Oral mucosal morphea: a new variant

Morphea is a cutaneous disorder characterized by an excessive collagen deposition. While in almost all cases the sclerosing process exclusively affects the skin, there are anecdotal cases in which associated mucosal involvement has been described. We here report the case of a woman developing a whitish indurated plaque over the left upper vestibular mucosa and hard palate leading to dental mobility and exposure of the roots of several teeth. Cone beam computed tomography of the left maxilla showed bone resorption involving the upper cuspid to the second molar region with widened periodontal ligament spaces, while light microscopy studies demonstrated epithelial atrophy and fibrosis of the dermis extending into the submucosa with hyalinization of subepithelial collagen. Our observation expands the spectrum of clinical presentations of morphea and provides the first example of isolated oral morphea. Its recognition is important to avoid significant local complications.

L51P - A BMP2 variant with osteoinductive activity via inhibition of Noggin

Bone morphogenetic proteins (BMP) have to be applied at high concentrations to stimulate bone healing. The limited therapeutic efficacy may be due to the local presence of BMP antagonists such as Noggin. Thus, inhibiting BMP antagonists is an attractive therapeutic option. We hypothesized that the engineered BMP2 variant L51P stimulates osteoinduction by antagonizing Noggin-mediated inhibition of BMP2. Primary murine osteoblasts (OB) were treated with L51P, BMP2, and Noggin. OB proliferation and differentiation were quantified with XTT and alkaline phosphatase (ALP) assays. BMP receptor dependent intracellular signaling in OB was evaluated with Smad and p38 MAPK phosphorylation assays. BMP2, Noggin, BMP receptor Ia/Ib/II, osteocalcin, and ALP mRNA expressions were analyzed with real-time PCR. L51P stimulated OB differentiation by blocking Noggin mediated inhibition of BMP2. L51P did not induce OB differentiation directly and did not activate BMP receptor dependent intracellular signaling via the Smad pathway. Treatment of OB cultures with BMP2 but not with L51P resulted in an increased expression of ALP, BMP2, and Noggin mRNA. By inhibiting the BMP antagonist Noggin, L51P enhances BMP2 activity and stimulates osteoinduction without exhibiting direct osteoinductive function. Indirect osteoinduction with L51P seems to be advantageous to osteoinduction with BMP2 as BMP2 stimulates the expression of Noggin thereby self-limiting its own osteoinductive activity. Treatment with L51P is the first protein-based approach available to augment BMP2 induced bone regeneration through inhibition of BMP antagonists. The described strategy may help to decrease the amounts of exogenous BMPs currently required to stimulate bone healing.

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 new variant of Actinobacillus pleuropneumoniae serotype 3 lacking the entire apxII operon

Actinobacillus pleuropneumoniae is an important respiratory pathogen causing pleuropneumonia in pig. The species is genetically characterized by the presence of 4 RTX (Repeats in the Structural ToXin) toxin genes: apxI, apxII, and apxIII genes are differentially present in various combinations among the different serotypes, thereby defining pathogenicity; the apxIV gene is present in all serotypes. Polymerase chain reaction (PCR)-based apx gene typing is done in many veterinary diagnostic laboratories, especially reference laboratories. The present report describes the isolation of atypical A. pleuropneumoniae from 4 independent cases from 2 countries. All isolates were beta-nicotinamide adenine dinucleotide (beta-NAD) dependent and nonhemolytic but showed strong co-hemolysis with the sphingomyelinase of Staphylococcus aureus on sheep blood agar. Classical biochemical tests as well as Matrix-assisted laser desorption ionization time-of-flight mass spectrometry and sequence-based analysis (16S ribosomal RNA [rRNA] and rpoB genes) identified them as A. pleuropneumoniae. Apx-toxin gene typing using 2 different PCR systems showed the presence of apxIV and only the apxIII operon (apxIIICABD). None of the apxI or apxII genes were present as confirmed by Southern blot analysis. The 16S rRNA and rpoB gene analyses as well as serotype-specific PCR indicate that the isolates are variants of serotype 3. Strains harboring only apxIV and the apxIII operon are possibly emerging types of A. pleuropneumoniae and should therefore be carefully monitored for epidemiological reasons.

A locus on chromosome 5 is associated with dilated cardiomyopathy in Doberman Pinschers

Dilated cardiomyopathy (DCM) is a heterogeneous group of heart diseases with a strong genetic background. Currently, many human DCM cases exist where no causative mutation can be identified. DCM also occurs with high prevalence in several large dog breeds. In the Doberman Pinscher a specific DCM form characterized by arrhythmias and/or echocardiographic changes has been intensively studied by veterinary cardiologists. We performed a genome-wide association study in Doberman Pinschers. Using 71 cases and 70 controls collected in Germany we identified a genome-wide significant association to DCM on chromosome 5. We validated the association in an independent cohort collected in the United Kingdom. There is no known DCM candidate gene under the association signal. Therefore, DCM in Doberman Pinschers offers the chance of identifying a novel DCM gene that might also be relevant for human health.

