Stromal cells differentially regulate neutrophil and lymphocyte recruitment through the endothelium

Stromal fibroblasts modify the initial recruitment of leucocytes by endothelial cells (EC), but their effects on subsequent transendothelial migration remain unclear. Here, EC and dermal or synovial fibroblasts were cultured on opposite surfaces of 3-lm pore filters and incorporated in static or flow-based migration assays. Fibroblasts had little effect on tumour necrosis factor-a-induced transendothelial migration of neutrophils,but tended to increase the efficiency of migration away from the endothelium.Surprisingly, similar close contact between EC and fibroblasts strongly reduced lymphocyte migration in static assays, and nearly abolished stable lymphocyte adhesion from flow. Fibroblasts did not alter endothelial surface expression of adhesion molecules or messenger RNA for chemokines. Inhibition of attachment did not occur when EC-fibroblast contact was restricted by using 04-lm pore filters, but under these conditions pre-treatment with heparinase partially inhibited adhesion. In the 3-lm pore co-cultures, inhibition of metalloproteinase activity partially recovered lymphocyte adhesion, but addition of CXCL12 (SDF-1a) to the endothelial surface did not. Hence, the ability of EC to present activating chemokines for lymphocytes may have been enzymatically inhibited by direct contact with fibroblasts. To avoid contact, we cultured EC and fibroblasts on separate 3-lm pore filters one above the other. Here,fibroblasts promoted the transendothelial migration of lymphocytes. Fibroblasts generate CXCL12, but blockade of CXCL12 receptor had no effect on lymphocyte migration. While stromal cells can provide signal(s)promoting leucocyte migration away from the sub-endothelial space,direct cell contact (which might occur in damaged tissue) may cause disruption of chemokine signalling, specifically inhibiting lymphocyte rather than neutrophil recruitment

The MHC2TA -168A>G gene polymorphism is not associated with rheumatoid arthritis in Austrian patients

An association between susceptibility to rheumatoid arthritis (RA) and a common -168A>G polymorphism in the MHC2TA gene with differential major histocompatibility complex (MHC) II molecule expression was recently reported in a Swedish population. The objective of the present study was to replicate this finding by examining the -168A>G polymorphism in an Austrian case–control study. Three hundred and sixty-two unrelated RA cases and 351 sex-matched and age-matched controls as well as 1,709 Austrian healthy individuals were genotyped. All participants were from the same ethnic background. Genotyping was performed using 5' allelic discrimination assays. The association between susceptibility to RA and the -168A>G single nucleotide polymorphism was examined by chi-square test. Comparison was made assuming a dominant effect (AG + GG genotypes versus AA genotype). In contrast to the primary report, the frequency of MHC2TA -168G allele carriers was not significantly different between patients and controls in the Austrian cohort. The homozygous MHC2TA -168 GG genotype was more frequent in matched controls than in Austrian RA patients. There was no association between the presence of RA-specific autoantibodies and the MHC2TA -168 GG genotype. In this cohort of Austrian patients, no association between the MHC2TA polymorphism and RA was found

An Experimental Study of Effects of Step Roughness in Skimming Flows on Stepped Chutes

On a spillway chute, a stepped design increases the rate of energy dissipation on the chute itself and reduces the size of a downstream energy dissipator. Up to date, the effects of step roughness on the flow properties remain unknown despite the practical relevance to damaged concrete steps, rock chutes and gabions weirs. New measurements were conducted in a large-size laboratory facility with two step conditions (smooth and rough) and three types of step roughness. Detailed air-water flow properties were measured systematically for several flow rates. The results showed faster flow motion on rough step chutes. Although the finding is counter-intuitive, it is linked with the location of the inception point of free-surface aeration being located further downstream than for a smooth stepped chute for an identical flow rate. In the aerated flow region, the velocities on rough-step chutes were larger than those of smooth chute flows for a given flow rate and dimensionless location from the inception point of free-surface aeration both at step edges and between step edges. The results suggest that design guidelines for smooth (concrete) stepped spillway may not be suitable to rough stepped chutes including gabion stepped weirs, and older stepped chutes with damaged steps

Pneumococcal polysaccharide vaccination in adults undergoing immunosuppressive treatment for inflammatory diseases - a longitudinal study.

INTRODUCTION: Patients undergoing immunosuppressive therapy are at increased risk of infection. Community-acquired pneumonia and invasive pneumococcal disease account for substantial morbidity and mortality in this population and may be prevented by vaccination. Ideally, immunization to pneumococcal antigens should take place before the start of immunosuppressive treatment. Often, however, the treatment cannot be delayed. Little is known about the efficacy of pneumococcal vaccines during immunosuppressive treatment. The objectives of this study were to determine the percentage of vaccine-naïve, immunosuppressed adults with inflammatory diseases seroprotected against Streptococcus pneumoniae and to assess factors associated with the immunogenicity, clinical impact and safety of 23-valent pneumococcal polysaccharide vaccine (PPV) in seronegative subjects. METHODS: This observational study included patients 18 years of age and older who were receiving prednisone ≥20 mg/day or other immunosuppressive drugs. Exclusion criteria were PPV administration in the previous 5 years, intravenous immunoglobulins and pregnancy. Serum immunoglobulin G (IgG) antibody levels against six pneumococcal serotypes were measured. Seropositivity was defined as IgG of 0.5 μg/ml or greater for at least four of six serotypes. Seronegative patients received PPV, and seropositive patients were included as a comparison group. Vaccine response and tolerance were assessed after 4-8 weeks. Disease activity was evaluated on the basis of the Physician Global Assessment scores. Serology was repeated after 1 year, and information on any kind of infection needing medical attention was collected. Outcomes were the proportion of seropositivity and infections between vaccinated and unvaccinated patients. RESULTS: Of 201 included patients, 35 received high-dose corticosteroids and 181 were given immunosuppressive drugs. Baseline seronegativity in 60 (30 %) patients was associated with corticotherapy and lower total IgG. After PPV, disease activity remained unchanged or decreased in 81 % of patients, and 87 % became seropositive. After 1 year, 67 % of vaccinated compared with 90 % of observed patients were seropositive (p < 0.001), whereas the rate of infections did not differ between groups. Those still taking prednisone ≥10 mg/day tended to have poorer serological responses and had significantly more infections. CONCLUSIONS: PPV was safe and moderately effective based on serological response. Seropositivity to pneumococcal antigens significantly reduced the risk of infections. Sustained high-dose corticosteroids were associated with poor vaccine response and more infections

