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Recent documents in eScholarship@UMMS
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Autoimmune and other acquired neutropenias

Fri, 08/11/2017 - 11:43am

This educational review addresses the diagnostic evaluation of patients for autoimmune and other forms of acquired neutropenia, including the futility of deconstructing the overlap of chronic "autoimmune," "benign," and "idiopathic" categories. Isolated neutropenias caused by infection, drugs, and immunologic disorders are also addressed. Discussion of management options emphasizes a conservative approach, with largely supportive care for these mostly benign and self-limited disorders.

Correlation of endoscopic disease severity with pediatric ulcerative colitis activity index score in children and young adults with ulcerative colitis

Fri, 08/11/2017 - 11:43am

AIM: To investigate of pediatric ulcerative colitis activity index (PUCAI) in ulcerative colitis correlate with mucosal inflammation and endoscopic assessment of disease activity (Mayo endoscopic score).

METHODS: We reviewed charts from ulcerative colitis patients who had undergone both colonoscopy over 3 years. Clinical assessment of disease severity within 35 d (either before or after) the colonoscopy were included. Patients were excluded if they had significant therapeutic interventions (such as the start of corticosteroids or immunosuppressive agents) between the colonoscopy and the clinical assessment. Mayo endoscopic score of the rectum and sigmoid were done by two gastroenterologists. Inter-observer variability in Mayo score was assessed.

RESULTS: We identified 99 patients (53% female, 74% pancolitis) that met inclusion criteria. The indications for colonoscopy included ongoing disease activity (62%), consideration of medication change (10%), assessment of medication efficacy (14%), and cancer screening (14%). Based on PUCAI scores, 33% of patients were in remission, 39% had mild disease, 23% had moderate disease, and 4% had severe disease. There was "moderate-substantial" agreement between the two reviewers in assessing rectal Mayo scores (kappa = 0.54, 95%CI: 0.41-0.68).

CONCLUSION: Endoscopic disease severity (Mayo score) assessed by reviewing photographs of pediatric colonoscopy has moderate inter-rater reliability, and agreement was less robust in assessing patients with mild disease activity. Endoscopic disease severity generally correlates with clinical disease severity as measured by PUCAI score. However, children with inflamed colons can have significant variation in their reported clinical symptoms. Thus, assessment of both clinical symptoms and endoscopic disease severity may be required in future clinical studies.

One More Controversy: Adeno-Associated Virus in Stem Cells

Fri, 08/11/2017 - 11:43am

Among the earliest treatment paradigms envisioned for gene therapy was the ex vivo genetic modification of autologous stem cells, particularly in the context of bone-marrow transplantation. Many early applications of gammaretrovirus and lentivirus vectors have used this platform, with the primary focus being on hematopoietic stem cells (HSCs). In the April 2017 special issue of Human Gene Therapy, a number of crucial issues in the field of adeno-associated virus (AAV) gene therapy were discussed. However, the issue of the utility of recombinant AAV (rAAV) gene therapy for stem-cell transduction was not directly addressed in that issue.

The problem is now grappled with directly in this June issue of Human Gene Therapy.

The BRG1 ATPase of human SWI/SNF chromatin remodeling enzymes as a driver of cancer

Fri, 08/11/2017 - 11:43am

Mammalian SWI/SNF enzymes are ATP-dependent remodelers of chromatin structure. These multisubunit enzymes are heterogeneous in composition; there are two catalytic ATPase subunits, BRM and BRG1, that are mutually exclusive, and additional subunits are incorporated in a combinatorial manner. Recent findings indicate that approximately 20% of human cancers contain mutations in SWI/SNF enzyme subunits, leading to the conclusion that the enzyme subunits are critical tumor suppressors. However, overexpression of specific subunits without apparent mutation is emerging as an alternative mechanism by which cellular transformation may occur. Here we highlight recent evidence linking elevated expression of the BRG1 ATPase to tissue-specific cancers and work suggesting that inhibiting BRG1 may be an effective therapeutic strategy.

Peripubertal blood lead levels and growth among Russian boys

Fri, 08/11/2017 - 11:42am

BACKGROUND: Childhood blood lead levels (BLL) have been associated with growth impairment.

OBJECTIVES: We assessed associations of peripubertal BLL with adolescent growth and near adult height in a longitudinal cohort of Russian boys.

