Molecular Characterization of Three Canine Models of Human Rare Bone Diseases: Caffey,van den Ende-Gupta,and Raine Syndromes

One to two percent of all children are born with a developmental disorder requiring pediatric hospital admissions. For many such syndromes, the molecular pathogenesis remains poorly characterized. Parallel developmental disorders in other species could provide complementary models for human rare diseases by uncovering new candidate genes, improving the understanding of the molecular mechanisms and opening possibilities for therapeutic trials. We performed various experiments, e.g. combined genome-wide association and next generation sequencing, to investigate the clinico-pathological features and genetic causes of three developmental syndromes in dogs, including craniomandibular osteopathy (CMO), a previously undescribed skeletal syndrome, and dental hypomineralization, for which we identified pathogenic variants in the canine SLC37A2 (truncating splicing enhancer variant), SCARF2 (truncating 2-bp deletion) and FAM20C (missense variant) genes, respectively. CMO is a clinical equivalent to an infantile cortical hyperostosis (Caffey disease), for which SLC37A2 is a new candidate gene. SLC37A2 is a poorly characterized member of a glucose-phosphate transporter family without previous disease associations. It is expressed in many tissues, including cells of the macrophage lineage, e.g. osteoclasts, and suggests a disease mechanism, in which an impaired glucose homeostasis in osteoclasts compromises their function in the developing bone, leading to hyperostosis. Mutations in SCARF2 and FAM20C have been associated with the human van den Ende-Gupta and Raine syndromes that include numerous features similar to the affected dogs. Given the growing interest in the molecular characterization and treatment of human rare diseases, our study presents three novel physiologically relevant models for further research and therapy approaches, while providing the molecular identity for the canine conditions.     

Mechanical efficiency of locomotion in females during different kinds of muscle action

The mechanical efficiencies (ME) of pure positive and pure negative work as well as of stretch-shortening cycle (SSC) exercise were investigated with a special sledge apparatus. The subjects were 20 young females who performed six different types of submaximal exercise: two of pure concentric exercise (positive work), two of pure eccentric exercise (negative work) and two SSC exercises. The work intensities were determined individually, from the recordings of distance obtained during a single maximal concentric exercise. Each exercise involved 60 muscle actions lasting a total of 3 min per testing condition. The MEs of pure positive work with intensities of 30% and 60% maximum (C30 and C60 respectively) were 15.5%, SD 2.6% and 14.3%, SD 1.9%, respectively. In pure negative work, when the dropping heights were 20 cm (E20) and 80 cm (E80), MEs were 28.4%, SD 6.9% and 47.9%, SD 10.1%, respectively. In SSC-exercise, the MEs during the positive phase of the take-off were 31.3%, SD 6.3% (E20/C90) and 35.0%, SD 7.0% (E80/C69). The total MEs in SSC-exercise were 29.1%, SD 4.0% (E20/C90) and 40.1%, SD 5.2% (E80/C60 x 100). In pure negative work, the increased stretching velocity increased the value of ME. In the concentric phase of SSC-exercise, the integrated electromyographic activity (iEMG) of vastus lateralis (VL) and vastus medialis (VM) muscles were lower (P less than 0.05) than in pure concentric work, when the mechanical work was the same (C60 vs E80/C60). During pure eccentric work, iEMGs were lower in comparison to the eccentric phase of SSC-exercise.(ABSTRACT TRUNCATED AT 250 WORDS)     

The Effect of Exercise on Blood Parameters in Standardbred and Finnish-Bred Horses

Serum enzyme activities, albumin, protein, urea, cholesterol, triglyceride, free fatty acid, glucose and lactate concentrations as well as hematocrit values were measured in standardbred and Finnish-bred horses at rest and after (i) a short controlled exercise and (ii) a trotting competition. There were no breed differences in the enzyme activities at rest and the 2 breeds responded in the same manner to the exercise. Only after the race proper significant increases in the enzyme activities were found. The activities rose more in the standardbred horses than in the Finnish-bred horses. Urea and cholesterol concentrations did not change after either exercise. Protein and albumin concentrations as well as hematocrit values increased significantly after the exercise. At rest hematocrit values were significantly higher in the standardbred horses and the difference persisted throughout the exercise. After the race proper also albumin and protein concentrations were higher in the standard-bred than in the Finnish-bred horses. Free fatty acid and triglyceride concentrations increased significantly during the exercise. Although glucose and lactate concentrations increased in both breeds, the behaviour of these parameters differed. Glucose concentrations remained increased for a longer period and the recovery from the increased lactate level was faster in the standardbred than in the Finnish-bred horses. The observed differences suggest that the standardbred horses have higher anaerobic capacity than the Finnish-bred horses.     

A century of community ecology: How much progress?

