Rituximab rescue therapy for autoimmune pulmonary alveolar proteinosis.

Autoimmune pulmonary alveolar proteinosis (aPAP) is a rare lung disease characterised by abnormal alveolar surfactant accumulation due to macrophage dysfunction. Whole lung lavage (WLL) is the cornerstone of first-line aPAP therapy, but effective rescue treatments have not yet been well established. We report a case of a 41-year-old man with aPAP in whom further WLL is contraindicated. His diagnosis was established using a combination of classical radiological findings, positive serum GM-CSF IgG antibodies and bronchoalveolar lavage (BAL) findings. Following a literature review of emerging therapies, a decision was made to treat with a course of rituximab to suppress GM-CSF autoantibody production and restore alveolar surfactant-macrophage homeostasis. A significant clinical response was demonstrated within 6 months with improvements in arterial oxygenation, respiratory membrane gas diffusion, six-minute walk test and radiological findings.

An unusual case of invasive pleuritis and miliary Mycoplasma pneumonia during check-point inhibitor therapy.

A growing body of evidence suggests that check-point inhibitors not only increase the overall risk of infections, but, due to an altered immune response, may also result in atypical manifestations. We report a case of a 38-year-old man with pleuritic chest pain, dyspnoea, fevers and a dry cough receiving combination ipilimumab and nivolumab immunotherapy for metastatic melanoma. Radiological findings demonstrated a diffuse increased fluorodeoxyglucose avidity of the thoracic pleura in addition to a disseminated miliary pattern of pulmonary nodularities. A subsequent bronchoscopy was macroscopically normal with unremarkable washings. In the context of a significantly elevated Mycoplasma serology, a diagnosis of Mycoplasma pneumoniae pneumonia (MPP) was made. The patient was successfully treated with a course of azithromycin and amoxicillin-clavulanic acid. We suggest an awareness of diffuse pleuritis and a disseminated miliary nodular pattern as atypical manifestations of MPP, potentially attributable to immune modulation in the context of immunotherapy.

Feasibility and Participant Experiences of a Written Emotional Disclosure Intervention for Parental Caregivers of People with Psychosis.

Caregivers are at increased risk of poor health but often cannot engage with health care because of practical constraints. Writing therapies such as written emotional disclosure (WED) might represent an accessible therapy for this population. This study aimed to determine whether WED is a suitable therapy for caregivers of people with psychosis. Data were collected from a feasibility trial comparing WED with a neutral writing task. Twenty four participants completed writing tasks; the content was analysed using the Linguistic Inquiry Word Count (LIWC) programme. Twenty one participants provided feedback about the writing task, and data were analysed using Burnard's thematic content analysis. WED was feasible to implement in caregivers of people with psychosis. All participants ascribed benefit to writing tasks, the majority describing a cathartic effect, enjoying time to oneself or distraction from caregiving. Quantitative comparison indicated differences in emotional content between intervention and control writing. However, qualitative analysis showed that control participants found it challenging to write neutrally, the majority citing the care-recipient in their writing. Community-based WED is a feasible therapy for caregivers of people with psychosis. The need for refinement of the control writing task for use in caregivers is discussed. Copyright © 2015 John Wiley & Sons, Ltd.

Immune derived opioidergic inhibition of viscerosensory afferents is decreased in Irritable Bowel Syndrome patients.

