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IKKalpha negatively regulates ASC-dependent inflammasome activation

Mon, 12/08/2014 - 4:39pm

The inflammasomes are multiprotein complexes that activate caspase-1 in response to infections and stress, resulting in the secretion of pro-inflammatory cytokines. Here we report that IkappaB kinase alpha (IKKalpha) is a critical negative regulator of apoptosis-associated specklike protein containing a C-terminal caspase-activation-andrecruitment (CARD) domain (ASC)-dependent inflammasomes. IKKalpha controls the inflammasome at the level of the adaptor ASC, which interacts with IKKalpha in the nucleus of resting macrophages in an IKKalpha kinase-dependent manner. Loss of IKKalpha kinase activity results in inflammasome hyperactivation. Mechanistically, the downstream nuclear effector IKK-related kinase (IKKi) facilitates translocation of ASC from the nucleus to the perinuclear area during inflammasome activation. ASC remains under the control of IKKalpha in the perinuclear area following translocation of the ASC/IKKalpha complex. Signal 2 of NLRP3 activation leads to inhibition of IKKalpha kinase activity through the recruitment of PP2A, allowing ASC to participate in NLRP3 inflammasome assembly. Taken together, these findings reveal a IKKi-IKKalpha-ASC axis that serves as a common regulatory mechanism for ASC-dependent inflammasomes.

Innate sensing of malaria parasites

Mon, 12/08/2014 - 4:39pm

Innate immune receptors have a key role in immune surveillance by sensing microorganisms and initiating protective immune responses. However, the innate immune system is a classic 'double-edged sword' that can overreact to pathogens, which can have deleterious effects and lead to clinical manifestations. Recent studies have unveiled the complexity of innate immune receptors that function as sensors of Plasmodium spp. in the vertebrate host. This Review highlights the cellular and molecular mechanisms by which Plasmodium infection is sensed by different families of innate immune receptors. We also discuss how these events mediate both host resistance to infection and the pathogenesis of malaria.

Genomic occupancy of Runx2 with global expression profiling identifies a novel dimension to control of osteoblastogenesis

Thu, 12/04/2014 - 4:08pm

BACKGROUND: Osteogenesis is a highly regulated developmental process and continues during the turnover and repair of mature bone. Runx2, the master regulator of osteoblastogenesis, directs a transcriptional program essential for bone formation through genetic and epigenetic mechanisms. While individual Runx2 gene targets have been identified, further insights into the broad spectrum of Runx2 functions required for osteogenesis are needed.

RESULTS: By performing genome-wide characterization of Runx2 binding at the three major stages of osteoblast differentiation--proliferation, matrix deposition and mineralization--we identify Runx2-dependent regulatory networks driving bone formation. Using chromatin immunoprecipitation followed by high-throughput sequencing over the course of these stages, we identify approximately 80,000 significantly enriched regions of Runx2 binding throughout the mouse genome. These binding events exhibit distinct patterns during osteogenesis, and are associated with proximal promoters and also non-promoter regions: upstream, introns, exons, transcription termination site regions, and intergenic regions. These peaks were partitioned into clusters that are associated with genes in complex biological processes that support bone formation. Using Affymetrix expression profiling of differentiating osteoblasts depleted of Runx2, we identify novel Runx2 targets including Ezh2, a critical epigenetic regulator; Crabp2, a retinoic acid signaling component; Adamts4 and Tnfrsf19, two remodelers of the extracellular matrix. We demonstrate by luciferase assays that these novel biological targets are regulated by Runx2 occupancy at non-promoter regions.

CONCLUSIONS: Our data establish that Runx2 interactions with chromatin across the genome reveal novel genes, pathways and transcriptional mechanisms that contribute to the regulation of osteoblastogenesis.

Confirmatory factor analysis of the thyroid-related quality of life questionnaire ThyPRO

Thu, 12/04/2014 - 4:08pm

Background and aim. Thyroid diseases are prevalent and chronic. With treatment, quality of life is restored in most, but not all patients. Construct validity of the thyroid-related quality of life questionnaire, ThyPRO, has been established by multi-trait scaling, but not evaluated with more elaborate methods. The purpose of the present study was to evaluate dimensionality of the ThyPRO scales and to attempt to understand possible item misfit through structural equation modeling for categorical data.

