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Muscle atrophy is a complex process that is in common with many different catabolic diseases including disuse/inactivity and ageing. The signalling pathways that control the atrophy program in the different disuse/inactivity conditions have not yet been completely dissected. It has been recently reported that inhibition of FoxO only partially spared muscle mass after denervation. The purposes of this study were: (i) to determine the involvement of FoxOs in hindlimb suspension disuse model, (ii) to define whether the molecular events of protein breakdown are shared among different unloaded muscles and finally (iii) to compare the data obtained in this model with another model of inactivity such as denervation. Both wild type and muscle-specific FoxO1,3,4 knockout (FoxO1,3,4−/−) mice were unloaded for 3 and 14 days and muscles were characterized by functional, morphological, biochemical and molecular assays. Our data show that FoxOs are required for muscle loss and force drop during unloading. Moreover, we found that FoxO-dependent atrogenes vary in different unloaded muscles and that they diverge from denervation. Our findings clearly indicate that the signalling network that controls the atrophy program is specific for each catabolic condition. This article is protected by copyright. All rights reserved

Purpose Recent studies and our preliminary data demonstrate that muscle-specific ablation of the autophagy-related protein Atg7, leads to block of autophagy, sarcopenia and destabilization of the neuro-muscular junction (NMJ). In addition, Atg7 knock-out (Atg7 KO) muscle fibers release exosomes containing the muscle specific, miR-206, which is consistently elevated in the plasma. Interestingly, we found that miR-206 content was elevated in the heart, suggesting cardiac uptake of the miR-carrying circulating exosomes. We thus aimed at defining the effects of miR-206 on heart homeostasis. Methods Here, we analyzed the cardiac phenotype of adult (12mo.) and aged (24mo.) Atg7 KO mice, as well as of adult C57BL/6J mice injected, via tail vein, with scramble- or miR-206-loaded exosomes. Exosomes were isolated from EDL muscle of control and Atg7 KO mice, as well as from HEK293 cells. Heart function was assessed by echocardiography and ECG-telemetry. Confocal IF, whole-mount IF on heart blocks and multiphoton imaging were used to assess heart structure and sympathetic innervation. Bioinformatics, molecular and biochemical analyses were performed to identify novel targets of miR-206. IF, BRET assay and imaging of TrKA translocation were performed in cultured sympathetic neurons (SNs). Results We demonstrate that circulating exosomes, containing miR-206, are taken up by the heart leading to sympathetic dysinnervation, accompanied to reduction in the neurogenic control of cardiac rhythm and increased arrhythmogenesis. In vitro assays demonstrated that exosome-carried miR-206 targets cardiac SNs (cSNs), compromising cell structure and function. Indeed, increased miR-206 expression is accompanied by cSN atrophy, irregular axonal distribution of the active neurotransmitter release sites, and reduction in axonal sprouting. These effects are likely attributed to the miR-206-mediated down-regulation of the NGF receptor p75, as demonstrated by bioinformatics, luciferase assay, molecular and biochemical analyses in vitro and ex vivo. BRET assay, performed in cultured SNs treated with exosomes carrying miR-206, showed reduced formation of p75/TrkA complexes, which generate high-affinity binding sites for NGF and enhance neurotrophin responsiveness. Consistent with impaired NGF retrograde transport, miR-206 over-expressing SNs displayed reduced NGF protein content and decreased phosphorylation of Akt, which is an NGF downstream target, regulating neuronal survival. Interestingly, these latter results were confirmed in the stellate ganglia from Atg KO and miR-206 treated mice. miR-206 causes heart dysinnervation Conclusions We identify miR-206 as a key molecular player in the “muscle-to-heart” communication. miR-206 may participate to the pathogenesis of secondary cardiac dysfunction in skeletal muscle diseases associated to increased circulating levels of miR-206, ranging from ageing to neurodegenerative disorders (i.e. ALS, DMD).

A recent multi-center study showed how estimated glomerular filtration rate (eGFR) and cancer-specific mortality (CSM) are linearly and inversely related in organ-confined renal cell carcinoma (RCC) whenever the eGFR decreases below specific thresholds. We addressed our previous work limitations related to heterogeneity and missing data, and explored the relationship between eGFR and CSM also in locally advanced RCC.All patients with RCC treated with either partial or radical nephrectomy from 1990 to 2018 at a single institution and with complete data on renal function were included. eGFR was managed as a time-dependent variable. The relationship between eGFR and CSM was analyzed using a Fine and Gray multivariable competing risks framework. Subdistribution hazard ratios (SHRs) were calculated accounting for deaths from other causes.Multivariable competing risks analysis showed a "piecewise" relationship between eGFR and CSM, with an inverse linear correlation for eGFR values below 85 mL/min. Below this breakpoint, a significant relationship existed between eGFR and CSM in both clinical (SHR, 1.27; P .001) and pathologic (SHR, 1.27; P = .001) models in stage I to II RCC subgroup. Conversely, no significance was recorded in this subgroup when considering eGFR values above 85 mL/min. In the stage III to IV subgroup, no significant relationships were recorded, regardless of eGFR values. The retrospective design with inherent biases in data collection represents a limitation.In patients undergoing surgery for stage I to II RCC, preservation of renal function over "safety limits" is protective from CSM.

