Role of Glutathione in Buffering Excess Intracellular Copper in Streptococcus pyogenes.

Copper (Cu) is an essential metal for bacterial physiology but in excess it is bacteriotoxic. To limit Cu levels in the cytoplasm, most bacteria possess a transcriptionally responsive system for Cu export. In the Gram-positive human pathogen Streptococcus pyogenes (group A Streptococcus [GAS]), this system is encoded by the copYAZ operon. This study demonstrates that although the site of GAS infection represents a Cu-rich environment, inactivation of the copA Cu efflux gene does not reduce virulence in a mouse model of invasive disease. In vitro, Cu treatment leads to multiple observable phenotypes, including defects in growth and viability, decreased fermentation, inhibition of glyceraldehyde-3-phosphate dehydrogenase (GapA) activity, and misregulation of metal homeostasis, likely as a consequence of mismetalation of noncognate metal-binding sites by Cu. Surprisingly, the onset of these effects is delayed by ∼4 h even though expression of copZ is upregulated immediately upon exposure to Cu. Further biochemical investigations show that the onset of all phenotypes coincides with depletion of intracellular glutathione (GSH). Supplementation with extracellular GSH replenishes the intracellular pool of this thiol and suppresses all the observable effects of Cu treatment. These results indicate that GSH buffers excess intracellular Cu when the transcriptionally responsive Cu export system is overwhelmed. Thus, while the copYAZ operon is responsible for Cu homeostasis, GSH has a role in Cu tolerance and allows bacteria to maintain metabolism even in the presence of an excess of this metal ion.IMPORTANCE The control of intracellular metal availability is fundamental to bacterial physiology. In the case of copper (Cu), it has been established that rising intracellular Cu levels eventually fill the metal-sensing site of the endogenous Cu-sensing transcriptional regulator, which in turn induces transcription of a copper export pump. This response caps intracellular Cu availability below a well-defined threshold and prevents Cu toxicity. Glutathione, abundant in many bacteria, is known to bind Cu and has long been assumed to contribute to bacterial Cu handling. However, there is some ambiguity since neither its biosynthesis nor uptake is Cu-regulated. Furthermore, there is little experimental support for this physiological role of glutathione beyond measuring growth of glutathione-deficient mutants in the presence of Cu. Our work with group A Streptococcus provides new evidence that glutathione increases the threshold of intracellular Cu availability that can be tolerated by bacteria and thus advances fundamental understanding of bacterial Cu handling.

Frontal fibrosing alopecia demographics: a survey of 29 patients.

Frontal fibrosing alopecia (FFA) is a form of scarring alopecia whose diagnosis is increasing globally. Although its etiology is unknown, FFA is thought to be a clinical subset of lichen planopilaris (LPP) that primarily affects postmenopausal women. Patients diagnosed with FFA between January 2006 and December 2013 at clinics of the Washington University Division of Dermatology (St. Louis, Missouri) were studied using patient surveys and chart notes to assess demographics, clinical features, medical history, and treatment. Twenty-nine patients were enrolled in the study, including 28 women and 1 man. The average age of disease onset was 55.4 years (range, 29-75 years). Many patients (55%) had a history of autoimmune diseases, including hypothyroidism (35%), mucocutaneous lichen planus (28%), psoriasis (7%), vitiligo (3%), systemic lupus erythematosus (3%), iritis (3%), Sjögren syndrome (3%), and ulcerative colitis (3%). Patients often identified a stressful inciting event prior to onset of hair loss. Patients tried an average of 3 different treatments for hair loss, with topical and intralesional steroids, hydroxychloroquine, topical calcineurin inhibitors, and excimer laser therapy being the most efficacious at limiting hair loss.

Melanoma in individuals with neurofibromatosis type 1: a retrospective study.