Methods. The current 84-item version of ThyPRO consists of 13 scales, covering domains of physical (4 scales) and mental (2 scales) symptoms, function and well-being (3 scales) and participation/social function (4 scales). The data were collected from a cross-sectional sample of 907 thyroid patients. One-factor confirmatory models were fitted to each scale, and evaluated by model fit statistics (comparative fit index > 0.95, root mean square error of approximation < 0.08), magnitude of factor loadings, model residual correlations and modification indices (MI). Indications of multi-dimensionality were tested in bi-factor models. Possible item misfit was evaluated in a combined, investigational model.

Results. Each ThyPRO scale was adequately represented by a unidimensional model after minor revisions. Eleven items were identified in the unidimensional models as potentially misfitting and were investigated further by multidimensional modeling.

Conclusion. Elaborate psychometric modeling supported the construct validity of the ThyPRO. However, 11 potentially misfitting items and 18 items with local dependence to other items are candidates for removal in future item reduction processes.

The global translation profile in a ribosomal protein mutant resembles that of an eIF3 mutant

Thu, 12/04/2014 - 4:08pm

BACKGROUND: Genome-wide assays performed in Arabidopsis and other organisms have revealed that the translation status of mRNAs responds dramatically to different environmental stresses and genetic lesions in the translation apparatus. To identify additional features of the global landscape of translational control, we used microarray analysis of polysomal as well as non-polysomal mRNAs to examine the defects in translation in a poly(A) binding protein mutant, pab2 pab8, as well as in a mutant of a large ribosomal subunit protein, rpl24b/shortvalve1.

RESULTS: The mutation of RPL24B stimulated the ribosome occupancy of mRNAs for nuclear encoded ribosomal proteins. Detailed analysis yielded new insights into the translational regulon containing the ribosomal protein mRNAs. First, the ribosome occupancy defects in the rpl24b mutant partially overlapped with those in a previously analyzed initiation factor mutant, eif3h. Second, a group of mRNAs with incomplete coding sequences appeared to be uncoupled from the regulon, since their dependence on RPL24B differed from regular mRNAs. Third, different sister paralogs of the ribosomal proteins differed in their translation state in the wild-type. Some sister paralogs also differed in their response to the rpl24b mutation. In contrast to rpl24b, the pab2 pab8 mutant revealed few gene specific translational defects, but a group of seed storage protein mRNAs were stimulated in their ribosome occupancy. In the course of this work, while optimizing the statistical analysis of ribosome occupancy data, we collected 12 biological replicates of translation states from wild-type seedlings. We defined 20% of mRNAs as having a high variance in their translation state. Many of these mRNAs were functionally associated with responses to the environment, suggesting that subtle variation in the environmental conditions is sensed by plants and transduced to affect the translational efficiency of hundreds of mRNAs.

CONCLUSIONS: These data represent the first genome-wide analysis of translation in a eukaryote defective in the large ribosomal subunit. RPL24 and eIF3h play similar but non-identical roles in eukaryotic translation. The data also shed light on the fine structure of the regulon of ribosomal protein mRNAs.

Single point mutation in Rabenosyn-5 in a female with intractable seizures and evidence of defective endocytotic trafficking

Thu, 12/04/2014 - 4:08pm

Background. We report a 6.5 year-old female with a homozygous missense mutation in ZFYVE20, encoding Rabenosyn-5 (Rbsn-5), a highly conserved multi-domain protein implicated in receptor-mediated endocytosis. The clinical presentation includes intractable seizures, developmental delay, microcephaly, dysostosis, osteopenia, craniofacial dysmorphism, macrocytosis and megaloblastoid erythropoiesis. Biochemical findings include transient cobalamin deficiency, severe hypertriglyceridemia upon ketogenic diet, microalbuminuria and partial cathepsin D deficiency.

Methods and results. Whole exome sequencing followed by Sanger sequencing confirmed a rare (frequency:0.003987) homozygous missense mutation, g.15,116,371 G inverted question mark > inverted question markA (c.1273G inverted question mark > inverted question markA), in ZFYVE20 resulting in an amino acid change from Glycine to Arginine at position 425 of the Rbsn protein (p.Gly425Arg), as the only mutation segregating with disease in the family. Studies in fibroblasts revealed expression and localization of Rbsn-5G425R in wild-type manner, but a 50% decrease in transferrin accumulation, which is corrected by wild-type allele transfection. Furthermore, the patient inverted question marks fibroblasts displayed an impaired proliferation rate, cytoskeletal and lysosomal abnormalities.