To compare the safety of on- vs off-clamp robotic partial nephrectomy (RAPN).302 patients with RENAL masses ≤ 10 were randomized to undergo on-clamp (150) vs off-clamp (152) RAPN (CLOCK trial-ClinicalTrials.gov NCT02287987) at seven institutions by one experienced surgeon per institution. Intra-operative data, complications, and positive surgical margins were compared.Due to a relevant rate of shift from the assigned treatment, the per-protocol analysis only was considered and the data from 129 on-clamp vs 91 off-clamp RAPNs analyzed. Tumor size (off-clamp vs on-clamp, 2.2 vs 3.0 cm, p 0.001) and RENAL score (5 vs 6, p 0.001) significantly differed. At univariate analysis, no differences were found regarding intra-operative estimated blood loss (off- vs on-clamp, 100 vs 100 ml, p = 0.7), post-operative complications rate (19% vs 26%, p = 0.2), post-operative anemia (Hb decrease 2.5 g/dl 26% vs 27%, p = 0.9; transfusion rate 3.4% vs 6.3%, p = 0.5; re-intervention due to bleeding 1.1% vs 4%, p = 0.4), acute kidney injury (4% vs 6%, p = 0.8), and positive surgical margins (3.5% vs 8.2%, p = 0.1). At multivariate analysis accounting for tumor diameter and complexity, considering the on-clamp group as the reference category, a significant difference was noted in the off-clamp group exclusively for blood loss (OR 0.3, 95% CI 0.09-0.52, p = 0.008).The on-clamp and off-clamp approaches for RAPN showed a comparable safety profile.

Spinal and bulbar muscular atrophy (SBMA) is caused by polyglutamine (polyQ) expansions in the androgen receptor (AR) gene. Although clinical and experimental evidence highlight a primary role for skeletal muscle in the onset, progression, and outcome of disease, the pathophysiological and molecular processes underlying SBMA muscle atrophy are poorly understood. Here we show that polyQ-expanded AR alters intrinsic muscle force generation before denervation. Reduced muscle force was associated with a switch in fiber-type composition, disrupted muscle striation, altered calcium (Ca++) dynamics in response to muscle contraction, and aberrant expression of excitation-contraction coupling (ECC) machinery genes in transgenic, knock-in and inducible SBMA mice and patients. Importantly, treatment to suppress polyQ-expanded AR toxicity restored ECC gene expression back to normal. Suppression of AR activation by surgical castration elicited similar ECC gene expression changes in normal mice, suggesting that AR regulates the expression of these genes in physiological conditions. Bioinformatic analysis revealed the presence of androgen-responsive elements on several genes involved in muscle function and homeostasis, and experimental evidence showed AR-dependent regulation of expression and promoter occupancy of the most up-regulated gene from transcriptomic analysis in SBMA muscle, i.e. sarcolipin, a key ECC gene. These observations reveal an unpredicted role for AR in the regulation of expression of genes involved in muscle contraction and Ca++ dynamics, a level of muscle function regulation that is disrupted in SBMA muscle, yet restored by pharmacologic treatment.

The maintenance of muscle mass and its ability to function relies on a bioenergetic efficient mitochondrial network. This network is highly impacted by fusion and fission events. We have recently shown that the acute deletion of the fusion protein Opa1 induces muscle atrophy, systemic inflammatory response, precocious epithelial senescence, and premature death that are caused by muscle-dependent secretion of FGF21. However, both fusion and fission machinery are suppressed in aging sarcopenia, cancer cachexia, and chemotherapy-induced muscle wasting. We generated inducible muscle-specific Opa1 and Drp1 double-knockout mice to address the physiological relevance of the concomitant impairment of fusion and fission machinery in skeletal muscle. Here we show that acute ablation of Opa1 and Drp1 in adult muscle causes the accumulation of abnormal and dysfunctional mitochondria, as well as the inhibition of autophagy and mitophagy pathways. This ultimately results in ER stress, muscle loss, and the reduction of force generation. However, the simultaneous inhibition of the fission protein Drp1 when Opa1 is absent alleviates FGF21 induction, oxidative stress, denervation, and inflammation rescuing the lethal phenotype of Opa1 knockout mice, despite the presence of any muscle weakness. Thus, the simultaneous inhibition of fusion and fission processes mitigates the detrimental effects of unbalanced mitochondrial fusion and prevents the secretion of pro-senescence factors.

