Connective tissue changes in surgically overloaded muscle

Summary The effects of overload on the connective tissue component of the soleus muscle of the rat have been investigated. Three weeks after tenotomy of its synergistic muscles the soleus underwent considerable increase in weight. This was shown to have resulted from an increase in size of the predominant fibre type. Whilst occasional groups of fibres appeared to have resulted from the splitting of large single fibres, there was no significant increase in the number of fibres in cross-section of the muscle belly. The connective tissue content of the overloaded muscles was investigated using both histological and biochemical techniques. It was found that muscle fibre hypertrophy was accompanied by an increase in the connective tissue component. Furthermore, there was an increase in the proportion of collagen to muscle fibre tissue.The author wish to acknowledge the expert technical assistance given by Mr P. Prentis. This investigation was supported by a grant from the Medical Research Council.     

Transformation of a novel direct-repeat repressor element into a promoter and enhancer by multimerisation.

Studies on the regulation of interferon (IFN) responsive genes have mainly been centred on the highly conserved IFN stimulated responsive elements (ISREs) which can mediate type I and II IFN inducibility. To date little is known about other functional cis-acting regulatory motifs in IFN responsive genes. We report here on the identification of a repressor element in the human MxA gene defined to a 19 base pair (bp) region which houses a 9 bp direct repeat. DNA-specific protein binding on this element is not affected by IFN treatment and is distinct from ISRE binding proteins. Remarkably, contrary to expectations, when the repressor element is multimerised and spliced, in either orientation, to a reporter gene it behaves like a functional, constitutive promoter. Positioning the multimerised element in front of the SV40 enhancerless promoter also led to enhanced expression. The same protein(s) seem to bind to both the single repressor element and its multimerised form. This discovery of phenotypic reversal on a repressor element via multimerisation may have important implications in vivo.     

Preliminary observations on the heavy metal content of four species of freshwater fish in NW England

The concentrations of zinc, lead and cadmium in four species of fish from several lacustrine sites were determined. Cyprinids were found to have significantly higher levels of the three metals studied than perch, Perca fluviatilis . Whilst zinc thresholds may be species specific, levels of lead and cadmium vary both within species and sites. There is some evidence that lead and cadmium levels increase with age in roach, Rutilus rutilus . Additionally both cadmium and lead appear to accumulate in the heart, liver, kidney and brain as well as in the bone. Relatively high levels of lead and cadmium were recorded in the eyes of bream, Abrama bramis .     

The effects of calcium on protein turnover in skeletal muscles of the rat.

Several experimental procedures were used to increase the intracellular concentration of Ca2+ and determine its effects on protein turnover in isolated extensor digitorum longus and soleus muscle. These methods included the use of ionophore A23187, caffeine, dibucaine, thymol and procaine, all agents known to induce the release of calcium by acting either on the sarcolemma and/or on the sarcoplasmic reticulum. Another approach involved varying the external concentration of Ca2+ in the media in which the muscles were incubated. The changes in muscle Ca2+ concentrations after exposure to the various calcium-releasing agents were in keeping with accepted modes of action of these agents on muscle membranes. The findings suggest that increasing the sarcoplasmic concentration of Ca2+ inhibits protein synthesis and enhances protein breakdown. These catabolic effects of Ca2+ are compared with the changes induced in muscle protein turnover after exposure to insulin or cyclic nucleotides, and in myopathic muscle and situations of work overload. Attention is also drawn to some of the difficulties involved in definitively implicating Ca2+ as a factor involved in the normal regulation of protein turnover.     