BACKGROUND: Neurofibromatosis type 1 (NF1) is a cancer syndrome associated with many different cancer types. There are limited studies examining melanoma risk in this population. OBJECTIVE: To identify melanoma cases in NF1 patients and compare melanoma incidence rates relative to a general population sample. METHODS: A retrospective single institution case review of 857 NF1 patients (seen from 7/1997 to 7/2017) was conducted. We calculated age- and calendar period-adjusted standardized incidence ratios (SIRs) for white patients >20 years old overall (N=282) and for females (N=156) at their last visit date. We obtained general population melanoma reference rates from the Surveillance, Epidemiology, and End Results (SEER) 9 database. RESULTS: Among 857 patients, 52.2% were female, 54% were <20 (mean±sd=10.9±4.6) years old, and 46% were >20 (40.4±14.9) years old at their last visit date. One white female patient had a malignant melanoma diagnosed at 47 years old. The adjusted SIR was 0.97 (95% CI 0.05-4.78) overall (N=282) and 1.62 (95% CI 0.08-7.98) for females (N=156). CONCLUSIONS: We did not find statistical evidence for an increased melanoma risk in adults with NF1. However, additional large studies are warranted to clarify whether melanoma risk is increased in NF1 patients.

Developmental transcription factor NFIB is a putative target of oncofetal miRNAs and is associated with tumour aggressiveness in lung adenocarcinoma.

Genes involved in fetal lung development are thought to play crucial roles in the malignant transformation of adult lung cells. Consequently, the study of lung tumour biology in the context of lung development has the potential to reveal key developmentally relevant genes that play critical roles in lung cancer initiation/progression. Here, we describe for the first time a comprehensive characterization of miRNA expression in human fetal lung tissue, with subsequent identification of 37 miRNAs in non-small cell lung cancer (NSCLC) that recapitulate their fetal expression patterns. Nuclear factor I/B (NFIB), a transcription factor essential for lung development, was identified as a potential frequent target for these 'oncofetal' miRNAs. Concordantly, analysis of NFIB expression in multiple NSCLC independent cohorts revealed its recurrent underexpression (in ∼40-70% of tumours). Interrogation of NFIB copy number, methylation, and mutation status revealed that DNA level disruption of this gene is rare, and further supports the notion that oncofetal miRNAs are likely the primary mechanism responsible for NFIB underexpression in NSCLC. Reflecting its functional role in regulating lung differentiation, low expression of NFIB was significantly associated with biologically more aggressive subtypes and, ultimately, poorer survival in lung adenocarcinoma patients. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Defence against methylglyoxal in Group A Streptococcus: a role for Glyoxylase I in bacterial virulence and survival in neutrophils?

Methylglyoxal is a dicarbonyl compound that acts as a toxic electrophile in biological systems. Methylglyoxal is produced in certain bacteria as a byproduct of glycolysis through methylglyoxal synthase. Like many bacteria, Group A Streptococcus (GAS), a Gram-positive human pathogen responsible for a wide spectrum of diseases, uses a two-step glyoxalase system to remove methylglyoxal. However, bioinformatic analysis revealed that no homologue of methylglyoxal synthase is present in GAS, suggesting that the role of the glyoxalase system is to detoxify methylglyoxal produced by the host. In this study, we investigated the role of methylglyoxal detoxification in the pathogenesis of GAS. A mutant (5448ΔgloA), deficient in glyoxylase I (S-lactoylglutathione lyase), was constructed and tested for susceptibility to methylglyoxal, human neutrophil survival and virulence in a murine model of infection. 5448ΔgloA was more sensitive to methylglyoxal and was also more susceptible to human neutrophil killing. Inhibition of neutrophil myeloperoxidase rescued the gloA-deficient mutant indicating that this enzyme was required for methylglyoxal production. Furthermore, the 5448ΔgloA mutant was slower at disseminating into the blood in the murine model. These data suggest that neutrophils produce methylglyoxal as an antimicrobial agent during bacterial infection, and the glyoxalase system is part of the GAS defence against the innate immune system during pathogenesis.

Ponatinib-induced neutrophilic panniculitis.