Introduction Accurate prediction of extraprostatic extension (EPE) of prostate cancer (PCa) is the keystone for deciding whether to perform a neurovascular bundle sparing (NVBs) radical prostatectomy or not, which will subsequently affect the postoperative functional outcomes especially potency. Partin tables are the most commonly used predictive tools (PTs) for prediction of EPE. Moreover, they are the most commonly externally validated. In these settings, the aim of our work is to perform a systematic review of the literature and a meta-analysis for the discriminative performance of the different versions of Partin tables for EPE prediction. Evidence acquisition A systematic search of Medline, Scopus and Cochrane library was performed to include all the external validation (EV) studies that reported the discriminative performance (area under the curve [AUC]) of the different versions of Partin tables as a PT for EPE. Different versions of Partin tables (1997, 2001, 2007, 2010, and 2013) were included in separate meta-analyses. The pooled AUC with 95% CI were calculated to determine the weighted summary AUC using the random effect model. Evidence synthesis Twenty-six studies carried out in different countries including the USA, Korea, Germany, Ireland, China, Austria, France, Italy, the UK, and India were included. Considering the small number and generally low quality of the EV studies in literature, most of the included studies showed some sort of bias especially in the sample size & missing data domain. The pooled EPE AUC were 0.642 (95% CI; 0.601-0.682), 0.672 (95% CI; 0.617-0.727), 0.659 (95% CI; 0.623-0.695), 0.669 (95% CI; 0.623-0.715) and 0.644 (95% CI; 0.545-0.742) for the 1997, 2001, 2007, 2010 and 2013 versions, respectively. Conclusions Despite being the most commonly used predictive tool for prediction of EPE, the pooled EPE AUC for different versions of Partin tables showed poor discriminative performance. Thus, surgeons must be cautious when referring to Partin tables for prediction of EPE. Further EV studies are required to confirm these results.

INTRODUCTION AND OBJECTIVES:Lymph node dissection (LND) during radical prostatectomy (RP) for prostate cancer (PCa) aims to provide information for staging and prognosis, while oncological and surv...

INTRODUCTION AND OBJECTIVES:Prediction of the presence of extracapsular extension (ECE) of prostate cancer (PCa) before surgery is of paramount importance to tailor the amount of nerve-sparing during radical prostatectomy (RP). A novel nomogram to predict ECE has been recently developed with the integration of a multiparametric magnetic resonance imaging (mpMRI) derived variable (Martini et al, BJUI 2018, 1-9). Authors defined the “presence of ECE” as the loss or irregularity of the capsule, whereas contact, bulge or abutment are considered as negative for ECE.We aimed to externally validate this nomogram on 137 prostatic lobes from 106 patients undergoing mpMRI-targeted biopsy plus saturation sampling.METHODS:We applied the model from Martini to the most recent cases of PCa patients (n=106) with a positive mpMRI submitted to RP. PCa was diagnosed in all cases by mpMRI-targeted plus systematic saturation biopsy. According to Martini′s model, we considered only lobes with a positive biopsy (137). The prima...

Addiction is accompanied by attentional biases (AB), wherein drug-related cues grab attention independently of their perceptual salience. AB have emerged in different flavours depending on the experimental approach, and their clinical relevance is still debated. In chronic smokers we sought evidence for dissociable attention abnormalities that may play distinct roles in the clinical manifestations of the disorder. Fifty smokers performed a modified visual probe-task measuring two forms of AB and their temporal dynamics, and data on their personality traits and smoking history/ status were collected. Two fully dissociable AB effects were found: A Global effect, reflecting the overall impact of smoke cues on attention, and a Location-specific effect, indexing the impact of smoke cues on visuospatial orienting. Importantly, the two effects could be neatly separated from one another as they: (i) unfolded with dissimilar temporal dynamics, (ii) were accounted for by different sets of predictors associated with personality traits and smoking history and (iii) were not correlated with one another. Importantly, the relevance of each of these two components in the single individual depends on a complex blend of personality traits and smoking habits, a result that future efforts addressing the clinical relevance of addiction-related AB should take into careful consideration., This study was supported by funding provided by the University of Verona to CDL, CC and LC

Background: The hypothesis that renal function could influence oncological outcomes is supported by anecdotal literature. Objective: To determine whether estimated glomerular filtration rate (eGFR) is related to cancer-specific mortality (CSM) in patients who had undergone surgery for renal cell carcinoma (RCC). Design, setting, and participants: A retrospective analysis of 3457 patients who underwent radical (39%) or partial nephrectomy (61%) for cT1–2 RCC between 1990 and 2015. Outcome measurements and statistical analysis: The eGFR was calculated by the Chronic Kidney Disease Epidemiology Collaboration equation. CSM was analyzed in a multivariable competing-risk framework, estimating the subdistribution hazard ratio (SHR) accounting for deaths from other causes. The relationship between eGFR and CSM was investigated from multiple statistical approaches—extended Cox regression with eGFR incorporated as a time-dependent covariate, landmark analysis, and joint modeling. Other predictors were selected by competing-risk random forest method and backward elimination. Results and limitations: The relationship between eGFR and CSM was graphically described by a linear spline, i.e. a continuous piecewise linear function with two lines joined by a knot. For eGFR treated as a time-dependent covariate, the knot was located at 65 ml/min; at landmark analysis with eGFR at the baseline, 12 mo, and last functional follow-up, the knots were 85, 60, and 65 ml/min, respectively. In multivariable competing-risk analysis, CSM was associated with eGFR only for values of eGFR below these cutoffs, with SHRs for every 10 ml/min of reduction in eGFR of 1.25 (p = 0.003), 1.16 (p = 0.028), 1.44 (p = 0.02), and 1.16 (p = 0.042), corresponding to time-dependent eGFR, and eGFR at baseline, 12 mo, and last functional follow-up, respectively. Joint modeling confirmed these results. A retrospective design with inherent biases in data collection represents a limitation. Conclusions: In patients undergoing surgery for RCC, renal function should be preserved in order to improve cancer-related survival. Patient summary: The relationship between renal function and probability of dying due to renal cancer is complex. The present study found a correlation between glomerular filtration rate and cancer specific mortality that could reconsider the oncological role of renal function in patients undergoing surgery for renal cancer. We investigate the correlation between preserved renal function during surgery and cancer-specific survival for renal cancer. Analyzing 3457 patients submitted to surgery, we found a linear and inverse correlation between estimated glomerular filtration rate (eGFR) and cancer-related mortality only for values of eGFR below certain limits.