The influence of body size on measurements of overall cardiac function

The purpose of this study was to determine the best scaling method to account for the effects of body size on measurements of overall cardiac function and subsequently the interpretation of data based on cardiac power output (CPO). CPO was measured at rest (CPO(rest)) and at maximal exercise (CPO(max)) on 88 and 103 healthy but untrained men and women, respectively, over the age range of 20-70 yr. Cardiac reserve (CR) was calculated as CPO(max) - CPO(rest). CPO(rest), CPO(max), and CR were all significantly related to body mass (BM), body surface area (BSA), and lean body mass (LBM). The linear regression model failed to completely normalize these measurements. In contrast, the allometric model produced size-independent values of CPO. Furthermore, all the assumptions associated with the allometric model were achieved. For CPO(rest), mean body size exponents were BM(0.33), BSA(0.60), and LBM(0.47). For CPO(max), the exponents were BM(0.41), BSA(0.81), and LBM(0.71). For CR, mean body size exponents were BM(0.44), BSA(0.87), and LBM(0.79). LBM was identified (from the root-mean-squares errors of the separate regression models) as the best physiological variable (based on its high metabolic activity) to be scaled in the allometric model. Scaling of CPO to LBM(b) (where b is the scaling exponent) dramatically reduced the between-gender differences with only a 7% difference in CPO(rest) and CPO(max) values. In addition, the gender difference in CR was completely removed. To avoid erroneous interpretations and conclusions being made when comparing data between men and women of different ages, the allometric scaling of CPO to LBM(b) would seem crucial.     

Mechanical signals and IGF-I gene splicing in vitro in relation to development of skeletal muscle

It has been shown that the insulin-like growth factor (IGF-I) gene is spliced in response to mechanical signals producing forms of IGF-I which have different actions. In order to study how mechanical signals influence this gene splicing in developing muscle, C2C12 cells were grown in three-dimensional (3D) culture and subjected to different regimens of mechanical strain. IGF-IEa which initiates the fusion of myoblasts to form myotubes was found to be constitutively expressed in myoblasts and myotubes (held under endogenous tension) and its expression upregulated by a single ramp stretch of 1-h duration but reduced by repeated cyclical stretch. In contrast, mechano growth factor (MGF), which is involved in the proliferation of mononucleated myoblasts that are required for secondary myotube formation and to establish the muscle satellite (stem) cell pool, showed no significant constitutive expression in static cultures, but was upregulated by a single ramp stretch and by cycling loading. The latter types of force simulate those generated in myoblasts by the first contractions of myotubes. These data indicate the importance of seeking to understand the physiological signals that determine the ratios of splice variants of some growth factor/tissue factor genes in the early stages of development of skeletal muscle.     

The effects of denervation on protein turnover of the soleus and extensor digitorum longus muscles of adult mice.

1. Changes in protein turnover of the soleus and EDL muscles of adult mice have been studied 1, 7 and 80 days after denervation. 2. Increased rates of protein degradation 7 and 80 days post-denervation correlated with the atrophy and loss of protein from these muscles. 3. Rates of protein synthesis in the EDL decreased 24 hr after nerve section. However, these synthetic rates increased again to become higher in the 7 day denervated muscles compared with their controls. These latter anabolic changes are inconsistent with the concept of a denervated muscle being inactive. 4. These findings have been compared with a similar study on muscles of growing rats. Any passive stretching of the denervated muscles by continued bone growth appears unlikely to be a crucial factor explaining the increased rates of protein synthesis 7 days after denervation.     

Distributional patterns and community structure of Caribbean coral reef fishes within a river-impacted bay

This study examined how riverine inputs, in particular sediment, influenced the community structure and trophic composition of reef fishes within Rio Bueno, north Jamaica. Due to river discharge a distinct gradient of riverine inputs existed across the study sites. Results suggested that riverine inputs (or a factor associated with them) had a structuring effect on fish community structure. Whilst fish communities at all sites were dominated by small individuals (<20 cm), diversity and total biomass were reduced with increased proximity to the river mouth. The abundance of all fishes, but particularly small-bodied, juvenile and herbivorous fishes was reduced in turbid water when compared to clear-water sites. Results strongly suggest that fluvial sediment inputs may play an important role in structuring fish assemblages even under intense fishing pressure.     

