Efficacy and safety of tanezumab monotherapy or combined with non-steroidal anti-inflammatory drugs in the treatment of knee or hip osteoarthritis pain.

OBJECTIVE: To evaluate whether subjects with knee or hip osteoarthritis (OA) pain on non-steroidal anti-inflammatory drugs (NSAIDs) received greater benefit when tanezumab monotherapy replaced or was coadministered with NSAIDs. METHODS: Subjects (N=2700) received intravenous tanezumab (5 or 10 mg) or placebo every 8 weeks with or without oral naproxen 500 mg twice daily or celecoxib 100 mg twice daily. Efficacy was assessed as change from baseline to week 16 in three co-primary endpoints: Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) Pain, WOMAC Physical Function and Patient's Global Assessment (PGA) of OA. Safety assessments included adverse events, physical and neurological examinations, laboratory tests and vital signs. RESULTS: Although all tanezumab treatments provided significant improvements in WOMAC Pain and Physical Function over either NSAID alone, only tanezumab+NSAIDs were significant versus NSAIDs with PGA and met the prespecified definition of superiority. Combination treatment did not substantially improve pain or function over tanezumab monotherapy. Adverse event frequency was higher with tanezumab than with NSAIDs and highest with combination therapy. Higher incidence of all-cause total joint replacements occurred with tanezumab+NSAID versus tanezumab monotherapy or NSAIDs. Rapidly progressive OA incidence was significantly greater versus NSAID in all tanezumab groups except tanezumab 5 mg monotherapy. CONCLUSIONS: Subjects receiving partial symptomatic relief of OA pain with NSAIDs may receive greater benefit with tanezumab monotherapy. While only coadministration of tanezumab with NSAIDs met the definition of superiority, combination treatment did not provide important benefits over tanezumab monotherapy; small differences in efficacy were negated by treatment-limiting or irreversible safety outcomes. TRIAL REGISTRATION NUMBER: NCT00809354.

Resolution of Discoid Lupus Alopecia With Systemic Hydroxychloroquine and Topical Pimecrolimus Combination Therapy.

Discoid lupus erythematosus (DLE) is an autoimmune skin condition that is typically part of the cutaneous manifestation of systemic lupus erythematosus (SLE). DLE is characterized by erythematous patches that can progress to depigmentation and alopecia, leading to scarring and permanent hair loss if left untreated. Herein, we present a unique case of localized DLE on the scalp in a 46-year-old female with no prior history of autoimmune disorders. The patient underwent several medication trials, including intralesional corticosteroids, topical calcineurin inhibitors, topical corticosteroids, and systemic hydroxychloroquine, with limited success in treating her discoid alopecia. Subsequently, a combination therapy of oral hydroxychloroquine and topical pimecrolimus significantly improved her scalp lesion. This case highlights the efficacy of combination therapy in managing localized DLE, providing valuable insights for future research focused on DLE alopecia management and optimizing treatment strategies for similar cases.

How confident are UK radiographers at performing paediatric computed tomography trauma scans?

INTRODUCTION: Approximately 45% of paediatric deaths in the United Kingdom (UK) were as a result of trauma. Computed tomography (CT) provides time efficient and accurate diagnosis, increasing chances of survival. Whilst use of CT in evaluating paediatric trauma has been invaluable it carries significant radiation risks, largely because children have greater radiation sensitivity than adults. Although national paediatric trauma workload in the UK is proportionately low, the majority of paediatric patients are conveyed to hospitals which predominantly undertake CT scans on adult patients. This research aimed to determine the confidence levels of radiographers when performing paediatric CT trauma scans in three public hospitals in the UK, and whether a teaching intervention improved their perceived self-confidence. METHODS: Individual questionnaires containing eight qualitative and quantitative questions were used to ascertain radiographers' perceived confidence levels. A teaching intervention was developed based on responses. A post-intervention questionnaire was used to determine whether radiographers' confidence levels had improved. RESULTS: Radiographers (n = 45) reported a mean confidence score of 5.6 (standard deviation 2.2) and 8.0 (standard deviation 1.7) scanning paediatric trauma patients pre- and post-intervention respectively. A paired two group t-test found this difference to be statistically significant at p < .00001. Radiographers reported several factors which negatively influenced confidence levels, including limited experience and postgraduate education. CONCLUSION: Radiographers reported to be less confident scanning paediatric CT trauma patients compared to adults, pre- and post-intervention, however this research does not clarify whether this is as a result of an increase in competence. Further research regarding this concept warrants investigation. IMPLICATIONS FOR PRACTICE: Results suggest further training based on negative factors reported by radiographers can increase confidence when performing this type of scan, assisting radiographers in optimising paediatric patient doses.