Background Apolipoprotein CIII (apo CIII ) is a crucial player in triglyceride‐rich lipoprotein metabolism, but may also act pleiotropically, provoking inflammatory responses and stimulating coagulation. Elevated apo CIII plasma levels have been associated with increased activity of coagulation factors. Since these features of prothrombotic diathesis are linked with venous thromboembolism ( VTE ), we hypothesized that apo CIII plays a role in VTE . Methods and Results We recorded nonfatal VTE events in 1020 patients (age 63.3±11.4 years; 29.1% women) with or without coronary artery disease (79.1% with coronary artery disease and 20.9% without coronary artery disease) during a long follow‐up. Complete plasma lipid and apolipoproteins were available for all patients. Forty‐five patients (4.4%) experienced nonfatal VTE events during a median follow‐up period of 144 months. Apo CIII plasma concentration at enrollment was higher in patients with VTE compared with patients without VTE (12.2 [95% CI, 11.10–13.5] mg/dL vs 10.6 [95% CI, 10.4–10.9] mg/dL, respectively; P =0.011). Patients with apo CIII levels above the median value (10.6 mg/dL) exhibited an increased risk of VTE (incidence rate, 6.0 [95% CI , 4.0–8.0] vs 1.8 [95% CI, 0.7–2.9] VTE events/1000 person‐years; unadjusted hazard ratio [ HR ], 3.42 [95% CI , 1.73–6.75]; P HR , 2.66; 95% CI , 1.31–5.37 [ P =0.007]), with inclusion of lipid parameters in the Cox model (HR, 3.74; 95% CI , 1.24–11.33 [ P =0.019]), and even with exclusion of patients who died at follow‐up ( HR, 3.92; 95% CI , 1.68–9.14 [ P =0.002]) or patients taking anticoagulants ( HR , 3.39; 95% CI , 1.72–6.69 [ P Conclusions Our results suggest that high plasma apo CIII concentrations may predict an increased risk of VTE in patients with cardiovascular disease.

To outline the design and rationale of clock, a large, phase III randomized controlled trial which examines the role vascular clamping during robot-assisted partial nephrectomy (RAPN) for localized renal cancer. To be eligible for study entry, patients must have normal coagulative function, preoperative estimated glomerular filtration rate (eGFR) >60 mL/min, normal contralateral kidney and a single renal mass with RENAL Score ≤10. Eligible patients are to be randomized in a 1:1 ratio between two arms. Randomized allocation was assigned by a permuted block design, stratified by center. Allocation arm was notified by the study internet-based e-form, managed by an independent software house. Arm A=RAPN with ischemia; Arm B=RAPN without ischemia. At any moment, from randomization to the end of the procedure, the investigators could amend the indication given by randomization and shift to the alternative clamping option, detailing the timing and reasons of their decision. CLOCK is a randomized controlled trial, which addresses two questions relating to the management of localized renal cancer treated by RAPN with or without ischemia: 1) what is the impact of the surgical technique on long-term renal function; 2) what are the factors influencing the shift from one technique to the other. The study will be completed 24 months after the last enrollment.

Sarcopenia is a loss of muscle mass and function in the elderly that reduces mobility, diminishes quality of life, and can lead to fall-related injuries, which require costly hospitalization and extended rehabilitation. This review focuses on the aging-related structural changes and mechanisms at cellular and subcellular levels underlying changes in the individual motor unit: specifically, the perikaryon of the α-motoneuron, its neuromuscular junction(s), and the muscle fibers that it innervates. Loss of muscle mass with aging, which is largely due to the progressive loss of motoneurons, is associated with reduced muscle fiber number and size. Muscle function progressively declines because motoneuron loss is not adequately compensated by reinnervation of muscle fibers by the remaining motoneurons. At the intracellular level, key factors are qualitative changes in posttranslational modifications of muscle proteins and the loss of coordinated control between contractile, mitochondrial, and sarcoplasmic reticulum protein expression. Quantitative and qualitative changes in skeletal muscle during the process of aging also have been implicated in the pathogenesis of acquired and hereditary neuromuscular disorders. In experimental models, specific intervention strategies have shown encouraging results on limiting deterioration of motor unit structure and function under conditions of impaired innervation. Translated to the clinic, if these or similar interventions, by saving muscle and improving mobility, could help alleviate sarcopenia in the elderly, there would be both great humanitarian benefits and large cost savings for health care systems.