Routes of Hendra Virus Excretion in Naturally-Infected Flying-Foxes: Implications for Viral Transmission and Spillover Risk

Pteropid bats or flying-foxes (Chiroptera: Pteropodidae) are the natural host of Hendra virus (HeV) which sporadically causes fatal disease in horses and humans in eastern Australia. While there is strong evidence that urine is an important infectious medium that likely drives bat to bat transmission and bat to horse transmission, there is uncertainty about the relative importance of alternative routes of excretion such as nasal and oral secretions, and faeces. Identifying the potential routes of HeV excretion in flying-foxes is important to effectively mitigate equine exposure risk at the bat-horse interface, and in determining transmission rates in host-pathogen models. The aim of this study was to identify the major routes of HeV excretion in naturally infected flying-foxes, and secondarily, to identify between-species variation in excretion prevalence. A total of 2840 flying-foxes from three of the four Australian mainland species (Pteropus alecto, P. poliocephalus and P. scapulatus) were captured and sampled at multiple roost locations in the eastern states of Queensland and New South Wales between 2012 and 2014. A range of biological samples (urine and serum, and urogenital, nasal, oral and rectal swabs) were collected from anaesthetized bats, and tested for HeV RNA using a qRT-PCR assay targeting the M gene. Forty-two P. alecto (n = 1410) had HeV RNA detected in at least one sample, and yielded a total of 78 positive samples, at an overall detection rate of 1.76% across all samples tested in this species (78/4436). The rate of detection, and the amount of viral RNA, was highest in urine samples (>serum, packed haemocytes >faecal >nasal >oral), identifying urine as the most plausible source of infection for flying-foxes and for horses. Detection in a urine sample was more efficient than detection in urogenital swabs, identifying the former as the preferred diagnostic sample. The detection of HeV RNA in serum is consistent with haematogenous spread, and with hypothesised latency and recrudesence in flying-foxes. There were no detections in P. poliocephalus (n = 1168 animals; n = 2958 samples) or P. scapulatus (n = 262 animals; n = 985 samples), suggesting (consistent with other recent studies) that these species are epidemiologically less important than P. alecto in HeV infection dynamics. The study is unprecedented in terms of the individual animal approach, the large sample size, and the use of a molecular assay to directly determine infection status. These features provide a high level of confidence in the veracity of our findings, and a sound basis from which to more precisely target equine risk mitigation strategies.     

The E-domain region of mechano-growth factor inhibits cellular apoptosis and preserves cardiac function during myocardial infarction

Insulin-like growth factor-1 (IGF-1) isoforms are expressed via alternative splicing. Expression of the minor isoform IGF-1Eb [also known as mechano-growth factor (MGF)] is responsive to cell stress. Since IGF-1 isoforms differ in their E-domain regions, we are interested in determining the biological function of the MGF E-domain. To do so, a synthetic peptide analog was used to gain mechanistic insight into the actions of the E-domain. Treatment of H9c2 cells indicated a rapid cellular uptake mechanism that did not involve IGF-1 receptor activation but resulted in a nuclear localization. Peptide treatment inhibited the intrinsic apoptotic pathway in H9c2 cells subjected to cell stress with sorbitol by preventing the collapse of the mitochondrial membrane potential and inhibition of caspase-3 activation. Therefore, we administered the peptide at the time of myocardial infarction (MI) in mice. At 2 weeks post-MI cardiac function, gene expression and cell death were assayed. A significant decline in both systolic and diastolic function was evident in untreated mice based on PV loop analysis. Delivery of the E-peptide ameliorated the decline in function and resulted in significant preservation of cardiac contractility. Associated with these changes were an inhibition of pathologic hypertrophy and significantly fewer apoptotic nuclei in the viable myocardium of E-peptide-treated mice post-MI. We conclude that administration of the MGF E-domain peptide may provide a means of modulating local tissue IGF-1 autocrine/paracrine actions to preserve cardiac function, prevent cell death, and pathologic remodeling in the heart.