Sustained survival of human hepatocytes in mice: A model for in vivo infection with human hepatitis B and hepatitis delta viruses.

Persistence of hepatocytes transplanted into the same or related species has been established. The long-term engraftment of human hepatocytes into rodents would be useful for the study of human viral hepatitis, where it might allow the species, technical and size limitations of the current animal models to be overcome. Although transgenic mice expressing the hepatitis B virus (HBV) genome produce infectious virus in their serum, the viral life cycle is not complete, in that the early stages of viral binding and entry into hepatocytes and production of an episomal transcriptional DNA template do not occur. As for hepatitis delta virus (HDV), another cause of liver disease, no effective therapy exists to eradicate infection, and it remains resistant even to recent regimens that have considerably changed the treatment of HBV (ref. 13). Here, we demonstrate long-term engraftment of primary human hepatocytes transplanted in a matrix under the kidney capsule of mice with administration of an agonistic antibody against c-Met. These mice were susceptible to HBV infection and completion of the viral life cycle. In addition, we demonstrate super-infection of the HBV-infected mice with HDV. Our results describe a new xenotransplant model that allows study of multiple aspects of human hepatitis viral infections, and may enhance studies of human liver diseases.

αvß8 integrin-expression by BATF3-dependent dendritic cells facilitates early IgA responses to Rotavirus.

Secretory intestinal IgA can protect from re-infection with rotavirus (RV), but very little is known about the mechanisms that induce IgA production during intestinal virus infections. Classical dendritic cells (cDCs) in the intestine can facilitate both T cell-dependent and -independent secretory IgA. Here, we show that BATF3-dependent cDC1, but not cDC2, are critical for the optimal induction of RV-specific IgA responses in the mesenteric lymph nodes. This depends on the selective expression of the TGFß-activating integrin αvß8 by cDC1. In contrast, αvß8 on cDC1 is dispensible for steady state immune homeostasis. Given that cDC2 are crucial in driving IgA during steady state but are dispensable for RV-specific IgA responses, we propose that the capacity of DC subsets to induce intestinal IgA responses reflects the context, as opposed to an intrinsic property of individual DC subsets.

Lymphocyte CC chemokine receptor 9 and epithelial thymus-expressed chemokine (TECK) expression distinguish the small intestinal immune compartment: Epithelial expression of tissue-specific chemokines as an organizing principle in regional immunity.

The immune system has evolved specialized cellular and molecular mechanisms for targeting and regulating immune responses at epithelial surfaces. Here we show that small intestinal intraepithelial lymphocytes and lamina propria lymphocytes migrate to thymus-expressed chemokine (TECK). This attraction is mediated by CC chemokine receptor (CCR)9, a chemoattractant receptor expressed at high levels by essentially all CD4(+) and CD8(+) T lymphocytes in the small intestine. Only a small subset of lymphocytes in the colon are CCR9(+), and lymphocytes from other tissues including tonsils, lung, inflamed liver, normal or inflamed skin, inflamed synovium and synovial fluid, breast milk, and seminal fluid are universally CCR9(-). TECK expression is also restricted to the small intestine: immunohistochemistry reveals that intense anti-TECK reactivity characterizes crypt epithelium in the jejunum and ileum, but not in other epithelia of the digestive tract (including stomach and colon), skin, lung, or salivary gland. These results imply a restricted role for lymphocyte CCR9 and its ligand TECK in the small intestine, and provide the first evidence for distinctive mechanisms of lymphocyte recruitment that may permit functional specialization of immune responses in different segments of the gastrointestinal tract. Selective expression of chemokines by differentiated epithelium may represent an important mechanism for targeting and specialization of immune responses.

Variation in antagonism of the interferon response to rotavirus NSP1 results in differential infectivity in mouse embryonic fibroblasts.

