98 research outputs found
Analysis and characterization of microbial biofilm associated with the roots of Phragmites australis and Typha latifolia
Searching for Mechanisms Underlying the Assembly of Calcium Entry Units: The Role of Temperature and pH
Store-operated Ca2+ entry (SOCE) is a mechanism that allows muscle fibers to recover external Ca2+, which first enters the cytoplasm and then, via SERCA pump, also refills the depleted intracellular stores (i.e., the sarcoplasmic reticulum, SR). We recently discovered that SOCE is mediated by Calcium Entry Units (CEUs), intracellular junctions formed by: (i) SR stacks containing STIM1; and (ii) I-band extensions of the transverse tubule (TT) containing Orai1. The number and size of CEUs increase during prolonged muscle activity, though the mechanisms underlying exercise-dependent formation of new CEUs remain to be elucidated. Here, we first subjected isolated extensor digitorum longus (EDL) muscles from wild type mice to an ex vivo exercise protocol and verified that functional CEUs can assemble also in the absence of blood supply and innervation. Then, we evaluated whether parameters that are influenced by exercise, such as temperature and pH, may influence the assembly of CEUs. Results collected indicate that higher temperature (36 °C vs. 25 °C) and lower pH (7.2 vs. 7.4) increase the percentage of fibers containing SR stacks, the n. of SR stacks/area, and the elongation of TTs at the I band. Functionally, assembly of CEUs at higher temperature (36 °C) or at lower pH (7.2) correlates with increased fatigue resistance of EDL muscles in the presence of extracellular Ca2+. Taken together, these results indicate that CEUs can assemble in isolated EDL muscles and that temperature and pH are two of the possible regulators of CEU formation. © 2023 by the authors
Retrospective Observational Study on Microbial Contamination of Ulcerative Foot Lesions in Diabetic Patients
According to recent studies, there are almost 435 million people worldwide with diabetes mellitus. It is estimated that of these 148 million will develop Diabetic foot ulcers (DFUs) during their lifetime, of which 35 to 50% will be infected. In this scenario, the presence and frequency of pathogenic microorganisms and their level of susceptibility to the most frequent classes of antibiotics used to treat this pathological condition from patients with DFUs admitted to the outpatient clinic of vascular surgery of the Federico II University Hospital of Naples from January 2019 to March 2021 were investigated. Furthermore, the diabetic population characteristics under study (i.e., general, clinical, and comorbidities) and the pathogenic bacteria isolated from lesions were also considered. Bacterial strains poorly susceptible to antibiotics were more frequent in polymicrobial infections than in monomicrobial infections. β-Lactams showed the highest levels of resistance, followed by fluoroquinolones, aminoglycosides, and finally macrolides. The main findings of the study demonstrated that the occurrence of resistant microorganisms is the dominant factor in ulcer healing; thus it is essential to investigate the antibiotics’ susceptibility before setting antibiotic therapy to avoid inappropriate prescriptions that would affect the treatment and increase the development and spread of antibiotic resistanc
Myomir dysregulation and reactive oxygen species in aged human satellite cells
AbstractSatellite cells that reside on the myofibre surface are crucial for the muscle homeostasis and regeneration. Aging goes along with a less effective regeneration of skeletal muscle tissue mainly due to the decreased myogenic capability of satellite cells. This phenomenon impedes proper maintenance and contributes to the age-associated decline in muscle mass, known as sarcopenia. The myogenic potential impairment does not depend on a reduced myogenic cell number, but mainly on their difficulty to complete a differentiation program. The unbalanced production of reactive oxygen species in elderly people could be responsible for skeletal muscle impairments. microRNAs are conserved post-transcriptional regulators implicated in numerous biological processes including adult myogenesis. Here, we measure the ROS level and analyze myomiR (miR-1, miR-133b and miR-206) expression in human myogenic precursors obtained from Vastus lateralis of elderly and young subjects to provide the molecular signature responsible for the differentiation impairment of elderly activated satellite cells
Improvement of muscle strength in a mouse model for congenital myopathy treated with HDAC and DNA methyltransferase inhibitors
