Antibody-independent protection against heterologous SARS-CoV-2 challenge conferred by prior infection or vaccination.

Vaccines have reduced severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) morbidity and mortality, yet emerging variants challenge their effectiveness. The prevailing approach to updating vaccines targets the antibody response, operating under the presumption that it is the primary defense mechanism following vaccination or infection. This perspective, however, can overlook the role of T cells, particularly when antibody levels are low or absent. Here we show, through studies in mouse models lacking antibodies but maintaining functional B cells and lymphoid organs, that immunity conferred by prior infection or mRNA vaccination can protect against SARS-CoV-2 challenge independently of antibodies. Our findings, using three distinct models inclusive of a novel human/mouse ACE2 hybrid, highlight that CD8+ T cells are essential for combating severe infections, whereas CD4+ T cells contribute to managing milder cases, with interferon-γ having an important function in this antibody-independent defense. These findings highlight the importance of T cell responses in vaccine development, urging a broader perspective on protective immunity beyond just antibodies.

Power Grids and Instrument Transformers up to 150 kHz: A Review of Literature and Standards.

The phenomenon of high-frequency distortion (HFD) in the electric grids, at both low-voltage (LV) and medium-voltage (MV) levels, is gaining increasing interest within the scientific and technical community due to its growing occurrence and the associated impact. These disturbances are mainly injected into the grid by new installed devices, essential for achieving decentralized generation based on renewable sources. In fact, these generation systems are connected to the grid through power converters, whose switching frequencies are significantly increasing, leading to a corresponding rise in the frequency of the injected disturbances. HFD represents a quite recent issue, but numerous scientific papers have been published in recent years on this topic. Furthermore, various international standards have also covered it, to provide guidance on instrumentation and related algorithms and indices for the measurement of these phenomena. When measuring HFD in MV grids, it is necessary to use instrument transformers (ITs) to scale voltages and currents to levels fitting with the input stages of power quality (PQ) instruments. In this respect, the recently released Edition 2 of the IEC 61869-1 standard extends the concept of the IT accuracy class up to 500 kHz; however, the IEC 61869 standard family provides guidelines on how to test ITs only at power frequency. This paper provides an extensive review of literature, standards, and the main outputs of European research projects focusing on HFD and ITs. This preliminary study of the state-of-the-art represents an essential starting point for defining significant waveforms to test ITs and, more generally, to achieve a comprehensive understanding of HFD. In this framework, this paper provides a summary of the most common ranges of amplitude and frequency variations of actual HFD found in real grids, the currently adopted measurement methods, and the normative open challenges to be addressed.

Characterization of Instrument Transformers under Realistic Conditions: Impact of Single and Combined Influence Quantities on Their Wideband Behavior.

Instrument transformers (ITs) play a key role in electrical power systems, facilitating the accurate monitoring and measurement of electrical quantities. They are essential for measurement, protection, and metering in transmission and distribution grids and accurately reducing the grid voltage and current for low-voltage input instrumentation. With the increase in renewable energy sources, electronic converters, and electric vehicles connected to power grids, ITs now face challenging distorted conditions that differ from the nominal ones. The study presented in this paper is a collaborative work between national metrology institutes and universities that analyzes IT performance in measuring distorted voltages and currents in medium-voltage grids under realistic conditions. Both current and voltage measuring transformers are examined, considering influence quantities like the temperature, mechanical vibration, burden, adjacent phases, and proximity effects. The study provides detailed insights into measurement setups and procedures, and it quantifies potential errors arising from IT behavior in measuring distorted signals in the presence of the various considered influence quantities and their combinations. The main findings reveal that the temperature has the most evident impact on the inductive voltage transformer performance, as well as the burden, causing significant changes in ratio error and phase displacement at the lower temperatures. As for low-power ITs, establishing a priori the effects of adjacent phases and proximity on the frequency responses of low-power ITs is a complex matter, because of their different characteristics and construction solutions.

Impact of DC Transient Disturbances on Harmonic Performance of Voltage Transformers for AC Railway Applications.

