Environmental stress influences mitochondrial metabolism in vascular cells: consequences for angiogenesis

While the important and varied roles that vascular cells play in both health and disease is well recognised, the focus on potential therapeutic targets continually shifts as new players emerge. Here, we outline how mitochondria may be viewed as more than simply energygenerating organelles, but instead as important sentinels of metabolic health and effectors of appropriate responses to physiological challenges

Electrical excitation of the heart in a basal vertebrate, the european river lamprey (Lampetra fluviatilis)

Hagfishes and lampreys (order Cyclostomata) are living representatives of an ancient group of jawless vertebrates (class Agnatha). Studies on cyclostome hearts may provide insights into the evolution of the vertebrate heart and thereby increase our understanding of cardiac function in higher vertebrates, including mammals. To this end, electrical ex-citability of the heart in a basal vertebrate, the European river lamprey (Lampetra fluviatilis), was examined. Ion currents of cardiac myocytes, action potentials (APs) of atrial and ven-tricular muscle, and electrocardiogram (in vivo) were mea-sured using the patch-clamp method, intracellular micro-electrodes, and trailing wires, respectively. The characteristic features of fairly high heart rate (28.4 5 3 beats min−1) and short AP duration (550 ± 44 and 122.1 ± 28.5 for ventricle and atrium, respectively) at low ambient temperature (5°C) are shared with cold-active teleost fishes. However, the ion current basis of the ventricular AP differs from that of other fishes. For inward currents, sodium current density (INa) is lower and calcium current density (ICa) higher than in teleost ventricles, while the kinetics of INa is slow and that of ICa is fast in comparison. Among the ventricular repolarizing currents, the delayed rectifier K+ current is smaller than in myocytes of several teleost species. Unlike mammalian hearts, ATP-sensitive K+ channels are constitutively open under normoxic conditions, thus contributing to negative resting membrane potential and repolarization of APs. Upstroke velocity of AP (5.4± and 6.3±0.6 V s−1 for ventricular and atrial myocytes, respectively) is slower than in teleost hearts. Ex-citability of the lamprey heart seems to possess both primitive and advanced characteristics. Short APs are appropriate to support brief and vigorous contractions (in common with. higher vertebrates), while relatively low AP upstroke velocities enable only relatively slow propagation of contraction over the heart

Integrated method for quantitative morphometry and oxygen transport modelling in striated muscle

Identifying structural limitations in O2 transport is primarily restricted by current methods employed to characterise the nature of physiological remodelling. Inadequate resolution or breadth of available data has impaired development of routine diagnostic protocols and effective therapeutic strategies. Understanding O2 transport within striated muscle faces major challenges, most notably in quantifying how well individual fibres are supplied by the microcirculation, which has necessitated exploring tissue O2 supply using theoretical modelling of diffusive exchange. Having identified capillary domains as a suitable model for the description of local O2 supply, and requiring less computation than numerically calculating the trapping regions that are supplied by each capillary via biophysical transport models, we sought to design a high throughput method for histological analysis. We present an integrated package that identifies optimal protocols for identification of important input elements, processing of digitised images with semi-automated routines, and incorporation of these data into a mathematical modelling framework with computed output visualised as the tissue partial pressure of O2 (PO2) distribution across a biopsy sample. Worked examples are provided using muscle samples from experiments involving rats and humans

Effects of prolonged anoxia on electrical activity of the heart in Crucian carp (Carassius carassius)

The effects of sustained anoxia on cardiac electrical excitability were examined in the anoxia-tolerant Crucian carp (Carassius carassius). The electrocardiogram (ECG) and expression of excitation-contraction coupling genes were studied in fish acclimatised to normoxia in summer (+18°C) or winter (+2°C), and in winter fish after 1, 3 and 6 weeks of anoxia. Anoxia induced a sustained bradycardia from a heart rate of 10.3±0.77 to 4.1±0.29 bpm (P<0.05) after 5 weeks, and heart rate slowly recovered to control levels when oxygen was restored. Heart rate variability greatly increased under anoxia, and completely recovered under re-oxygenation. The RT interval increased from 2.8±0.34 s in normoxia to 5.8±0.44 s under anoxia (P<0.05), which reflects a doubling of the ventricular action potential (AP) duration. Acclimatisation to winter induced extensive changes in gene expression relative to summer-acclimatised fish, including depression in those coding for the sarcoplasmic reticulum calcium pump (Serca2-q2) and ATP-sensitive K(+) channels (Kir6.2) (P<0.05). Genes of delayed rectifier K(+) (kcnh6) and Ca(2+) channels (cacna1c) were up-regulated in winter fish (P<0.05). In contrast, the additional challenge of anoxia caused only minor changes in gene expression, e.g. depressed expression of Kir2.2b K(+) channel gene (kcnj12b), whereas expression of Ca(2+) (cacna1a, -c and -g) and Na(+) channel genes (scn4a and scn5a) were not affected. These data suggest that low temperature pre-conditions the Crucian carp heart for winter anoxia, whereas sustained anoxic bradycardia and prolongation of AP duration are directly induced by oxygen shortage without major changes in gene expression