Alterations in the neuro-immune axis contribute toward viscerosensory nerve sensitivity and symptoms in Irritable Bowel Syndrome (IBS). Inhibitory factors secreted from immune cells inhibit colo-rectal afferents in health, and loss of this inhibition may lead to hypersensitivity and symptoms. We aimed to determine the immune cell type(s) responsible for opioid secretion in humans and whether this is altered in patients with IBS. The ß-endorphin content of specific immune cell lineages in peripheral blood and colonic mucosal biopsies were compared between healthy subjects (HS) and IBS patients. Peripheral blood mononuclear cell (PBMC) supernatants from HS and IBS patients were applied to colo-rectal sensory afferent endings in mice with post-inflammatory chronic visceral hypersensitivity (CVH). ß-Endorphin was identified predominantly in monocyte/macrophages relative to T or B cells in human PBMC and colonic lamina propria. Monocyte derived ß-endorphin levels and colonic macrophage numbers were lower in IBS patients than healthy subjects. PBMC supernatants from healthy subjects had greater inhibitory effects on colo-rectal afferent mechanosensitivity than those from IBS patients. The inhibitory effects of PBMC supernatants were more prominent in CVH mice compared to healthy mice due to an increase in µ-opioid receptor expression in dorsal root ganglia neurons in CVH mice. Monocyte/macrophages are the predominant immune cell type responsible for ß-endorphin secretion in humans. IBS patients have lower monocyte derived ß-endorphin levels than healthy subjects, causing less inhibition of colonic afferent endings. Consequently, altered immune function contributes toward visceral hypersensitivity in IBS.

PI16 is expressed by a subset of human memory Treg with enhanced migration to CCL17 and CCL20.

The peptidase inhibitor PI16 was shown previously by microarray analysis to be over-expressed by CD4-positive/CD25-positive Treg compared with CD4-positive/CD25-negative Th cells. Using a monoclonal antibody to the human PI16 protein, we found that PI16-positive Treg have a memory (CD45RO-positive) phenotype and express higher levels of FOXP3 than PI16-negative Treg. PI16-positive Treg are functional in suppressor assays in vitro with potency similar to PI16-negative Treg. Further phenotyping of the PI16-positive Treg revealed that the chemokine receptors CCR4 and CCR6 are expressed by more of the PI16-positive/CD45RO-positive Treg compared with PI16-negative/CD45RO-positive Treg or Th cells. PI16-positive Treg showed enhanced in vitro migration towards the inflammatory chemokines CCL17 and CCL20, suggesting they can migrate to sites of inflammation. We conclude that PI16 identifies a novel distinct subset of functional memory Treg which can migrate to sites of inflammation and regulate the pro-inflammatory response at those sites.

Mutation of two Mycoplasma arthritidis surface lipoproteins with divergent functions in cytadherence.

Mycoplasma arthritidis is a natural pathogen of rats, causing an acute polyarthritis. Previous studies identified two membrane-bound lipoproteins, Maa1 and Maa2, thought to be associated with cytadherence of M. arthritidis strain 158p10p9. We have since confirmed that Maa1 is a major adhesin, although the role of Maa2 has proven more elusive. Both proteins were capable of eliciting protective immunity in rats against challenge with the virulent strain 158p10p9, suggesting that they may be important in pathogenesis. The purpose of this study was to better understand the roles of Maa1 and Maa2 in cytadherence in vitro. Insertion mutants were created for both genes by transposon mutagenesis. In vitro adherence of the Maa1 mutant KOMaa1 to rat L2 lung cells was reduced to the level previously reported for a spontaneous low-adherence mutant of 158p10p9 in which Maa1 is truncated and nonfunctional. Surprisingly, adherence of the Maa2 mutant KOMaa2 was approximately fivefold greater than that of the wild type. Complementation of KOMaa1 and KOMaa2 with wild-type alleles of maa1 and maa2, respectively, returned adherence to wild-type levels. This work confirms our earlier observation that Maa1 is a major adhesin for M. arthritidis strain 158p10p9. Maa2, on the other hand, may play a suppressive or modulatory role, possibly serving to release organisms from microcolonies at certain stages of infection.

Escherichia coli biotin protein ligase: characterization and development of a high-throughput assay.

