygenation, determined using polarographic needle electrodes and oxygen partial pressure histography, was poorer in anemic animals than in normal animals. This reduction could be reversed partially, but not compensated fully by rhEPO treatment in smaller tumors (< or = 1.4 ml). These changes suggest that rhEPO, by improving tumor oxygenation, may increase the efficacy of standard radiotherapy in anemic animals and may be of use in anemic tumor patients in whom the success of radiotherapy or O2-dependent chemotherapy might be limited by tumor hypoxia.

In vivo oxygen consumption rate of DS sarcoma cells on inhibition of DNA synthesis.

The effect of inhibiting DNA synthesis on the cellular O2 consumption rate of tumor cells (DS sarcoma) in vivo was analyzed using a photometric technique. Five days after DS-sarcoma ascites was induced in SD rats, animals were treated either with fludarabine (400 mg/kg i.p., 6 h prior to measurements) or lovastatin (3 x 20 mg/kg i.p., 24, 15, and 3 h prior to measurements), drugs that can inhibit tumor cell proliferation. In addition to cellular O2 consumption, the cell cycle distribution and the fraction of DNA-synthesizing cells in the tumor ascites were measured. Both drugs lowered DNA synthesis significantly, an effect that was more pronounced with fludarabine. The cellular O2 consumption rate following lovastatin application was significantly impaired (approximately 33%), whereas fludarabine had practically no effect on the respiration rate of tumor cells. From these data, it is concluded that a reduction in DNA synthesis does not necessarily result in a decrease in the O2 consumption rate of tumor cells in vivo.

Erythropoietin restores the anemia-induced reduction in cyclophosphamide cytotoxicity in rat tumors.

The aim of this study was to examine the impact of anemia prevention by recombinant human erythropoietin (rHuEPO) treatment on the cytotoxicity of cyclophosphamide in solid experimental tumors. Anemia was induced using a single dose of carboplatin (50 mg/kg i.v.) resulting in a long-lasting reduction (30%) of the hemoglobin concentration. In a second group, the development of anemia was prevented by rHuEPO (1000 IU/kg) administered s.c. three times/week starting 7 days before carboplatin application. Four days after carboplatin treatment, tumors (DS-sarcoma of the rat) were implanted s.c. onto the hind food dorsum. Neither carboplatin nor rHuEPO treatment influenced tumor growth rate per se. When tumors were treated with a single dose of cyclophosphamide (60 mg/kg i.p.) 5 days after implantation, a growth delay with a subsequent regrowth of the tumors was observed. In the anemia group, the growth delay was significantly shorter compared with nonanemic controls (13.3 days versus 8.6 days). In the group where anemia was prevented by rHuEPO treatment, growth delay was comparable with that of nonanemic controls (13.3 days). These results suggest that chemotherapy-induced anemia reduces cytotoxicity of cyclophosphamide in tumors, whereas correction of anemia by rHuEPO treatment (epoetin alpha) increases the sensitivity, probably as a result of an improved oxygen supply to tumor tissue.

Enhancement of oxidative cell injury and antitumor effects of localized 44 degrees C hyperthermia upon combination with respiratory hyperoxia and xanthine oxidase.

The effects of respiratory hyperoxia (RH) and xanthine oxidase (XO) during localized hyperthermia (HT) were investigated by determining markers of oxidative damage to lipids and proteins and tumor growth. Anesthetized rats with s.c. DS-sarcomas underwent one of the following treatments: (a) localized saline-bath HT (60 min, 44 degrees C); (b) HT + RH (100% O2); and (c) HT + RH + XO (15 units/kg i.v.). Sham-treated animals served as controls. Tumors were investigated for: (a) thiobarbituric acid-reactive substance formation and protein-bound 4-hydroxynonenal, as indicators of lipid peroxidation; (b) reactive oxygen-mediated protein modifications; (c) apoptosis; and (d) tumor volume growth. Upon treatment, increases in thiobarbituric acid-reactive substances, protein-bound 4-hydroxynonenal, protein-associated carbonyl functions, and number of cells undergoing apoptosis were found in tumor tissue, together with an inhibition of tumor growth. When treatment groups were compared, effects in the group HT + RH + XO were generally most pronounced. These findings indicate that the antitumor effect of HT is at least partially mediated through the selective induction of lipid peroxidation and oxidative injury in tumor cells, leading to apoptosis. This effect was enhanced by adding RH or RH + XO, presumably due to enhanced tissue damage following an increased formation of reactive oxygen species, with higher levels of lipid peroxidation and protein oxidation.

Adhesion molecules in nonrheumatic aortic valve disease: endothelial expression, serum levels and effects of valve replacement.

OBJECTIVES: We studied the expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and endothelial selectin (E-selectin) on aortic valve endothelium in patients undergoing valve replacement. We also assessed the relation between serum levels and endothelial expression and also the changes in serum levels following surgery. BACKGROUND: Nonrheumatic aortic valve disease is believed to be a degenerative condition. Increased tissue and soluble adhesion molecule levels are described in inflammatory conditions. METHODS: Aortic valves from 22 surgical (16 bicuspid, 6 tricuspid) and 6 autopsy (4 normal, 2 thickened) cases were studied by immunohistochemistry. Soluble adhesion molecules were measured in peripheral blood preoperatively, and at 6 and 18 months postoperatively, and compared with controls. RESULTS: The majority of the surgically removed tricuspid and bicuspid valves expressed adhesion molecules (E-selectin, 75% and 100%; ICAM-1, 75% and 80%; VCAM-1, 69% and 60%, respectively). The normal postmortem valves did not express these, while the diseased ones did. Endothelial expression of E-selectin correlated strongly with serum levels (r = 0.695, p = 0.004). Soluble E-selectin levels were significantly higher at baseline compared with controls (p = 0.017) and fell significantly at 18 months postoperatively (p = 0.005). CONCLUSIONS: Adhesion molecule expression on diseased valves supports an inflammatory component in "degenerative" aortic valve disease. The diseased valves may be the main source of elevated soluble E-selectin in this condition as blood levels correlate with endothelial expression and blood levels fall at 18 months postoperatively.