Hyperglycaemia up-regulates placental growth factor (PlGF) expression and secretion in endothelial cells via suppression of PI3 kinase-Akt signalling and activation of FOXO1

Placenta growth factor (PlGF) is a pro-inflammatory angiogenic mediator that promotes many pathologies including diabetic complications and atherosclerosis. Widespread endothelial dysfunction precedes the onset of these conditions. As very little is known of the mechanism(s) controlling PlGF expression in pathology we investigated the role of hyperglycaemia in the regulation of PlGF production in endothelial cells. Hyperglycaemia stimulated PlGF secretion in cultured primary endothelial cells, which was suppressed by IGF-1-mediated PI3K/Akt activation. Inhibition of PI3K activity resulted in significant PlGF mRNA up-regulation and protein secretion. Similarly, loss or inhibition of Akt activity significantly increased basal PlGF expression and prevented any further PlGF secretion in hyperglycaemia. Conversely, constitutive Akt activation blocked PlGF secretion irrespective of upstream PI3K activity demonstrating that Akt is a central regulator of PlGF expression. Knock-down of the Forkhead box O-1 (FOXO1) transcription factor, which is negatively regulated by Akt, suppressed both basal and hyperglycaemia-induced PlGF secretion, whilst FOXO1 gain-of-function up-regulated PlGF in vitro and in vivo. FOXO1 association to a FOXO binding sequence identified in the PlGF promoter also increased in hyperglycaemia. This study identifies the PI3K/Akt/FOXO1 signalling axis as a key regulator of PlGF expression and unifying pathway by which PlGF may contribute to common disorders characterised by endothelial dysfunction, providing a target for therapy

Adrenergic and adenosinergic regulation of the cardiovascular system in an Antarctic icefish: Insight into central and peripheral determinants of cardiac output.

Icefishes characteristically lack the oxygen-binding protein haemoglobin and therefore are especially reliant on cardiovascular regulation to augment oxygen transport when oxygen demand increases, such as during activity and warming. Using both in vivo and in vitro experiments, we evaluated the roles for adrenaline and adenosine, two well-established cardio- and vasoactive molecules, in regulating the cardiovascular system of the blackfin icefish, Chaenocephalus aceratus. Despite increasing cardiac contractility (increasing twitch force and contraction kinetics in isometric myocardial strip preparations) and accelerating heart rate (ƒH), adrenaline (5 nmol kg-1 bolus intra-arterial injection) did not significantly increase cardiac output (Q̇) in vivo because it elicited a large decrease in vascular conductance (Gsys). In contrast, and despite preliminary data suggesting a direct negative inotropic effect of adenosine on isolated atria and little effect on isolated ventricle strips, adenosine (500 nmol kg-1) generated a large increase in Q̇ by increasing Gsys, a change reminiscent of that previously reported during both acute warming and invoked activity. Our data thus illustrate how Q̇ in C. aceratus may be much more dependent on peripheral control of vasomotor tone than direct regulation of the heart

Modelling oxygen capillary supply to striated muscle tissues

The ability to characterise functional capillary supply (FCS) plays a key role in developing effective therapeutic interventions for numerous pathological conditions, such as chronic ischaemia in skeletal or cardiac muscle. Detailed tissue geometry, such as muscle fibre size, has been incorporated into indices of FCS by considering the distribution of Voronoi tessellations (‘capillary domains’) generated from vessel locations in a plane perpendicular to muscle fibre orientation, implicitly assuming that each Voronoi polygon represents the area of supply of its enclosed capillary. However, to assess the capacity of FCS in muscle, we are naturally led to use a modelling framework that can account for the local anatomic and metabolic heterogeneities of muscle fibres. Such a framework can be used to explore the validity of the Voronoi polygon representation of FCS regions while also providing a general platform for robust predictions of FCS

