Numerous stimulatory growth factors that can influence muscle regeneration are known. Recently, it has been demonstrated that neutralization of muscle growth inhibitory factors, such as myostatin (Mstn; also known as growth differentiation factor 8, Gdf8), also leads to increased muscle regeneration in mdx mice that are known to have cycles of degeneration. However, the precise mechanism by which Mstn regulates muscle regeneration has not yet been fully determined. To investigate the role of Mstn in adult skeletal muscle regeneration, wild-type and myostatin-null (Mstn-/-) mice were injured with notexin. Forty-eight hours after injury, accelerated migration and enhanced accretion of myogenic cells (MyoD1+) and macrophages (Mac-1+) was observed at the site of regeneration in Mstn-/- muscle as compared with wild-type muscle. Inflammatory cell numbers decreased more rapidly in the Mstn-/- muscle, indicating that the whole process of inflammatory cell response is accelerated in Mstn-/- mice. Consistent with this result, the addition of recombinant Mstn reduced the activation of satellite cells (SCs) and chemotactic movements of both myoblasts and macrophages ex vivo. Examination of regenerated muscle (28 days after injury) also revealed that Mstn-/- mice showed increased expression of decorin mRNA, reduced fibrosis and improved healing as compared with wild-type mice. On the basis of these results, we propose that Mstn negatively regulates muscle regeneration not only by controlling SC activation but also by regulating the migration of myoblasts and macrophages to the site of injury. Thus, antagonists of Mstn could potentially be useful as pharmacological agents for the treatment of disorders of overt degeneration and regeneration.

During somatic cell nuclear transfer the gene expression profile of the donor cell has to be changed or reprogrammed extensively to reflect that of a normal embryo. In this study we focused on the switching on of embryonic genes by screening with a microarray consisting of 5000 independent cDNA isolates derived from a bovine blastocyst library which we constructed for this purpose. Expression profiling was performed using linearly amplified RNA from individual day 7 nuclear transfer (NT) and genetically half-identical in vitro produced (IVP) blastocysts. We identified 92 genes expressed at lower levels in NT embryos whereas transcripts of 43 genes were more abundant in NT embryos (P % 0.05, R 1.5-fold change). A range of functional categories was represented among the identified genes, with a preponderance of constitutively expressed genes required for the maintenance of basal cellular function. Using a stringent quantitative SYBR-green real time RT-PCR based approach we found, when comparing the means of the expression levels of a larger set of individual embryos, that differences were small (! 2-fold) and only significant for two of the seven analysed genes (KRT18, SLC16A1). Notably, examination of transcript levels of a single gene in individual embryos could not distinguish an NT from a control embryo. This unpredictability of individual gene expression on a global background of multiple gene expression changes argues for a predominantly stochastic nature of reprogramming errors.

The nature of human-animal interactions is an important factor contributing to animal welfare and productivity. Reducing stress during routine husbandry procedures is likely to improve animal welfare. We examined how the type of early handling of calves affected responses to two common husbandry procedures, ear-tagging and disbudding. Forty Holstein-Friesian calves (n 5 20/treatment) were exposed to one of two handling treatments daily from 1 to 5 weeks of age: (1) positive (n 5 20), involving gentle handling (soft voices, slow movements, patting), and (2) negative (n 5 20), involving rough handling (rough voices, rapid movements, pushing). Heart rate (HR), respiration rate (RR) and behaviour (activity, tail flicking) were measured before and after ear-tagging and disbudding (2 days apart). Cortisol was measured at 220 (baseline), 20 and 40 min relative to disbudding time. There were no significant treatment differences in HR, RR or behaviour in response to either procedure. However, the following changes occurred across both treatment groups. HR increased after disbudding (by 14.7 6 4.0 and 18.6 6 3.8 bpm, positive and negative, respectively; mean 6 s.e.m.) and ear-tagging (by 8.7 6 3.1 and 10.3 6 3.0 bpm, positive and negative, respectively). After disbudding, there was an increase in RR (by 8.2 6 3.4 and 9.3 6 3.4 breaths/min, positive and negative, respectively), overall activity (by 9.4 6 1.2 and 9.9 6 1.3 frequency/min, positive and negative, respectively) and tail flicking (by 13.2 6 2.8 and 11.2 6 3.0 frequency/min, positive and negative, respectively), and cortisol increased from baseline at 20 min post procedure (by 10.3 6 1.1 and 12.3 6 1.1 nmol/l positive and negative, respectively). Although we recorded significant changes in calf responses during ear-tagging and disbudding, the type of prior handling had no effect on responses. The effects of handling may have been overridden by the degree of pain and/or stress associated with the procedures. Further research is warranted to understand the welfare impact and interaction between previous handling and responses to husbandry procedures.

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