This paper analyzes the impact of typical DC transient events occurring in railway grids on the frequency performance of instrument transformers (ITs) installed onboard trains and in AC substations for power quality (PQ) applications. PQ monitoring in railway systems is an issue of great interest because it plays a key role in the improvement of energy efficiency. The measurement chain for the PQ measurements, at 15 kV at 16.7 Hz and 25 kV at 50/60 Hz, commonly includes ITs to scale the voltage to levels fitting the input of the measurement units. Nevertheless, the behavior of ITs in the presence of PQ phenomena represents an open issue from a normative point of view, even for those installed in conventional AC power supply systems. In this context, the paper presents a possible definition of DC transient disturbances test waveforms, a measurement procedure, and a setup to assess the impact of these disturbances on the harmonic performances of ITs for railway systems. Preliminary experimental tests carried out on two commercial ITs under wide ranges of variation for the amplitude and the time duration of DC disturbances show that, in some cases, the error introduced in harmonic measurements can exceed 100%.

How Instrument Transformers Influence Power Quality Measurements: A Proposal of Accuracy Verification Tests.

The integration of renewable energy sources on a large scale in the electrical energy distribution systems, as well as the widespread of non-linear loads, has led to a significant increase in power quality (PQ) disturbances. For this reason, PQ monitoring is also becoming a key task in medium voltage (MV) grids. The measurement of PQ at MV levels can only be performed using instrument transformers (ITs) to scale down the level of voltage and current to levels suitable for the input stage of PQ instruments. However, no international standards currently require the verification of the errors introduced by ITs in the measurement of PQ phenomena. Moreover, this issue is only partially addressed in the scientific literature, where papers dealing with specific and limited aspects of the problem can be found. For this reason, this paper aims to comprehensively assess the issue, proposing IT accuracy verification tests for different PQ parameters. First, a set of PQ phenomena relevant for IT testing is chosen, as well as the associated ranges of variation, based on a review of the enforced standards and the scientific literature. For each selected PQ phenomenon, possible performance indices and test waveforms are proposed. Finally, the proposed procedure is validated by applying it to the characterization of two different types of commercial voltage transformers.

Dorsogluteal intramuscular injection depth needed to reach muscle tissue according to body mass index and gender: A systematic review.

AIMS AND OBJECTIVE(S): The objective of this systematic review was to determine the needle length required to reach the dorsogluteal muscle based on body mass index and sex. Our aim was to provide evidence-based recommendations to current intramuscular injection guidelines from the result(s) of this review. BACKGROUND: Studies worldwide are documenting reduced medication effectiveness due to improperly placed dorsogluteal intramuscular injections because of incorrect needle length, wrong site selection and/or obesity. Current intramuscular injection guidelines lack specific instructions according to weight or sex. While there are similar concerns with other injectable sites, this review focuses solely on adult dorsogluteal intramuscular injections. DESIGN: A systematic review of relevant literature of dorsogluteal intramuscular injections based on body mass index and sex. METHODS: This systematic review was reported using the PRISMA checklist 2020. The review protocol was registered with Center for Open Science (OSF). We analysed 1,412 articles from nine databases. We compared twelve studies that utilised computerised tomography or ultrasonography using The Johns Hopkins Evidence-Based Practice Model and Guidelines. RESULTS: A significant number of dorsogluteal intramuscular injections are administered into subcutaneous tissue rather than muscle because needles are too short for populations with body mass indexes over 25, especially women. Poor landmarking often results in improperly placed injections. CONCLUSIONS: To prevent administering a dorsogluteal intramuscular injection into subcutaneous tissue, women with a BMI of 25 and over require needles longer than 38 mm (1.5 inches). Men have less subcutaneous tissue in the dorsogluteal area and only require longer needles if BMI is 35 and over. If skin-to-muscle depth is questionable in either sex, an ultrasound-guided intramuscular injection is warranted for accurate dorsogluteal placement. Landmarking and needle length are key to appropriately placed IM injections. RELEVANCE TO CLINICAL PRACTICE: Dorsogluteal injections are often injected into subcutaneous tissue rather than muscle because needles are not long enough to reach muscle, especially in women. Critical elements that determine placement of intramuscular injections into muscle versus subcutaneous tissue are sex, BMI, needle length and landmarking. Medications delivered into subcutaneous tissue may have reduced bioavailability.

