Preclinical Research in McArdle Disease: A Review of Research Models and Therapeutic Strategies

McArdle disease; Glycogen phosphorylase; Research modelsEnfermedad de McArdle; Glucógeno fosforilasa; Modelos de investigaciónMalaltia de McArdle; Glicogen fosforilasa; Models de recercaMcArdle disease is an autosomal recessive disorder of muscle glycogen metabolism caused by pathogenic mutations in the PYGM gene, which encodes the skeletal muscle-specific isoform of glycogen phosphorylase. Clinical symptoms are mainly characterized by transient acute “crises” of early fatigue, myalgia and contractures, which can be accompanied by rhabdomyolysis. Owing to the difficulty of performing mechanistic studies in patients that often rely on invasive techniques, preclinical models have been used for decades, thereby contributing to gain insight into the pathophysiology and pathobiology of human diseases. In the present work, we describe the existing in vitro and in vivo preclinical models for McArdle disease and review the insights these models have provided. In addition, despite presenting some differences with the typical patient’s phenotype, these models allow for a deep study of the different features of the disease while representing a necessary preclinical step to assess the efficacy and safety of possible treatments before they are tested in patients.The present manuscript was funded by grants received from the Fondo de Investigaciones Sanitarias (FIS, grant PI19/01313 and PI17/2052) and co-funded by “Fondos FEDER”

Chapter Managing Heat Transfer Issues in Thermoelectric Microgenerators

This chapter deals with heat transfer challenges in the microdomain. It focuses on practical issues regarding this matter when attempting the fabrication of small footprint thermoelectric generators (μTEGs). Thermoelectric devices are designed to bridge a heat source (e.g. hot surface) and a heat sink (e.g. ambient) assuring that a significant fraction of the available temperature difference is captured across the active thermoelectric materials. Coexistence of those contrasted temperatures in small devices is challenging. It requires careful decisions about the geometry and the intrinsic thermal properties of the materials involved. The geometrical challenges lead to micromachined architectures, which silicon technologies provide in a controlled way, but leading to fragile structures, too. In addition, extracting heat from small systems is problematic because of the high thermal resistance associated to heat exchanged by natural convection between the surrounding air and small bare surfaces. Forced convection or the application of a cold finger clearly shows the usefulness of assembling a heat exchanger in a way that is effective and compliant with the mechanical constraints of micromachined devices. Simulations and characterization of fabricated structures illustrate the effectiveness of this element integration and its impact on the trade-off between electrical and thermal behavior of the active materials in device performance

Growth analysis and blood profile in piglets born by embryo transfer

Assisted reproductive technologies (ART), besides solving several reproductive problems, it has also been used as a tool to improve the animal productivity that is required for feeding the human population. One of these techniques, the embryo transfer (ET), has presented limitations in the porcine species, which could constrain its use in the porcine industry. To clarify the potential of this technique, we aimed to compare the impact of using ET or artificial insemination (AI) on the phenotype of the offspring during its first days of age, in terms of growth and blood parameters. At birth, the body weight was higher for ET-females than AI-females, but this difference was no longer observed at day 15. On day 3, it was observed a higher concentration of red blood cells, haemoglobin, and haematocrit in females-ET and a higher concentration of white blood cells in both ET-derived piglets (males and females) when compared to AI groups. On day 3, the biochemical analysis showed a higher level of albumin for ET-derived males, and a lower level of bilirubin for ET-females than AI controls. However, all values were within the normal ranges. Our results indicate that piglets derived from ET seem to be phenotypically similar to those born by AI, which provides preliminary evidence that the ET procedure is a safe technique, but additional studies beyond 15 days of life are requested to conclude its global impact. Furthermore, the presented reference values of blood parameters in this species are interesting data for the pig industry

Generation of Calpain-3 knock-out porcine embryos by CRISPR-Cas9 electroporation and intracytoplasmic microinjection of oocytes before insemination