Plexiform hybrid granular cell tumor/perineurioma: a novel variant of benign peripheral nerve sheath tumor with divergent differentiation

The descriptive term hybrid peripheral nerve sheath tumor refers to any neoplasm of the neurilemmal apparatus composed of more than one pathologically defined tumoral equivalent derived from its constituent cells. Within this uncommon nosological category, participation of granular cell tumor - a neoplasm of modified Schwann cells - has been reported only exceptionally. We describe a hitherto not documented variant composed of an organoid mixture of granular cell tumor and perineurioma with plexiform growth. A solitary subcutaneous nodule of 1.5 cm diameter was excised from the right ring finger of a 19-year-old female with no antecedents of neurofibromatosis or relevant trauma. Histology revealed a monotonous, yet cytologically dimorphic proliferation of classical granular cells intermingled with flattened, inconspicuous perineurial cells. Immunohistochemical double labeling detected expression of S100 protein in the former and of EMA and GLUT-1 in the latter. While the respective staining patterns for S100 protein and EMA or GLUT-1 tended to be mutually exclusive, a minority of cells exhibited transitional granular cell/perineurial immunophenotype. Electron microscopy permitted direct visualization of a plethora of lysosomes in the granular cell moiety, and of pinocytotic vesicles and tight junctions in perineurial cells. Intratumoral axons were not detected. Expanding intraneurally, the lesion showed discrete encapsulation by the local perineurium, and resulted in plexiform growth. The MIB-1 labeling index averaged 1%. We interpret our findings as supporting evidence for the dual cell lineage to have arisen through metaplasia, with the tumor's dynamics probably having been driven by the granular cell component.

Inhibition of endogenous antagonists with an engineered BMP-2 variant increases BMP-2 efficacy in rat femoral defect healing

Bone morphogenetic proteins (BMP) have been used successfully by orthopedic clinicians to augment bone healing. However, these osteoinductive proteins must be applied at high concentrations to induce bone formation. The limited therapeutic efficacy may be due to the local expression of BMP antagonists such as Noggin that neutralize exogenous and endogenous BMPs. If so, inhibiting BMP antagonists may provide an attractive option to augment BMP induced bone formation. The engineered BMP-2 variant L51P is deficient in BMP receptor type I binding, but maintains its affinity for BMP receptor type II and BMP antagonists including Noggin, Chordin and Gremlin. This modification makes L51P a BMP receptor-inactive inhibitor of BMP antagonists. We implanted β-tricalcium phosphate ceramics loaded with BMP-2 and/or L51P into a critical size defect model in the rat femur to investigate whether the inhibition of BMP antagonist with L51P enhances the therapeutic efficacy of exogenous BMP-2. Our study reveals that L51P reduces the demand of exogenous BMP-2 to induce bone healing markedly, without promoting bone formation directly when applied alone.

A common variant of PNPLA3 (p.I148M) is not associated with alcoholic chronic pancreatitis

Background Chronic pancreatitis (CP) is an inflammatory disease that in some patients leads to exocrine and endocrine dysfunction. In industrialized countries the most common aetiology is chronic alcohol abuse. Descriptions of associated genetic alterations in alcoholic CP are rare. However, a common PNPLA3 variant (p.I148M) is associated with the development of alcoholic liver cirrhosis (ALC). Since, alcoholic CP and ALC share the same aetiology PNPLA3 variant (p.I148M) possibly influences the development of alcoholic CP. Methods Using melting curve analysis we genotyped the variant in 1510 patients with pancreatitis or liver disease (961 German and Dutch alcoholic CP patients, 414 German patients with idiopathic or hereditary CP, and 135 patients with ALC). In addition, we included in total 2781 healthy controls in the study. Results The previously published overrepresentation of GG-genotype was replicated in our cohort of ALC (p-value <0.0001, OR 2.3, 95% CI 1.6–3.3). Distributions of genotype and allele frequencies of the p.I148M variant were comparable in patients with alcoholic CP, idiopathic and hereditary CP and in healthy controls. Conclusions The absence of an association of PNPLA3 p.I148M with alcoholic CP seems not to point to a common pathway in the development of alcoholic CP and alcoholic liver cirrhosis.

Gain of chromosome 8q is associated with metastases and poor survival of patients with clear cell renal cell carcinoma

The aim of this study was to evaluate the prevalence of chromosome 8q gain in clear cell renal cell carcinoma (CCRCC) and to correlate the findings with tumor phenotype and disease-specific survival (DSS).