Rotavirus NSP1 has been shown to function as an E3 ubiquitin ligase that mediates proteasome-dependent degradation of interferon (IFN) regulatory factors (IRF), including IRF3, -5, and -7, and suppresses the cellular type I IFN response. However, the effect of rotavirus NSP1 on viral replication is not well defined. Prior studies used genetic analysis of selected reassortants to link NSP1 with host range restriction in the mouse, suggesting that homologous and heterologous rotaviruses might use their different abilities to antagonize the IFN response as the basis of their host tropisms. Using a mouse embryonic fibroblast (MEF) model, we demonstrate that heterologous bovine (UK and NCDV) and porcine (OSU) rotaviruses fail to effectively degrade cellular IRF3, resulting in IRF3 activation and beta IFN (IFN-beta) secretion. As a consequence of this failure, replication of these viruses is severely restricted in IFN-competent wild-type, but not in IFN-deficient (IFN-alpha/beta/gamma receptor- or STAT1-deficient) MEFs. On the other hand, homologous murine rotaviruses (ETD or EHP) or the heterologous simian rotavirus (rhesus rotavirus [RRV]) efficiently degrade cellular IRF3, diminish IRF3 activation and IFN-beta secretion and are not replication restricted in wild-type MEFs. Genetic reassortant analysis between UK and RRV maps the distinctive phenotypes of IFN antagonism and growth restriction in wild-type MEFs to NSP1. Therefore, there is a direct relationship between the replication efficiencies of different rotavirus strains in MEFs and strain-related variations in NSP1-mediated antagonism of the type I IFN response.

Assessment of oral midazolam limited sampling strategies to predict area under the concentration time curve (AUC) during cytochrome P450 (CYP) 3A baseline, inhibition and induction or activation.

UNLABELLED: A previous study reported a 2- and 3-timepoint limited sampling strategy (LSS) model accurately predicted oral midazolam area under the concentration time curve (AUC), and thus cytochrome P450 (CYP) 3A activity. OBJECTIVE: This study evaluated whether the LSS models predict midazolam AUC during CYP3A baseline, inhibition and induction/activation. MATERIALS AND METHODS: Plasma midazolam concentrations from 106 healthy adults from 6 published studies were obtained where oral midazolam was co-administered alone or with ketoconazole, double-strength grapefruit juice, Ginkgo biloba extract, pleconaril, or rifampin. Observed and predicted midazolam AUCs were determined. Bias and precision of the LSS models were determined. RESULTS: Contrasting results were observed for the 2- and 3-timepoint LSS models in accurately predicting midazolam AUC during baseline CYP3A conditions. With the exception of 1 study (single dose, double-strength grapefruit juice), the 2- and 3-timepoint LSS models did not accurately predict midazolam AUC during conditions of CYP3A inhibition and induction/activation. CONCLUSION: The previously reported 2- and 3-timepoint oral midazolam LSS models are not applicable to the evaluated conditions of CYP3A baseline, inhibition, and induction/ activation.

Role of interferon in homologous and heterologous rotavirus infection in the intestines and extraintestinal organs of suckling mice.

Recent studies demonstrated that viremia and extraintestinal rotavirus infection are common in acutely infected humans and animals, while systemic diseases appear to be rare. Intraperitoneal infection of newborn mice with rhesus rotavirus (RRV) results in biliary atresia (BA), and this condition is influenced by the host interferon response. We studied orally inoculated 5-day-old suckling mice that were deficient in interferon (IFN) signaling to evaluate the role of interferon on the outcome of local and systemic infection after enteric inoculation. We found that systemic replication of RRV, but not murine rotavirus strain EC, was greatly enhanced in IFN-alpha/beta and IFN-gamma receptor double-knockout (KO) or STAT1 KO mice but not in mice deficient in B- or T-cell immunity. The enhanced replication of RRV was associated with a lethal hepatitis, pancreatitis, and BA, while no systemic disease was observed in strain EC-infected interferon-deficient mice. In IFN-alpha/beta receptor KO mice the extraintestinal infection and systemic disease were only moderately increased, while RRV infection was not augmented and systemic disease was not present in IFN-gamma receptor KO mice. The increase of systemic infection in IFN-deficient mice was also observed during simian strain SA11 infection but not following bovine NCDV, porcine OSU, or murine strain EW infection. Our data indicate that the requirements for the interferon system to inhibit intestinal and extraintestinal viral replication in suckling mice vary among different heterologous and homologous rotavirus strains, and this variation is associated with lethal systemic disease.

Three-dimensional structure of rhesus rotavirus by cryoelectron microscopy and image reconstruction.