Conclusion. These results are consistent with a functional defect in the early endocytic pathway resulting from mutation in Rbsn-5, which secondarily disrupts multiple cellular functions dependent on endocytosis, leading to a severe multi-organ disorder.

Preliminary evaluation of near infrared spectroscopy as a method to detect plasma leakage in children with dengue hemorrhagic fever

Thu, 12/04/2014 - 4:08pm

BACKGROUND: Dengue viral infections are prevalent in the tropical and sub-tropical regions of the world, resulting in substantial morbidity and mortality. Clinical manifestations range from a self-limited fever to a potential life-threatening plasma leakage syndrome (dengue hemorrhagic fever). The objective of this study was to assess the utility of near infrared spectroscopy (NIRS) measurements of muscle oxygen saturation (SmO2) as a possible continuous measure to detect plasma leakage in children with dengue.

METHODS: Children ages 6 months to 15 years of age admitted with suspected dengue were enrolled from the dengue ward at Queen Sirikit National Institute for Child Health. Children were monitored daily until discharge. NIRS data were collected continuously using a prototype CareGuide Oximeter 1100 with sensors placed on the deltoid or thigh. Daily ultrasound of the chest and a right lateral decubitus chest x-ray the day after defervescence were performed to detect and quantitate plasma leakage in the pleural cavity.

RESULTS: NIRS data were obtained from 19 children with laboratory-confirmed dengue. Average minimum SmO2 decreased for all subjects prior to defervescence. Average minimum SmO2 subsequently increased in children with no ultrasound evidence of pleural effusion but remained low in children with pleural effusion following defervescence. Average minimum SmO2 was inversely correlated with pleural space fluid volume. ROC analysis revealed a cut-off value for SmO2 which yielded high specificity and sensitivity.

CONCLUSIONS: SmO2 measured using NIRS may be a useful guide for real-time and non-invasive identification of plasma leakage in children with dengue. Further investigation of the utility of NIRS measurements for prediction and management of severe dengue syndromes is warranted.

NOTCH1 inhibition in vivo results in mammary tumor regression and reduced mammary tumorsphere-forming activity in vitro

Thu, 12/04/2014 - 4:08pm

INTRODUCTION: NOTCH activation has been recently implicated in human breast cancers, associated with a poor prognosis, and tumor-initiating cells are hypothesized to mediate resistance to treatment and disease relapse. To address the role of NOTCH1 in mammary gland development, transformation, and mammary tumor-initiating cell activity, we developed a doxycycline-regulated mouse model of NOTCH1-mediated mammary transformation.

METHODS: Mammary gland development was analyzed by using whole-mount analysis and by flow cytometry in nulliparous transgenic mice maintained in the presence/absence of doxycycline (or intracellular NOTCH1). Mammary tumors were examined histologically and immunophenotyped by staining with antibodies followed by flow cytometry. Tumors were transplanted into mammary fat pads under limiting dilution conditions, and tumor-initiating cell frequency was calculated. Mammary tumor cells were also plated in vitro in a tumorsphere assay in the presence/absence of doxycycline. RNA was isolated from mammary tumor cell lines cultured in the presence/absence of doxycycline and used for gene-expression profiling with Affymetrix mouse arrays. NOTCH1-regulated genes were identified and validated by using quantitative real-time polymerase chain reaction (PCR). Mammary tumor-bearing mice were treated with doxycycline to suppress NOTCH1 expression, and disease recurrence was monitored.

RESULTS: Similar to published studies, we show that constitutive expression of human intracellular NOTCH1 in the developing mouse mammary gland inhibits side branching and promotes luminal cell fate. These mice develop mammary adenocarcinomas that express cytokeratin (CK) 8/18. In vivo limiting-dilution analyses revealed that these mammary tumors exhibit functional heterogeneity and harbor a rare (1/2,978) mammary tumor-initiating cell population. With this dox-regulated NOTCH1 mammary tumor model, we demonstrate that NOTCH1 inhibition results in mammary tumor regression in vivo and prevents disease recurrence in four of six tumors tested. Consistent with the in vivo data, NOTCH1 inhibition reduces mammary tumorsphere activity in vitro. We also identify the embryonic stem cell transcription factor Nanog as a novel NOTCH1-regulated gene in tumorspheres and in mouse and human breast cancer cell lines.