A high-resolution comparative radiation hybrid map of equine chromosome 4q12-q22

In this study, we present a comprehensive 5000-rad radiation hybrid map of a 40-cM region on equine chromosome 4 (ECA4) that contains quantitative trait loci for equine osteochondrosis. We mapped 29 gene-associated sequence tagged site markers using primers designed from equine expressed sequence tags or BAC clones in the ECA4q12-q22 region. Three blocks of conserved synteny, showing two chromosomal breakpoints, were identified in the segment of ECA4q12-q22. Markers from other segments of HSA7q mapped to ECA13p and ECA4p, and a region of HSA7p was homologous to ECA13p. Therefore, we have improved the resolution of the human-equine comparative map, which allows the identification of candidate genes underlying traits of interest.

Single linkage group per chromosome genetic linkage map for the horse, based on two three-generation, full-sibling, crossbred horse reference families

A genetic linkage map of the horse consisting of 742 markers, which comprises a single linkage group for each of the autosomes and the X chromosome, is presented. The map has been generated from two three-generation full-sibling reference families, sired by the same stallion, in which there are 61 individuals in the F2 generation. Each linkage group has been assigned to a chromosome and oriented with reference to markers mapped by fluorescence in situ hybridization. The average interval between markers is 3.7 cM and the linkage groups collectively span 2772 cM. The 742 markers comprise 734 microsatellite and 8 gene-based markers. The utility of the microsatellite markers for comparative mapping has been significantly enhanced by comparing their flanking sequences with the human genome sequence; this enabled conserved segments between human and horse to be identified. The new map provides a valuable resource for genetically mapping traits of interest in the horse.

Molecular characterization of the equine collagen, type IX, alpha 2 (COL9A2) gene on horse chromosome 2p16-->p15

The mammalian collagen, type IX, alpha 2 gene (COL9A2) encodes the alpha-2 chain of type IX collagen and is located on horse chromosome 2p16-->p14 harbouring a quantitative trait locus for osteochondrosis. We isolated a bacterial artificial chromosome (BAC) clone containing the equine COL9A2 gene and determined the complete genomic sequence of this gene. Cloning and characterization of equine COL9A2 revealed that the equine gene consists of 32 exons spanning approximately 15 kb. The COL9A2 transcript encodes a single protein of 688 amino acids. Thirty two single nucleotide polymorphisms (SNPs) equally distributed in the gene were detected in a mutation scan of eight unrelated Hanoverian warmblood stallions, including one SNP that affects the amino acid sequence of COL9A2. Comparative analyses between horse, human, mouse and rat indicate that the chromosomal location of equine COL9A2 is in agreement with known chromosomal synteny relationships. The comparison of the gene structure and transcript revealed a high degree of conservation towards the other mammalian COL9A2 genes. We chose three informative SNPs for association and linkage disequilibrium tests in three to five paternal half-sib families of Hanoverian warmblood horses consisting of 44 to 75 genotyped animals. The test statistics did not reach the significance threshold of 5% and so we could not show an association of COL9A2 with equine osteochondrosis.

Age at diagnosis and loss of heterozygosity on chromosome 1p and 19q in oligodendroglial tumors

The age distribution and incidence of loss of heterozygosity (LOH) of 1p and 19q was analyzed in 85 oligodendroglial tumors WHO II and III. The peak of tumor manifestation was in the age group of 35 to 55 years. There was no association between age at diagnosis and LOH incidence. We conclude that the prognostic effect of age on survival is not mediated by LOH 1p/19q.

A degradation-sensitive anionic trypsinogen (PRSS2) variant protects against chronic pancreatitis

Chronic pancreatitis is a common inflammatory disease of the pancreas. Mutations in the genes encoding cationic trypsinogen (PRSS1) and the pancreatic secretory trypsin inhibitor (SPINK1) are associated with chronic pancreatitis. Because increased proteolytic activity owing to mutated PRSS1 enhances the risk for chronic pancreatitis, mutations in the gene encoding anionic trypsinogen (PRSS2) may also predispose to disease. Here we analyzed PRSS2 in individuals with chronic pancreatitis and controls and found, to our surprise, that a variant of codon 191 (G191R) is overrepresented in control subjects: G191R was present in 220/6,459 (3.4%) controls but in only 32/2,466 (1.3%) affected individuals (odds ratio 0.37; P = 1.1 x 10(-8)). Upon activation by enterokinase or trypsin, purified recombinant G191R protein showed a complete loss of trypsin activity owing to the introduction of a new tryptic cleavage site that renders the enzyme hypersensitive to autocatalytic proteolysis. In conclusion, the G191R variant of PRSS2 mitigates intrapancreatic trypsin activity and thereby protects against chronic pancreatitis.