Protein Kinase A Activation Promotes Cancer Cell Resistance to Glucose Starvation and Anoikis.

Cancer cells often rely on glycolysis to obtain energy and support anabolic growth. Several studies showed that glycolytic cells are susceptible to cell death when subjected to low glucose availability or to lack of glucose. However, some cancer cells, including glycolytic ones, can efficiently acquire higher tolerance to glucose depletion, leading to their survival and aggressiveness. Although increased resistance to glucose starvation has been shown to be a consequence of signaling pathways and compensatory metabolic routes activation, the full repertoire of the underlying molecular alterations remain elusive. Using omics and computational analyses, we found that cyclic adenosine monophosphate-Protein Kinase A (cAMP-PKA) axis activation is fundamental for cancer cell resistance to glucose starvation and anoikis. Notably, here we show that such a PKA-dependent survival is mediated by parallel activation of autophagy and glutamine utilization that in concert concur to attenuate the endoplasmic reticulum (ER) stress and to sustain cell anabolism. Indeed, the inhibition of PKA-mediated autophagy or glutamine metabolism increased the level of cell death, suggesting that the induction of autophagy and metabolic rewiring by PKA is important for cancer cellular survival under glucose starvation. Importantly, both processes actively participate to cancer cell survival mediated by suspension-activated PKA as well. In addition we identify also a PKA/Src mechanism capable to protect cancer cells from anoikis. Our results reveal for the first time the role of the versatile PKA in cancer cells survival under chronic glucose starvation and anoikis and may be a novel potential target for cancer treatment.

ECM Remodeling in Breast Cancer with Different Grade: Contribution of 2D-DIGE Proteomics.

Tumor extracellular matrix (ECM) plays a pivotal role in outcome of breast cancer (BC) patients. Overexpression of 58 genes, encoding 43 structural ECM proteins, has been identified to determine a specific cluster of BC with accelerated metastatic potential only in the undifferentiated (grade III) phenotype. The scope of this study is to characterize protein repertoire able to predict patient outcome in BC according to ECM gene expression pattern and histological grade. The differential proteomic analysis is based on 2D-differential gel electrophoresis, MALDI-MS, bioinformatics, and immunoblotting. Results suggest a relationship among ECM remodeling, signal mechanotransduction, and metabolic rewiring in BCs characterized by a specific mRNA ECM signature and identified a set of dysregulated proteins characteristic of hormone receptors expression as fibrinogen-ß chain, collagen α-1(VI) chain, and α-1B-glycoprotein. Furthermore, in triple negative tumors with ECM signature, the FGG and α5ß1/αvß3 integrins increase whereas detyrosinated α-tubulin and mimecan decrease leading to unorganized integrin presentation involving focal adhesion kinase, activation of Rho GTPases associated to epithelial mesenchymal transition. In hormone receptors negative BCs characterized by a specific ECM gene cluster, the differentially regulated proteins, identified in the present study, can be potentially relevant to predict patient's outcome.

Immunizations challenge healthcare personnel and affects immunization rates.

PURPOSE: This study measured 1. medical office immunization rates and 2. health care personnel competency in managing vaccine practices before and after evidence-based immunization education was provided. METHOD: This descriptive study compared 32 family medicine and pediatric offices and 178 medical assistants, licensed practical nurses, registered nurses, nurse practitioners, and physicians in knowledge-based testing pre-education, post-education, and 12-months post-education. Immunization rates were assessed before and 18-months post-education. RESULTS: Immunization rates increased 10.3% - 18months post-education; knowledge increased 7.8% - 12months post-education. Family medicine offices, licensed practical nurses, and medical assistants showed significant knowledge deficits before and 12-months post-education. All demographic groups scored less in storage/handling 12-months post-education. CONCLUSION: This study is one of the first studies to identify competency challenges in effective immunization delivery among medical assistants, licensed practical nurses, and family medicine offices. Formal and continuous education in immunization administration and storage/handling is recommended among these select groups.