CONCLUSIONS: These data indicate that NOTCH1 inhibition results in mammary tumor regression in vivo and interferes with disease recurrence. We demonstrate that NOTCH1-transformed mouse mammary tumors harbor a rare mammary tumor-initiating population and that NOTCH1 contributes to mammary tumor-initiating activity. This work raises the possibility that NOTCH therapeutics may target mammary tumor-initiating cells in certain human breast cancer subtypes.

Cancer-testis gene expression is associated with the methylenetetrahydrofolate reductase 677 C>T polymorphism in non-small cell lung carcinoma

Thu, 12/04/2014 - 4:08pm

BACKGROUND: Tumor-specific, coordinate expression of cancer-testis (CT) genes, mapping to the X chromosome, is observed in more than 60% of non-small cell lung cancer (NSCLC) patients. Although CT gene expression has been unequivocally related to DNA demethylation of promoter regions, the underlying mechanism leading to loss of promoter methylation remains elusive. Polymorphisms of enzymes within the 1-carbon pathway have been shown to affect S-adenosyl methionine (SAM) production, which is the sole methyl donor in the cell. Allelic variants of several enzymes within this pathway have been associated with altered SAM levels either directly, or indirectly as reflected by altered levels of SAH and Homocysteine levels, and altered levels of DNA methylation. We, therefore, asked whether the five most commonly occurring polymorphisms in four of the enzymes in the 1-carbon pathway associated with CT gene expression status in patients with NSCLC.

METHODS: Fifty patients among a cohort of 763 with NSCLC were selected based on CT gene expression status and typed for five polymorphisms in four genes known to affect SAM generation by allele specific q-PCR and RFLP.

RESULTS: We identified a significant association between CT gene expression and the MTHFR 677 CC genotype, as well as the C allele of the SNP, in this cohort of patients. Multivariate analysis revealed that the genotype and allele strongly associate with CT gene expression, independent of potential confounders.

CONCLUSIONS: Although CT gene expression is associated with DNA demethylation, in NSCLC, our data suggests this is unlikely to be the result of decreased MTHFR function.

GRAIDs: a framework for closing the gap in the availability of health promotion programs and interventions for people with disabilities

Thu, 12/04/2014 - 4:07pm

BACKGROUND: Evidence-based health promotion programs developed and tested in the general population typically exclude people with disabilities. To address this gap, a set of methods and criteria were created to adapt evidence-based health promotion programs for people with disabilities. In this first study, we describe a framework for adapting evidence-based obesity prevention strategies for people with disabilities. We illustrate how the framework has been used to adapt the U.S. Centers for Disease Control and Prevention's (CDC) obesity prevention strategies for individuals with physical and developmental disabilities.

METHODS: The development of inclusion guidelines, recommendations and adaptations for obesity prevention (referred to as GRAIDs--Guidelines, Recommendations, Adaptations Including Disability) consists of five components: (i) a scoping review of the published and grey literature; (ii) an expert workgroup composed of nationally recognized leaders in disability and health promotion who review, discuss and modify the scoping review materials and develop the content into draft GRAIDs; (iii) focus groups with individuals with disabilities and their family members (conducted separately) who provide input on the potential applicability of the proposed GRAIDs in real world settings; (iv) a national consensus meeting with 21 expert panel members who review and vote on a final set of GRAIDs; and (v) an independent peer review of GRAIDs by national leaders from key disability organizations and professional groups through an online web portal.

RESULTS: This is an ongoing project, and to date, the process has been used to develop 11 GRAIDs to coincide with 11 of the 24 CDC obesity prevention strategies.

CONCLUSION: A set of methods and criteria have been developed to allow researchers, practitioners and government agencies to promote inclusive health promotion guidelines, strategies and practices for people with disabilities. Evidence-based programs developed for people without disabilities can now be adapted for people with disabilities using the GRAIDs framework.

Modeling T cell receptor recognition of CD1-lipid and MR1-metabolite complexes

Thu, 12/04/2014 - 4:07pm

BACKGROUND: T cell receptors (TCRs) can recognize diverse lipid and metabolite antigens presented by MHC-like molecules CD1 and MR1, and the molecular basis of many of these interactions has not been determined. Here we applied our protein docking algorithm TCRFlexDock, previously developed to perform docking of TCRs to peptide-MHC (pMHC) molecules, to predict the binding of alphabeta and gammadelta TCRs to CD1 and MR1, starting with the structures of the unbound molecules.

