Different modulation by histamine of IL-4 and interferon-gamma (IFN-γ) release according to the phenotype of human Th0, Th1 and Th2 clones

Histamine, an important inflammatory mediator in allergic diseases and asthma, has been reported to have modulator effects on T cells, suggesting that the bronchial microenvironment may regulate the function of resident T cells. We examined the effect of histamine on the release of the Th2-associated cytokines IL-4 and IL-5 and the Th1-associated cytokine IFN-γ by 30 CD4⁺ T cell clones from peripheral blood or bronchial biopsy of one atopic subject. Based on the IL-4/IFN-γ ratio, the clones were ascribed to the Th2 (ratio >1), Th0 (ratio 0.1 and 1) or Th1 (ratio <0.1) phenotype. Histamine inhibited IFN-γ production by Th1-like cells (P<0.02, Kruskall–Wallis), especially from bronchial biopsy, but had no effect on IL-4 release. Regarding Th0 clones, histamine inhibited IL-4 production (P<0.02) in a dose-dependent manner and slightly inhibited IFN-γ production, but had no effect on Th2-like cells. Histamine had a heterogeneous and insignificant effect on IL-5 production. The H₂-receptor antagonist ranitidine completely reversed the inhibition of IL-4 and IFN-γ production, whereas the agonist dimaprit mimicked this effect. In contrast, H₁- and H₃-receptor agonists and antagonists had no significant effect. These data demonstrate that histamine has different effects on IL-4 and IFN-γ release by T helper cells according to their phenotype via H₂-receptors. This study extends the immunomodulatory effects of histamine which may contribute to the perpetuation of airway inflammation in asthma.     

The role of size, sequence and haplotype in the stability of FRAXA and FRAXE alleles during transmission

Factors involved in the stability of trinucleotide repeats during transmission were studied in 139 families in which a full mutation, premutation or intermediate allele at either FRAXA or FRAXE was segregating. The transmission of alleles at FRAXA, FRAXE and four microsatellite loci were recorded for all individuals. Instability within the minimal and common ranges (0-40 repeats for FRAXA, 0-30 repeats for FRAXE) was extremely rare; only one example was observed, an increased in size at FRAXA from 29 to 39 repeats. Four FRAXA and three FRAXE alleles in the intermediate range (41-60) repeats for FRAXA, 31-60 for FRAXE) were unstably transmitted. Instability was more frequent for FRAXA intermediate alleles that had a tract of pure CGG greater than 37 although instability only occurred in two of 13 such transmissions: the changes observed were limited to only one or two repeats. Premutation FRAXA alleles over 100 repeats expanded to a full mutation during female transmission in 100% of cases, in agreement with other published series. There was no clear correlation between haplotype and probability of expansion of FRAXA premutations. Instability at FRAXA or FRAXE was more often observed in conjunction with a second instability at an independent locus suggesting genomic instability as a possible mechanism by which at least some FRAXA and FRAXE mutations arise.      

CYP2D6 polymorphism in systemic lupus erythematosus patients

Objectives: To determine whether patients with idiopathic systemic lupus erythematosus (SLE) are associated with impaired CYP2D6 activity and to gain insight into whether there is an association between particular CYP2D6 genotypes and susceptibility to SLE, and whether CYP2D6 polymorphism is linked to any specific clinical features of SLE. Methods: Debrisoquine sulfate (10 mg p.o.) was given to 159 healthy volunteers and 39 idiopathic SLE patients. Genotypic assay was carried out in 80 healthy volunteers and 32 patients. A 10-ml blood sample was drawn for genotypic assay. Debrisoquine and 4-hydroxydebrisoquine were determined in 8-h urine samples. Blood samples were analysed for the presence of mutations in the CYP2D6 gene, by using polymerase chain reaction (PCR) specific for CYP2D6*3 and CYP2D6*4 alleles. Results: The metabolic ratio of debrisoquine to 4-hydroxydebrisoquine ranged from 0.01 to 86.98 in healthy subjects and from 0.02 to 96 in SLE patients. We observed the poor metabolizer(PM) debrisoquine phenotype in three of 39 patients with idiopathic SLE (7.6%) and five of 159 healthy subjects (3.1%). There was no significant difference in the frequency of PM phenotypes between idiopathic SLE and healthy subjects (Fisher's exact test, P = 0.19). No significant difference in the distribution of overall genotypes and allele frequencies were observed between the two groups. No significant relationships were found between specific clinical features and the overall genotype. Conclusion: The results of this study confirm that CYP2D6 activity is not impaired in SLE and that there is no association between SLE and phenotypic CYP2D6 status. The results also showed that there was no difference in the frequency of CYP2D6A and CYP2D6B alleles between controls and patients with SLE. Received: 14 May 1998 / Accepted in revised form: 19 October 1998     

