Unbiased Quantitative Proteomics Reveals a Crucial Role of the Allergen Context for the Activation of Human Dendritic Cells.

Worldwide, more than 1 billion people suffer from allergic diseases. However, until now it is not fully understood how certain proteins can induce allergic immune responses, while others cannot. Studies suggest that allergenicity is a process not only determined by properties of the allergen itself but also by costimulatory factors, that are not classically associated with allergic reactions. To investigate the allergenicity of the major birch pollen allergen Bet v 1 and the impact of adjuvants associated with pollen, e.g. lipopolysaccharide (LPS), we performed quantitative proteome analysis to study the activation of monocyte-derived dendritic cells (moDCs). Thus, we treated cells with birch pollen extract (BPE), recombinant Bet v 1, and LPS followed by proteomic profiling via high-performance liquid chromatography and tandem mass spectrometry (HPLC-MS/MS) using isobaric labelling. Enrichment and pathway analysis revealed the influence of regulated proteins especially in cytokine signalling and dendritic cell activation. We found highly regulated, but differentially expressed proteins after treatment with BPE and LPS, whereas the cellular response to Bet v 1 was limited. Our findings lead to the conclusion that Bet v 1 needs a specific "allergen context" involving cofactors apart from LPS to induce an immune response in human moDCs.

Waning protection of influenza vaccination during four influenza seasons, 2011/2012 to 2014/2015.

BACKGROUND: Concerns have been raised about intraseasonal waning of the protection conferred by influenza vaccination. METHODS: During four influenza seasons, we consecutively recruited individuals aged 18years or older who had received seasonal influenza vaccine and were subsequently admitted to the hospital for influenza infection, asassessed by reverse transcription polymerase chain reaction. We estimated the adjusted odds ratio (aOR) of influenza infection by date of vaccination, defined by tertiles, as early, intermediate or late vaccination. We used a test-negative approach with early vaccination as reference to estimate the aOR of hospital admission with influenza among late vaccinees. We conducted sensitivity analyses by means of conditional logistic regression, Cox proportional hazards regression, and using days between vaccination and hospital admission rather than vaccination date. RESULTS: Among 3615 admitted vaccinees, 822 (23%) were positive for influenza. We observed a lower risk of influenza among late vaccinees during the 2011/2012 and 2014/2015A(H3N2)-dominant seasons: aOR=0.68 (95% CI: 0.47-1.00) and 0.69 (95% CI: 0.50-0.95). We found no differences in the risk of admission with influenza among late versus early vaccinees in the 2012/2013A(H1N1)pdm09-dominant or 2013/2014B/Yamagata lineage-dominant seasons: aOR=1.18 (95% CI: 0.58-2.41) and 0.98 (95% CI: 0.56-1.72). When we restricted our analysis to individuals aged 65years or older, we found a statistically significant lower risk of admission with influenza among late vaccinees during the 2011/2012 and 2014/2015A(H3N2)-dominant seasons: aOR=0.61 (95% CI: 0.41-0.91) and 0.69 (95% CI: 0.49-0.96). We observed 39% (95% CI: 9-59%) and 31% (95% CI: 5-50%) waning of vaccine effectiveness among participants aged 65years or older during the two A(H3N2)-dominant seasons. Similar results were obtained in the sensitivity analyses. CONCLUSION: Waning of vaccine protection was observed among individuals aged 65years old or over in two A(H3N2)-dominant influenza seasons.

GRAND-4: the German retrospective analysis of long-term persistence in women with osteoporosis treated with bisphosphonates or denosumab.