RESULTS: Evaluating against TCR-CD1d complexes with crystal structures, we achieved near-native structures in the top 20 models for two out of four cases, and an acceptable-rated prediction for a third case. We also predicted the structure of an interaction between a MAIT TCR and MR1-antigen that has not been structurally characterized, yielding a top-ranked model that agreed remarkably with a characterized TCR-MR1-antigen structure that has a nearly identical TCR alpha chain but a different beta chain, highlighting the likely dominance of the conserved alpha chain in MR1-antigen recognition. Docking performance was improved by re-training our scoring function with a set of TCR-pMHC complexes, and for a case with an outlier binding mode, we found that alternative docking start positions improved predictive accuracy. We then performed unbound docking with two mycolyl-lipid specific TCRs that recognize lipid-bound CD1b, which represent a class of interactions that is not structurally characterized. Highly-ranked models of these complexes showed remarkable agreement between their binding topologies, as expected based on their shared germline sequences, while differences in residue-level interactions with their respective antigens point to possible mechanisms underlying their distinct specificities.

CONCLUSIONS: Together these results indicate that flexible docking simulations can provide accurate models and atomic-level insights into TCR recognition of MHC-like molecules presenting lipid and other small molecule antigens.

Institution specific risk factors for 30 day readmission at a community hospital: a retrospective observational study

Thu, 12/04/2014 - 4:07pm

BACKGROUND: As of October 1, 2012, hospitals in the United States with excess readmissions based on the Centers for Medicare and Medicaid Services (CMS) risk-adjusted ratio began being penalized. Given the impact of high readmission rates to hospitals nationally, it is important for individual hospitals to identify which patients may be at highest risk of readmission. The objective of this study was to assess the association of institution specific factors with 30-day readmission.

METHODS: The study is a retrospective observational study using administrative data from January 1, 2009 through December 31, 2010 conducted at a 257 bed community hospital in Massachusetts. The patients included inpatient medical discharges from the hospitalist service with the primary diagnoses of congestive heart failure, pneumonia or chronic obstructive pulmonary disease. The outcome was 30-day readmission rates. After adjusting for known factors that impact readmission, provider associated factors (i.e. hours worked and census on the day of discharge) and hospital associated factors (i.e. floor of discharge, season) were compared.

RESULTS: Over the study time period, there were 3774 discharges by hospitalists, with 637 30-day readmissions (17% readmission rate). By condition, readmission rates were 19.6% (448/2284) for congestive heart failure, 13.0% (141/1083) for pneumonia, and 14.7% (200/1358) for chronic obstructive lung disease. After adjusting for known risk factors (gender, age, length of stay, Elixhauser sum score, admission in the previous year, insurance, disposition, primary diagnosis), we found that patients discharged in the winter remained significantly more likely to be readmitted compared to the summer (OR 1.54, p = 0.0008). Patients discharged from the cardiac floor had a trend toward decreased readmission compared a medical/oncology floor (OR 0.85, p = 0.08). Hospitalist work flow factors (census and hours on the day of discharge) were not associated with readmission.

CONCLUSIONS: We found that 30 day hospital readmissions may be associated with institution specific risk factors, even after adjustment for patient factors. These institution specific risk factors may be targets for interventions to prevent readmissions.

Design characteristics of the CORRONA CERTAIN study: a comparative effectiveness study of biologic agents for rheumatoid arthritis patients

Thu, 12/04/2014 - 4:07pm

BACKGROUND: Comparative effectiveness research has recently attracted considerable attention. The Comparative Effectiveness Registry to study Therapies for Arthritis and Inflammatory Conditions (CERTAIN) is an ongoing prospective cohort study of adult patients with Rheumatoid Arthritis (RA).