Ponatinib is a bcr-abl tyrosine-kinase inhibitor (TKI) used to treat resistant and refractory chronic myeloid leukemia and Philadelphia chromosome-positive acute lymphoblastic leukemia that express bcr-abl. Neutrophilic panniculitis has been described in rare cases of patients on other TKIs in the same class as ponatinib. We present the first case of neutrophilic panniculitis following treatment with ponatinib and summarize the other cases of panniculitis caused by TKIs.

DNA methylation is globally disrupted and associated with expression changes in chronic obstructive pulmonary disease small airways.

DNA methylation is an epigenetic modification that is highly disrupted in response to cigarette smoke and involved in a wide spectrum of malignant and nonmalignant diseases, but surprisingly not previously assessed in small airways of patients with chronic obstructive pulmonary disease (COPD). Small airways are the primary sites of airflow obstruction in COPD. We sought to determine whether DNA methylation patterns are disrupted in small airway epithelia of patients with COPD, and evaluate whether changes in gene expression are associated with these disruptions. Genome-wide methylation and gene expression analysis were performed on small airway epithelial DNA and RNA obtained from the same patient during bronchoscopy, using Illumina's Infinium HM27 and Affymetrix's Genechip Human Gene 1.0 ST arrays. To control for known effects of cigarette smoking on DNA methylation, methylation and gene expression profiles were compared between former smokers with and without COPD matched for age, pack-years, and years of smoking cessation. Our results indicate that aberrant DNA methylation is (1) a genome-wide phenomenon in small airways of patients with COPD, and (2) associated with altered expression of genes and pathways important to COPD, such as the NF-E2-related factor 2 oxidative response pathway. DNA methylation is likely an important mechanism contributing to modulation of genes important to COPD pathology. Because these methylation events may underlie disease-specific gene expression changes, their characterization is a critical first step toward the development of epigenetic markers and an opportunity for developing novel epigenetic therapeutic interventions for COPD.

Survey of current enteral nutrition practices in treatment of amyotrophic lateral sclerosis.

BACKGROUND AND AIMS: Enteral nutrition (EN) is commonly prescribed for dysphagia and weight loss in amyotrophic lateral sclerosis (ALS), but there are currently no ALS-specific EN guidelines. We aimed to survey current practices prescribing EN to ALS patients. METHODS: An online survey was distributed using list servers administered by the Academy of Nutrition and Dietetics (AND), Muscular Dystrophy Association (MDA), and ALS Association (ALSA). RESULTS: A total of 148 dietitians, nurses, and physicians participated in the survey, of whom 50% were dietitians and 68% were associated with an ALS clinic. Only 47% of respondents reported their patients to be fully compliant with EN recommendations. Side effects (fullness, diarrhea, constipation, and bloating) were the most important reason for patient noncompliance, followed by dependence on caregivers. By contrast, only 3% of providers rated depression/hopelessness as the most important reason for noncompliance. Half of those surveyed reported that more than 25% of patients continued to lose weight after starting EN. CONCLUSIONS: Our survey results show a high frequency of gastrointestinal side effects and weight loss in ALS patients receiving EN. These findings may be limited by sampling error and non-response bias. Prospective studies are needed to help establish EN guidelines for ALS.

Uric acid levels predict survival in men with amyotrophic lateral sclerosis.

Elevated uric acid levels have recently been found to be associated with slower disease progression in Parkinson's disease, Huntington's disease, multiple system atrophy, and mild cognitive impairment. The aim of this study is to determine whether serum uric acid levels predict survival in amyotrophic lateral sclerosis (ALS). A total of 251 people with ALS enrolled in two multicenter clinical trials were included in our analysis. The main outcome measure was survival time, which was calculated as time to death, tracheostomy, or permanent assistive ventilation, with any event considered a survival endpoint. Cox proportional hazards models were used to estimate the hazard ratio (HR) of reaching a survival endpoint according to baseline uric acid levels after adjusting for markers of disease severity (FVC, total ALSFRS-R score, time since symptom onset, diagnostic delay, BMI, bulbar vs. spinal onset, age, and riluzole use). There was a dose-dependent survival advantage in men, but not women, with higher baseline uric acid levels (logrank test: p = 0.018 for men, p = 0.81 for women). There was a 39% reduction in risk of death during the study for men with each 1 mg/dl increase in uric acid levels (adjusted HR: 0.61, 95% CI 0.39-0.96, p = 0.03). This is the first study to demonstrate that serum uric acid is associated with prolonged survival in ALS, after adjusting for markers of disease severity. Similar to previous reports in Parkinson's disease, this association was seen in male subjects only.