UNLABELLED: This retrospective database study assessed 2-year persistence with bisphosphonates or denosumab in a large German cohort of women with a first-time prescription for osteoporosis treatment. Compared with intravenous or oral bisphosphonates, 2-year persistence was 1.5-2 times higher and risk of discontinuation was significantly lower (P < 0.0001) with denosumab. INTRODUCTION: Persistence with osteoporosis therapies is critical for fracture risk reduction. Detailed data on long-term persistence (≥2 years) with bisphosphonates and denosumab are sparse. METHODS: From the German IMS® database, we included women aged 40 years or older with a first-time prescription for bisphosphonates or denosumab between July 2010 and August 2014; patients were followed up until December 2014. The main outcome was treatment discontinuation, with a 60-day permissible gap between filled prescriptions. Two-year persistence was estimated using Kaplan-Meier survival curves, with treatment discontinuation as the failure event. Denosumab was compared with intravenous (i.v.) and oral bisphosphonates separately. Cox proportional hazard ratios (HRs) for the 2-year risk of discontinuation were calculated, with adjustment for age, physician specialty, health insurance status, and previous medication use. RESULTS: Two-year persistence with denosumab was significantly higher than with i.v. or oral bisphosphonates (39.8 % [n = 21,154] vs 20.9 % [i.v. ibandronate; n = 20,472] and 24.8 % [i.v. zoledronic acid; n = 3966] and 16.7-17.5 % [oral bisphosphonates; n = 114,401]; all P < 0.001). Patients receiving i.v. ibandronate, i.v. zoledronic acid, or oral bisphosphonates had a significantly increased risk of treatment discontinuation than did those receiving denosumab (HR = 1.65, 1.28, and 1.96-2.02, respectively; all P < 0.0001). CONCLUSIONS: Two-year persistence with denosumab was 1.5-2 times higher than with i.v. or oral bisphosphonates, and risk of discontinuation was significantly lower with denosumab than with bisphosphonates. A more detailed understanding of factors affecting medication-taking behavior may improve persistence and thereby reduce rates of fracture.

NOAC and intracerebral bleeding--presentation of four cases and review of the literature.

AF might be a life threatening disease. Patients have been under oral antithrombotic treatment in order to avoid thrombotic events. Although this treatment proved to be effective in the last decades there was always the inconvenience of a regular blood control. In the last months NOACs have been flooding the market promising to be as effective as their older concurrents in certain circumstances and highlighting the fact that the control of INR has become obsolete. However, as there is no specific antidote up to date, NOACs might present a life threatening event in case of an intracerebral haemorrhage. The brain surgeons might find themselves in a difficult situation when they have to decide whether to operate on a patient with a compromised haemostasis or not. We present four patients who were treated with NOACs for AF. Three of them were admitted with intracerebral haemorrhage in our neurosurgical unit from January to October 2013. The fourth patient bled one week after stopping his treatment with NOAC. Furthermore we take a closer look to the existing literature and try to portray the issue from a neurosurgical point of view.

Changes in BMI and management of patients with childhood onset growth hormone deficiency in the transition phase.

OBJECTIVE: We aimed at evaluating BMI changes during GH therapy pause, to assess the transfer pattern between children and adult medical services in our clinic and to confirm the previous growth hormone deficiency (GHD) diagnosis. DESIGN: Retrospective cohort study. METHODS: We identified 75 transition patients (age at first visit <25 years) with pituitary deficiency (ICD-10:E.23) and GHD referred to our clinic between 2000-2009. RESULTS: Out of 75 patients with GHD (45 males, 30 females), 20 subjects suffered from an idiopathic GHD (iGHD) and 55 from an organic GHD (oGHD). During the GH therapy pause (26.4±34.8 months), we observed a significant BMI increase (23.6±4.4 to 27.1±7.2, p=0.02). Most males with iGHD discontinued endocrinologic control and GH substitution completely (5 patients out of 20) or after the first contact (3 patients out of 20). Most females with GHD continued medical control after transferral (22 patients). We retested 34/75 patients with GHD (45.3%). The preferred test was the growth hormone-releasing hormone-arginine (GHRH-arginine) (20/34 patients, 58.9%), followed by the insulin hypoglycemia test (IHT) alone (9 patients). 4 patients received both, the GHRH-arginine and the IHT. Seven retested patients with iGHD (63.6%) and all oGHD retested patients were still deficient. CONCLUSIONS: Our results provide information on negative effects of the discontinuation of GH treatment during the transition phase and should help to improve the compliance with treatment in this group of patients. Paediatric and adult endocrinologists participating together in a transition programme should emphasize on the positive effect of GH substitution in adulthood. Efforts should be made to particularly improve the transferral of male adolescents with iGHD, since they seem to escape medical care.

