Cardiovascular development: towards biomedical applicability: Epicardium-derived cells in cardiogenesis and cardiac regeneration

During cardiogenesis, the epicardium grows from the proepicardial organ to form the outermost layer of the early heart. Part of the epicardium undergoes epithelial-mesenchymal transformation, and migrates into the myocardium. These epicardium- derived cells differentiate into interstitial fibroblasts, coronary smooth muscle cells, and perivascular fibroblasts. Moreover, epicardium-derived cells are important regulators of formation of the compact myocardium, the coronary vasculature, and the Purkinje fiber network, thus being essential for proper cardiac development. The fibrous structures of the heart such as the fibrous heart skeleton and the semilunar and atrioventricular valves also depend on a contribution of these cells during development. We hypothesise that the essential properties of epicardium-derived cells can be recapitulated in adult diseased myocardium. These cells can therefore be considered as a novel source of adult stem cells useful in clinical cardiac regeneration therapy

Immunolocalization of dystrophin and neurofilament protein in muscle spindles of normal, mdx-dystrophic, and denervated mice

Dystrophin is a high molecular weight protein localized under the sarcolemma of normal extrafusal muscle fibers but absent in skeletal muscles of Duchenne muscular dystrophy patients and in the mdx mouse. Muscle spindles in the soleus of 32-week-old normal and age-matched mdx mice were examined by immunocytochemical methods to determine the localization of dystrophin in polar and equatorial regions of the intrafusal fibers. Spindles were serially-sectioned in transverse and longitudinal planes, and they were double-labelled with an antibody to dystrophin and with a 200 kD neurofilament protein antibody which revealed their sensory innervation. By fluorescence microscopy, intrafusal fibers in the soleus of mdx mice were deficient in dystrophin throughout their lengths, whereas their sensory nerve-terminals stained intensely with the nerve-specific antibody and appeared unaltered in dystrophy. In the normal soleus, polar regions of bag and chain fibers exhibited a peripheral rim of sarcolemmal staining equivalent to that seen in the neighboring extrafusal fibers. Dystrophin labelling in equatorial regions of normal intrafusal fibers, however, showed dystrophin-deficient segments alternating in a spiral fashion with positive-staining domains along the sarcolemma. Double-labelling for dystrophin and neurofilament protein showed that these dystrophin-deficient sites were subjacent to the annulospiral sensory-nerve wrappings terminating on the intrafusal fibers. Additionally, it was found that chronic denervation of muscle spindles in normal mice did not affect the expression of dystrophin either at these sites or at the non-sensory regions of the sarcolemma. The results of this study suggest that dystrophin is not an integral part of the subsynaptic sensory-membrane in equatorial regions of normal intrafusal fibers, and, that the neurotrophic effect of sensory innervation is not the principle cause of this unique arrangement of dystrophin in equatorial regions. In dystrophy, intrafusal fibers display the same primary defect in muscular dystrophy as seen in the extrafusal fibers. However, because of their small-diameters, capsular investment, and relatively low tension outputs, dystrophic intrafusal fibers may be less prone to the sarcolemmal membrane disruption that is characteristic of extrafusal fibers in this disorder.Medicine, Faculty ofGraduat

Morphological quantitative and ultracytochemical studies on the internal membrane systems of normal and mdx-dystrophic murine skeletal muscle fibers

Mitochondria and the sarcotubular system in skeletal muscle fibers are specialized intracellular membranes that participate in the generation of energy (ATP) and i n the regulation of contraction and relaxation. This thesis comprises four parts dealing with morphological, histochemical, stereological and biochemical approaches to study these organelles in muscle fibers of adult normal and dystrophic (mdx) mice. The purpose of this research was to test the hypothesis that the internal membrane systems of skeletal muscle fibers are altered in the mdx mouse. Mitochondrial oxidative capacities were assessed in muscle fibers of the extensor digitorum longus (EDL), soleus and diaphragm of normal and mdx mice by histochemical and microphotometric methods in Chapter 1. Relative to controls, significant shifts i n oxidative staining patterns of mdx muscle fibers were observed including a 20% overall lower oxidative capacity in the mdx EDL and soleus and a contrasting 7% higher oxidative capacity in the mdx diaphragm. Transmission electron microscopy (TEM) was combined with stereology in Chapter 2 to determine the ultrastructural arrangement and volume density of the sarcotubular system in muscle fibers in these muscles. Three fiber types (white, intermediate and red) were distinguished in the normal and mdx EDL based on mitochondrial volume in fiber cores and Z line thickness. A mixture of two fiber types, intermediate and red fibers, occupied the soleus and diaphragm. The majority of mdx muscle fibers displayed a similar ultrastructure and disposition of these organelles in comparison to their normal counterparts with nonsignificant differences in sarcotubular volume density between normal and mdx fiber types. Degenerative fibers in mdx muscles exhibited dilated SR, sarcolemmal and myofibril disruption, and mononuclear cell invasion. High-resolution scanning electron microscopy (HRSEM) was utilized in Chapter 3 to reveal the spatial distribution of mitochondria and the sarcotubular system by extracting non-membranous components of the sarcoplasm in freeze- fractured specimens. Finally, in Chapter 4, a cerium-based, ultracytochemical staining method was used for the detection and measurement of the calcium and magnesium-dependent paranitrophenylphosphatase (Ca2 +-Mg2 + pNPPase) activity associated with the sarcoplasmic reticulum (SR) membrane. By TEM, the majority of Ca2+-Mg2+pNPPase activity was localized to the terminal cisternae regions of the SR membrane with moderate phosphatase activity in mid-A band and I band regions of the SR. Comparative spectrophotometry of reaction media revealed a significant 25% higher phosphatase activity in mdx muscle tissue when compared to normal muscle suggesting that Ca2+-sequestering by the SR is enhanced in muscle fibers of the mdx-dystrophic mouse. Overall, these data suggest that mitochondria and SR are altered in the skeletal muscle fibers of mdx mice and point to specific differences in oxidative capacity and Ca2+-sequestering properties of these organelles. [Scientific formulae used in this abstract could not be reproduced.]Medicine, Faculty ofGraduat