To date there are no therapies for patients with congenital myopathies, muscle disorders causing poor quality of life of affected individuals. In approximately 30% of the cases, patients with congenital myopathies carry either dominant or recessive mutations in the ryanodine receptor 1 (RYR1) gene; recessive RYR1 mutations are accompanied by reduction of RyR1 expression and content in skeletal muscles and are associated with fiber hypotrophy and muscle weakness. Importantly, muscles of patients with recessive RYR1 mutations exhibit increased content of class II histone deacetylases and of DNA genomic methylation. We recently created a mouse model knocked-in for the p.Q1970fsX16+ p.A4329D RyR1 mutations, which are isogenic to those carried by a severely affected child suffering from a recessive form of RyR1-related multi-mini core disease. The phenotype of the RyR1 mutant mice recapitulates many aspects of the clinical picture of patients carrying recessive RYR1 mutations. We treated the compound heterozygous mice with a combination of two drugs targeting DNA methylases and class II histone deacetylases. Here, we show that treatment of the mutant mice with drugs targeting epigenetic enzymes improves muscle strength, RyR1 protein content, and muscle ultrastructure. This study provides proof of concept for the pharmacological treatment of patients with congenital myopathies linked to recessive RYR1 mutations
Extracellular GTP is a Potent Water- Transport Regulator via Aquaporin 5 Plasma-Membrane Insertion in M1-CCD Epithelial Cortical Collecting Duct Cells
Background/Aims: Extracellular GTP is able to modulate some specific functions in neuron, glia and muscle cell models as it has been demonstrated over the last two decades. In fact, extracellular GTP binds its specific plasma membrane binding sites and induces signal transduction via [Ca(2+)]i increase. We demonstrate, for the first time, that extracellular GTP is able to modulate cell swelling in M1-CCD cortical collecting duct epithelial cells via upregulation of aquaporin 5 (AQP5) expression. Methods: We used videoimaging, immunocitochemistry, flow cytometry, confocal techniques, Western blotting and RT-PCR for protein and gene expression analysis, respectively. Results: We demonstrate that AQP5 mRNA is up-regulated 7 h after the GTP exposure in the cell culture medium, and its protein level is increased after 12-24 h. We show that AQP5 is targeted to the plasma membrane of M1-CCD cells, where it facilitates cell swelling, and that the GTP-dependent AQP5 up-regulation occurs via [Ca(2+)]i increase. Indeed, GTP induces both oscillating and transient [Ca(2+)]i increase, and specifically the oscillating kinetic appears to be responsible for blocking cell cycle in the S-phase while the [Ca(2+)]i influx, with whatever kinetic, seems to be responsible for inducing AQP5 expression. Conclusion: The role of GTP as a regulator of AQP5-mediated water transport in renal cells is of great importance in the physiology of renal epithelia, due to its possible physiopathological implications. GTP-dependent AQP5 expression could act as osmosensor. In addition, the data presented here suggest that GTP might play the same role in other tissues where rapid water transport is required for cell volume regulation and maintenance of the homeostasis. © 2014 S. Karger AG, Basel. ispartof: Cellular Physiology and Biochemistry vol:33 issue:3 pages:731-46 ispartof: location:Germany status: publishe
Multicenter observational study on the reliability of the HEART score
Objective To rapidly and safely identify the risk of developing acute coronary syndrome in patients with chest pain who present to the emergency department, the clinical use of the History, Electrocardiogram, Age, Risk Factors, and Troponin (HEART) scoring has recently been proposed. This study aimed to assess the inter-rater reliability of the HEART score calculated by a large number of Italian emergency physicians.
Methods The study was conducted in three academic emergency departments using clinical scenarios obtained from medical records of patients with chest pain. Twenty physicians, who took the HEART score course, independently assigned a score to different clinical scenarios, which were randomly administered to the participants, and data were collected and recorded in a spreadsheet by an independent investigator who was blinded to the study\u2019s aim.
Results After applying the exclusion criteria, 53 scenarios were finally included in the analysis. The general inter-rater reliability was good (kappa statistics [\u3ba], 0.63; 95% confidence interval, 0.57 to 0.70), and a good inter-rater agreement for the high- and low-risk classes (HEART score, 7 to 10 and 0 to 3, respectively; \u3ba, 0.60 to 0.73) was observed, whereas a moderate agreement was found for the intermediate-risk class (HEART score, 4 to 6; \u3ba, 0.51). Among the different items of the HEART score, history and electrocardiogram had the worse agreement (\u3ba, 0.37 and 0.42, respectively).
Conclusion The HEART score had good inter-rater reliability, particularly among the high- and low-risk classes. The modest agreement for history suggests that major improvements are needed for objectively assessing this component.