BAX 855, a PEGylated rFVIII product with prolonged half-life. Development, functional and structural characterisation.

A longer acting recombinant FVIII is expected to serve patients' demand for a more convenient prophylactic therapy. We have developed BAX 855, a PEGylated form of Baxter's rFVIII product ADVATE™ based on the ADVATE™ manufacturing process. The conjugation process for preparing BAX 855 uses a novel PEG reagent. The production process was adjusted to yield a rFVIII conjugate with a low PEGylation degree of about 2 moles PEG per FVIII molecule. This optimised modification degree resulted in an improved PK profile for rFVIII without compromising its specific activity. PEGylation sites were identified by employing various HPLC- and MS-based methods. These studies not only indicated that about 60% of the PEG chains are localised to the B-domain, which is cleaved off upon physiological activation during the coagulation process, but also demonstrated an excellent lot to lot consistency with regard to PEGylation site distribution. Detailed biochemical characterization further showed that PEGylated FVIII retained all the physiological functions of the FVIII molecule with the exception of binding to the LRP clearance receptor which was reduced for BAX 855 compared to ADVATE™. This might provide an explanation for the prolonged circulation time of BAX 855 as reduced receptor binding might slow-down clearance. Preclinical studies showed improved pharmacokinetic behaviour and clinically relevant prolonged efficacy compared to ADVATE™ without any signs of toxicity or elevated immunogenicity. The comprehensive preclinical data package formed the basis for approval of the phase 1 clinical study by European authorities which started in 2011.

The transcription factor Grainy head and the steroid hormone ecdysone cooperate during differentiation of the skin of Drosophila melanogaster.

The arthropod epidermis is an epithelium that deposits the apical cuticle, which is a stratified extracellular matrix (ECM) protecting the animal against pathogens, preventing dehydration and also serving as an exoskeleton. Differentiation of the cuticle conceivably implies coordinated production, secretion and localization of its components. The underlying molecular mechanisms are poorly explored. In this work, we show that the transcription factor Grainy head and the steroid hormone ecdysone drive the production of two partially overlapping sets of cuticle factors. Nevertheless, Grainy head is needed to modulate the expression of ecdysone signalling factors; the significance of this cross-talk is yet unclear. In addition, we found that ecdysone signalling negatively regulates its own impact. In conclusion, our findings suggest that at least two independently triggered pathways have evolved in parallel to cooperatively ensure the stereotypic implementation of the cuticle. As Grainy head is also essential for epithelial differentiation in vertebrates, we speculate that it acts to decode the ancient skin programme common to all animals. Full differentiation of the skin necessitates a second, complementing taxon-specific programme that requires its own decoder, which is represented by ecdysone in arthropods, whereas the vertebrate specific one remains to be identified.

A computational study on the intriguing mechanisms of the gas-phase thermal activation of methane by bare [Ni(H)(OH)]+.

A detailed computational study on the reaction mechanisms of the thermal activation of methane by the bare complex [Ni(H)(OH)](+) has been conducted. The experimentally observed reaction features, i.e. the ligand exchange Ni(H) → Ni(CH(3)), the H/D scrambling between the incoming methane and the hydrido ligand of the nickel complex, the spectator-like behavior of the OH ligand, and the relatively moderate reaction efficiency of 6% relative to the collision rate of the ion/molecule reaction, can be explained by considering three competing mechanisms, and a satisfactory agreement between experiment and theory has been found.

DTI of the visual pathway in cerebral lesions.

Diffusion tensor imaging (DTI) can be used to localise the visual pathway (VP). In the service of the neurosurgery we have been working since the beginning of this year to develop a protocol which is suitable for the every day clinical routine to show the tracts of the white matter. Many lesions of the brain concern the white matter. Up to date it is still difficult to portray the visual pathway. Many centers all around the world are actually trying to localize the visual pathway, yet it is still used for the research. The application of the DTI-data for surgical interventions remains still a rarity. We believe that using this technique it would reduce the intraoperative risk and improve the postoperative outcome. From the beginning of this year we have been able to localize the visual pathway in 14 patients with different illnesses and we performed also postoperative controls. Using this new technique we were able to minimize the intraoperative risk in our patients.