METHODS: 481 boys were enrolled at ages 8-9years and followed annually to age 18. At enrollment, BLL was measured, and height, weight, and pubertal staging were obtained annually during 10years of follow-up. Mixed effects models were used to assess the associations of BLL with longitudinal age-adjusted World Health OrganizationZ-scores for height (HT-Z) and body mass index (BMI-Z), and annual height velocity (HV). Interactions between boys' age and BLL on growth outcomes were evaluated.

RESULTS: The median (range) BLL was 3.0 (0.5-31.0) mug/dL. At age 18years, 79% of boys had achieved near adult height (HV < 1.0cm/year), and means (SD) for HT-Z and BMI-Z were 0.15 (0.92) and -0.32 (1.24). Over 10years of follow-up, after covariate adjustment, boys with higher ( > /=5mug/dL) BLL compared with lower BLL were shorter (adjusted mean difference in HT-Z=-0.43, 95% CI -0.60, -0.25, p-value < 0.001), translating to a 2.5cm lower height at age 18years. The decrement in height for boys with higher BLL was most pronounced at 12 to 15years of age (interaction p=0.03). Boys with higher BLL were leaner (adjusted mean difference in BMI-Z=-0.22, 95% CI: -0.45, 0.01, p=0.06).

CONCLUSIONS: Higher peripubertal BLLs were associated with shorter height through age 18years, suggesting a persistent effect of lead on linear growth.

A C9ORF72 BAC mouse model recapitulates key epigenetic perturbations of ALS/FTD

Fri, 08/11/2017 - 11:42am

BACKGROUND: Amyotrophic Lateral Sclerosis (ALS) is a fatal and progressive neurodegenerative disorder with identified genetic causes representing a significant minority of all cases. A GGGGCC hexanucleotide repeat expansion (HRE) mutation within the C9ORF72 gene has recently been identified as the most frequent known cause of ALS. The expansion leads to partial heterochromatinization of the locus, yet mutant RNAs and dipeptide repeat proteins (DPRs) are still produced in sufficient quantities to confer neurotoxicity. The levels of these toxic HRE products positively correlate with cellular toxicity and phenotypic severity across multiple disease models. Moreover, the degree of epigenetic repression inversely correlates with some facets of clinical presentation in C9-ALS patients. Recently, bacterial artificial chromosomes (BAC) have been used to generate transgenic mice that harbor the HRE mutation, complementing other relevant model systems such as patient-derived induced pluripotent stem cells (iPSCs). While epigenetic features of the HRE have been investigated in various model systems and post-mortem tissues, epigenetic dysregulation at the expanded locus in C9-BAC mice remains unexplored.

METHODS AND RESULTS: Here, we sought to determine whether clinically relevant epigenetic perturbations caused by the HRE are mirrored in a C9-BAC mouse model. We used complementary DNA methylation assessment and immunoprecipitation methods to demonstrate that epigenetic aberrations caused by the HRE, such as DNA and histone methylation, are recapitulated in the C9-BAC mice. Strikingly, we found that cytosine hypermethylation within the promoter region of the human transgene occurred in a subset of C9-BAC mice similar to what is observed in patient populations. Moreover, we show that partial heterochromatinization of the C9 HRE occurs during the first weeks of the mouse lifespan, indicating age-dependent epigenetic repression. Using iPSC neurons, we found that preventing R-loop formation did not impede heterochromatinization of the HRE.

CONCLUSIONS: Taken together, these observations provide further insight into mechanism and developmental time-course of epigenetic perturbations conferred by the C9ORF72 HRE. Finally, we suggest that epigenetic repression of the C9ORF72 HRE and nearby gene promoter could impede or delay motor neuron degeneration in C9-BAC mouse models of ALS/FTD.

Safe and Efficient Silencing with a Pol II, but Not a Pol lII, Promoter Expressing an Artificial miRNA Targeting Human Huntingtin

