An unusual splice defect in the mitofusin 2 gene (MFN2) is associated with degenerative axonopathy in Tyrolean Grey cattle

Tyrolean Grey cattle represent a local breed with a population size of approximately 5000 registered cows. In 2003, a previously unknown neurological disorder was recognized in Tyrolean Grey cattle. The clinical signs of the disorder are similar to those of bovine progressive degenerative myeloencephalopathy (weaver syndrome) in Brown Swiss cattle but occur much earlier in life. The neuropathological investigation of an affected calf showed axonal degeneration in the central nervous system (CNS) and femoral nerve. The pedigrees of the affected calves suggested a monogenic autosomal recessive inheritance. We localized the responsible mutation to a 1.9 Mb interval on chromosome 16 by genome-wide association and haplotype mapping. The MFN2 gene located in this interval encodes mitofusin 2, a mitochondrial membrane protein. A heritable human axonal neuropathy, Charcot-Marie-Tooth disease-2A2 (CMT2A2), is caused by MFN2 mutations. Therefore, we considered MFN2 a positional and functional candidate gene and performed mutation analysis in affected and control Tyrolean Grey cattle. We did not find any non-synonymous variants. However, we identified a perfectly associated silent SNP in the coding region of exon 20 of the MFN2 gene. This SNP is located within a putative exonic splice enhancer (ESE) and the variant allele leads to partial retention of the entire intron 19 and a premature stop codon in the aberrant MFN2 transcript. Thus we have identified a highly unusual splicing defect, where an exonic single base exchange leads to the retention of the preceding intron. This splicing defect represents a potential explanation for the observed degenerative axonopathy. Marker assisted selection can now be used to eliminate degenerative axonopathy from Tyrolean Grey cattle.

A deletion in the N-myc downstream regulated gene 1 (NDRG1) gene in Greyhounds with polyneuropathy

The polyneuropathy of juvenile Greyhound show dogs shows clinical similarities to the genetically heterogeneous Charcot-Marie-Tooth (CMT) disease in humans. The pedigrees containing affected dogs suggest monogenic autosomal recessive inheritance and all affected dogs trace back to a single male. Here, we studied the neuropathology of this disease and identified a candidate causative mutation. Peripheral nerve biopsies from affected dogs were examined using semi-thin histology, nerve fibre teasing and electron microscopy. A severe chronic progressive mixed polyneuropathy was observed. Seven affected and 17 related control dogs were genotyped on the 50k canine SNP chip. This allowed us to localize the causative mutation to a 19.5 Mb interval on chromosome 13 by homozygosity mapping. The NDRG1 gene is located within this interval and NDRG1 mutations have been shown to cause hereditary motor and sensory neuropathy-Lom in humans (CMT4D). Therefore, we considered NDRG1 a positional and functional candidate gene and performed mutation analysis in affected and control Greyhounds. A 10 bp deletion in canine NDRG1 exon 15 (c.1080_1089delTCGCCTGGAC) was perfectly associated with the polyneuropathy phenotype of Greyhound show dogs. The deletion causes a frame shift (p.Arg361SerfsX60) which alters several amino acids before a stop codon is encountered. A reduced level of NDRG1 transcript could be detected by RT-PCR. Western blot analysis demonstrated an absence of NDRG1 protein in peripheral nerve biopsy of an affected Greyhound. We thus have identified a candidate causative mutation for polyneuropathy in Greyhounds and identified the first genetically characterized canine CMT model which offers an opportunity to gain further insights into the pathobiology and therapy of human NDRG1 associated CMT disease. Selection against this mutation can now be used to eliminate polyneuropathy from Greyhound show dogs.

Severe phenotype with cis-acting heterozygous PMP22 mutations

We report on a 20-year-old male with severe Charcot-Marie-Tooth (CMT) disease and a de novo deletion (c.281delG, p.G94AfsX17) on the paternal PMP22 allele harboring c.353C>T (p.T118M). RNA-based sequence analysis confirmed the absence of nonsense-mediated decay and the presence of the mutant transcripts in Epstein-Barr virus-transformed lymphoblastoid cells of our patient. His clinical findings included early onset of polyneuropathy, loss of muscle mass with distal pareses, hammer toes, and progressive scoliosis. There was no neuropsychological alteration. Our results suggest that the deletion c.281delG alone is responsible for the severe CMT phenotype. To the best of our knowledge, this is the second report on a proven paternal origin of a de novo single-base mutation in the PMP22 gene.