Limb girdle muscular dystrophy type R1 (LGMDR1) is an autosomal recessive myopathy described in humans resulting from a deficiency of calpain-3 protein (CAPN3). This disease lacks effective treatment and an appropriate model, so the generation of KO pigs by CRISPR-Cas9 offers a way to better understand disease ethology and to develop novel therapies. Microinjection is the main method described for gene editing by CRISPR-Cas9 in porcine embryo, but electroporation, which allows handling more embryos faster and easier, has also recently been reported. The objective of the current study was to optimize porcine oocyte electroporation to maximize embryo quality and mutation rate in order to efficiently generate LGMDR1 porcine models. We found that the efficiency of generating CAPN3 KO embryos was highest with 4 electroporation pulses and double sgRNA concentration than microinjection. Direct comparison between microinjection and electroporation demonstrated similar rates of embryo development and mutation parameters. The results of our study demonstrate that oocyte electroporation, an easier and faster method than microinjection, is comparable to standard approaches, paving the way for democratization of transgenesis in pigs

Expression of Glycogen Phosphorylase Isoforms in Cultured Muscle from Patients with McArdle's Disease Carrying the p.R771PfsX33 PYGM Mutation

Mutations in the PYGM gene encoding skeletal muscle glycogen phosphorylase (GP) cause a metabolic disorder known as McArdle's disease. Previous studies in muscle biopsies and cultured muscle cells from McArdle patients have shown that PYGM mutations abolish GP activity in skeletal muscle, but that the enzyme activity reappears when muscle cells are in culture. The identification of the GP isoenzyme that accounts for this activity remains controversial

Managing Heat Transfer Issues in Thermoelectric Microgenerators

This chapter deals with heat transfer challenges in the microdomain. It focuses on practical issues regarding this matter when attempting the fabrication of small footprint thermoelectric generators (μTEGs). Thermoelectric devices are designed to bridge a heat source (e.g. hot surface) and a heat sink (e.g. ambient) assuring that a significant fraction of the available temperature difference is captured across the active thermoelectric materials. Coexistence of those contrasted temperatures in small devices is challenging. It requires careful decisions about the geometry and the intrinsic thermal properties of the materials involved. The geometrical challenges lead to micromachined architectures, which silicon technologies provide in a controlled way, but leading to fragile structures, too. In addition, extracting heat from small systems is problematic because of the high thermal resistance associated to heat exchanged by natural convection between the surrounding air and small bare surfaces. Forced convection or the application of a cold finger clearly shows the usefulness of assembling a heat exchanger in a way that is effective and compliant with the mechanical constraints of micromachined devices. Simulations and characterization of fabricated structures illustrate the effectiveness of this element integration and its impact on the trade-off between electrical and thermal behavior of the active materials in device performance

Semen extenders used in the artificial insemination of swine: Review

Nuestro objetivo fue revisar a la luz de los últimos trabajos publicados los requisitos de un diluyente de inseminación, la importancia de la elección del mismo en las condiciones de producción actual y apuntar hacia dónde van dirigidas las investigaciones para su aplicación en un futuro próximo. Asumiendo que no hay un diluyente perfecto, su elección debe ir asociada al tipo de uso que se vaya a hacer de él. Cuando el tiempo de conservación sea inferior a tres días, la elección más racional sería la utilización de un diluyente de corta duración con unos costes menores y con unos resultados equivalentes a los de diluyentes de larga duración. Cuando lo que se pretende es conservar dosis seminales más allá de 4 días (debido a las largas distancias, evaluaciones sanitarias del semen, etc) se deben utilizar diluyentes de larga duración y aumentar la concentración de la dosis para compensar las pérdidas por envejecimiento de los espermatozoides. En cualquier caso la elección del diluyente debe realizarse con el objetivo de optimizar los resultados de fertilidad y prolificidad en las condiciones particulares de cada explotación porcina, ya que su repercusión en el rendimiento económico de la explotación es crucia

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