Fri, 08/11/2017 - 11:42am

Huntington's disease is a devastating, incurable neurodegenerative disease affecting up to 12 per 100,000 patients worldwide. The disease is caused by a mutation in the Huntingtin (Htt) gene. There is interest in reducing mutant Huntingtin by targeting it at the mRNA level, but the maximum tolerable dose and long-term effects of such a treatment are unknown. Using a self-complementary AAV9 vector, we delivered a mir-155-based artificial miRNA under the control of the chicken beta-actin or human U6 promoter. In mouse brain, the artificial miRNA reduced the human huntingtin mRNA by 50%. The U6, but not the CbetaA promoter, produced the artificial miRNA at supraphysiologic levels. Embedding the antisense strand in a U6-mir-30 scaffold reduced expression of the antisense strand but increased the sense strand. In mice treated with scAAV9-U6-mir-155-HTT or scAAV9-CbetaA-mir-155-HTT, activated microglia were present around the injection site 1 month post-injection. Six months post-injection, mice treated with scAAV9-CbetaA-mir-155-HTT were indistinguishable from controls. Those that received scAAV9-U6-mir-155-HTT showed behavioral abnormalities and striatal damage. In conclusion, miRNA backbone and promoter can be used together to modulate expression levels and strand selection of artificial miRNAs, and in brain, the CbetaA promoter can provide an effective and safe dose of a human huntingtin miRNA.

Patient safety during procedural sedation using capnography monitoring: a systematic review and meta-analysis

Fri, 08/11/2017 - 11:42am

OBJECTIVE: To evaluate the effect of capnography monitoring on sedation-related adverse events during procedural sedation and analgesia (PSA) administered for ambulatory surgery relative to visual assessment and pulse oximetry alone.

DESIGN AND SETTING: Systematic literature review and random effects meta-analysis of randomised controlled trials (RCTs) reporting sedation-related adverse event incidence when adding capnography to visual assessment and pulse oximetry in patients undergoing PSA during ambulatory surgery in the hospital setting. Searches for eligible studies published between 1 January 1995 and 31 December 2016 (inclusive) were conducted in PubMed, the Cochrane Library and EMBASE without any language constraints. Searches were conducted in January 2017, screening and data extraction were conducted by two independent reviewers, and study quality was assessed using a modified Jadad scale.

INTERVENTIONS: Capnography monitoring relative to visual assessment and pulse oximetry alone.

PRIMARY AND SECONDARY OUTCOME MEASURES: Predefined endpoints of interest were desaturation/hypoxaemia (the primary endpoint), apnoea, aspiration, bradycardia, hypotension, premature procedure termination, respiratory failure, use of assisted/bag-mask ventilation and death during PSA.

RESULTS: The literature search identified 1006 unique articles, of which 13 were ultimately included in the meta-analysis. Addition of capnography to visual assessment and pulse oximetry was associated with a significant reduction in mild (risk ratio (RR) 0.77, 95% CI 0.67 to 0.89) and severe (RR 0.59, 95% CI 0.43 to 0.81) desaturation, as well as in the use of assisted ventilation (OR 0.47, 95% CI 0.23 to 0.95). No significant differences in other endpoints were identified.

CONCLUSIONS: Meta-analysis of 13 RCTs published between 2006 and 2016 showed a reduction in respiratory compromise (from respiratory insufficiency to failure) during PSA with the inclusion of capnography monitoring. In particular, use of capnography was associated with less mild and severe oxygen desaturation, which may have helped to avoid the need for assisted ventilation.

An Unusual Manifestation of Celiac Disease in an Adolescent With Down Syndrome and Graves Disease

Fri, 08/11/2017 - 11:42am

We describe an adolescent girl with Down syndrome and Graves disease presenting with polymenorrhea as the initial symptom of Celiac disease.

A role for the mevalonate pathway in the induction of subtype cross-reactive immunity to influenza A virus by human gammadelta T lymphocytes

Thu, 08/10/2017 - 3:02pm

The major gammadelta T cell subset in the human peripheral blood expresses the Vgamma9delta2 TCR and recognizes non-peptidic prenyl pyrophosphate antigens such as isopentylpyrophosphate (IPP). Upon activation the gammadelta T cells rapidly secrete antiviral cytokines similar to classical memory alphabeta T cells. Here we have investigated the ability of gammadelta T lymphocytes from human PBMC to become activated by influenza A virus infection. Vgamma9Vdelta2 T lymphocytes rapidly upregulate expression of CD25 and CD69 and produce IFN-gamma following influenza infection of PBMC. Moreover, the recognition is cross-reactive between various subtypes of influenza, but not with vaccinia virus. Vgamma9Vdelta2 T cell responses are potently reduced by the HMG-CoA reductase inhibitor mevastatin, which inhibits the mevalonate pathway and IPP synthesis. Our results indicate that influenza virus infection induces the rapid activation and function of Vgamma9Vdelta2 T lymphocytes in the peripheral blood via a mechanism that depends on the mevalonate pathway.