Fully automated immunoassay for quantitative determination of FXIII.

UNLABELLED: Coagulation factor XIII (FXIII) is essential for clot stabilization. Deficiency of FXIII is associated with a risk of bleeding and impaired wound healing. Substitution therapy with FXIII remedies for patients with low plasma levels of FXIII requires diagnostic quantification of the factor before and during therapy. Here, we describe a prototype of a preliminary research immunoassay for quantification of FXIII antigen on automated coagulation instruments. The prototype assay is based on a monoclonal antibody (mAb) directed against FXIII A chain, whereas the mAbs are coupled to latex particles. FXIII in a plasma specimen causes agglutination of the latex particles, which can be quantified turbidimetrically. Performance data of the assay prototype processed on BCS® XP and Sysmex® CA-1500 instruments demonstrate a good correlation to the Berichrom® factor XIII activity assay1 from Siemens Healthcare Diagnostics (r = 0.94). RESULTS: Comparability of instruments was excellent (r = 0.98). Coefficients of variation of total imprecision measurements ranged from 2.2 to 3.4%. Linearity was excellent over the range tested (12-121% FXIII). Analytical sensitivity was 0.51% FXIII on BCS XP and 0.44% FXIII on Sysmex CA-1500, respectively. No interference (>10% bias) was observed with haemoglobin (up to 400 mg/dl), cholesterol (up to 300 mg/dl), bilirubin (up to 60 mg/dl) or triglycerides (up to 3000 mg/dl). CONCLUSION: The preliminary research assay prototype has the potential for excellent analytical sensitivity, precision, and dynamic range suitable to measure reliably FXIII antigen levels in human plasma.

Growth pattern of untreated boys with simple virilizing congenital adrenal hyperplasia indicates relative androgen insensitivity during the first six months of life.

CONTEXT: Mild forms of simple virilizing congenital adrenal hyperplasia (CAH) may be missed in newborn screening. In the pre-newborn-screening era, missed diagnosis of simple virilizing CAH was not infrequent in boys. Elevated adrenal androgens lead to accelerated growth and bone maturation. Traditional treatment of CAH consists of the suppression of ACTH through glucocorticoid replacement, in an attempt to reduce excessive androgen production. OBJECTIVE: To retrospectively analyze early growth pattern and bone maturation in untreated boys with simple virilizing CAH. PATIENTS: In the pre-newborn screening era, 13 boys had a late diagnosis of simple virilizing classical CAH. Diagnosis of 21-hydroxylase deficiency was confirmed by mutation analysis of the CYP21A2 gene in all patients. Growth data were retrospectively collected from standarized preventive medical checkups at the regular pediatrician until the time of diagnosis of CAH. RESULTS: Length was 0.1 ± 0.8 SDS (mean ± SD) at birth, 0.2 ± 1 SDS at 3 months, 0.2 ± 0.9 SDS at 6 months, 0.7 ± 1 SDS at 1 year, +1.1 ± 0.9 SDS at 2 years and +1.8 ± 1.2 SDS at 4 years. At diagnosis, mean chronological age was 4.4 ± 1.6 years and height SDS was 2 ± 1.7. Bone age was accelerated (9.4 ± 4 years) at diagnosis. Signs that had led to diagnosis were pubic hair (n = 11), accelerated growth rate (n = 6) and birth of an affected sister (n = 3). Despite late start of hydrocortisone treatment, mean final height was -1 ± 0.9 SDS. Seven of 18 patients had a final height within 1 SD of target height. CONCLUSION: Height velocity is not markedly increased in untreated boys with simple virilizing CAH in the first 6 months of life, indicating that infants are relatively androgen insensitive during that period. After the first 6 months of life, growth velocity increases significantly and elevated androgens lead to advanced skeletal maturation. This observation has implications for lower hydrocortisone doses to be used in CAH children during the first 6 months of life. In addition, staying alert for clinical symptoms and signs of simple virilizing CAH is still warranted, since mild forms may be missed in newborn screening.

Growth patterns in the first three years of life in children with classical congenital adrenal hyperplasia diagnosed by newborn screening and treated with low doses of hydrocortisone.