METHODS/DESIGN: CERTAIN uses the existing Consortium of Rheumatology Researchers of North America (CORRONA) network of participating private and academic sites in order to recruit patients fulfilling the 1987 ACR criteria that have at least moderate disease activity. Patients starting or switching biologic agents either anti-TNF therapy or a non anti-TNF biologic are eligible for enrollment, depending on the treatment selected by their physician. Enrollment is expected to be completed by March of 2014, and 2711 patients will participate in the study. As of October 7th 2013, 2234 patients have been enrolled. Patient visits and laboratory blood work are mandated every three months for one year. Safety data is collected through one year and beyond. The primary comparative effectiveness endpoint is attainment of low RA disease activity at one year among patients who have been exposed to at least one prior TNF-alpha inhibitor agent prior to enrollment. Multiple secondary effectiveness and safety endpoints will be addressed by investigating the entire population enrolled (naive and biologic experienced).

DISCUSSION: The unique design features of CERTAIN will inform comparative effectiveness and safety questions for choosing biologic agents for the management of RA.

An anti-inflammatory diet as treatment for inflammatory bowel disease: a case series report

Thu, 12/04/2014 - 4:07pm

BACKGROUND: The Anti-Inflammatory Diet (IBD-AID) is a nutritional regimen for inflammatory bowel disease (IBD) that restricts the intake of certain carbohydrates, includes the ingestion of pre- and probiotic foods, and modifies dietary fatty acids to demonstrate the potential of an adjunct dietary therapy for the treatment of IBD.

METHODS: Forty patients with IBD were consecutively offered the IBD-AID to help treat their disease, and were retrospectively reviewed. Medical records of 11 of those patients underwent further review to determine changes in the Harvey Bradshaw Index (HBI) or Modified Truelove and Witts Severity Index (MTLWSI), before and after the diet.

RESULTS: Of the 40 patients with IBD, 13 patients chose not to attempt the diet (33%). Twenty-four patients had either a good or very good response after reaching compliance (60%), and 3 patients' results were mixed (7%). Of those 11 adult patients who underwent further medical record review, 8 with CD, and 3 with UC, the age range was 19-70 years, and they followed the diet for 4 or more weeks. After following the IBD-AID, all (100%) patients were able to discontinue at least one of their prior IBD medications, and all patients had symptom reduction including bowel frequency. The mean baseline HBI was 11 (range 1-20), and the mean follow-up score was 1.5 (range 0-3). The mean baseline MTLWSI was 7 (range 6-8), and the mean follow-up score was 0. The average decrease in the HBI was 9.5 and the average decrease in the MTLWSI was 7.

CONCLUSION: This case series indicates potential for the IBD-AID as an adjunct dietary therapy for the treatment of IBD. A randomized clinical trial is warranted.

Efficiency of bridging-sheet recruitment explains HIV-1 R5 envelope glycoprotein sensitivity to soluble CD4 and macrophage tropism

Thu, 12/04/2014 - 4:07pm

HIV-1 R5 viruses vary extensively in their capacity to infect macrophages. R5 viruses that confer efficient infection of macrophages are able to exploit low levels of CD4 for infection and predominate in brain tissue, where macrophages are a major target for infection. HIV-1 R5 founder viruses that are transmitted were reported to be non-macrophage-tropic. Here, we investigated the sensitivities of macrophage-tropic and non-macrophage-tropic R5 envelopes to neutralizing antibodies. We observed striking differences in the sensitivities of Env(+) pseudovirions to soluble CD4 (sCD4) and to neutralizing monoclonal antibodies (MAbs) that target the CD4 binding site. Macrophage-tropic R5 Envs were sensitive to sCD4, while non-macrophage-tropic Envs were significantly more resistant. In contrast, all Envs were sensitive to VRC01 regardless of tropism, while MAb b12 conferred an intermediate neutralization pattern where all the macrophage-tropic and about half of the non-macrophage-tropic Envs were sensitive. CD4, b12, and VRC01 share binding specificities on the outer domain of gp120. However, these antibodies differ in their ability to induce conformational changes on the trimeric envelope and in specificity for residues on the V1V2 loop stem and beta20-21 junction that are targets for CD4 in recruiting the bridging sheet. These distinct specificities of CD4, b12, and VRC01 likely explain the observed differences in Env sensitivity to inhibition by these reagents and provide an insight into the envelope mechanisms that control macrophage tropism. We present a model where the efficiency of bridging-sheet recruitment by CD4 is a major determinant of HIV-1 R5 envelope sensitivity to soluble CD4 and macrophage tropism.

