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.

Immunohistochemical localization of osteoclastogenic cell mediators in feline tooth resorption and healthy teeth

Tooth resorption is among the most common and most challenging problems in feline dentistry It is a progressive disease eventually leading to tooth loss and often root replacement. The etiology of moth resorption remains obscure and to date no effective therapeutic approach is known. The present study is aimed at assessing the reliability of radiographic imaging and addressing the possible involvement of receptor activator of NF kappa B (RANK), its ligand (RANKL), and osteoprotegerin (OPG) in the process of tooth resorption. Teeth from 8 cats were investigated by means of radiographs and paraffin sections followed by immunolabeling. Six cats were diagnosed with tooth resorption based on histopathologic and radiographic findings. Samples were classified according to a four-stage diagnostic system. Radiologic assessment of tooth resorption correlated very strongly with histopathologic findings. Tooth resorption was accompanied by a strong staining with all three antibodies used, especially with anti-RANK and anti-RANKL antibodies. The presence of OPG and RANKL at the resorption site is indicative of repair attempts by fibroblasts and stromal cells. These findings should be extended by further investigations in order to elucidate the pathophysiologic processes underlying tooth resorption that might lead to prophylactic and/or therapeutic measures. J Vet Dent 27(2); 75 - 83, 2010

Complication and failure rates of fixed dental prostheses in patients treated for periodontal disease

Objectives: To evaluate the biological and technical complication rates of fixed dental prostheses (FDP) with end abutments or cantilever extensions on teeth (FDP-tt/cFDP-tt) on implants (FDP-ii/cFDP-ii) and tooth-implant-supported (FDP-ti/cFDP-ti) in patients treated for chronic periodontitis. Material and methods: From a cohort of 392 patients treated between 1978 and 2002 by graduate students, 199 were re-examined in 2005. Of these, 84 patients had received ceramo-metal FDPs (six groups). Results: At the re-evaluation, the mean age of the patients was 62 years (36.2–83.4). One hundred and seventy-five FDPs were seated (82 FDP-tt, 9 FDP-ii, 20 FDP-ti, 39 cFDP-tt, 15 cFDP-ii, 10 cFDP-ti). The mean observation time was 11.3 years; 21 FDPs were lost, and 46 technical and 50 biological complications occurred. Chances for the survival of the three groups of FDPs with end abutments were very high (risk for failure 2.8%, 0%, 5.6%). The probability to remain without complications and/or failure was 70.3%, 88.9% and 74.7% in FDPs with end abutments, but 49.8–25% only in FDPs with extensions at 10 years. Conclusions: In patients treated for chronic periodontitis and provided with ceramo-metal FDPs, high survival rates, especially for FDPs with end abutments, can be expected. The incidence rates of any negative events were increased drastically in the three groups with extension cFDPs (tt, ii, ti). Strategic decisions in the choice of a particular FDP design and the choice of teeth/implants as abutments appear to influence the risks for complications to be expected with fixed reconstruction. If possible, extensions on tooth abutments should be avoided or used only after a cautious clinical evaluation of all options.

[Multiple tooth resorption in an Italian greyhound]

An Italian greyhound was presented three times during a two-year period for dental prophylaxis due to periodontal disease. Clinical examination revealed lesions on several teeth. Radiographs revealed extensive resorptive root lesions. On histological examination, the presence of odontoclasts and signs of boney remodeling of the roots confirmed the resorptive nature of the lesions. Given the extent of the lesions, and poor prognosis with conservative treatment alone, teeth affected by the most severe resorption were extracted at each visit using a flap technique combined with alveolar vestibular osteotomy. Dental resorptive lesions are rarely detected in the dog but may be more frequent than previously thought. The routine use of dental radiographs can be used to reveal these lesions in the dog.