Oxygen transport kinetics underpin rapid and robust diaphragm recovery following chronic spinal cord injury

Months after spinal cord injury (SCI), respiratory deficits remain the primary cause of morbidity and mortality for patients. It is possible to induce partial respiratory motor functional recovery in chronic SCI following 2 weeks of spinal neuroplasticity. However, the peripheral mechanisms underpinning this recovery are largely unknown, limiting development of new clinical treatments with potential for complete functional restoration. Utilizing a rat hemisection model, diaphragm function and paralysis was assessed and recovered at chronic time points following trauma through chondroitinase ABC induced neuroplasticity. We simulated the diaphragm's in vivo cyclical length change and activity patterns using the work loop technique at the same time as assessing global and local measures of the muscles histology to quantify changes in muscle phenotype, microvascular composition, and oxidative capacity following injury and recovery. These data were fed into a physiologically informed model of tissue oxygen transport. We demonstrate that hemidiaphragm paralysis causes muscle fibre hypertrophy, maintaining global oxygen supply, although it alters isolated muscle kinetics, limiting respiratory function. Treatment induced recovery of respiratory activity normalized these effects, increasing oxygen supply, restoring optimal diaphragm functional properties. However, metabolic demands of the diaphragm were significantly reduced following both injury and recovery, potentially limiting restoration of normal muscle performance. The mechanism of rapid respiratory muscle recovery following spinal trauma occurs through oxygen transport, metabolic demand and functional dynamics of striated muscle. Overall, these data support a systems‐wide approach to the treatment of SCI, and identify new targets to mediate complete respiratory recovery

Advances and challenges in skeletal muscle angiogenesis.

The role of capillaries is to serve as the interface for delivery of oxygen and removal of metabolites to/from tissues. During the past decade there has been a proliferation of studies that have advanced our understanding of angiogenesis demonstrating tissue capillary supply is under strict control during health, but poorly controlled in disease - resulting in either excessive capillary growth (pathological angiogenesis) or losses in capillarity (rarefaction). Given that skeletal muscle comprises nearly 40% of body mass in humans, skeletal muscle capillary density has a significant impact on metabolism, endocrine function, and locomotion, and is tightly regulated at many different levels. Skeletal muscle is also high adaptable, and thus one of the few organ systems which can be experimentally manipulated (e.g. by exercise) to study physiologic regulation of angiogenesis. This review will focus on 1) the methodological concerns that have arisen in determining skeletal muscle capillarity, and 2) highlight the concepts that are reshaping our understanding of the angio-adaptation process. We also summarize selected new findings (physical influences, molecular changes and ultrastructural rearrangement of capillaries) that identify areas of future research with the greatest potential to expand our understanding of how angiogenesis is normally regulated, and that may also help to better understand conditions of uncontrolled (pathologic) angiogenesis

Firearms-related skeletal muscle trauma: pathophysiology and novel approaches for regeneration

One major cause of traumatic injury is firearm-related wounds (i.e., ballistic trauma), common in both civilian and military populations, which is increasing in prevalence and has serious long-term health and socioeconomic consequences worldwide. Common primary injuries of ballistic trauma include soft-tissue damage and loss, haemorrhage, bone fracture, and pain. The majority of injuries are of musculoskeletal origin and located in the extremities, such that skeletal muscle offers a major therapeutic target to aid recovery and return to normal daily activities. However, the underlying pathophysiology of skeletal muscle ballistic trauma remains poorly understood, with limited evidence-based treatment options. As such, this review will address the topic of firearm-related skeletal muscle injury and regeneration. We first introduce trauma ballistics and the immediate injury of skeletal muscle, followed by detailed coverage of the underlying biological mechanisms involved in regulating skeletal muscle dysfunction following injury, with a specific focus on the processes of muscle regeneration, muscle wasting and vascular impairments. Finally, we evaluate novel approaches for minimising muscle damage and enhancing muscle regeneration after ballistic trauma, which may have important relevance for primary care in victims of violence