Specific protein changes contribute to the differential muscle mass loss during ageing.

In the skeletal muscle, the ageing process is characterized by a loss of muscle mass and strength, coupled with a decline of mitochondrial function and a decrease of satellite cells. This profile is more pronounced in hindlimb than in forelimb muscles, both in humans and in rodents. Utilizing light and electron microscopy, myosin heavy chain isoform distribution, proteomic analysis by 2D-DIGE, MALDI-TOF MS and quantitative immunoblotting, this study analyzes the protein levels and the nuclear localization of specific molecules, which can contribute to a preferential muscle loss. Our results identify the molecular changes in the hindlimb (gastrocnemius) and forelimb (triceps) muscles during ageing in rats (3- and 22-month-old). Specifically, the oxidative metabolism contributes to tissue homeostasis in triceps, whereas respiratory chain disruption and oxidative-stress-induced damage imbalance the homeostasis in gastrocnemius muscle. High levels of dihydrolipoyllysine-residue acetyltransferase (Dlat) and ATP synthase subunit alpha (Atp5a1) are detected in triceps and gastrocnemius, respectively. Interestingly, in triceps, both molecules are increased in the nucleus in aged rats and are associated to an increased protein acetylation and myoglobin availability. Furthermore, autophagy is retained in triceps whereas an enhanced fusion, decrement of mitophagy and of regenerative potential is observed in aged gastrocnemius muscle.

Changes in muscle proteomics in the course of the Caudwell Research Expedition to Mt. Everest.

This study employed differential proteomic and immunoassay techniques to elucidate the biochemical mechanisms utilized by human muscle (vastus lateralis) in response to high altitude hypoxia exposure. Two groups of subjects, participating in a medical research expedition (A, n = 5, 19 d at 5300 m altitude; B, n = 6, 66 d up to 8848 m) underwent a ≈ 30% drop of muscular creatine kinase and of glycolytic enzymes abundance. Protein abundance of most enzymes of the tricarboxylic acid cycle and oxidative phosphorylation was reduced both in A and, particularly, in B. Restriction of α-ketoglutarate toward succinyl-CoA resulted in increased prolyl hydroxylase 2 and glutamine synthetase. Both A and B were characterized by a reduction of elongation factor 2 alpha, controlling protein translation, and by an increase of heat shock cognate 71 kDa protein involved in chaperone-mediated autophagy. Increased protein levels of catalase and biliverdin reductase occurred in A alongside a decrement of voltage-dependent anion channels 1 and 2 and of myosin-binding protein C, suggesting damage to the sarcomeric structures. This study suggests that during acclimatization to hypobaric hypoxia the muscle behaves as a producer of substrates activating a metabolic reprogramming able to support anaplerotically the tricarboxylic acid cycle, to control protein translation, to prevent energy expenditure and to activate chaperone-mediated autophagy.

hnRNPA2/B1 and nELAV proteins bind to a specific U-rich element in CDK5R1 3'-UTR and oppositely regulate its expression.

Cyclin-dependent kinase 5 regulatory subunit 1 (CDK5R1) encodes p35, a specific activator of cyclin-dependent kinase 5 (CDK5). CDK5 and p35 have a fundamental role in neuronal migration and differentiation during CNS development. Both the CDK5R1 3'-UTR's remarkable size and its conservation during evolution strongly indicate an important role in post-transcriptional regulation. We previously validated different regulatory elements in the 3'-UTR of CDK5R1, which affect transcript stability, p35 levels and cellular migration through the binding with nELAV proteins and miR-103/7 miRNAs. Interestingly, a 138 bp-long region, named C2.1, was identified as the most mRNA destabilizing portion within CDK5R1 3'-UTR. This feature was maintained by a shorter region of 73 bp, characterized by two poly-U stretches. UV-CL experiments showed that this region interacts with protein factors. UV-CLIP assays and pull-down experiments followed by mass spectrometry analysis demonstrated that nELAV and hnRNPA2/B1 proteins bind to the same U-rich element. These RNA-binding proteins (RBPs) were shown to oppositely control CDK5R1 mRNA stability and p35 protein content at post-trascriptional level. While nELAV proteins have a positive regulatory effect, hnRNPA2/B1 has a negative action that is responsible for the mRNA destabilizing activity both of the C2.1 region and of the full-length 3'-UTR. In co-expression experiments of hnRNPA2/B1 and nELAV RBPs we observed an overall decrease of p35 content. We also demonstrated that hnRNPA2/B1 can downregulate nELAV protein content but not vice versa. This study, by providing new insights on the combined action of different regulatory factors, contributes to clarify the complex post-transcriptional control of CDK5R1 gene expression.

