Distribution of mutations in the PEX gene in families with X-linked hypophosphataemic rickets (HYP)

Mutations in the PEX gene at Xp22.1 (phosphate-regulating gene with homologies to endopeptidases, on the X-chromosome), are responsible for X-linked hypophosphataemic rickets (HYP). Homology of PEX to the M13 family of Zn2+ metallopeptidases which include neprilysin (NEP) as prototype, has raised important questions regarding PEX function at the molecular level. The aim of this study was to analyse 99 HYP families for PEX gene mutations, and to correlate predicted changes in the protein structure with Zn2+ metallopeptidase gene function. Primers flanking 22 characterised exons were used to amplify DNA by PCR, and SSCP was then used to screen for mutations. Deletions, insertions, nonsense mutations, stop codons and splice mutations occurred in 83% of families screened for in all 22 exons, and 51% of a separate set of families screened in 17 PEX gene exons. Missense mutations in four regions of the gene were informative regarding function, with one mutation in the Zn2+-binding site predicted to alter substrate enzyme interaction and catalysis. Computer analysis of the remaining mutations predicted changes in secondary structure, N-glycosylation, protein phosphorylation and catalytic site molecular structure. The wide range of mutations that align with regions required for protease activity in NEP suggests that PEX also functions as a protease, and may act by processing factor(s) involved in bone mineral metabolism.      

Investigating transmission of Mycobacterium bovis in the United Kingdom in 2005 to 2008

Due to an increase in bovine tuberculosis in cattle in the United Kingdom, we investigated the characteristics of Mycobacterium bovis infection in humans and assessed whether extensive transmission of M. bovis between humans has occurred. A cross-sectional study linking demographic, clinical, and DNA fingerprinting (using 15-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat [MIRU-VNTR] typing) data on cases reported between 2005 and 2008 was undertaken. A total of 129 cases of M. bovis infection in humans were reported over the period, with a decrease in annual incidence from 0.065 to 0.047 cases per 100,000 persons. Most patients were born pre-1960, before widespread pasteurization was introduced (73%), were of white ethnicity (83%), and were born in the United Kingdom (76%). A total of 102 patients (79%) had MIRU-VNTR typing data. A total of 31 of 69 complete MIRU-VNTR profiles formed eight distinct clusters. The overall clustering proportion determined using the n - 1 method was 33%. The largest cluster, comprising 12 cases, was indistinguishable from a previously reported West Midlands outbreak strain cluster and included those cases. This cluster was heterogeneous, having characteristics supporting recent zoonotic and human-to-human transmission as well as reactivation of latent disease. Seven other, smaller clusters identified had demographics supporting recrudescence rather than recent infection. A total of 33 patients had incomplete MIRU-VNTR profiles, of which 11 may have yielded 2 to 6 further small clusters if typed to completion. The incidence of M. bovis in humans in the United Kingdom remains low, and the epidemiology is predominantly that of reactivated disease.     

Overview and update on molecular testing in non-small cell lung carcinoma utilizing endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) samples

Lung carcinoma remains one of the most frequent and aggressive human neoplasms. Fortunately, in the last decades, the increasing knowledge of the molecular mechanisms leading to cancer development has allowed the use of targeted therapies with improvement of prognosis in many patients. Clinical management has also changed after the introduction of endobronchialultrasonographic bronchoscopy that allows a conservative staging of lung tumors, avoiding the need of mediastinoscopy for lymph node staging. Lung pathologists and cytopathologists are facing the challenge of giving the more comprehensive prognostic and predictive information with ever smaller tissue or cytological samples. The aim of this review is to summarize the molecular testing for non-small cell lung carcinoma and how pathologists can contribute to the patient's outcome with a conscious management of biological samples.     

Coadministration of Interleukin 12 Expression Vector with Antigen 2 cDNA Enhances Induction of Protective Immunity against Coccidioides immitis