Nosocomial respiratory syncytial virus infection in Canadian pediatric hospitals: a Pediatric Investigators Collaborative Network on Infections in Canada Study

OBJECTIVE: To determine nosocomial transmission of respiratory syncytial virus (RSV) in Canadian pediatric hospitals, outcomes associated with nosocomial disease, and infection control practices. DESIGN: A prospective cohort study in the 1992 to 1994 winter respiratory seasons. SETTING: Nine Canadian pediatric university-affiliated hospitals. PARTICIPANTS: Hospitalized children with symptoms of lower respiratory tract infection (at least one of cough, wheezing, dyspnea, tachypnea, and apnea) and RSV antigen identified in a nasopharyngeal aspirate. RESULTS: Of 1516 children, 91 (6%) had nosocomial RSV (NRSV), defined as symptoms of lower respiratory tract infection and RSV antigen beginning >72 hours after admission. The nosocomial ratio (NRSV/[com-munity-acquired RSV {CARSV})] + NRSV) varied by site from 2.8% to 13%. The median length of stay attributable to RSV for community-acquired illness was 5 days, but 10 days for nosocomial illness. Four children with NRSV (4. 4%) died within 2 weeks of infection, compared with 6 (0.42%) with CARSV (relative risk = 10.4, 95% confidence interval: 3.0, 36.4). All sites isolated RSV-positive patients in single rooms or cohorted them. In a multivariate model, no particular isolation policy was associated with decreased nosocomial ratio, but gowning to enter the room was associated with increased risk of RSV transmission (incidence rate ratio 2.81; confidence interval: 1.65, 4.77). CONCLUSIONS: RSV transmission risk in Canadian pediatric hospitals is generally low. Although use of barrier methods varies, all sites cohort or isolate RSV-positive patients in single rooms. Children with risk factors for severe disease who acquire infection nosocomially have prolonged stays and excess mortality.     

A monoclonal antibody directed against a granule membrane glycoprotein (GMP-140/PADGEM, P-selectin, CD62P) inhibits ristocetin-induced platelet aggregation

P-selectin (also called CD62, GMP-140, PADGEM, CD62P) is a recently described member of a family of vascular adhesion receptors expressed by activated platelets and endothelial cells that are involved in leucocyte cell adhesion. The aim of this study was to characterize a new monoclonal antibody (LYP7) directed against activated human blood platelets that inhibits ristocetin-induced platelet aggregation. Immunoadsorbent affinity chromatography and immunoprecipitation studies showed that LYP7 (IgG₁) bound a surface-labelled glycoprotein (GP) which changed its apparent molecular mass (M_r) on reduction from 138 kD (situated below GPIIb) to 148 kD (above GPIIbα). LYP7 and S12, a monoclonal antibody directed against P-selectin immunoprecipitated the same band. Using ELISA assay, purified P-selectin was shown to bind LYP7 and S12 monoclonal antibodies. Binding sites of ¹²⁵I-labelled LYP7, which was greatly increased on thrombin-stimulated (2 U/ml) washed platelets (10825±2886, mean ±SD) (K_d=1.5±0.5 nm ) compared to resting platelets (2801±1278, mean ±SD) (K_d=1.5±0.6 nm ), was found to be normal on thrombin-stimulated platelets taken from a patient with grey platelet syndrome or a patient with Glanzmann thrombasthenia. LYP7 (IgG₁, F(ab′)₂ or Fab fragments) inhibited ristocetin-induced platelet aggregation of platelets in a dose-dependent fashion without affecting the binding of von Willebrand (vWf ) factor. However, agglutination of formaldehyde-fixed platelets induced by ristocetin was not affected by monoclonal antibody LYP7. In addition, the binding of thrombin-activated platelets to neutrophils was inhibited by monoclonal antibody LYP7. These results strongly suggest that P-selectin, by promoting cell–cell contact, may play an active role in platelet–platelet interactions.     