An ARHGEF10 deletion is highly associated with a juvenile-onset inherited polyneuropathy in Leonberger and Saint Bernard dogs

An inherited polyneuropathy (PN) observed in Leonberger dogs has clinical similarities to a genetically heterogeneous group of peripheral neuropathies termed Charcot-Marie-Tooth (CMT) disease in humans. The Leonberger disorder is a severe, juvenile-onset, chronic, progressive, and mixed PN, characterized by exercise intolerance, gait abnormalities and muscle atrophy of the pelvic limbs, as well as inspiratory stridor and dyspnea. We mapped a PN locus in Leonbergers to a 250 kb region on canine chromosome 16 (Praw = 1.16×10-10, Pgenome, corrected = 0.006) utilizing a high-density SNP array. Within this interval is the ARHGEF10 gene, a member of the rho family of GTPases known to be involved in neuronal growth and axonal migration, and implicated in human hypomyelination. ARHGEF10 sequencing identified a 10 bp deletion in affected dogs that removes four nucleotides from the 3'-end of exon 17 and six nucleotides from the 5'-end of intron 17 (c.1955_1958+6delCACGGTGAGC). This eliminates the 3'-splice junction of exon 17, creates an alternate splice site immediately downstream in which the processed mRNA contains a frame shift, and generates a premature stop codon predicted to truncate approximately 50% of the protein. Homozygosity for the deletion was highly associated with the severe juvenile-onset PN phenotype in both Leonberger and Saint Bernard dogs. The overall clinical picture of PN in these breeds, and the effects of sex and heterozygosity of the ARHGEF10 deletion, are less clear due to the likely presence of other forms of PN with variable ages of onset and severity of clinical signs. This is the first documented severe polyneuropathy associated with a mutation in ARHGEF10 in any species.

Defective propagation of signals generated by sympathetic nerve stimulation in the liver of connexin32-deficient mice.

The gap junctional protein connexin32 is expressed in hepatocytes, exocrine pancreatic cells, Schwann cells, and other cell types. We have inactivated the connexin32 gene by homologous recombination in the mouse genome and have generated homozygous connexin32-deficient mice that were viable and fertile but weighed on the average approximately 17% less than wild-type controls. Electrical stimulation of sympathetic nerves in connexin32-deficient liver triggered a 78% lower amount of glucose mobilization from glycogen stores, when compared with wild-type liver. Thus, connexin32-containing gap junctions are essential in mouse liver for maximal intercellular propagation of the noradrenaline signal from the periportal (upstream) area, where it is received from sympathetic nerve endings, to perivenous (downstream) hepatocytes. In connexin32-defective liver, the amount of connexin26 protein expressed was found to be lower than in wild-type liver, and the total area of gap junction plaques was approximately 1000-fold smaller than in wild-type liver. In contrast to patients with connexin32 defects suffering from X chromosome-linked Charcot-Marie-Tooth disease (CMTX) due to demyelination in Schwann cells of peripheral nerves, connexin32-deficient mice did not show neurological abnormalities when analyzed at 3 months of age. It is possible, however, that they may develop neurodegenerative symptoms at older age.

Septin6 and Septin7 GTP binding proteins regulate AP-3- and ESCRT-dependent multivesicular body biogenesis

Septins (SEPTs) form a family of GTP-binding proteins implicated in cytoskeleton and membrane organization, cell division and host/pathogen interactions. The precise function of many family members remains elusive. We show that SEPT6 and SEPT7 complexes bound to F-actin regulate protein sorting during multivesicular body (MVB) biogenesis. These complexes bind AP-3, an adapter complex sorting cargos destined to remain in outer membranes of maturing endosomes, modulate AP-3 membrane interactions and the motility of AP-3-positive endosomes. These SEPT-AP interactions also influence the membrane interaction of ESCRT (endosomal-sorting complex required for transport)-I, which selects ubiquitinated cargos for degradation inside MVBs. Whereas our findings demonstrate that SEPT6 and SEPT7 function in the spatial, temporal organization of AP-3- and ESCRT-coated membrane domains, they uncover an unsuspected coordination of these sorting machineries during MVB biogenesis. This requires the E3 ubiquitin ligase LRSAM1, an AP-3 interactor regulating ESCRT-I sorting activity and whose mutations are linked with Charcot-Marie-Tooth neuropathies.