Keywords HEART score; HEART pathway; Chest pain; Acute coronary syndrome; Emergency service, hospita
Single delivery of an adeno-associated viral construct to transfer the CASQ2 gene to knock-in mice affected by catecholaminergic polymorphic ventricular tachycardia is able to cure the disease from birth to advanced age
Background. Catecholaminergic polymorphic ventricular tachycardia is an inherited arrhythmogenic disorder characterized by sudden cardiac death in children. Drug therapy is still insufficient to provide full protection against cardiac arrest, and the use of implantable defibrillators in the pediatric population is limited by side effects. There is therefore a need to explore the curative potential of gene therapy for this disease. We investigated the efficacy and durability of viral gene transfer of the calsequestrin 2 (CASQ2) wild-type gene in a catecholaminergic polymorphic ventricular tachycardia knock-in mouse model carrying the CASQ2R33Q/R33Q (R33Q) mutation.
Methods and Results. We engineered an adeno-associated viral vector serotype 9 (AAV9) containing cDNA of CASQ2wild-type (AAV9-CASQ2) plus the green fluorescent protein (GFP) gene to infect newborn R33Q mice studied by in vivo and in vitro protocols at 6, 9, and 12 months to investigate the ability of the infection to prevent the disease and adult R33Q mice studied after 2 months to assess whether the AAV9-CASQ2 delivery could revert the catecholaminergic polymorphic ventricular tachycardia phenotype. In both protocols, we observed the restoration of physiological expression and interaction of CASQ2, junctin, and triadin; the rescue of electrophysiological and ultrastructural abnormalities in calcium release units present in R33Q mice; and the lack of life-threatening arrhythmias.
Conclusions. Our data demonstrate that viral gene transfer of wild-type CASQ2 into the heart of R33Q mice prevents and reverts severe manifestations of catecholaminergic polymorphic ventricular tachycardia and that this curative effect lasts for 1 year after a single injection of the vector, thus posing the rationale for the design of a clinical trial.Facultad de Ciencias MédicasCentro de Investigaciones Cardiovasculare
Local expression of SOD1G93A mutant protein triggers neuromuscular junction dismantlement
The alteration of Reactive Oxygen Species (ROS) homeostasis plays a causal role in several chronic pathology such as aging and neurodegenerative diseases like Amyotrophic Lateral Sclerosis (ALS). Although it is recognized that axon and synapses are first cellular sites of degeneration in ALS disease, controversy exists on whether pathological events initially begin at the NMJs and then, in a dying back phenomena, contribute to motor neuron degeneration. Moreover, the precise molecular mechanisms of pathology-associated deterioration in neuromuscular system have remained elusive (1). Here we provide evidences that muscle specific accumulation of SOD1G93A in the transgenic mice model MLC/SOD1G93A (2) induces mitochondria dysfunction and triggers NMJ dismantlement. Further, we demonstrate that treatment of MLC/SOD1G93A mice with Trolox, a potent antioxidant, is sufficient to rescue mitochondria and NMJ defects in the MLC/SOD1G93A mice, stabilizing muscle-nerve connection. The analysis of potential molecular mechanisms that mediate the toxic activity of SOD1 revealed the activation of specific Protein Kinase as a downstream player of NMJ dismantlement. Overall our data demonstrate that muscle specific expression of SOD1G93A mutation causes mitochondrial impairment and NMJ dismantlement, suggesting that muscle defects and NMJs alteration precede motor neuron degeneration rather than resulting from it
Multicenter observational study on the reliability of the HEART score
Objective To rapidly and safely identify the risk of developing acute coronary syndrome in patients with chest pain who present to the emergency department, the clinical use of the History, Electrocardiogram, Age, Risk Factors, and Troponin (HEART) scoring has recently been proposed. This study aimed to assess the inter-rater reliability of the HEART score calculated by a large number of Italian emergency physicians. Methods The study was conducted in three academic emergency departments using clinical scenarios obtained from medical records of patients with chest pain. Twenty physicians, who took the HEART score course, independently assigned a score to different clinical scenarios, which were randomly administered to the participants, and data were collected and recorded in a spreadsheet by an independent investigator who was blinded to the study’s aim. Results After applying the exclusion criteria, 53 scenarios were finally included in the analysis. The general inter-rater reliability was good (kappa statistics [κ], 0.63; 95% confidence interval, 0.57 to 0.70), and a good inter-rater agreement for the high- and low-risk classes (HEART score, 7 to 10 and 0 to 3, respectively; κ, 0.60 to 0.73) was observed, whereas a moderate agreement was found for the intermediate-risk class (HEART score, 4 to 6; κ, 0.51). Among the different items of the HEART score, history and electrocardiogram had the worse agreement (κ, 0.37 and 0.42, respectively). Conclusion The HEART score had good inter-rater reliability, particularly among the high- and low-risk classes. The modest agreement for history suggests that major improvements are needed for objectively assessing this component
- …