The receptor for urokinase-type plasminogen activator and urokinase is translocated from two distinct intracellular compartments to the plasma membrane on stimulation of human neutrophils

The cellular receptor for urokinase-type plasminogen activator (uPAR) binds pro-urokinase (pro-uPA) and facilitates its conversion to enzymatically active urokinase (uPA). uPA in turn activates surface- bound plasminogen to plasmin, a process of presumed importance for a number of biologic processes including cell migration and resolution of thrombi. We have previously shown that uPAR is expressed on the plasma membrane of circulating neutrophils, and we now report that stimulation with phorbol myristate acetate (PMA), FMLP, or tumor necrosis factor- alpha results in a rapid increase in the expression of uPAR. This process is accompanied by an increased cell-associated plasminogen activation after preincubation of neutrophils with pro-uPA in vitro. By subcellular fractionation of unstimulated neutrophils, 50% of uPAR is recovered in fractions containing latent alkaline phosphatase, corresponding to an intracellular compartment of easily mobilizable secretory vesicles distinct from both primary and specific granules, whereas the remaining 50% of uPAR is associated with a compartment eluting close to the specific granules. In contrast, the ligand pro-uPA is primarily (approximately 80%) found in the specific granules, but small amounts of pro-uPA/uPA (approximately 20%) coelute with latent alkaline phosphatase. Stimulation of neutrophils with FMLP results in translocation of uPAR as well as of pro-uPA from the secretory vesicles, whereas stimulation with PMA is required to translocate material from specific granules. Flow cytometry of neutrophils saturated with exogenous diisopropyl fluorophosphate-uPA shows a large excess (approximately 90%) of unoccupied uPAR on resting as well as FMLP- and PMA-stimulated neutrophils, suggesting a possible role for exogenous pro-uPA in providing neutrophils with a potential for plasminogen activation. These processes may be important for neutrophil extravasation and migration through extracellular matrix and for the contribution of neutrophils to resolution of thrombi.     

Evidence that large granular lymphocytes from B-CLL patients with hypogammaglobulinemia down-regulate B-cell immunoglobulin synthesis [published erratum appears in Blood 1989 Jun;73(8):2232]

B-chronic lymphocytic leukemia (CLL) patients frequently suffer from moderate to severe hypogammaglobulinemia. This complication is a serious cause of morbidity and mortality in this disorder. There is recent evidence that natural killer (NK) cells modulate B-cell immunoglobin (Ig) synthesis/secretion. The authors therefore evaluated the circulating NK cells from B-CLL patients on their ability to regulate mitogen-induced B-cell Ig synthesis. Blood, NK cells (CD16+, CD3-) from three B-CLL patients with hypogammaglobulinemia were able to clearly down-regulate the pokeweed mitogen (PWM)-induced-B-cell Ig secretion. In contrast, CD16+, CD3- cells from age-sex-matched controls or B-CLL patients with normal Ig were either nonregulatory or enhanced mitogen-induced B-cell Ig secretion. An alternative explanation for hypogammaglobulinemia in B-CLL patients is the immunomodulation of B- cell Ig production/secretion by CD16+, CD3- blood cells.     

