Clinical impact of real-time evaluation of the biological activity and degradation of hepatocyte growth factor

Hepatocyte growth factor (HGF) is essential for injury repair. Despite high HGF levels in chronic ulcers, up-regulation of HGF receptor in ulcer tissue and decreased biological activity of HGF in ulcer secretions have been observed. With a surface plasmon resonance-based method, we assessed the binding of HGF to antibodies, receptors, and the basement membrane and identified binding interactions that are indispensable for the biological activity of HGF. Recombinant HGF (rHGF) lots were tested for activity, structural integrity, and degradation, and the results were verified in an in vitro model of cell injury. Biologically active rHGF, as well as plasma from healthy volunteers, bound to heparan sulphate proteoglycan (HSPG) and to anti-HGF antibodies. Decreased binding to HSPG was the first event in rHGF degradation. This study established the feasibility of identifying patients with chronic inflammation who need exogenous HGF and of using ligand-binding assessment to evaluate rHGF lots for biological activity

An Update on MyoD Evolution in Teleosts and a Proposed Consensus Nomenclature to Accommodate the Tetraploidization of Different Vertebrate Genomes

DJM was supported by a Natural Environment Research Council studentship (NERC/S/A/2004/12435).Background: MyoD is a muscle specific transcription factor that is essential for vertebrate myogenesis. In several teleost species, including representatives of the Salmonidae and Acanthopterygii, but not zebrafish, two or more MyoD paralogues are conserved that are thought to have arisen from distinct, possibly lineage-specific duplication events. Additionally, two MyoD paralogues have been characterised in the allotetraploid frog, Xenopus laevis. This has lead to a confusing nomenclature since MyoD paralogues have been named outside of an appropriate phylogenetic framework. Methods and Principal Findings: Here we initially show that directly depicting the evolutionary relationships of teleost MyoD orthologues and paralogues is hindered by the asymmetric evolutionary rate of Acanthopterygian MyoD2 relative to other MyoD proteins. Thus our aim was to confidently position the event from which teleost paralogues arose in different lineages by a comparative investigation of genes neighbouring myod across the vertebrates. To this end, we show that genes on the single myod-containing chromosome of mammals and birds are retained in both zebrafish and Acanthopterygian teleosts in a striking pattern of double conserved synteny. Further, phylogenetic reconstruction of these neighbouring genes using Bayesian and maximum likelihood methods supported a common origin for teleost paralogues following the split of the Actinopterygii and Sarcopterygii. Conclusion: Our results strongly suggest that myod was duplicated during the basal teleost whole genome duplication event, but was subsequently lost in the Ostariophysi ( zebrafish) and Protacanthopterygii lineages. We propose a sensible consensus nomenclature for vertebrate myod genes that accommodates polyploidization events in teleost and tetrapod lineages and is justified from a phylogenetic perspective.Publisher PDFPeer reviewe

ccdc80-l1 Is Involved in Axon Pathfinding of Zebrafish Motoneurons

Axon pathfinding is a subfield of neural development by which neurons send out axons to reach the correct targets. In particular, motoneurons extend their axons toward skeletal muscles, leading to spontaneous motor activity. In this study, we identified the zebrafish Ccdc80 and Ccdc80-like1 (Ccdc80-l1) proteins in silico on the basis of their high aminoacidic sequence identity with the human CCDC80 (Coiled-Coil Domain Containing 80). We focused on ccdc80-l1 gene that is expressed in nervous and non-nervous tissues, in particular in territories correlated with axonal migration, such as adaxial cells and muscle pioneers. Loss of ccdc80-l1 in zebrafish embryos induced motility issues, although somitogenesis and myogenesis were not impaired. Our results strongly suggest that ccdc80-l1 is involved in axon guidance of primary and secondary motoneurons populations, but not in their proper formation. ccdc80-l1 has a differential role as regards the development of ventral and dorsal motoneurons, and this is consistent with the asymmetric distribution of the transcript. The axonal migration defects observed in ccdc80-l1 loss-of-function embryos are similar to the phenotype of several mutants with altered Hedgehog activity. Indeed, we reported that ccdc80-l1 expression is positively regulated by the Hedgehog pathway in adaxial cells and muscle pioneers. These findings strongly indicate ccdc80-l1 as a down-stream effector of the Hedgehog pathway

Development temperature has persistent effects on muscle growth responses in gilthead sea bream

Initially we characterised growth responses to altered nutritional input at the transcriptional and tissue levels in the fast skeletal muscle of juvenile gilthead sea bream. Fish reared at 21–22°C (range) were fed a commercial diet at 3% body mass d−1 (non-satiation feeding, NSF) for 4 weeks, fasted for 4d (F) and then fed to satiation (SF) for 21d. 13 out of 34 genes investigated showed consistent patterns of regulation between nutritional states. Fasting was associated with a 20-fold increase in MAFbx, and a 5-fold increase in Six1 and WASp expression, which returned to NSF levels within 16h of SF. Refeeding to satiation was associated with a rapid (<24 h) 12 to 17-fold increase in UNC45, Hsp70 and Hsp90α transcripts coding for molecular chaperones associated with unfolded protein response pathways. The growth factors FGF6 and IGF1 increased 6.0 and 4.5-fold within 16 h and 24 h of refeeding respectively. The average growth in diameter of fast muscle fibres was checked with fasting and significant fibre hypertrophy was only observed after 13d and 21d SF. To investigate developmental plasticity in growth responses we used the same experimental protocol with fish reared at either 17.5–18.5°C (range) (LT) or 21–22°C (range) (HT) to metamorphosis and then transferred to 21–22°C. There were persistent effects of development temperature on muscle growth patterns with 20% more fibres of lower average diameter in LT than HT group of similar body size. Altering the nutritional input to the muscle to stimulate growth revealed cryptic changes in the expression of UNC45 and Hsp90α with higher transcript abundance in the LT than HT groups, whereas there were no differences in the expression of MAFbx and Six1. It was concluded that myogenesis and gene expression patterns during growth are not fixed, but can be modified by temperature during the early stages of the life cycle.Publisher PDFPeer reviewe