Muscle proteomics reveals novel insights into the pathophysiological mechanisms of collagen VI myopathies.

Mutations in the collagen VI genes cause the Ullrich congenital muscular dystrophy (UCMD), with severe phenotype, and Bethlem myopathy (BM) with mild to moderate phenotype. Both, UCMD and BM patients show dystrophic features with degeneration/regeneration and replacement of muscle with fat and fibrous connective tissue. At molecular level, UCMD patients show autophagic impairment and increased PTP opening; these features are less severe in BM. To elucidate the biochemical mechanisms adopted by the muscle to adapt to collagen VI deficiency in BM and UCMD patients, a proteome analysis was carried out on human muscle biopsies. Qualitative and quantitative differences were assessed by 2D-DIGE coupled to MALDI-ToF/ToF MS. Proteomics results, coupled with immunoblotting, indicate changes in UPR, hexosamine pathway, and amino acid and fatty acid metabolism, suggesting an association of ER stress, metabolic dysregulation, autophagic impairment, and alteration in mechanotransduction signaling. Overall, these results indicate that despite the common downregulation of hexosamine pathway in UCMD and BM, in BM the protein quality control system is sustained by a metabolic adaptation supporting energy requirements for the maintenance of autophagy, counteracting ER misfolded protein overload. In UCMD, this multilayered system may be disrupted and worsened by the metabolic rewiring, which leads to lipotoxicity.

Contraceptive Counseling: Construction and Validation of Instrument-"5C Contraceptive Counseling".

INTRODUCTION: Contraceptive illiteracy leads to non-adherence, discontinuation, and dissatisfaction with the method. Person-centered contraceptive counseling is based on quality care on a communicative basis that promotes shared decision-making, leading to a choice adapted to the woman's needs, lifestyle, and health condition. We intend to build and validate an instrument that serves as a guide for quality contraceptive counseling, facilitating decision-making about contraceptive methods. METHODS: We used the Delphi method in a total of two rounds. The content was validated through a panel of eighteen experts with experience in teaching, research in contraceptive counseling, and obstetric nursing. To assess the consensus and stability of the responses, two questionnaires were administered and the Content Validity Index and Content Validity Ratio were calculated. RESULTS: The initial version of the guide, consisting of six indicators and thirty-five items, was submitted to the panel of experts to obtain consensus and stability from respondents (first round). The results showed a response rate of 66.0%; thirty-four indicators reached consensus and one did not reach consensus. Suggestions for modifying the indicator were received by the experts and incorporated in the next round. In the second round, the response rate increased to 78.0%. Two indicators were resubmitted, of which one was accepted. This resulted in the final version of the instrument, with six points and thirty-five items. DISCUSSION: The guide proved to be a valid tool for nurse-midwives to provide quality contraceptive advice to women, allowing them to make autonomous and informed choices regarding their sexual and reproductive health.

Nursing students' knowledge regarding sexuality, sex, and gender diversity in a multicenter study.