BACKGROUND: Linear growth is the best clinical parameter for monitoring metabolic control in classical congenital adrenal hyperplasia (CAH). OBJECTIVE: to analyze growth patterns in children with CAH diagnosed by newborn screening and treated with relatively low doses of hydrocortisone during the first year of life. PATIENTS AND METHODS: 51 patients (27 females) were diagnosed with classical CAH by newborn screening. All patients were treated with relatively low doses of hydrocortisone (9-15 mg/m(2) body surface area). 47 patients were additionally treated with fludrocortisone. RESULTS: at birth, height SDS (H-SDS) was 1.1 ± 1 in girls and 0.9 ± 1.5 in boys. After 3 months, H-SDS decreased to 0.4 ± 0.9 in girls and to 0.1 ± 1.3 in boys. Over the 3-year period, H-SDS further decreased to -0.4 ± 1.8 in girls and to -0.8 ± 1 in boys and approached the genetic height potential (target H-SDS of girls -0.5 ± 0.3 and target H-SDS of boys -0.9 ± 0.7). During the first 9 months of age, growth velocity was slightly decreased in girls (18.2 ± 1.9 cm) and boys (17.3 ± 1.6 cm) when compared to a healthy reference population (girls 19.0 ± 3.9 cm and boys 18.7 ± 4.7 cm). At the age of 3 years, bone age was appropriate for chronological age in both girls (2.7 ± 0.5 years) and boys (2.9 ± 0.5 years). CONCLUSION: birth length is above average in children with classical CAH, which might be the result of untreated hyperandrogenism in utero. With relatively low doses of hydrocortisone treatment, growth velocity decreases slightly during the first 9 months and H-SDS then approaches the genetic height potential.

Biochemical, molecular and preclinical characterization of a double-virus-reduced human butyrylcholinesterase preparation designed for clinical use.

BACKGROUND AND OBJECTIVES: A human plasma-derived butyrylcholinesterase preparation manufactured on the industrial scale is described. MATERIAL AND METHODS: The human butyrylcholinesterase (hBChE) product was extensively investigated for its purity using immunological and electrophoretic methods and characterized by thorough glycoproteomic approaches. A comprehensive preclinical testing programme addressing safety and pharmacokinetic parameters supplemented the biochemical characterization. RESULTS: The high-purity hBChE preparation is tetrameric and has high specific activity and molecular integrity of the protein backbone. Acute toxicity studies and in vivo thrombogenicity studies provided evidence of a sufficient safety margin for use in humans. CONCLUSION: Extensive preclinical safety and pharmacokinetic testing confirmed that this hBChE preparation can be used for further efficacy testing as a bioscavenger for toxic organophosphate compounds in appropriate animal models and ultimately in humans.

Modulation of factor VIII-specific memory B cells.

The development of inhibitory antibodies against factor VIII (FVIII) is the major complication in patients with haemophilia A who are treated with FVIII products. Memory B cells play an essential role in maintaining established antibody responses. Upon re-exposure to the same antigen, they are rapidly re-stimulated to proliferate and differentiate into antibody-secreting plasma cells (ASC) that secrete high-affinity antibodies. It is, therefore, reasonable to believe that memory B cells have to be eradicated or inactivated for immune tolerance induction therapy to be successful in patients with haemophilia A and FVIII inhibitors. The aim of our studies was the development of strategies to prevent FVIII-specific memory B cells from becoming re-stimulated. We established a 6-day in vitro culture system that enabled us to study the regulation of FVIII-specific murine memory-B-cell re-stimulation. We tested the impact of the blockade of co-stimulatory interactions, of different concentrations of FVIII and of ligands for toll-like receptors (TLR). The blockade of B7-CD28 and CD40-CD40 ligand interactions prevented FVIII-specific murine memory B cells from becoming re-stimulated by FVIII in vitro and in vivo. Furthermore, high concentrations of FVIII blocked re-stimulation of FVIII-specific murine memory B cells. Triggering of TLR7 amplified re-stimulation by low concentrations of FVIII and prevented blockade by high concentrations of FVIII. We conclude that we defined modulators that either amplify or inhibit the re-stimulation of FVIII-specific murine memory B cells. Currently, we are investigating whether the same modulators operate in patients with haemophilia A and FVIII inhibitors.