This study tested the effect of two block setting conditions i.e., the usual block setting [US] and an anthropometry-driven block setting [AS] on the kinematic and kinetic parameters of the sprint start. Furthermore, we verified whether this effect is influenced by the relative lengths of the sprinter’s trunk and lower limbs i.e., the Cormic Index by subdividing sprinters into brachycormic, metricormic and macrocormic groups. Forty-two sprinters performed 6 maximal-effort 10 m sprints using the US and AS conditions. Dynamometric starting blocks measured forces generated by the sprinters. The times at 5 m and 10 m in the sprint trials were measured with photocells. Results showed that the anteroposterior block distances were significantly different between the two conditions (P

Our aim was to review and externally validate all the available predictive tools (PTs) predicting extraprostatic extension (EPE) using the area under the curve, calibration plots, and scaled Brier score. A literature search was performed showing 19 models predicting EPE. External validation was carried out on 6360 prostate cancer patients submitted to RP. Most of the PTs showed poor discrimination and unsatisfactory calibration. The majority of the available PTs are not reliable for the prediction of EPE in populations other than the development one; thus, they may not be completely appropriate for patients' counselling or for surgical strategy preplanning.

Mitochondrial quality control is essential in highly structured cells such as neurons and muscles. In skeletal muscle the mitochondrial fission proteins are reduced in different physiopathological conditions including ageing sarcopenia, cancer cachexia and chemotherapy-induced muscle wasting. However, whether mitochondrial fission is essential for muscle homeostasis is still unclear. Here we show that muscle-specific loss of the pro-fission dynamin related protein (DRP) 1 induces muscle wasting and weakness. Constitutive Drp1 ablation in muscles reduces growth and causes animal death while inducible deletion results in atrophy and degeneration. Drp1 deficient mitochondria are morphologically bigger and functionally abnormal. The dysfunctional mitochondria signals to the nucleus to induce the ubiquitin-proteasome system and an Unfolded Protein Response while the change of mitochondrial volume results in an increase of mitochondrial Ca2+ uptake and myofiber death. Our findings reveal that morphology of mitochondrial network is critical for several biological processes that control nuclear programs and Ca2+ handling., Muscle loss is associated with altered expression of proteins involved in mitochondrial homeostasis, but whether this is causative remains unclear. Here, the authors show that genetic ablation of the pro-fission protein DRP1 leads to accumulation of abnormal mitochondria that induce muscle atrophy by altering Ca2+ homeostasis and cellular stress responses.

The progress of the grapevine genomics and the development of high-throughput technologies for gene expression analysis stimulated the investigation of the physical, biochemical and physiological changes of grape berry growth and maturation at transcriptomic level. The molecular information generated in the last decade is however still fragmented since it relies upon detailed analysis of few stages and thus lacks continuity over grape development. To identify the molecular events associated with berry development at a higher temporal resolution and define a transcriptomic map, we performed RNA-seq analysis of berry samples collected every week from fruit-set to maturity in Pinot noir and Cabernet Sauvignon for three consecutive years, resulting in 219 samples. Using the most variable portion of the transcriptome, we built a preliminary transcriptomic model of berry development based on the Cabernet Sauvignon samples. The Pinot noir samples were then aligned onto this preliminary ripening map to investigate its performance in describing the development of another grape variety. A further step for testing the model was the projection of RNA-seq samples of fruit development of five red-skin Italian cultivars. For all these surveys, the transcriptomic route allowed a precise definition of the progression of berry development during both formation and ripening phases.

Electron paramagnetic resonance (EPR), coupled with site-directed spin labeling, has been proven to be a particularly suitable technique to extract information on the fraction of myosin heads strongly bound to actin upon muscle contraction. The approach can be used to investigate possible structural changes occurring in myosin of fiber s altered by diseases and aging. In this work, we labeled myosin at position Cys707, located in the SH1-SH2 helix in the myosin head cleft, with iodoacetamide spin label, a spin label that is sensitive to the reorientational motion of this protein during the ATPase cycle and characterized the biochemical states of the labeled myosin head by means of continuous wave EPR. After checking the sensitivity and the power of the technique on different muscles and species, we investigated whether changes in the fraction of strongly bound myosin heads might explain the contractile alterations observed in atrophic and hypertrophic murine muscles. In both conditions, the difference in contractile force could not be justified simply by the difference in muscle mass. Our results showed that in atrophic muscles the decrease in force generation was attributable to a lower fraction of strongly bound cross bridges during maximal activation. In contrast in hypertrophic muscles, the increase in force generation was likely due to several factors, as pointed out by the comparison of the EPR experiments with the tension measurements on single skinned fibers.

Bone and skeletal muscle are currently considered a unified functional unit, showing complementary regulation at mechanical, biochemical, paracrine and metabolic levels. This functional unit undergoes a central hormonal regulation which is mainly ascribed to sex steroids and, in particular, androgens. However, recent evidence suggest that another testicular hormone lines the classical anabolic effect of testosterone on bone and muscle, the insulin-like peptide 3 (INSL3) acting on its specific receptor RXFP2. This minireview focuses on the most recent findings describing the role of INSL3/RXFP2 axis on the muscolo-skeletal system, from the mechanistic insights to the phenotypic consequences. Pathophysiological and therapeutic widenings deriving from available data are also discussed.