Immune parameters of QX-resistant and wild caught Saccostrea glomerata hemocytes in relation to Marteilia sydneyi infection.

Sydney rock oysters (SRO) Saccostrea glomerata suffer mass mortalities during summer and autumn as a result of infection by a protozoan parasite Marteilia sydneyi (QX disease). Mass selected disease resistant (QXR) lines have been used with some success in affected estuaries in recent years, with resistance attributed to oxidative defense systems. However, the role of hemocytes in resistance to QX by SRO has not been fully explored. In the present study, fifty QXR and fifty wild caught (WC) oysters were collected from a lease at Pimpama River during a QX outbreak in January 2011. Hemocytes characteristics (type, morphology) and functions (mortality, phagocytosis and oxidative activity) from both oyster lines were analyzed by flow cytometry in the context of infection intensity and parasite viability (determined histologically). Amongst the QXR oysters, 20% were diseased containing viable parasite, 74% had killed M. sydneyi and 6% were uninfected. In contrast, 86% of WC oysters were diseased, 2% had killed M. sydneyi and 12% were healthy. Significant differences in hemocyte number and physiology between the two oyster lines were found (ANOVA). Phagocytosis rate and the mean oxidative activity per cell were similar between both oyster lines. Higher numbers of infiltrating and circulating hemocytes, higher percentage of circulating granulocytes, their higher size and complexity in QXR oysters, and the production of reactive oxygen species were associated with the ability to kill the parasite. High abundance of M. sydneyi in the digestive tubule epithelium of both oyster lines implied inability to kill the parasite at the beginning of the infection. However, QXR oysters had the ability to kill M. sydneyi at the stage of sporangiosorae in the epithelium of digestive tubules. The similar phagocytic ability of hemocytes from both oyster lines, the size of the parasite at this infection stage, and its localization suggested that encapsulation is likely to be the main process involved in the eradication of M. sydneyi by QXR oysters.

Genetic changes in the evolution of multidrug resistance for cultured human ovarian cancer cells.

The multidrug resistant (MDR) phenotype is often attributed to the activity of ATP-binding cassette (ABC) transporters such as P-glycoprotein (ABCB1). Previous work has suggested that modulation of MDR may not necessarily be a single gene trait. To identify factors that contribute to the emergence of MDR, we undertook integrative genomics analysis of the ovarian carcinoma cell line SKOV3 and a series of MDR derivatives of this line (SKVCRs). As resistance increased, comparative analysis of gene expression showed conspicuous activation of a network of genes in addition to ABCB1. Functional annotation and pathway analysis revealed that many of these genes were associated with the extracellular matrix and had previously been implicated in tumor invasion and cell proliferation. Further investigation by whole genome tiling-path array CGH suggested that changes in gene dosage were key to the activation of several of these overexpressed genes. Remarkably, alignment of whole genome profiles for SKVCR lines revealed the emergence and decline of specific segmental DNA alterations. The most prominent alteration was a novel amplicon residing at 16p13 that encompassed the ABC transporter genes ABCC1 and ABCC6. Loss of this amplicon in highly resistant SKVCR lines coincided with the emergence of a different amplicon at 7q21.12, which harbors ABCB1. Integrative analysis suggests that multiple genes are activated during escalation of drug resistance, including a succession of ABC transporter genes and genes that may act synergistically with ABCB1. These results suggest that evolution of the MDR phenotype is a dynamic, multi-genic process in the genomes of cancer cells.