Protein kinase C and the cytoskeleton.

The protein kinase C family of serine-threonine kinases are important signal transducers participating in many different agonist-induced signalling cascades. PKC is activated by increases in diacylglycerol produced in response to agonist-induced hydrolysis of inositol phospholipids. PKC is thought to reside in the cytosol in an inactive conformation and translocate to the plasma membrane upon cell activation where it modifies various cellular functions through phosphorylation of target substrates. Increasing evidence has illustrated that this family of enzymes is capable of translocating to other subcellular sites than the plasma membrane. A key to understanding the functions of the members of this family is identifying their physiological substrates and their relationship with those target substrates. The idea that PKC may be an important regulator of cytoskeletal function has been suggested by numerous studies. Activation of PKC in a variety of different cell types leads to changes in the cell cytoskeleton including lymphocyte surface receptor capping, smooth muscle contraction and actin rearrangement in T cells and neutrophils. Given the ubiquitous expression of PKC and the diversity of cytoskeletons in different cell types it is not surprising that PKC has been shown to be associated with and/or phosphorylate a wide range of cytoskeletal components. This review examines the interaction of PKC with the cytoskeleton and discusses some of the cytoskeletal functions ascribed to PKC to date.

The effects of conjugated linoleic acid supplementation on immune function in healthy volunteers.

OBJECTIVE: To assess the effects of dietary supplementation using two isomeric blends of conjugated linoleic acid (CLA) on immune function in healthy human volunteers. DESIGN: Double-blind, randomised, placebo-controlled intervention trial. SUBJECTS AND INTERVENTION: A total of 55 healthy volunteers (n=20 males, n=35 females) were randomised into one of three study groups who received 3 g/day of a fatty acid blend containing a 50:50 cis-9, trans-11: trans-10, cis-12 CLA isomer blend (2 g CLA), and 80:20 cis-9, trans-11: trans-10, cis-12 (80:20) CLA isomer blend (1.76 g CLA) or linoleic acid (control, 2 g linoleic acid) for 8 weeks. RESULTS: Supplementation with the 80:20 CLA isomer blend significantly (P< or =0.05) enhanced PHA-induced lymphocyte proliferation. CLA decreased basal interleukin (IL)-2 secretion (P< or =0.01) and increased PHA-induced IL-2 and tumor necrosis factor alpha (TNF(alpha)) production (P< or =0.01). However, these effects were not solely attributable to CLA as similar results were observed with linoleic acid. CLA supplementation had no significant effect on peripheral blood mononuclear cells IL-4 production, or on serum-soluble intercellular adhesion molecule-1 (sICAM-1) or plasma prostaglandin E2 (PGE2) or leukotreine B4 (LTB4) concentrations. CONCLUSIONS: This study shows that CLA supplementation had a minimal effect on the markers of human immune function. Furthermore, supplementation with CLA had no immunological benefit compared with linoleic acid.

Leukocyte function-associated antigen 1 (LFA-1) and CD44 are signalling molecules for cytoskeleton-dependent morphological changes in activated T cells.

Signaling through the leukocyte function-associated antigen 1 (LFA-1) molecule has previously been shown to induce homotypic aggregation in T cells and to induce cytoskeletal changes in T lymphoma cells. In this study we describe the induction of a dendritic phenotype associated with cytoskeletal rearrangement in activated human peripheral blood T cells stimulated with monoclonal antibody SPV-L7 to LFA-1 alpha. Maximal expression of this phenotype required 72 h preactivation with phorbol myristate acetate and expression was abolished using the protein kinase C inhibitor staurosporine. Monoclonal antibody to CD18, the beta-chain of LFA-1, did not induce this phenotype. Monoclonal antibody MEM 83 to presumably a discrete epitope on LFA-1 alpha did not induce this phenotype but induced homotypic aggregation. However, a monoclonal antibody to CD44 induced a similar phenotype in activated lymphocytes. Induction of both homotypic in activated lymphocytes. Induction of both homotypic aggregation and the dendritic phenotype was abolished by preincubation with soluble intracellular adhesion molecule 1 (ICAM-1). Cytoskeletal inhibitors prevented the morphological changes in SPV-L7-activated lymphocytes. Preincubation with tyrosine kinase inhibitor, protein kinase C inhibitors, and inhibitors of new protein synthesis also prevented these morphological changes. These data suggest that discrete epitopes on LFA-1 alpha may be capable of inducing discrete signals either for homotypic aggregation or for a dendritic phenotype. As both LFA-1 and CD44 are involved in the migration of lymphocytes through high endothelial venules, these data could suggest that these molecules transduce signals resulting in cytoskeletal modification necessary for lymphocyte transmigration.