Long-term clinical outcomes of biodegradable polymer biolimus-eluting stents versus durable polymer sirolimus-eluting stents in patients with coronary artery disease (LEADERS): 4 year follow-up of a randomised non-inferiority trial

Background The effectiveness of durable polymer drug-eluting stents comes at the expense of delayed arterial healing and subsequent late adverse events such as stent thrombosis (ST). We report the 4 year follow-up of an assessment of biodegradable polymer-based drug-eluting stents, which aim to improve safety by avoiding the persistent inflammatory stimulus of durable polymers. Methods We did a multicentre, assessor-masked, non-inferiority trial. Between Nov 27, 2006, and May 18, 2007, patients aged 18 years or older with coronary artery disease were randomly allocated with a computer-generated sequence to receive either biodegradable polymer biolimus-eluting stents (BES) or durable polymer sirolimus-eluting stents (SES; 1:1 ratio). The primary endpoint was a composite of cardiac death, myocardial infarction, or clinically-indicated target vessel revascularisation (TVR); patients were followed-up for 4 years. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00389220. Findings 1707 patients with 2472 lesions were randomly allocated to receive either biodegradable polymer BES (857 patients, 1257 lesions) or durable polymer SES (850 patients, 1215 lesions). At 4 years, biodegradable polymer BES were non-inferior to durable polymer SES for the primary endpoint: 160 (18·7%) patients versus 192 (22·6%) patients (rate ratios [RR] 0·81, 95% CI 0·66–1·00, p for non-inferiority <0·0001, p for superiority=0·050). The RR of definite ST was 0·62 (0·35–1·08, p=0·09), which was largely attributable to a lower risk of very late definite ST between years 1 and 4 in the BES group than in the SES group (RR 0·20, 95% CI 0·06–0·67, p=0·004). Conversely, the RR of definite ST during the first year was 0·99 (0·51–1·95; p=0·98) and the test for interaction between RR of definite ST and time was positive (pinteraction=0·017). We recorded an interaction with time for events associated with ST but not for other events. For primary endpoint events associated with ST, the RR was 0·86 (0·41–1·80) during the first year and 0·17 (0·04–0·78) during subsequent years (pinteraction=0·049). Interpretation Biodegradable polymer BES are non-inferior to durable polymer SES and, by reducing the risk of cardiac events associated with very late ST, might improve long-term clinical outcomes for up to 4 years compared with durable polymer SES. Funding Biosensors Europe SA, Switzerland.

A Systematic Review of the Clinical Performance of Tooth-Retained and Implant-Retained Double Crown Prostheses with a Follow-Up of ≥3 Years

Purpose: The objective of this review was to systematically screen the literature for data related to the survival and complication rates observed with dental or implant double crown abutments and removable prostheses under functional loading for at least 3 years. Materials and Methods: A systematic review of the dental literature from January 1966 to December 2009 was performed in electronic databases (PubMed and Embase) as well as by an extensive hand search to investigate the clinical outcomes of double crown reconstructions. Results: From the total of 2412 titles retrieved from the search, 65 were selected for full-text review. Subsequently, 17 papers were included for data extraction. An estimation of the cumulative survival and complication rates was not feasible due to the lack of detailed information. Tooth survival rates for telescopic abutment teeth ranged from 82.5% to 96.5% after an observation period of 3.4 to 6 years, and for tooth-supported double crown retained dentures from 66.7% to 98.6% after an observation period of 6 to 10 years. The survival rates of implants were between 97.9% and 100% and for telescopic-retained removable dental prostheses with two mandibular implants, 100% after 3.0 and 10.4 years. The major biological complications affecting the tooth abutments were gingival inflammation, periodontal disease, and caries. The most frequent technical complications were loss of cementation and loss of facings. Conclusions: The main findings of this review are: (I) double crown tooth abutments and dentures demonstrated a wide range of survival rates. (II) Implant-supported mandibular overdentures demonstrated a favorable long-term prognosis. (III) A greater need for prosthetic maintenance is required for both tooth-supported and implant-supported reconstructions. (IV) Future areas of research would involve designing appropriate longitudinal studies for comparisons of survival and complication rates of different reconstruction designs.

Free-Ranging Wild Boar: A Disease Threat to Domestic Pigs in Switzerland?