Introduction: Sexuality is an integral part of development and personality, and is important in healthcare. Nurses are among the most representative healthcare professionals. For holistic and inclusive nursing care practice and to improve equality, human rights, well-being, and health of individuals, the curricula of nursing courses must integrate broad knowledge about sexuality and its diversity. This study aimed to identify and analyze nursing students' knowledge of sexuality, sex, and gender diversity. The present study was part of a multicenter study conducted in Europe. Methods: Questionnaires were administered in three nursing schools to assess nursing students' knowledge (n = 75). Data processing was performed using Excel® software version 20 and IRaMuTeQ (R Interface pour les Analysis Multidimensionnelles de Textes et de Questionnaires) 0.7 alpha 2, allowing organization by category and subsequent thematic analysis using content analysis. Results: The textual corpus "Nursing students' knowledge about sexuality in its diversity," was divided into two sub-corpus: "Students' perception of sexuality" and "Students' perception of gender identity," originating Class 6 "Eroticism" (14.23%) and Classes 4 "Sexual Orientation" (16.07%) and 3 "Heteronormative" (16.07%), the latter with greater proximity to each other and consequently to Class 6. Similarly, Classes 1 "Gender" (20.36%) and 5 "Cisgender" (12.14%) also presented a greater interrelationship between themselves and consecutively with Class 2 "Gender Identity" (15.36%). Discussion: The analyses revealed that though nursing students possessed knowledge about sexuality and its diversity, this knowledge was elementary and did not reveal a sustained appropriation of concepts related to sexuality, sexual orientation, and gender diversity. For some questions, the absence of students' answers were noteworthy, and may be associated with their personal reservation in expressing themselves on this sensitive and intimate theme. To ensure diversity, inclusivity, and impartiality in nursing practice, it is imperative to change the curriculum plans of nursing courses to address the theme of sexuality during the training process of nurses in Europe.

Are IM injections IM in obese and overweight females? A study in injection technique.

If given incorrectly, intramuscular injections may result in poor absorption of drug, reduced drug effectiveness, or irritation to surrounding tissues. In this study, IM injection techniques were observed and documented for needle length, injection site, needle insertion, and stretching or bunching of the skin during injection in a population of adult females. The patients' weights and BMIs were recorded to determine the amount of subcutaneous fat at the injection site. In 22 patients of varied weights, 90% of injections were given within current Advisory Committee on Immunization Practice (ACIP) guidelines in normal and underweight patients, and 17% were given within ACIP guidelines in overweight and obese patients. The study concluded that the needle length used is often too short in overweight and obese individuals.

Educating to sexuality care: the nurse educator's experience in a multicenter study.

Background: Sexuality is an issue inherent in the lives of all human beings. Education for Sexuality takes place informally, through relationships with the environment, with the family as a model, and formally, as a pedagogical practice in Teaching. Education for sexuality is recognized as an instrument of social transformation that leads to changes in behaviors and norms related to sexuality. Objectives: Knowing the perception of nursing professors about sexuality education in professional training, recognizing attitudes of these professors in relation to sexual education and identifying barriers in education for sexuality. Methods: Exploratory and descriptive study, using qualitative methodology. Data collection was carried out from semi-structured interviews and thematic analysis. Results: The interviewees consider sexuality education to be very important, being taught in the nursing course, addressing different themes. In general, they reported feeling comfortable teaching these topics. The identified barriers to the level of education students are in, students' knowledge and reactions to the topic, religious and cultural issues, and the time available to talk about the topic and professional aspects. Conclusion: Sexuality is a fundamental theme in nursing education and needs to be further explored to overcome the barriers associated with its approach.

Biofortified Beverage with Chlorogenic Acid from Stressed Carrots: Anti-Obesogenic, Antioxidant, and Anti-Inflammatory Properties.

Using wounding stress to increase the bioactive phenolic content in fruits and vegetables offers a promising strategy to enhance their health benefits. When wounded, such phenolics accumulate in plants and can provide antioxidant, anti-inflammatory, and anti-obesogenic properties. This study investigates the potential of using wounding stress-treated carrots biofortified with phenolic compounds as a raw material to extract carrot juice with increased nutraceutical properties. Fresh carrots were subjected to wounding stress via slicing and then stored at 15 °C for 48 h to allow phenolic accumulation. These phenolic-enriched slices were blanched, juiced, and blended with orange juice (75:25 ratio) and 15% (w/v) broccoli sprouts before pasteurization. The pasteurized juice was characterized by its physicochemical attributes and bioactive compound content over 28 days of storage at 4 °C. Additionally, its antioxidant, anti-inflammatory, and anti-obesogenic potentials were assessed using in vitro assays, both pre- and post-storage. The results reveal that juice derived from stressed carrots (SJ) possessed 49%, 83%, and 168% elevated levels of total phenolics, chlorogenic acid, and glucosinolates, respectively, compared to the control juice (CJ) (p < 0.05). Both juices reduced lipid accumulation in 3T3-L1 cells and nitric oxide production in Raw 264.7 cells, without significant differences between them. SJ further displayed a 26.4% increase in cellular antioxidant activity. The juice's bioactive characteristics remained stable throughout storage time. In conclusion, the utilization of juice obtained from stressed carrots in a blend with orange juice and broccoli sprouts offers a promising method to produce a beverage enriched in bioactive compounds and antioxidant potential.