Do Psychosocial Interventions Improve Quality of Life and Wellbeing in Adults with Neuromuscular Disorders? A Systematic Review and Narrative Synthesis

Background: Quality of life and well-being are frequently restricted in adults with neuromuscular disorders. As such, identification of appropriate interventions is imperative. Objective: The objective of this paper was to systematically review and critically appraise quantitative studies (RCTs, controlled trials and cohort studies) of psychosocial interventions designed to improve quality of life and well-being in adults with neuromuscular disorders. Method: A systematic review of the published and unpublished literature was conducted. Studies meeting inclusion criteria were appraised using a validated quality assessment tool and results presented in a narrative synthesis. Results: Out of 3,136 studies identified, ten studies met criteria for inclusion within the review. Included studies comprised a range of interventions including: cognitive behavioural therapy, dignity therapy, hypnosis, expressive disclosure, gratitude lists, group psychoeducation and psychologically informed rehabilitation. Five of the interventions were for patients with Amyotrophic Lateral Sclerosis (ALS). The remainder were for patients with post-polio syndrome, muscular dystrophies and mixed disorders, such as Charcot-Marie-Tooth disease, myasthenia gravis and myotonic dystrophy. Across varied interventions and neuromuscular disorders, seven studies reported a short-term beneficial effect of intervention on quality of life and well-being. Whilst such findings are encouraging, widespread issues with the methodological quality of these studies significantly compromised the results. Conclusion: There is no strong evidence that psychosocial interventions improve quality of life and well-being in adults with neuromuscular disorders, due to a paucity of high quality research in this field. Multi-site, randomised controlled trials with active controls, standardised outcome measurement and longer term follow-ups are urgently required.

Does psychological stress mediate social deprivation in tooth loss?

It is unclear which theoretical dimension of psychological stress affects health status. We hypothesized that both distress and coping mediate the relationship between socio-economic position and tooth loss. Cross-sectional data from 2915 middle-aged adults evaluated retention of < 20 teeth, behaviors, psychological stress, and sociodemographic characteristics. Principal components analysis of the Perceived Stress Scale (PSS) extracted 'distress' (a = 0.85) and 'coping' (a =0.83) factors, consistent with theory. Hierarchical entry of explanatory variables into age- and sex-adjusted logistic regression models estimated odds ratios (OR) and 95% confidence intervals [95% CI] for retention of < 20 teeth. Analysis of the separate contributions of distress and coping revealed a significant main effect of coping (OR = 0.7 [95% CI = 0.7-0.8]), but no effect for distress (OR = 1.0 [95% CI = 0.9-1.1]) or for the interaction of coping and distress. Behavior and psychological stress only modestly attenuated socio-economic inequality in retention of < 20 teeth, providing evidence to support a mediating role of coping.

Celebrating difference/censoring difference? : Marie Chouinard’s bODY rEMIX

This paper examines performances that defy established representations of disease, deformity and bodily difference. Historically, the ‘deformed’ body has been cast – onstage and in sideshows – as flawed, an object of pity, or an example of the human capacity to overcome. Such representations define the boundaries of the ‘normal’ body by displaying its Other. They bracket the ‘abnormal’ body off as an example of deviance from the ‘norm’, thus, paradoxically, decreasing the social and symbolic visibility (and agency) of disabled people. Yet, in contemporary theory and culture, these representations are reappropriated – by disabled artists, certainly, but also as what Carrie Sandahl has called a ‘master trope’ for representing a range of bodily differences. In this paper, I investigate this phenomenon. I analyse French Canadian choreographer Marie Chouinard’s bODY rEMIX/gOLDBERG vARIATIONS, in which 10 able-bodied dancers are reborn as bizarre biotechnical mutants via the use of crutches, walkers, ballet shoes and barres as prosthetic pseudo-organs. These bodies defy boundaries, defy expectations, develop new modes of expression, and celebrate bodily difference. The self-inflicted pain dancers experience during training is cast as a ‘disablement’ that is ultimately ‘enabling’. I ask what effect encountering able bodies celebrating ‘dis’ or ‘diff’ ability has on audiences. Do we see the emergence of a once-repressed Other, no longer silenced, censored or negated? Or does using ‘disability’ to express the dancers’ difference and self-determination usurp a ‘trope’ by which disabled people themselves might speak back to the dominant culture, creating further censorship?

Tooth fracture risk analysis based on a new finite element dental structure models using micro-CT data

The finite element (FE) analysis is an effective method to study the strength and predict the fracture risk of endodontically-treated teeth. This paper presents a rapid method developed to generate a comprehensive tooth FE model using data retrieved from micro-computed tomography (μCT). With this method, the inhomogeneity of material properties of teeth was included into the model without dividing the tooth model into different regions. The material properties of the tooth were assumed to be related to the mineral density. The fracture risk at different tooth portions was assessed for root canal treatments. The micro-CT images of a tooth were processed by a Matlab software programme and the CT numbers were retrieved. The tooth contours were obtained with thresholding segmentation using Amira. The inner and outer surfaces of the tooth were imported into Solidworks and a three-dimensional (3D) tooth model was constructed. An assembly of the tooth model with the periodontal ligament (PDL) layer and surrounding bone was imported into ABAQUS. The material properties of the tooth were calculated from the retrieved CT numbers via ABAQUS user's subroutines. Three root canal geometries (original and two enlargements) were investigated. The proposed method in this study can generate detailed 3D finite element models of a tooth with different root canal enlargements and filling materials, and would be very useful for the assessment of the fracture risk at different tooth portions after root canal treatments.