The risk of transmission of pathogens from free-ranging wild boars (Sus scrofa scrofa) to outdoor domestic pigs (S. scrofa domesticus) is of increasing concern in many European countries. We assess this risk, using Switzerland as an example. We estimated 1) the prevalence of important pathogens in wild boars and 2) the risk of interactions between wild boars and outdoor pigs. First, we tested 252 wild boars from selected areas between 2008 and 2010 for infection with Brucella spp. Bacterial prevalence was estimated to 28.8% (confidence interval [CI] 23.0-34.0) when using bacterial culture (B. suis Biovar 2) and real-time polymerase chain reaction. Antibody prevalence was 35.8% (CI 30.0-42.0), which was significantly higher than in previous studies in Switzerland. We also tested 233 wild boars for porcine reproductive and respiratory syndrome virus (PRRSV). Antibody prevalence was 0.43% (CI 0.01-2.4) for EU-PRRSV and real-time reverse transcription polymerase chain reaction results were negative. These findings suggest that B. suis is increasingly widespread in wild boars and PRRSV is currently not of concern. Second, we documented the spatial overlap between free-ranging wild boars and outdoor piggeries by mapping data on their respective occurrence. Wild boars are most widespread in the mountain range along the western and northern Swiss borders, while most piggeries are located in central lowlands. A risk of interaction is mainly expected at the junction between these two bioregions. This risk may increase if wild boars expand eastward and southward beyond anthropogenic barriers believed to limit their range. Therefore, we evaluated the potential of expansion of the wild boar population. Population trends suggest a continuous increase of wild boars for the past 15 yr. Surveillance of selected wildlife passages using cameras on highways and main roads indicates that these barriers are permeable (average of up to 13 wild boar crossings per 100 days). Thus an increase of wild boar range should be considered. There may be a risk of B. suis spillover from wild boars in Switzerland, which could increase in the future. Data on the occurrence of interactions between pigs and wild boars are needed to assess this risk.

Renal tissue oxygenation in essential hypertension and chronic kidney disease

Animal studies suggest that renal tissue hypoxia plays an important role in the development of renal damage in hypertension and renal diseases, yet human data were scarce due to the lack of noninvasive methods. Over the last decade, blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI), detecting deoxyhemoglobin in hypoxic renal tissue, has become a powerful tool to assess kidney oxygenation noninvasively in humans. This paper provides an overview of BOLD-MRI studies performed in patients suffering from essential hypertension or chronic kidney disease (CKD). In line with animal studies, acute changes in cortical and medullary oxygenation have been observed after the administration of medication (furosemide, blockers of the renin-angiotensin system) or alterations in sodium intake in these patient groups, underlining the important role of renal sodium handling in kidney oxygenation. In contrast, no BOLD-MRI studies have convincingly demonstrated that renal oxygenation is chronically reduced in essential hypertension or in CKD or chronically altered after long-term medication intake. More studies are required to clarify this discrepancy and to further unravel the role of renal oxygenation in the development and progression of essential hypertension and CKD in humans.

Chlorhexidine and preservation of sound tooth structure in older adults. A placebo-controlled trial

The Trial to Enhance Elderly Teeth Health (TEETH) was designed to test the impact of regular rinsing with a 0.12% chlorhexidine (CHX) solution on tooth loss, and the causes of tooth loss (caries, periodontal disease and trauma) were also investigated. This paper reports on the effectiveness of a 0.12% CHX solution for controlling caries using a tooth surface (coronal and root) survival analysis. A total of 1,101 low income elders in Seattle (United States) and Vancouver (Canada), aged 60-75 years, were recruited for a double-blind clinical trial and assigned to either a CHX (n = 550) or a placebo (n = 551) mouth rinse. Subjects alternated between daily rinsing for 1 month, followed by weekly rinsing for 5 months. All sound coronal and root surfaces at baseline were followed annually for up to 5 years. At each follow-up examination, those tooth surfaces with caries, restored, or extracted were scored as 'carious'. The hazard ratio associated with CHX for a sound surface to become filled, decayed, or extracted was 0.87 for coronal surfaces (95% confidence interval: 0.71-1.14, p = 0.20) and 0.91 for root surfaces (95% confidence interval: 0.73-1.14, p = 0.41). These findings suggest that regular rinsing with CHX does not have a substantial effect on the preservation of sound tooth structure in older adults.