Novel use of a micro-computed tomography scanner to trace larvae of wood boring insects

Article first published online: 20 DEC 2010Tracing the tunnels of wood boring larvae in wood or other plant material is generally a difficult and destructive process. Here we present a novel non-destructive method using an in vivo micro-CT scanner to obtain tomograms of small tunnels. The tunnels were formed by larval xiphydriid woodwasps in the branches of the Tasmanian tree Anodopetalum biglandulosum (Cunoniaceae) and show that, as the larvae develop, the tunnels become progressively larger until pupation occurs just below the surface of the bark. The adult then chews its way through the remaining wood and bark before emerging. This non-destructive method is discussed more broadly in regard to its application for tracing the tunnels of wood boring insects, for detecting insects in grain and other plant material for quarantine purposes, and for tracking the tunnelling behaviour and development of live insects in wood. © 2010 The Authors. Journal compilation © 2010 Australian Entomological Society.John T. Jennings and Andrew D. Austi

Administration of epidural labor analgesia is not associated with a decreased risk of postpartum depression in an urban Canadian population of mothers: a secondary analysis of prospective cohort data

Marissa Nahirney,1 Amy Metcalfe,2 Katie H Chaput3 1O’Brien Centre, 2Obstetrics and Gynecology, 3Alberta Children’s Hospital Research Institute and Pediatrics, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada Background: Postpartum depression (PPD) is a common complication of pregnancy, affecting approximately 13% of mothers internationally. Previous research has examined whether epidural analgesia used for pain control during labor and birth is associated with a lower risk of PPD, but reports conflicting results and may have suffered from methodological shortcomings. Our study aimed to prospectively assess whether epidural analgesia is associated with a lower risk of PPD (at either 6 weeks or 6 months postpartum) after attempting to adequately adjust for selection bias and confounding variables. Methods: We conducted a secondary analysis of a prospective cohort of urban Canadian mothers who were recruited at birth in Calgary, Canada, in 2010, for a primary study on predictors of PPD. Mothers with full-term, singleton infants who did not require neonatal intensive care unit admission of >24 hours were included, and filled out questionnaires at birth, 6 weeks and 6 months postpartum including demographics, birth data, maternal and infant physical health, lifestyle, breastfeeding and maternal mental health. Descriptive statistics were calculated for participant characteristics and to identify potential confounder variables. Multivariable logistic regression analysis was conducted to assess whether epidural analgesia is associated with PPD after controlling for available confounding variables. Results: Our study included 206 mothers who had vaginal deliveries and were free of depression at delivery. We found an incidence of PPD of 13.3% (n=27) and no statistically significant association between epidural use and PPD, regardless of adjustment for potential confounding variables (unadjusted odds ratio [OR] 0.86, 95% confidence interval [CI] 0.69–1.22; adjusted OR (for body mass index 1.04, 95% CI 0.40–2.77). Conclusion: We did not observe a significant association between epidural use and PPD. While the CIs are wide, we do not believe that this masks a clinically relevant association, and as such, the risks and benefits of epidural analgesia communicated to women during labor and delivery should not be modified. Keywords: epidural analgesia, labor analgesia, postpartum depression, maternal mental healt

Structure, distribution and innervation of muscle spindles in avian fast and slow skeletal muscle

Muscle spindles in 2 synergistic avian skeletal muscles, the anterior (ALD) and posterior (PLD) latissimus dorsi, were studied by light and electron microscopy to determine whether morphological or quantitative differences existed between these sensory receptors. Differences were found in the density, distribution and location of muscle spindles in the 2 muscles. They also differed with respect to the morphology of their capsules and intracapsular components. The slow ALD possessed muscle spindles which were evenly distributed throughout the muscle, whereas in the fast PLD they were mainly concentrated around the single nerve entry point into the muscle. The muscle spindle index (number of spindles per gram wet muscle weight) in the ALD was more than double that of its fast-twitch PLD counterpart (130.5±2.0 vs 55.4±2.0 respectively, n=6). The number of intrafusal fibres per spindle ranged from 1 to 8 in the ALD and 2 to 9 in the PLD, and their diameters varied from 5.0 to 16.0 μm and 4.5 to 18.5 μm, respectively. Large diameter intrafusal fibres were more frequently encountered in spindles of the PLD. Unique to the ALD was the presence of monofibre muscle spindles (12.7% of total spindles observed in ALD) which contained a solitary intrafusal fibre. In muscle spindles of both the ALD and PLD, sensory nerve endings terminated in a spiral fashion on the intrafusal fibres in their equatorial regions. Motor innervation was restricted to either juxtaequatorial or polar regions of the intrafusal fibres. Outer capsule components were extensive in polar and juxtaequatorial regions of ALD spindles, whereas inner capsule cells of PLD spindles were more numerous in juxtaequatorial and equatorial regions. Overall, muscle spindles of the PLD exhibited greater complexity with respect to the number of intrafusal fibres per spindle, range of intrafusal fibre diameters and development of their inner capsules. It is postulated that the differences in muscle spindle density and structure observed in this study reflect the function of the muscles in which they reside