Sequential immunization with heterologous chimeric flaviviruses induces broad-spectrum cross-reactive CD8+ T cell responses

Thu, 08/10/2017 - 3:02pm

Flavivirus vaccines based on ChimeriVax technology contain the nonstructural genes of the yellow fever vaccine and the premembrane and envelope genes of heterologous flaviviruses, such as Japanese encephalitis and West Nile viruses. These chimeric vaccines induce both humoral and cell-mediated immunity. Mice were vaccinated with yellow fever, chimeric Japanese encephalitis virus (YF/JE), or chimeric West Nile virus (YF/WN) vaccines, followed by a secondary homologous or heterologous vaccination; the hierarchy and function of CD8(+) T cell responses to a variable envelope epitope were then analyzed and compared with those directed against a conserved immunodominant yellow fever virus NS3 epitope. Sequential vaccination with heterologous chimeric flaviviruses generated a broadly cross-reactive CD8(+) T cell response dependent on both the sequence of infecting viruses and epitope variant. The enhanced responses to variant epitopes after heterologous vaccination were not related to preexisting antibody or to higher virus titers. These results demonstrate that the sequence of vaccination affects the expansion of cross-reactive CD8(+) T cells after heterologous chimeric flavivirus challenge.

NIAID workshop on Flavivirus immunity

Thu, 08/10/2017 - 3:02pm

On September 16, 2009, the National Institute of Allergy and Infectious Diseases (NIAID), part of the U.S. National Institutes of Health, convened a workshop to discuss current knowledge of T- and B-cell immune epitopes for members of the Flavivirus genus (family Flaviviridae), and how this information could be used to increase our basic understanding of host-pathogen interactions and/or advance the development of new or improved vaccines and diagnostics for these pathogens. B-cell and T-cell responses to flaviviruses are critical components of protective immunity against these pathogens. However, they have also been linked to disease pathogenesis. A detailed understanding of the biological significance of immune epitope information may provide clues regarding the mechanisms governing the induction of protective versus pathogenic adaptive immune responses.

Dengue--how best to classify it

Thu, 08/10/2017 - 3:02pm

Dengue has emerged as a major public health problem worldwide. Dengue virus infection causes a wide range of clinical manifestations. Since the 1970s, clinical dengue has been classified according to the World Health Organization guideline as dengue fever and dengue hemorrhagic fever. The classification has been criticized with regard to its usefulness and its applicability. In 2009, the World Health Organization issued a new guideline that classifies clinical dengue as dengue and severe dengue. The 2009 classification differs significantly from the previous classification in both conceptual and practical levels. The impacts of the new classification on clinical practice, dengue research, and public health policy are discussed.

Complement-dependent lysis of influenza a virus-infected cells by broadly cross-reactive human monoclonal antibodies

Thu, 08/10/2017 - 3:02pm

We characterized human monoclonal antibodies (MAbs) cloned from influenza virus-infected patients and from influenza vaccine recipients by complement-dependent lysis (CDL) assay. Most MAbs active in CDL were neutralizing, but not all neutralizing MAbs can mediate CDL. Two of the three stalk-specific neutralizing MAbs tested were able to mediate CDL and were more cross-reactive to temporally distant H1N1 strains than the conventional hemagglutination-inhibiting and neutralizing MAbs. One of the stalk-specific MAbs was subtype cross-reactive to H1 and H2 hemagglutinins, suggesting a role for stalk-specific antibodies in protection against influenza illness, especially by a novel viral subtype which can cause pandemics.

T cells and pathogenesis of hantavirus cardiopulmonary syndrome and hemorrhagic fever with renal syndrome

Thu, 08/10/2017 - 3:02pm

We previously hypothesized that increased capillary permeability observed in both hantavirus cardiopulmonary syndrome (HCPS) and hemorrhagic fever with renal syndrome (HFRS) may be caused by hantavirus-specific cytotoxic T cells attacking endothelial cells presenting viral antigens on their surface based on clinical observations and in vitro experiments. In HCPS, hantavirus-specific T cell responses positively correlated with disease severity. In HFRS, in one report, contrary to HCPS, T cell responses negatively correlated with disease severity, but in another report the number of regulatory T cells, which are thought to suppress T cell responses, negatively correlated with disease severity. In rat experiments, in which hantavirus causes persistent infection, depletion of regulatory T cells helped infected rats clear virus without inducing immunopathology. These seemingly contradictory findings may suggest delicate balance in T cell responses between protection and immunopathogenesis. Both too strong and too weak T cell responses may lead to severe disease. It is important to clarify the role of T cells in these diseases for better treatment (whether to suppress T cell functions) and protection (vaccine design) which may need to take into account viral factors and the influence of HLA on T cell responses.