Chemical probes for the recognition of cannabinoid receptors in native systems.

Receptors made visible: The described biotin-tagged small-molecule probes with excellent affinities for the CB(1) and CB(2) cannabinoid receptors (CB(1)R and CB(2)R) enable direct visualization of these receptors in native cellular systems, including neurons, microglia, and immune cells. This method could overcome some of the limitations of current methodologies and may help to dissect the complexity of the endogenous cannabinoid system.

Comparative proteomic analyses of Duchenne muscular dystrophy and Becker muscular dystrophy muscles: changes contributing to preserve muscle function in Becker muscular dystrophy patients.

BACKGROUND: Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are characterized by muscle wasting leading to loss of ambulation in the first or third decade, respectively. In DMD, the lack of dystrophin hampers connections between intracellular cytoskeleton and cell membrane leading to repeated cycles of necrosis and regeneration associated with inflammation and loss of muscle ordered structure. BMD has a similar muscle phenotype but milder. Here, we address the question whether proteins at variance in BMD compared with DMD contribute to the milder phenotype in BMD, thus identifying a specific signature to be targeted for DMD treatment. METHODS: Proteins extracted from skeletal muscle from DMD/BMD patients and young healthy subjects were either reduced and solubilized prior two-dimensional difference in gel electrophoresis/mass spectrometry differential analysis or tryptic digested prior label-free liquid chromatography with tandem mass spectrometry. Statistical analyses of proteins and peptides were performed by DeCyder and Perseus software and protein validation and verification by immunoblotting. RESULTS: Proteomic results indicate minor changes in the extracellular matrix (ECM) protein composition in BMD muscles with retention of mechanotransduction signalling, reduced changes in cytoskeletal and contractile proteins. Conversely, in DMD patients, increased levels of several ECM cytoskeletal and contractile proteins were observed whereas some proteins of fast fibres and of Z-disc decreased. Detyrosinated alpha-tubulin was unchanged in BMD and increased in DMD although neuronal nitric oxide synthase was unchanged in BMD and greatly reduced in DMD. Metabolically, the tissue is characterized by a decrement of anaerobic metabolism both in DMD and BMD compared with controls, with increased levels of the glycogen metabolic pathway in BMD. Oxidative metabolism is severely compromised in DMD with impairment of malate shuttle; conversely, it is active in BMD supporting the tricarboxylic acid cycle and respiratory chain. Adipogenesis characterizes DMD, whereas proteins involved in fatty acids beta-oxidation are increased in BMD. Proteins involved in protein/amino acid metabolism, cell development, calcium handling, endoplasmic reticulum/sarcoplasmic reticulum stress response, and inflammation/immune response were increased in DMD. Both disorders are characterized by the impairment of N-linked protein glycosylation in the endoplasmic reticulum. Authophagy was decreased in DMD whereas it was retained in BMD. CONCLUSIONS: The mechanosensing and metabolic disruption are central nodes of DMD/BMD phenotypes. The ECM proteome composition and the metabolic rewiring in BMD lead to preservation of energy levels supporting autophagy and cell renewal, thus promoting the retention of muscle function. Conversely, DMD patients are characterized by extracellular and cytoskeletal protein dysregulation and by metabolic restriction at the level of α-ketoglutarate leading to shortage of glutamate-derived molecules that over time triggers lipogenesis and lipotoxicity.