The rapid rise in pathogenic bacteria resistant to current treatments, coupled with the paucity of new therapeutic agents in the pipeline, has resulted in a significant need for new antibiotics. One strategy to overcome resistance requires new chemical entities that inhibit key enzymes in essential metabolic processes that have not been previously targeted and for which there is no preexisting drug resistance. Biotin protein ligase (BPL), required to complete acetyl CoA carboxylase's capability for fatty acid biosynthesis, is one target that has not yet been fully explored. However, its application in large-scale compound screens has been limited due to the lack of a truly high-throughput assay for enzyme activity. Here we report a novel assay system for BPL from Escherichia coli (BirA). This assay employs fluorescence polarization technology together with a unique peptide substrate for BirA. Additionally, the multiple handling steps and requirement for radiolabeled ligands associated with previous assays have been eliminated. Kinetic analysis of MgATP (K(m) 0.25+/-0.01 mM) and biotin (K(m) 1.45+/-0.15 microM) binding produced results consistent with published data. Inhibition studies with end products of the BPL reaction, AMP and pyrophosphate, further validated the assay. Statistical analysis, performed upon both intraassay and interassay results (n = 30), showed the coefficient of variance to be <10% across all data sets. Furthermore, Z' factors between 0.5 and 0.8 demonstrated the utility of this technology in high-throughput applications.

Mechanism, selectivity promotion, and new ultraselective pathways in Ag-catalyzed heterogeneous epoxidation.

The selective oxidation of styrene on clean and modified Ag(100) surfaces has been studied by synchrotron fast XPS and temperature-programmed reaction spectroscopy. By following the time dependence of surface species, it is unequivocally demonstrated that the necessary and sufficient conditions for epoxide formation are oxygen adatoms and pi-adsorbed alkene molecules. Increased oxygen coverage and coadsorbed Cs have pronounced and opposite effects on epoxidation selectivity, consistent with the view that the valence charge density on O(a) is pivotal in determining this property. Submonolayer quantities of Cs nitrate generated in situ open a new, low-temperature ultraselective, epoxidation pathway thought to involve direct oxygen transfer from the oxyanion to the alkene.

Altered epithelial cell proportions in the fetal lung of glucocorticoid receptor null mice.

Glucocorticoids provide important signals for maturation of the fetal lung and antenatal glucocorticoids are used to reduce the respiratory insufficiency suffered by preterm infants. To further understand the role of glucocorticoids in fetal lung maturation, we have analyzed mice with a targeted null mutation for the glucocorticoid receptor (GR) gene, which severely retards lung development. The lungs of fetal GR-null mice have increased lung weight and DNA content, are condensed and hypercellular, with reduced septal thinning leading to a 6-fold increase in the airway to capillary diffusion distance. In fetal GR-null mice, mRNA levels of the type II epithelial cell surfactant protein genes A and C were reduced by approximately 50%. Analysis of epithelial cell types by electron microscopy revealed that the proportions of type II cells were increased by approximately 30%, whereas the proportions of type-I cells were markedly reduced (by approximately 50%). Similarly, we found a 50% reduction in mRNA levels for T1alpha and aquaporin-5, two type I cell-specific markers, and a 20% reduction in aquaporin-1 mRNA levels. This demonstrates that during murine embryonic development, receptor-mediated glucocorticoid signaling facilitates the differentiation of epithelial cells into type I cells, but is not obligatory for type II cell differentiation.

Restoration of cytoadherence to an adherence-deficient mutant of Mycoplasma arthritidis by genetic complementation.

Mycoplasma arthritidis causes a severe septic arthritis in rats under natural and experimental conditions. An earlier study implicated a membrane lipoprotein designated MAA1 in cytadherence of M. arthritidis. In addition, a spontaneous adherence-deficient mutant was shown to contain a nonsense mutation in the gene encoding MAA1, resulting in production of a truncated product, MAA1Delta. In the present study, a wild-type maa1 gene carried on transposon Tn4001T was introduced into the low-adherence mutant by polyethylene glycol-mediated transformation. The presence of the tranposon and the wild-type maa1 gene in the chromosome of transformants was confirmed by PCR and Southern hybridization. The latter procedure also confirmed that each transformant contained a single copy of the transposon. Western immunoblotting showed that transformants produced both wild-type MAA1 and MAA1Delta, indicating that the introduced wild-type maa1 gene was functional. This phenotype was stably maintained after multiple subcultures even in the absence of antibiotic selection. Finally, transformants were shown to adhere to rat L-2 lung cells in culture at wild-type levels, providing confirmation for an important role for MAA1 in adherence.