The degree of leukocytosis and urine GATA-3 mRNA levels are risk factors for severe acute kidney injury in Puumala virus nephropathia epidemica

Thu, 08/10/2017 - 3:02pm

Puumala hantavirus (PUUV) infection, also known as nephropathia epidemica, is the most common cause of hemorrhagic fever with renal syndrome (HFRS) in Europe. The pathogenesis of PUUV nephropathia epidemica is complex and multifactorial, and the risk factors for severe acute kidney injury (AKI) during acute PUUV infection are not well defined. We conducted a prospective study of hospitalized patients with PUUV infection in Tampere, Finland to identify acute illness risk factors for HFRS severity. Serial daily blood and urine samples were collected throughout acute illness and at 2 week and 6 month convalescent visits. By univariate analyses, the maximum white blood cell count during acute illness was a risk factor for severe AKI. There were no significant associations between PUUV-induced AKI severity and platelet counts, C-reactive protein, or alanine aminotransferase levels. Maximum plasma interleukin (IL)-6, urine IL-6, and urine IL-8 concentrations were positively associated with PUUV-induced AKI. Finally, the maximum urinary sediment GATA-3 mRNA level was positively correlated with the peak fold-change in serum creatinine, regardless of AKI severity classification. By multivariate analyses, we found that the maximum levels of leukocytes and urinary sediment GATA-3 mRNA during acute illness were independent risk factors for severe PUUV-induced AKI. We have identified novel acute illness risk factors for severe PUUV-induced AKI.

A human CD4+ T cell epitope in the influenza hemagglutinin is cross-reactive to influenza A virus subtypes and to influenza B virus

Thu, 08/10/2017 - 3:02pm

The hemagglutinin protein (HA) of the influenza virus family is a major antigen for protective immunity. Thus, it is a relevant target for developing vaccines. Here, we describe a human CD4(+) T cell epitope in the influenza virus HA that lies in the fusion peptide of the HA. This epitope is well conserved in all 16 subtypes of the HA protein of influenza A virus and the HA protein of influenza B virus. By stimulating peripheral blood mononuclear cells (PBMCs) from a healthy adult donor with peptides covering the entire HA protein based on the sequence of A/Japan/305/1957 (H2N2), we generated a T cell line specific to this epitope. This CD4(+) T cell line recognizes target cells infected with influenza A virus seasonal H1N1 and H3N2 strains, a reassortant H2N1 strain, the 2009 pandemic H1N1 strain, and influenza B virus in cytotoxicity assays and intracellular-cytokine-staining assays. It also lysed target cells infected with avian H5N1 virus. We screened healthy adult PBMCs for T cell responses specific to this epitope and found individuals who had ex vivo gamma interferon (IFN-gamma) responses to the peptide epitope in enzyme-linked immunospot (ELISPOT) assays. Almost all donors who responded to the epitope had the HLA-DRB1*09 allele, a relatively common HLA allele. Although natural infection or standard vaccination may not induce strong T and B cell responses to this highly conserved epitope in the fusion peptide, it may be possible to develop a vaccination strategy to induce these CD4(+) T cells, which are cross-reactive to both influenza A and B viruses.

Comparison of complement dependent lytic, hemagglutination inhibition and microneutralization antibody responses in influenza vaccinated individuals

Thu, 08/10/2017 - 3:02pm

Virus specific, non-neutralizing antibodies such as complement dependent lytic (CDL) antibodies may reduce morbidity following infection through the clearance of infectious virus particles and infected cells. We examined hemagglutination inhibition (HAI), microneutralization (MN) and CDL antibody titers to influenza A H1 and H3 virus strains in 23 healthy young adults who received the 2005-2006 trivalent inactivated influenza vaccine. Post vaccination, we detected statistically significant increases in MN and CDL antibodies but not in HAI antibodies. Statistically significantly higher fold increases in CDL antibodies post vaccination were seen compared with MN and HAI antibodies post vaccination. However, the overall fold increases were modest, likely related to the fact that most of the subjects had received influenza vaccination previously. This study showed that influenza vaccination is not only capable of increasing the level of antibodies that neutralize virus but also antibodies that can cause lysis of infected cells. The biological significance of these CDL antibodies merits further investigation in clinical studies.