CDCP1, a transmembrane noncatalytic receptor, the expression of which has been associated with a poor prognosis in certain epithelial cancers, was found to be expressed in highly aggressive triple-negative breast cancer (TNBC) cell models, in which it promoted aggressive activities-ie, migration, invasion, anchorage-independent tumor growth, and the formation of vascular-like structures in vitro. By immunohistochemical (IHC) analysis of 100 human TNBC specimens, CDCP1 was overexpressed in 57% of samples, 38% of which exhibited a gain in CDCP1 copy number by fluorescence in situ hybridization (FISH). CDCP1 positivity was significantly associated between FISH and IHC. CDCP1 expression and gains in CDCP1 copy number synergized with nodal (N) status in determining disease-free and distant disease-free survival. The hazard ratios (HRs) of the synergies between CDCP1 positivity by IHC and FISH and lymph node positivity in predicting relapse did not differ significantly, indicating that CDCP1 overexpression in human primary TNBCs, regardless of being driven by gains in CDCP1, is for a critical factor in the progression of N-positive TNBCs. Thus, CDCP1 is a novel marker of the most aggressive N-positive TNBCs and a potential therapeutic target.

BACKGROUND The aim of this study was to evaluate the effectiveness of ferric carboxymaltose (FCM) in patients with iron deficiency anemia (IDA) secondary to gastrointestinal chronic blood loss (CBL), who received chronic transfusion support. STUDY DESIGN AND METHODS We retrospectively evaluated 38 patients with IDA (hemoglobin [Hb]

Skeletal muscle is a highly adaptive tissue, capable of altering muscle fiber size, functional capacity and metabolism in response to physiological stimuli. However, pathological conditions such as cancer growth compromise the mechanisms that regulate muscle homeostasis, resulting in loss of muscle mass, functional impairment and compromised metabolism. This tumor-induced condition is characterized by enhanced muscle protein breakdown and amino acids release that sustain liver gluconeogenesis and tissue protein synthesis. Proteolysis is controlled by the two most important cellular degradation systems, the ubiquitin proteasome and autophagy lysosome. These systems are carefully regulated by different signalling pathways that determine protein and organelle turnover. In this review we will describe the involvement of the ubiquitin proteasome and autophagy lysosome systems in cancer cachexia and the principal signalling pathways that regulate tumor-induced protein breakdown in muscle.

Skeletal muscles and bones form a joined functional unit sharing a complex mechanical, biochemical and hormonal crosstalk. A number of factors, including sex hormones, physiologically regulate the musculoskeletal system. Striking gender differences in muscle and bone mass, and function are mainly caused by distinct actions exerted by oestrogens and androgens. However, relaxin and relaxin-related peptides, such as insulin-like peptide 3 (INSL3), might contribute to these sex-associated differences in physiological and pathological conditions (such as osteoporosis and sarcopenia). Relaxin is a 'pregnancy' hormone, but it is also produced from the prostate gland, and has recently attracted attention as a potential drug for cardiovascular disorders and fibrosis. In contrast, INSL3 is a male-specific hormone produced by the Leydig cells of the testis with a fundamental role in testicular descent during fetal life. Recent evidence suggests that both hormones have interesting roles in the musculoskeletal system. Relaxin and INSL3, by finely tuning bone formation and resorption, are involved in bone remodelling processes, and relaxin contributes to the healing of injured ligaments and promotes skeletal muscle regeneration. Here, we review the most recent findings on the effects of relaxin and INSL3 on skeletal muscle and the cell components of bone. In the light of the experimental evidence available and animal models, their clinical implications are also discussed. Linked articles This article is part of a themed section on Recent Progress in the Understanding of Relaxin Family Peptides and their Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.10/issuetoc.

KEY POINTS: Using an experimental rat intensive care unit (ICU) model, not limited by early mortality, we have previously shown that passive mechanical loading attenuates the loss of muscle mass an ...

Measuring biomarkers (e.g. volatile organic compounds [VOCs]) in exhaled breath is an attractive approach to monitor airway inflammation in asthma and other lung diseases. Olfactive technology by electronic nose (e-Nose) has been applied to identify VOCs in exhaled breath. We compared e-Nose respiratory patterns in a pediatric cohort with asthma classificate children with different asthma control. This cross-sectional study involved 38 children: 28 with asthma and 10 healthy controls . The asthmatic patients were categorized as having controlled (AC), partially controlled (APC) or uncontrolled asthma (ANC) based on level of asthma symptom control according to Global Initiative for Asthma (GINA). Clinical exams, exhaled breath collection for generating e-Nose VOC profiles, and spirometry were performed. Exhaled breath samples were obtained using a commercial electronic nose (Cyranose 320; Smith Detections, Pasadena, CA, USA). The discriminative ability of breathprints were investigated by principal component analysis and penalized logistic regression. The e-Nose was able to discriminate between the CON (controls) + AC and the ANC + APC group with an area under the curve [AUC] of 0.85 (95% confidence interval [CI] 0.72 to 0.98) and a cross-validated AUC of 0.80 (95% CI 0.70 to 0.85). Sensitivity and specificity calculated using the Youden index were 0.79 and 0.84, respectively. Exhaled biomarker patterns were easy to obtain with the device and were able to differentiate children with uncontrolled symptomatic asthma from asymptomatic controls.