Optimization, refinement and reduction of murine in vivo experiments to assess therapeutic approaches for haemophilia A.

The tail cut bleeding model (CUT) is routinely used in factor VIII-deficient mice to assess pharmacodynamic effects of therapeutic strategies for haemophilia A. Results from this model are highly variable, many modifications to the model are reported and at times the animals' wellbeing may be compromised by recording survival as an endpoint. We therefore investigated if the ferric chloride carotid occlusion model (COM) used for thrombosis research can be applied to enhance data quality and animal welfare in haemophilia A research. Relative dose effects and relative dose variations were calculated for the CUT and COM. The requisite sample sizes were estimated and the importance of survival rates to assess rebleeds during recovery was evaluated by correlating initial blood loss to mortality. Relative dose effects increased with higher doses in both models. The COM was more sensitive at lower doses than the CUT, had up to 82% less variation across doses and clearly showed superior accuracy. Only 5% of the sample size required for the CUT would be needed to establish non-inferiority between a specific therapeutic dose in haemophilia A mice and healthy wild-type animals. A strong statistically significant correlation was found between initial blood loss and mortality within 24 h. Our findings clearly suggest that the COM is a valid tool for assessing haemophilia A treatment in vivo. The highly reproducible data means that significantly fewer animals are required and a more humane endpoint can be used by directly assessing clot stability instead of survival rate.

No correlation between androgen receptor CAG and GGN repeat length and the degree of genital virilization in females with 21-hydroxylase deficiency.

CONTEXT: In 21-hydroxylase (CYP21A2) deficiency (21OHD), the level of in vitro enzymatic function allows for classification of mutation groups (null, A, B, C) and prediction of disease severity. However, genital virilization in affected females correlates only weakly with CYP21A2 mutation groups, suggesting the influence of genetic modifiers. OBJECTIVE: The objective of the study was to investigate the influence of the polymorphic CAG and GGn repeats of the androgen receptor (AR) gene on the degree of genital virilization in 21OHD females. DESIGN AND PATIENTS: Design of the study was the determination of CYP21A2 genotype, degree of genital virilization (Prader stage), and X-weighted biallelic mean of AR CAG and GGn repeat length in 205 females with 21OHD. OUTCOME MEASUREMENTS: Correlation of AR CAG and GGn repeat lengths with Prader stages using nested stepwise logistic regression analysis was measured. RESULTS: CYP21A2 mutation groups null and A showed significantly higher levels of genital virilization than groups B and C (P < 0.01). However, Prader stages varied considerably within mutation groups: null, Prader I-V (median IV); A, Prader I-V (median IV); B, Prader I-V (median III); C, 0-III (median I). Mean GGn repeat length of patients was not significantly associated with Prader stages, classified as low (0-I), intermediate (II-III), or severe (IV-V) (odds ratio per repeat: 0.98, 95% confidence interval 0.71-1.35). In contrast, patients with Prader 0-I showed a trend toward longer CAG repeats without reaching statistical significance (P = 0.07, odds ratio per repeat: 0.82, 95% confidence interval 0.65-1.02). CONCLUSION: Neither CAG nor GGn repeat lengths are statistically significant modifiers of genital virilization in females with 21OHD.

Hormone therapy affects plasma measures of factor VII-activating protease in younger postmenopausal women.

OBJECTIVES: Current reviews indicate that hormone therapy (HT) has a protective role in coronary heart disease (CHD) in younger postmenopausal women, whereas HT contributes to CHD in older women. Factor VII-activating protease (FSAP) is a serine protease that accumulates in unstable atherosclerotic plaques. FSAP is presumably involved in plaque stability and rupture. Reduced plasma concentration of FSAP may be associated with the development and expression of atherosclerosis and may thus contribute to precipitation of CHD. Here we address the potential influence of various HT regimens on plasma measures of FSAP in postmenopausal women treated for 1 year with different HT formulations or no HT. METHODS: Six groups of postmenopausal women (n = 139) were allocated to five different HT modalities or no HT. Samples were collected at baseline and after 12 months of treatment. Prototype assays were used for the determination of FSAP antigen and FSAP activity. RESULTS: The FSAP measures were comparable at baseline. No significant changes were observed in the control group after 12 months. HT in general induced a significant increase in FSAP antigen (7.7 microg/ml at baseline and 8.0 microg/ml after 12 months, p = 0.05), FSAP activity (1.54 PEU/ml at baseline and 1.68 PEU/ml after 12 months, p < 0.001) and FSAP ratio (202 mPEU/microg at baseline and 210 mPEU/microg after 12 months, p = 0.01). CONCLUSIONS: HT increases the plasma measures of FSAP. This increase may contribute to the protective effect on CHD induced by HT in younger postmenopausal women.