The hunt for protective correlates of immunity to Plasmodium falciparum malaria

Thu, 12/04/2014 - 4:07pm

Determining an immunologic correlate of protection against Plasmodium falciparum malaria has been the holy grail of natural infection studies, and sought after as an endpoint for malaria vaccine trials. An in vitro assay that provides an accurate and precise assessment of protective immunity to malaria would make smaller, short-duration studies feasible, rather than the currently powered study designs that use morbidity or mortality as outcomes. Such a biomarker would be especially desirable in situations where malaria control measures that result in decreases in clinical endpoints and putatively waning protective immunity have been implemented. In an article published in BMC Medicine, Osier and colleagues addressed this problem, and demonstrated that antibodies promoting opsonic phagocytosis of merozoites provide a functional link between antigen-specific responses and protection. Understanding the mechanisms conferring protection against malaria not only improves our knowledge of basic human immunology, but promises to help in the design of an effective malaria vaccine.

Pregnancy-associated breast cancers are driven by differences in adipose stromal cells present during lactation

Thu, 12/04/2014 - 4:07pm

INTRODUCTION: The prognosis of breast cancer is strongly influenced by the developmental stage of the breast when the tumor is diagnosed. Pregnancy-associated breast cancers (PABCs), cancers diagnosed during pregnancy, lactation, or in the first postpartum year, are typically found at an advanced stage, are more aggressive and have a poorer prognosis. Although the systemic and microenvironmental changes that occur during post-partum involution have been best recognized for their role in the pathogenesis of PABCs, epidemiological data indicate that PABCs diagnosed during lactation have an overall poorer prognosis than those diagnosed during involution. Thus, the physiologic and/or biological events during lactation may have a significant and unrecognized role in the pathobiology of PABCs.

METHODS: Syngeneic in vivo mouse models of PABC were used to examine the effects of system and stromal factors during pregnancy, lactation and involution on mammary tumorigenesis. Mammary adipose stromal cell (ASC) populations were isolated from mammary glands and examined by using a combination of in vitro and in vivo functional assays, gene expression analysis, and molecular and cellular assays. Specific findings were further investigated by immunohistochemistry in mammary glands of mice as well as in functional studies using ASCs from lactating mammary glands. Additional findings were further investigated using human clinical samples, human stromal cells and using in vivo xenograft assays.

RESULTS: ASCs present during lactation (ASC-Ls), but not during other mammary developmental stages, promote the growth of carcinoma cells and angiogenesis. ASCs-Ls are distinguished by their elevated expression of cellular retinoic acid binding protein-1 (crabp1), which regulates their ability to retain lipid. Human breast carcinoma-associated fibroblasts (CAFs) exhibit traits of ASC-Ls and express crabp1. Inhibition of crabp1in CAFs or in ASC-Ls abolished their tumor-promoting activity and also restored their ability to accumulate lipid.

CONCLUSIONS: These findings imply that (1) PABC is a complex disease, which likely has different etiologies when diagnosed during different stages of pregnancy; (2) both systemic and local factors are important for the pathobiology of PABCs; and (3) the stromal changes during lactation play a distinct and important role in the etiology and pathogenesis of PABCs that differ from those during post-lactational involution.

Phylogeographic evidence of cognate recognition site patterns and transformation efficiency differences in H. pylori: theory of strain dominance

Thu, 12/04/2014 - 4:07pm

BACKGROUND: Helicobacter pylori has diverged in parallel to its human host, leading to distinct phylogeographic populations. Recent evidence suggests that in the current human mixing in Latin America, European H. pylori (hpEurope) are increasingly dominant at the expense of Amerindian haplotypes (hspAmerind). This phenomenon might occur via DNA recombination, modulated by restriction-modification systems (RMS), in which differences in cognate recognition sites (CRS) and in active methylases will determine direction and frequency of gene flow. We hypothesized that genomes from hspAmerind strains that evolved from a small founder population have lost CRS for RMS and active methylases, promoting hpEurope's DNA invasion. We determined the observed and expected frequencies of CRS for RMS in DNA from 7 H. pylori whole genomes and 110 multilocus sequences. We also measured the number of active methylases by resistance to in vitro digestion by 16 restriction enzymes of genomic DNA from 9 hpEurope and 9 hspAmerind strains, and determined the direction of DNA uptake in co-culture experiments of hspAmerind and hpEurope strains.