We previously identified an extracellular matrix (ECM) gene expression pattern in breast cancer (BC), called ECM3, characterized by a high expression of genes encoding structural ECM proteins. Since ECM is reportedly implicated in response to therapy of BCs, the aim of this work is to investigate the prognostic and predictive value of ECM3 molecular classification in HER2-positive BCs. ECM3 resulted in a robust cluster that identified a subset of 25&ndash, 37% of HER2-positive tumors with molecular aggressive features. ECM3 was significantly associated with worse prognosis in two datasets of HER2-positive BCs untreated with adjuvant therapy. Analyses carried out on two of our cohorts of patients treated or not with adjuvant trastuzumab showed association of ECM3 with worse prognosis only in patients not treated with trastuzumab. Moreover, investigating a dataset that includes gene profile data of tumors treated with neoadjuvant trastuzumab plus chemotherapy or chemotherapy alone, ECM3 was associated with increased pathological complete response if treated with trastuzumab. In the in vivo experiments, increased diffusion and trastuzumab activity were found in tumors derived from injection of HER2-positive cells with Matrigel that creates an ECM-rich tumor environment. Taken together, these results indicate that HER2-positive BCs classified as ECM3 have an aggressive phenotype but they are sensitive to trastuzumab treatment.

BACKGROUND AND OBJECTIVES The role of pre-donation blood pressure (BP) as independent contributor to post-donation vasovagal reactions (VVRs) is still debated. Differences between a liberal (i.e., inclusion of hypotensive donors) and a restrictive policy (i.e., not accepting hypotensive donors) should be investigated. This study aims to investigate the consequences of a liberal policy in development of VVRs after whole-blood donations. MATERIALS AND METHODS We compared the incidence of VVRs between 2015 (restrictive policy) and 2016 (liberal policy) and the associated risk factors. We evaluated respectively 22 789 vs. 21 676 blood donations obtained from 18 001 blood donors (12 501 donated in both years). RESULTS Comparing the results we obtained between 2015 and 2016, donations showed an overlap of the cohorts. Two hundred fifteen VVRs (incidence rate 0·48%) were observed, 104 (0·46%) of which in 2015, and 111 (0·51%) in 2016. A preliminary univariate analysis showed that donors with systolic BP

Macroautophagy/autophagy is emerging as an important process in adult muscle stem cells functions: it regulates metabolic reprogramming during activation from a quiescent state, maintains stemness and prevents senescence. We now show that autophagy is specifically required for neonatal myogenesis and muscle development. Specific deletion of Atg7 in PAX7(+) (paired box 7) precursors led in mice to a dwarf phenotype, with an effect restricted to the neonatal phase of muscle development. Atg7 knockdown suppressed neonatal satellite cell (nSC) proliferation and differentiation, downregulating the GH-IGF1 functions. When we disrupted autophagy, NFE2L2/NRF2 (nuclear factor, erythroid 2 like 2) accumulated in muscle and nSCs and negatively modulated DDIT3/CHOP (DNA-damage inducible transcript 3) expression. Lower levels of DDIT3 were responsible for reduced GHR expression leading to impaired local production of IGF1. Our results conclusively identify a novel autophagy-dependent pathway that regulates nSC behavior and indicate that autophagy is required for skeletal muscle development in the neonatal phase. Abbreviations: AKT/protein kinase B: Thymoma viral proto-oncogene; ASCs: adult stem cells; ATF4: activating transcription factor 4; ATG7: autophagy related 7; BAT: brown adipose tissue; BMP: bone morphogenetic protein; CEBPB: CCAAT/enhancer binding protein (C/EBP), beta; CSA: cross sectional area; CTNNB1: catenin (cadherin associated protein), beta 1; DDIT3: DNA-damage inducible transcript 3; DM: differentiation medium; E: embryonic stage; EIF2AK3/PERK; EIF4EBP1: eukaryotic translation initiation factor 2 alpha kinase 3; eukaryotic translation initiation factor 4E binding protein 1; ER: endoplasmic reticulum; FGF21: fibroblast growth factor 21; GH: growth hormone; GHR: growth hormone receptor; HSCs: hematopoietic stem cells; IGF1: insulin-like growth factor 1; ITGAM: integrin alpha M; KEAP1: kelch-like ECH-associated protein 1; LY6A/Sca-1; MAP1LC3: lymphocyte antigen 6 complex, locus A; microtubule-associated protein 1 light chain 3; MAPK1/ERK2: mitogen-activated protein kinase 1; MAPK3/ERK1: mitogen-activated protein kinase 3; miRNAs: microRNAs; MSCs: mesenchymal stem cells; MTOR: mechanistic target of rapamycin kinase; mtUPR: mitochondrial unfolded protein response; MYF5: myogenic factor 5; MYH: myosin, heavy polypeptide; MYOD1: myogenic differentiation 1; MYOG: myogenin; NFE2L2: nuclear factor, erythroid derived 2, like 2; nSC: neonatal satellite cells; NSCs: neuronal stem cells; P: postnatal day; PAX7: paired box 7; PECAM1: platelet/endothelial cell adhesion molecule 1; PPARG: peroxisome proliferator activated receptor gamma; PTPRC: protein tyrosine phosphatase, receptor type, C; ROS: reactive oxygen species; RPS6: ribosomal protein S6; SCs: adult satellite cells; SQSTM1: sequestosome 1; STAT5: signal transducer and activator of transcription 5; TGFB1: transforming growth factor beta 1; WAT: white adipose tissue; WT: wild type.