Interleukin 12 (IL-12) plays an important role in the induction of protective immunity against cancer and infectious diseases. In this study we asked whether IL-12 cDNA could increase the protective capacity of the antigen 2 (Ag2) gene vaccine in experimental coccidioidomycosis. Coimmunization of BALB/c mice with a single-chain IL-12 cDNA (p40-L-p35) and Ag2 cDNA, both subcloned into the pVR1012 plasmid, significantly enhanced protection against systemic challenge with 2,500 arthroconidia, as evidenced by a greater-than-1.3-log-unit reduction in the fungal load in the lungs and spleens compared to mice receiving the pVR1012 vector alone, Ag2 cDNA alone, or IL-12 cDNA alone. The enhanced protection was associated with increased gamma interferon secretion; production of immunoglobulin G2a (IgG2a), IgG2b, and IgG3 antibodies to Coccidioides immitis antigen; and the influx of CD4(+) and CD8(+) T cells in lungs and spleens. When challenged by the pulmonary route, mice covaccinated with Ag2 cDNA and IL-12 cDNA were not protected at the lung level but did show a significant reduction in the fungal load in their livers and spleens compared to mice vaccinated with Ag2 cDNA or IL-12 cDNA alone. These results suggest that IL-12 acts as a therapeutic adjuvant to enhance Ag2 cDNA-induced protective immunity against experimental coccidioidomycosis through the induction of Th1-associated immune responses.     

PLP1 gene analysis in 88 patients with leukodystrophy

Pelizaeus–Merzbacher disease (PMD) is caused in most cases by either duplications or point mutations in the PLP1 gene. This disease, a dysmyelinating disorder affecting mainly the central nervous system, has a wide clinical spectrum and its causing mutations act through different molecular mechanisms. Eighty‐eight male patients with leukodystrophy were studied. PLP1 gene analysis was performed by the Multiplex Ligation‐dependent Probe Amplification technique and DNA sequencing, and, in duplicated cases of PLP1, gene dosage was completed by using array‐CGH. We have identified 21 patients with mutations in the PLP1 gene, including duplications, short and large deletions and several point mutations in our cohort. A customized array‐CGH at the Xq22.2 area identified several complex rearrangements within the PLP1 gene region. Mutations found in the PLP1 gene are the cause of PMD in around 20% of the patients in this series.     

The complementary role of music therapy in the detection of awareness in disorders of consciousness: An audit of concurrent SMART and MATADOC assessments

In the behavioural assessment of disorders of consciousness (DOC), best practice is for several different assessment tools to be used to encourage a variety of different responses indicative of awareness. Anecdotal evidence suggests a range of musical stimuli may be particularly effective in eliciting responses to guide the assessment process, although comparative data regarding behavioural domains is lacking. This study examined 42 concurrent records of patients assessed using the Sensory Modality Assessment and Rehabilitation Technique (SMART), and the Music Therapy Assessment Tool for Awareness in Disorders of Consciousness (MATADOC) to explore the relationship between diagnosis and behavioural characteristics of the cohort. Whilst the two tools produced a high level of agreement in diagnostic outcome (Spearman Rho .80), divergent diagnosis and weaker correlations between behavioural response items highlight contrasting sensitivities of the tools. Whilst MATADOC has higher sensitivity within auditory and visual domains relative to SMART, SMART has higher sensitivity in the motor domain. The significant contribution of musical response items in MATADOC, and the tactile response item in SMART, indicates both tools provide unique behavioural data predictive of awareness. Multidisciplinary assessment using SMART and MATADOC provides complementary data contributing to a fuller understanding of a patient's level of awareness.     

Music Therapy Assessment Tool for Awareness in Disorders of Consciousness (MATADOC): Standardisation of the principal subscale to assess awareness in patients with disorders of consciousness

Establishing valid and reliable measures for use with patients with disorders of consciousness (DOC) following profound brain injury is challenging due to a number of factors including the complex presentation of such patients and assessor variability. The auditory modality has been demonstrated to have greater sensitivity for detecting awareness in DOC patients. However, there are no measures developed to assess auditory responsiveness specifically. The objective of this study was to examine the psychometric properties of the principal subscale of a music therapy assessment tool (MATADOC) developed for use with adult DOC patients. The subscale assesses behavioural domains essential for diagnosis of awareness. Twenty-one adult patients were recruited from a specialist rehabilitation unit. In a prospective study with repeated measures, internal consistency, inter-rater and test–retest reliability and dimensionality were examined. The five-item scale showed satisfactory internal reliability (α = .76) and a strong first principal component. Corrected item-total correlations were all > .45. Inter-rater intra-class correlations (ICCs) ranged from 0.65–1.00 and intra-rater ICCs from 0.77–0.90. Rasch analysis confirmed these impressions of a reliable, unidimensional and homogenous scale. Diagnostic outcomes had 100% agreement with a validated external reference standard. The results indicate that the MATADOC principal subscale provides a new behavioural measure that can contribute to interdisciplinary assessment of awareness with DOC patients.