RESULTS: Most of the CRS were underrepresented with consistency between whole genomes and multilocus sequences. Although neither the frequency of CRS nor the number of active methylases differ among the bacterial populations (average 8.6 +/- 2.6), hspAmerind strains had a restriction profile distinct from that in hpEurope strains, with 15 recognition sites accounting for the differences. Amerindians strains also exhibited higher transformation rates than European strains, and were more susceptible to be subverted by larger DNA hpEurope-fragments than vice versa.

CONCLUSIONS: The geographical variation in the pattern of CRS provides evidence for ancestral differences in RMS representation and function, and the transformation findings support the hypothesis of Europeanization of the Amerindian strains in Latin America via DNA recombination.

Clinical disease activity and acute phase reactant levels are discordant among patients with active rheumatoid arthritis: acute phase reactant levels contribute separately to predicting outcome at one year

Thu, 12/04/2014 - 4:07pm

INTRODUCTION: Clinical trials of new treatments for rheumatoid arthritis (RA) typically require subjects to have an elevated acute phase reactant (APR), in addition to tender and swollen joints. However, despite the elevation of individual components of the Clinical Disease Activity Index (CDAI) (tender and swollen joint counts and patient and physician global assessment), some patients with active RA may have normal erythrocyte sedimentation rate (ESR) and/or C-reactive protein (CRP) levels and thus fail to meet entry criteria for clinical trials. We assessed the relationship between CDAI and APRs in the Consortium of Rheumatology Researchers of North America (CORRONA) registry by comparing baseline characteristics and one-year clinical outcomes of patients with active RA, grouped by baseline APR levels.

METHODS: This was an observational study of 9,135 RA patients who had both ESR and CRP drawn and a visit at which CDAI was > 2.8 (not in remission).

RESULTS: Of 9,135 patients with active RA, 58% had neither elevated ESR nor CRP; only 16% had both elevated ESR and CRP and 26% had either ESR or CRP elevated. Among the 4,228 patients who had a one-year follow-up visit, both baseline and one-year follow-up modified Health Assessment Questionnaire (mHAQ) and CDAI scores were lowest for patients with active RA but with neither APR elevated; both mHAQ and CDAI scores increased sequentially with the increase in number of elevated APR levels at baseline. Each individual component of the CDAI followed the same trend, both at baseline and at one-year follow-up. The magnitude of improvement in both CDAI and mHAQ scores at one year was associated positively with the number of APRs elevated at baseline.

CONCLUSIONS: In a large United States registry of RA patients, APR levels often do not correlate with disease activity as measured by joint counts and global assessments. These data strongly suggest that it is appropriate to obtain both ESR and CRP from RA patients at the initial visit. Requiring an elevation in APR levels as a criterion for inclusion of RA patients in studies of experimental agents may exclude some patients with active disease.

Emergency department patient safety incident characterization: an observational analysis of the findings of a standardized peer review process

Thu, 12/04/2014 - 4:07pm

BACKGROUND: Emergency Department (ED) care has been reported to be prone to patient safety incidents (PSIs). Improving our understanding of PSIs is essential to prevent them. A standardized, peer review process was implemented to identify and analyze ED PSIs. The primary objective of this investigation was to characterize ED PSIs identified by the peer review process. A secondary objective was to characterize PSIs that led to patient harm. In addition, we sought to provide a detailed description of the peer review process for others to consider as they conduct their own quality improvement initiatives.

METHODS: An observational study was conducted in a large, urban, tertiary-care ED. Over a two-year period, all ED incident reports were investigated via a standardized, peer review process. PSIs were identified and analyzed for contributing factors including systems failures and practitioner-based errors. The classification system for factors contributing to PSIs was developed based on systems previously reported in the emergency medicine literature as well as the investigators' experience in quality improvement and peer review. All cases in which a PSI was discovered were further adjudicated to determine if patient harm resulted.

RESULTS: In 24 months, 469 cases were investigated, identifying 152 PSIs. In total, 188 systems failures and 96 practitioner-based errors were found to have contributed to the PSIs. In twelve cases, patient harm was determined to have resulted from PSIs. Systems failures were identified in eleven of the twelve cases in which a PSI resulted in patient harm.

CONCLUSION: Systems failures were almost twice as likely as practitioner-based errors to contribute to PSIs, and systems failures were present in the majority of cases resulting in patient harm. To effectively reduce PSIs, ED quality improvement initiatives should focus on systems failure reduction.