The structure of rhesus rotavirus was examined by cryoelectron microscopy and image analysis. Three-dimensional reconstructions of infectious virions were computed at 26- and 37-A resolution from electron micrographs recorded at two different levels of defocus. The major features revealed by the reconstructions are (a) both outer and inner capsids are constructed with T = 13l icosahedral lattice symmetry; (b) 60 spikelike projections, attributed to VP4, extend at least 100 A from the outer capsid surface; (c) the outer capsid, attributed primarily to VP7, has a smoothly rippled surface at a mean radius of 377 A and is perforated by 132 aqueous holes ranging from 40-65 A in diameter; (d) the inner capsid has a "bristled" outer surface composed of 260 trimeric-shaped columns of density, attributed to VP6, which merge with a smooth, spherical shell of density at a lower, mean radius of 299 A, and which is perforated by holes in register with those in the outer capsid; (e) a "core" region contains a third, nonspherical shell of density at a mean radius of 225 A that encapsidates the double-stranded RNA genome; and (f) the space between the outer and inner capsids forms an open aqueous network that may provide pathways for the diffusion of ions and small regulatory molecules as well as the extrusion of RNA. The assignment of different viral structural proteins to specific features of the reconstruction has been tentatively made on the basis of excluded volume estimates and previous biochemical characterizations of rotavirus.

Protective effect of rotavirus VP6-specific IgA monoclonal antibodies that lack neutralizing activity.

Rotaviruses are the leading cause of severe gastroenteritis and dehydrating diarrhea in young children and animals worldwide. A murine model and "backpack tumor" transplantation were used to determine the protective effect of antibodies against VP4(an outer capsid viral protein) and VP6(a major inner capsid viral protein). Only two non-neutralizing immunoglobulin A (IgA) antibodies to VP6 were capable of preventing primary and resolving chronic murine rotavirus infections. These antibodies were not active, however, when presented directly to the luminal side of the intestinal tract. These findings support the hypothesis that in vivo intracellular viral inactivation by secretory IgA during transcytosis is a mechanism of host defense against rotavirus infection.

Human rotavirus type 2: cultivation in vitro.

A strain of type 2 human rotavirus (Wa) was grown to relatively high titer through 14 passages in primary cultures of African green monkey kidney (AGMK) cells. This passage series was initiated with virus that had been passaged 11 times serially in newborn gnotobiotic piglets. In contrast, virus present in the stool of patient Wa as well as virus from the first, second, or third passage in piglets could not be propagated successfully in African green monkey kidney cells. Prior to each passage in cell culture, the virus was treated with trypsin and the inoculated cultures were centrifuged at low speed. Cultivation of a type 2 human rotavirus should aid attempts to characterize this virus and to develop a means of immunoprophylaxis for a serious diarrheal disease of human infants.

Molecular basis of age-dependent gastric inactivation of rhesus rotavirus in the mouse.

Rotavirus requires specific proteolytic activation by trypsin for efficient replication in tissue culture. To observe the nature of intestinal proteolytic activation of rotavirus in vivo, metabolically labeled rhesus rotavirus (RRV) grown in the presence of trypsin inhibitors was administered to adult and 10-d-old suckling mice by gavage. In the adult stomach, vp4 was cleaved in a manner distinct from in vitro trypsin cleavage. In the suckling stomach, RRV vp4 remains largely uncleaved. The alternative cleavage in the adult stomach was associated with a profound decrease in viral infectivity. vp4 from RRV recovered from the suckling small intestinal lumen was cleaved in a pattern similar or identical to in vitro trypsin-activated virus with bands comigrating with vp5* and vp8*. In contrast, vp4 was not observed in any recognizable form in RRV recovered from adult intestines. Comparison of infectivity of virus recovered from suckling and adult intestines revealed a 10,000-fold decrease in titer in the virus recovered from the adult intestine. In vitro digestions of RRV revealed that pepsin digestion can cleave RRV vp4 and markedly enhance acid-induced loss of rotavirus infectivity. Subsequent digestion with chymotrypsin removes most of the pepsin cleavage products of vp4. Virus injected directly into jejunal loops of adult mice and virus administered orally to adult mice pretreated with antiacid drugs retained infectivity. These studies indicate the development of gastric acid and pepsin secretion may be an important host defense factor in rotavirus gastroenteritis.

Expression of mucosal homing receptor alpha4beta7 by circulating CD4+ cells with memory for intestinal rotavirus.

The integrin alpha4beta7 mediates lymphocyte binding to mucosal addressin cell adhesion molecule-1, and its expression defines lymphocytes capable of trafficking through the intestines and the intestinal lymphoid tissues. We examined the ability of discrete alpha4beta7(hi) and alpha4beta7- subsets of circulating memory phenotype (CD45RA-) CD4+ T cells to proliferate in response to rotavirus, a ubiquitous intestinal pathogen. alpha4beta7(hi) memory (CD45RA-) CD4+ T cells displayed much greater reactivity to rotavirus than alpha4beta7- memory or naive (CD45RA+) CD4+ T cells. In contrast, alpha4beta7- memory cells were the predominant population responsive to mumps antigen after intramuscular vaccination. Our results are consistent with the conclusion that natural rotavirus infection, an enteric pathogen, results in a specific circulating memory CD4+ response that is largely limited to the gut-homing alpha4beta7+ subpopulation. This phenotype is not shared with memory cells elicited by intramuscular immunization (shown here) or by skin contact allergens. The results support the hypothesis that gut trafficking memory CD4+ T cells comprise cellular memory for intestinal antigens and suggest that regulated expression of alpha4beta7 helps target and segregate intestinal versus systemic immune response.