Much of the apparent progress in community ecology amounts to little more than re-inventing the wheel, albeit with technical improvements. Many central ideas in the field were stated by A.K. Cajander at the turn of the century. Thereafter, community ecology has moved back and forth between competition-centered and individualistic views of community structure. The chief problems have probably been inappropriate methodology (lack of rigorous formulation and/or critical experimental testing of theories) and a tendency to work only on subcommunities that are delimited by taxonomic criteria. Although these problems are beginning to be remedied, a new one has emerged: ignorance of history and a tendency to re-invent old ideas under new, flashy names. This problem is potentially as dangerous as the old ones and must be tackled by improving our first-hand contact with the classics in our field.     

BVOC Emissions From a Subarctic Ecosystem,as Controlled by Insect Herbivore Pressure and Temperature

Ecosystems - The biogenic volatile organic compounds, BVOCs have a central role in ecosystem–atmosphere interactions. High-latitude ecosystems are facing increasing temperatures and insect...     

Glycosylation, transport, and complex formation of palmitoyl protein thioesterase 1 (PPT1) – distinct characteristics in neurons

Background

About 20 % of nemaline myopathies are thus far related to skeletal muscle alpha-actin. Seven actin mutants located in different parts of the actin molecule and linked to different forms of the disease were selected and expressed as EGFP-tagged constructs in differentiated C2C12 mytoubes. Results were compared with phenotypes in patient skeletal muscle fibres and with previous expression studies in fibroblasts and C2C12 myoblasts/myotubes.

Results

Whereas EGFP wt-actin nicely incorporated into endogenous stress fibres and sarcomeric structures, the mutants showed a range of phenotypes, which generally changed upon differentiation. Many mutants appeared delocalized in myoblasts but integrated into endogenous actin structures after 4–6 days of differentiation, demonstrating a poor correlation between the appearance in myotubes and the severity of the disease. However, for some mutants, integration into stress fibres induced aberrant structures in differentiated cells, like thickening or fragmentation of stress fibres. Other mutants almost failed to integrate but formed huge aggregates in the cytoplasm of myotubes. Those did not co-stain with alpha-actinin, a main component of nemaline bodies found in patient muscle. Interestingly, nuclear aggregates as formed by two of the mutants in myoblasts were found less frequently or not at all in differentiated cells.

Conclusion

Myotubes are a suitable system to study the capacity of a mutant to incorporate into actin structures or to form or induce pathological changes. Some of the phenotypes observed in undifferentiated myoblasts may only be in vitro effects. Other phenotypes, like aberrant stress fibres or rod formation may be more directly correlated with disease phenotypes. Some mutants did not induce any changes in the cellular actin system, indicating the importance of additional studies like functional assays to fully characterize the pathological impact of a mutant.     

Neuronostatin, a novel peptide encoded by somatostatin gene, regulates cardiac contractile function and cardiomyocyte survival

Neuronostatin, a recently discovered peptide encoded by somatostatin gene, is involved in regulation of neuronal function, blood pressure, food intake, and drinking behavior. However, the biological effects of neuronostatin on cardiac myocytes are not known, and the intracellular signaling mechanisms induced by neuronostatin remain unidentified. We analyzed the effect of neuronostatin in isolated perfused rat hearts and in cultured primary cardiomyocytes. Neuronostatin infusion alone had no effect on left ventricular (LV) contractile function or on isoprenaline- or preload-induced increase in cardiac contractility. However, infusion of neuronostatin significantly decreased the positive inotropic response to endothelin-1 (ET-1). This was associated with an increase in phosphorylation of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase (JNK). Treatment of both neonatal and adult cardiomyocytes with neuronostatin resulted in reduced cardiomyocyte viability. Inhibition of JNK further increased the neuronostatin-induced cell death. We conclude that neuronostatin regulates cardiac contractile function and cardiomyocyte survival. Receptors for neuronostatin need to be identified to further characterize the biological functions of the peptide.     

Distinct subcellular localization for constitutive and agonist-modulated palmitoylation of the human delta opioid receptor

Protein palmitoylation is a reversible lipid modification that plays important roles for many proteins involved in signal transduction, but relatively little is known about the regulation of this modification and the cellular location where it occurs. We demonstrate that the human delta opioid receptor is palmitoylated at two distinct cellular locations in human embryonic kidney 293 cells and undergoes dynamic regulation at one of these sites. Although palmitoylation could be readily observed for the mature receptor (Mr 55,000), [3H]palmitate incorporation into the receptor precursor (Mr 45,000) could be detected only following transport blockade with brefeldin A, nocodazole, and monensin, indicating that the modification occurs initially during or shortly after export from the endoplasmic reticulum. Blocking of palmitoylation with 2-bromopalmitate inhibited receptor cell surface expression, indicating that it is needed for efficient intracellular transport. However, cell surface biotinylation experiments showed that receptors can also be palmitoylated once they have reached the plasma membrane. At this location, palmitoylation is regulated in a receptor activation-dependent manner, as was indicated by the opioid agonist-promoted increase in the turnover of receptor-bound palmitate. This agonist-mediated effect did not require receptor-G protein coupling and occurred at the cell surface without the need for internalization or recycling. The activation-dependent modulation of receptor palmitoylation may thus contribute to the regulation of receptor function at the plasma membrane.