The use of tumour necrosis factor alpha-blockers in daily routine. An Austrian consensus project

To define relevant disease parameters and their respective limits indicating the initiation of TNF-α-blockers in individual patients. Subsequently, to analyze retrospectively patients with rheumatoid arthritis (RA), psoriatic arthritis (PsA) or ankylosing spondylitis (AS), who started TNF-α inhibition in 2006. Points to consider, regarded relevant for individual treatment decisions as well as their assessment methods, were ascertained by experts’ consensus applying the Delphi technique. Subsequently, these parameters’ thresholds with respect to the initiation of a TNF-α-blocker were identified. Thereafter, the rheumatologists representing 12 centres all over Austria agreed to retrospectively analyze their patients started on a TNF-α-blocker in 2006. Experts’ opinion regarding disease parameters relevant to initiate TNF-α-blockers in RA patients only slightly differed from those applied in clinical trials, but the parameters’ threshold values were considerably lower. For PsA patients, some differences and for AS patients, considerable differences between experts’ opinion and clinical studies appeared, which held also true for decisive parameters’ means and thresholds. Six hundred and fifty patients, started on TNF-blockers in 2006, could be analyzed retrospectively, 408 RA patients (53.3 years mean, 340 females), 93 PsA patients (48.9 years mean, 59 males) and 149 AS patients AS (42.2 years mean, 108 males), representing approximately 25% of all Austrian patients initiated on a TNF-blocker in this respective year. Far more individualized, patient-oriented treatment approaches, at least in part, are applied in daily routine compared with those derived from clinical trials or recommendations from investigative rheumatologists

High-Resolution Description of Antibody Heavy-Chain Repertoires in Humans

Antibodies' protective, pathological, and therapeutic properties result from their considerable diversity. This diversity is almost limitless in potential, but actual diversity is still poorly understood. Here we use deep sequencing to characterize the diversity of the heavy-chain CDR3 region, the most important contributor to antibody binding specificity, and the constituent V, D, and J segments that comprise it. We find that, during the stepwise D-J and then V-DJ recombination events, the choice of D and J segments exert some bias on each other; however, we find the choice of the V segment is essentially independent of both. V, D, and J segments are utilized with different frequencies, resulting in a highly skewed representation of VDJ combinations in the repertoire. Nevertheless, the pattern of segment usage was almost identical between two different individuals. The pattern of V, D, and J segment usage and recombination was insufficient to explain overlap that was observed between the two individuals' CDR3 repertoires. Finally, we find that while there are a near-infinite number of heavy-chain CDR3s in principle, there are about 3–9 million in the blood of an adult human being

Antibody Repertoires in Humanized NOD-scid-IL2Rγnull Mice and Human B Cells Reveals Human-Like Diversification and Tolerance Checkpoints in the Mouse

Immunodeficient mice reconstituted with human hematopoietic stem cells enable the in vivo study of human hematopoiesis. In particular, NOD-scid-IL2Rγnull engrafted mice have been shown to have reasonable levels of T and B cell repopulation and can mount T-cell dependent responses; however, antigen-specific B-cell responses in this model are generally poor. We explored whether developmental defects in the immunoglobulin gene repertoire might be partly responsible for the low level of antibody responses in this model. Roche 454 sequencing was used to obtain over 685,000 reads from cDNA encoding immunoglobulin heavy (IGH) and light (IGK and IGL) genes isolated from immature, naïve, or total splenic B cells in engrafted NOD-scid-IL2Rγnull mice, and compared with over 940,000 reads from peripheral B cells of two healthy volunteers. We find that while naïve B-cell repertoires in humanized mice are chiefly indistinguishable from those in human blood B cells, and display highly correlated patterns of immunoglobulin gene segment use, the complementarity-determining region H3 (CDR-H3) repertoires are nevertheless extremely diverse and are specific for each individual. Despite this diversity, preferential DH-JH pairings repeatedly occur within the CDR-H3 interval that are strikingly similar across all repertoires examined, implying a genetic constraint imposed on repertoire generation. Moreover, CDR-H3 length, charged amino-acid content, and hydropathy are indistinguishable between humans and humanized mice, with no evidence of global autoimmune signatures. Importantly, however, a statistically greater usage of the inherently autoreactive IGHV4-34 and IGKV4-1 genes was observed in the newly formed immature B cells relative to naïve B or total splenic B cells in the humanized mice, a finding consistent with the deletion of autoreactive B cells in humans. Overall, our results provide evidence that key features of the primary repertoire are shaped by genetic factors intrinsic to human B cells and are principally unaltered by differences between mouse and human stromal microenvironments