Liposome-mediated transfection of intact viral particles reveals that plasma membrane penetration determines permissivity of tissue culture cells to rotavirus.

Rotaviruses are an important cause of gastroenteritis in human infants. In vivo, rotavirus displays striking cell tropism with viral replication generally restricted to the villus tip enterocytes of the small intestine. We studied a panel of cell lines that vary significantly in their permissivity to rotavirus infection. L cells and HEp2 cells were relatively resistant to rotavirus infection compared with permissive Ma104 cells and HT29 cells. RNA transcription among the cell lines was proportional to antigen synthesis making a translational or posttranslational block an unlikely source of observed differences in susceptibility. All of the cell lines bound and internalized radiolabeled virus equally well, as measured by escape from surface protease treatment. Analysis of the escape of cell bound virus from neutralizing monoclonal antibody revealed that rotavirus did not immediately enter an eclipse phase in nonpermissive cells, but was internalized in an infectious form for several hours, possibly sequestered within endocytic vacuoles. L cells and HEp2 cells were as permissive as Ma104 and HT29 cells when rotavirus infection was mediated by transfection of single- or double-shelled rotavirus particles with cationic liposomes (Lipofectin). Rotavirus cell tropism in tissue culture cells is determined by the ability of infecting virions to traverse the plasma membrane of the cells into the cytoplasmic compartment.

The isolation and characterization of a Norwalk virus-specific cDNA.

Norwalk virus, an important cause of epidemic, acute, nonbacterial gastroenteritis in adults and children, has eluded adaptation to tissue culture, the development of an animal model, and molecular cloning. In this study, a portion of the Norwalk viral genome encoding an immunoreactive region was cloned from very small quantities of infected stool using sequence-independent single primer amplification. Six overlapping complementary DNA (cDNA) clones were isolated by immunologic screening. The expressed recombinant protein from a representative clone reacted with six of seven high titer. Norwalk-specific, postinfection sera but not with corresponding preinfection sera. Nucleic acid sequence for all clones defined a single open reading frame contiguous with the lambda gt11-expressed beta-galactosidase protein. Only oligonucleotide probes specific for the positive strand (defined by the open reading frame) hybridized to an RNaseA-sensitive, DNaseI-resistant nucleic acid sequence extracted from Norwalk-infected stool. Furthermore, RNA extracted from serial postinfection, but not preinfection, stools from three of five volunteers hybridized to a Norwalk virus cDNA probe. Clone-specific oligonucleotide probes hybridized with cesium chloride gradient fractions containing purified Norwalk virion. In conclusion, an antigenic, protein-coding region of the Norwalk virus genome has been identified. This epitope has potential utility in future sero- and molecular epidemiologic studies of Norwalk viral gastroenteritis.

Bonzo/CXCR6 expression defines type 1-polarized T-cell subsets with extralymphoid tissue homing potential.

Chemokine receptor expression is finely controlled during T-cell development. We show that newly identified chemokine receptor Bonzo/CXCR6 is expressed by subsets of Th1 or T-cytotoxic 1 (Tc1) cells, but not by Th2 or Tc2 cells, establishing Bonzo as a differential marker of polarized type 1 T cells in vitro and in vivo. Priming of naive T cells by dendritic cells induces expression of Bonzo on T cells. IL-12 enhances this dendritic cell-dependent upregulation, while IL-4 inhibits it. In blood, 35-56% of Bonzo+ CD4 T cells are Th1 cells, and 60-65% of Bonzo+ CD8 T cells are Tc1 cells, while few Bonzo+ cells are type 2 T cells. Almost all Bonzo+ Tc1 cells contain preformed granzyme A and display cytotoxic effector phenotype. Most Bonzo+ T cells lack L-selectin and/or CCR7, homing receptors for lymphoid tissues. Instead, Bonzo+ T cells are dramatically enriched among T cells in tissue sites of inflammation, such as rheumatoid joints and inflamed livers. Bonzo may be important in trafficking of effector T cells that mediate type 1 inflammation, making it a potential target for therapeutic modulation of inflammatory diseases.