CD137 enhances monocyte-ICAM-1 interactions in an E-selectin-dependent manner under flow conditions.

Expression of CD137, a member of the tumor necrosis factor receptor family, is inducible on vascular endothelial cells by proinflammatory stimuli. Its ligand is expressed as a transmembrane protein on the surface of monocytes, and transmits activating signals into monocytes (reverse signaling) inducing monocyte migration. These findings led to the hypothesis that CD137 expression on activated endothelial cells facilitates recruitment of monocytes into inflammatory tissues including atherosclerotic lesions. Data from this study demonstrate that CD137 expression is inducible on endothelial cells by TNF stimulation. Recombinant CD137 protein and CD137 expressed on activated endothelial cells enhance ICAM-1-mediated adhesion of monocytes under defined flow conditions in vitro. CD137 seems not to play a role in tethering of monocytes since this activity is completely E-selectin-dependent. In addition, LFA-1 affinity and clustering on monocytic cells is enhanced by CD137. In summary, CD137 expression is induced on vascular endothelial cells by proinflammatory mediators and strengthens ICAM-1 and LFA-1-mediated adhesion of monocytes. These data support a role for CD137 in the recruitment of monocytes to inflammatory tissues.

Effects of growth hormone treatment in juvenile idiopathic arthritis: bone and body composition.

BACKGROUND: Inflammation and glucocorticoid therapy are major factors influencing growth and bone maturation in patients with juvenile idiopathic arthritis (JIA). In addition to alterations in total bone mineral density and bone geometry, longitudinal data confirm that the main contributors to errant bone maturation in patients with JIA are reductions in muscle mass and force. Growth hormone (GH) therapy, which has shown efficacy in controlling disease, may also positively influence body composition. For several years, GH therapy has been used to treat growth retardation in patients with JIA receiving glucocorticoids. GH therapy normalizes growth velocity, increases height, bone mineral density and bone mass and changes bone geometry. Despite ongoing glucocorticoid therapy, muscle mass and bone size substantially increase with GH therapy. Increased bone size suggests improved bone stability, which may reduce fracture risk. Along with the increase in muscle mass, patients experience stabilized or slightly decreased fat mass during GH therapy. CONCLUSIONS: All these effects suggest an anabolic effect of GH therapy on bone and body composition.

Development of a plasma- and albumin-free recombinant von Willebrand factor.

Baxter has developed a recombinant therapy for treating von Willebrand's disease. Recombinant VWF is co-expressed with the rFVIII in CHO cells used to produce the rFVIII product Advate. This rVWF is used as a drug component for a rVWF-rFVIII complex drug product. CHO cells produce partially processed and partially un-processed rVWF still containing the pro-peptide. In order to make a consistent preparation containing mature and processed rVWF only rVWF is exposed to recombinant furin to remove the pro-peptide. Recombinant VWF and furin are produced under serum- and protein-free conditions. It is highly purified by a series of chromatographic steps and formulated in a protein-free buffer and has a homogeneous multimer distribution. The specific activity is higher in rVWF than in commercial plasma-derived VWF-FVIII complex products. SDS agarose electrophoretic analysis shows the presence of ultra-high molecular weight multimers. The FVIII-binding capacity and affinity of rVWF to FVIII is comparable to VWF in plasma. Carbohydrate analysis shows an intact glycosylation pattern. Recombinant VWF binds to collagen and promotes platelet adhesion under shear stress. It stabilizes endogenous FVIII in VWF-deficient knock-out mice as seen by a secondary rise in murine FVIII.