Effects of thrice weekly nocturnal hemodialysis on arterial stiffness

ObjectiveIn this study, we compared the changes in arterial stiffness in chronic hemodialysis patients treated with 8-h vs. 4-h thrice weekly in-center hemodialysis.MethodsSixty prevalent chronic hemodialysis patients assigned to 8-h nocturnal in-center thrice weekly HD (NHD) and 60 control cases assigned to 4-h thrice weekly conventional HD (CHD) were followed for one year. Radial–carotid pulse wave velocity, augmentation index and echocardiography were performed at baseline and 12^th month.ResultsMean age of the patients was 49 ± 11 years, 30.8% were female, 27.5% had diabetes mellitus and mean dialysis vintage was 57 ± 47 months. Baseline demographical, clinical and laboratory parameters were similar between groups. During a mean follow-up of 15.0 ± 0.1 months, blood pressure remained similar in both groups while the number of mean daily anti-hypertensive substances decreased in the NHD group. In the NHD group, time-averaged serum phosphorus and calcium–phosphorus product were lower than the CHD group. Pulse wave velocity and augmentation index decreased in the NHD group (from 11.02 ± 2.51 m/s to 9.61 ± 2.39 m/s and from 28.8 ± 10.3% to 26.2 ± 12.1%; p = 0.008 and p = 0.04, respectively). While augmentation index increased in the CHD group (28.0 ± 9.4 to 31.0 ± 10.7%, p = 0.02), pulse wave velocity did not change. Subendocardial viability ratio and ejection duration improved in the NHD group (from 135 ± 28 to 143 ± 25%, p = 0.01 and from 294 ± 34 ms to 281 ± 34 ms, p = 0.003, respectively), accompanied by regression of left ventricular mass index. In multiple stepwise linear regression analyses, NHD was associated with improvements in augmentation index, ejection duration and subendocardial viability ratio.ConclusionsThese data indicate that arterial stiffness is ameliorated by implementation of longer hemodialysis sessions.     

Organization of the gene for human platelet/endothelial cell adhesion molecule-1 shows alternatively spliced isoforms and a functionally complex cytoplasmic domain

Platelet endothelial cell adhesion molecule-1 (PECAM-1) is a cell-cell adhesion molecule that is expressed on circulating platelets, on leukocytes, and at the intercellular junctions of vascular endothelial cells and mediates the interactions of these cells during the process of transendothelial cell migration. The cDNA for PECAM-1 encodes an open reading frame of 738 amino acids (aa) that is organized into a 27- aa signal peptide, a 574-aa extracellular domain composed of 6 Ig homology units, and a relatively long cytoplasmic tail of 118 aa containing multiple sites for posttranslational modification and postreceptor signal transduction. To provide a molecular basis for the precise evaluation of the structure and function of this transmembrane glycoprotein, we have determined the organization of the human PECAM-1 gene. The PECAM-1 gene, which has been localized to human chromosome 17, is a single-copy gene of approximately 65 kb in length and is broken into 16 exons by introns ranging in size from 86 to greater than 12,000 bp in length. Typical of other members of the Ig superfamily, each of the extracellular Ig homology domains is encoded by a separate exon, consistent with PECAM-1 having arisen by gene duplication and exon shuffling of ancestral Ig superfamily genes. However, the cytoplasmic domain was found to be surprisingly complex, being encoded by seven short exons that may represent discrete functional entities. Alternative splicing of the cytoplasmic tail appears to generate multiple PECAM-1 isoforms that may regulate phosphorylation, cytoskeletal association, and affinity modulation of the mature protein. Finally, a processed pseudogene having 76% identity with PECAM- 1 cDNA was identified and localized to human chromosome 3. These findings should have important implications for structure/function analysis of PECAM-1 and its role in vascular adhesive interactions.