Developing an evidence-based clinical pathway for the assessment, diagnosis and management of acute Charcot Neuro-Arthropathy : a systematic review

Background: Charcot Neuro-Arthropathy (CN) is one of the more devastating complications of diabetes. To the best of the authors' knowledge, it appears that no clinical tools based on a systematic review of existing literature have been developed to manage acute CN. Thus, the aim of this paper was to systematically review existing literature and develop an evidence-based clinical pathway for the assessment, diagnosis and management of acute CN in patients with diabetes. Methods: Electronic databases (Medline, PubMed, CINAHL, Embase and Cochrane Library), reference lists, and relevant key websites were systematically searched for literature discussing the assessment, diagnosis and/or management of acute CN published between 2002-2012. At least two independent investigators then quality rated and graded the evidence of each included paper. Consistent recommendations emanating from the included papers were then fashioned in a clinical pathway. Results: The systematic search identified 267 manuscripts, of which 117 (44%) met the inclusion criteria for this study. Most manuscripts discussing the assessment, diagnosis and/or management of acute CN constituted level IV (case series) or EO (expert opinion) evidence. The included literature was used to develop an evidence-based clinical pathway for the assessment, investigations, diagnosis and management of acute CN. Conclusions: This research has assisted in developing a comprehensive, evidence-based clinical pathway to promote consistent and optimal practice in the assessment, diagnosis and management of acute CN. The pathway aims to support health professionals in making early diagnosis and providing appropriate immediate management of acute CN, ultimately reducing its associated complications such as amputations and hospitalisations.

Consent for third molar tooth extractions in Australia and New Zealand: A review of current practice

Background Informed consent is the legal requirement to educate a patient about a proposed medical treatment or procedure so that he or she can make informed decisions. The purpose of the study was to examine the current practice for obtaining informed consent for third molar tooth extractions (wisdom teeth) by Oral and Maxillofacial Surgeons in Australia and New Zealand. Methods An online survey was sent to 180 consultant Oral and Maxillofacial Surgeons in Australia and New Zealand. Surgeons were asked to answer (yes/no) whether they routinely warned of a specific risk of third molar tooth extraction in their written consent. Results 71 replies were received (39%). The only risks that surgeons agreed should be routinely included in written consent were a general warning of infection (not alveolar osteitis), inferior alveolar nerve damage (temporary and permanent) and lingual nerve damage (temporary and permanent). Conclusions There is significant variability among Australian and New Zealand Oral and Maxillofacial Surgeons regarding risk disclosure for third molar tooth extractions. We aim to improve consistency in consent for third molar extractions by developing an evidence-based consent form.

Estimation of gear tooth deflection by the finite element method

The finite element method (FEM) is used to determine for pitch-point, mid-point and tip loading, the deflection curve of a Image 1 diamentral pitch (DP) standard spur gear tooth corresponding to number of teeth of 14, 21, 26 and 34. In all these cases the deflection of the gear tooth at the point of loading obtained by FEM is in good agreement with the experimental value. The contraflexure in the deflection curve at the point of loading observed experimentally in the cases of pitch-point and mid-point loading, is predicted correctly by the FEM analysis.

Characterisation of the efficacy of endodontic medications using a three-dimensional fluorescent tooth model: An ex vivo study

The purpose of this study was to establish a three-dimensional fluorescent tooth model to investigate bacterial viability against intra-canal medicaments across the thickness and surface of root dentine. Dental microbial biofilms (Enterococcus faecalis and Streptococcus mutans) were established on the external root surface and bacterial kill was monitored over time against intra-canal medicament (Ca(OH)2 ) using fluorescent microscopy in conjunction with BacLight SYTO9 and propidium iodide stains. An Olympus digital camera fitted to SZX16 fluorescent microscope captured images of bacterial cells in biofilms on the external root surface. Viability of biofilm was measured by calculating the total pixel area of green (viable bacteria) and red (non-viable bacteria) for each image using ImageJ® software. All data generated were assessed for normality and then analysed using a Mann-Whitney t-test. The viability of S. mutans biofilm following Ca(OH)2 treatment showed a significant decline compared with the untreated group (P = 0.0418). No significant difference was seen for E. faecalis biofilm between the Ca(OH)2 and untreated groups indicating Ca(OH)2 medicament is ineffective against E. faecalis biofilm. This novel three-dimensional fluorescent biofilm model provides a new clinically relevant tool for testing of medicaments against dental biofilms.