VCP/p97 (valosin containing protein) is a key regulator of cellular proteostasis. It orchestrates protein turnover and quality control in vivo, processes fundamental for proper cell function. In humans, mutations in VCP lead to severe myo- and neuro-degenerative disorders such as inclusion body myopathy with Paget disease of the bone and frontotemporal dementia (IBMPFD), amyotrophic lateral sclerosis (ALS) or and hereditary spastic paraplegia (HSP). We analyzed here the in vivo role of Vcp and its novel interactor Washc4/Swip (WASH complex subunit 4) in the vertebrate model zebrafish (Danio rerio). We found that targeted inactivation of either Vcp or Washc4, led to progressive impairment of cardiac and skeletal muscle function, structure and cytoarchitecture without interfering with the differentiation of both organ systems. Notably, loss of Vcp resulted in compromised protein degradation via the proteasome and the macroautophagy/autophagy machinery, whereas Washc4 deficiency did not affect the function of the ubiquitin-proteasome system (UPS) but caused ER stress and interfered with autophagy function in vivo. In summary, our findings provide novel insights into the in vivo functions of Vcp and its novel interactor Washc4 and their particular and distinct roles during proteostasis in striated muscle cells.

Summary: Protein aggregates and cytoplasmic vacuolization are major hallmarks of multisystem proteinopathies (MSPs) that lead to muscle weakness. Here, we identify METTL21C as a skeletal muscle-specific lysine methyltransferase. Insertion of a β-galactosidase cassette into the Mettl21c mouse locus revealed that METTL21C is specifically expressed in MYH7-positive skeletal muscle fibers. Ablation of the Mettl21c gene reduced endurance capacity and led to age-dependent accumulation of autophagic vacuoles in skeletal muscle. Denervation-induced muscle atrophy highlighted further impairments of autophagy-related proteins, including LC3, p62, and cathepsins, in Mettl21c−/− muscles. In addition, we demonstrate that METTL21C interacts with the ATPase p97 (VCP), which is mutated in various human MSP conditions. We reveal that METTL21C trimethylates p97 on the Lys315 residue and found that loss of this modification reduced p97 hexamer formation and ATPase activity in vivo. We conclude that the methyltransferase METTL21C is an important modulator of protein degradation in skeletal muscle under both normal and enhanced protein breakdown conditions. : Wiederstein et al. describe the skeletal muscle methyltransferase Mettl21c. They found that ablation of Mettl21c in mice results in muscle weakness and disturbance of the protein degradation machinery. Those changes are hallmarks of multisystem proteinopathies. They demonstrate that Mettl21c modulates p97 activity, which is frequently mutated in human patients with muscle weakness. Keywords: methyltransferases, skeletal muscle, p97, atrophy, autophagy

Aim: Neuromuscular junction (NMJ) represents the morphofunctional interface between muscle and nerve. Several chronic pathologies such as aging and neurodegenerative diseases, including muscular dystrophy and amyotrophic lateral sclerosis, display altered NMJ and functional denervation. However, the triggers and the molecular mechanisms underlying the dismantlement of NMJ remain unclear. Results: Here we provide evidence that perturbation in redox signaling cascades, induced by muscle-specific accumulation of mutant SOD1G93A in transgenic MLC/SOD1G93A mice, is causally linked to morphological alterations of the neuromuscular presynaptic terminals, high turnover rate of acetylcholine receptor, and NMJ dismantlement. The analysis of potential molecular mechanisms that mediate the toxic activity of SOD1G93A revealed a causal link between protein kinase Cθ (PKCθ) activation and NMJ disintegration. Innovation: The study discloses the molecular mechanism that triggers functional denervation associated with the toxic activity of muscle SOD1G93A expression and suggests the possibility of developing a new strategy to counteract age- and pathology-associated denervation based on pharmacological inhibition of PKCθ activity. Conclusions: Collectively, these data indicate that muscle-specific accumulation of oxidative damage can affect neuromuscular communication and induce NMJ dismantlement through a PKCθ-dependent mechanism

In this commentary we discuss new findings presented by Shang et al. regarding the role of macrophage-derived glutamine in skeletal muscle repair. Loss-of-function of glutamate dehydrogenase in macrophages led to an upregulation of glutamine synthesis which sustained glutamine levels in muscle tissue and facilitated satellite cell proliferation and differentiation.

© 2018 Taylor and Francis Group, LLC. In yeast, Tom22, the central component of the TOMM (translocase of outer mitochondrial membrane) receptor complex, is responsible for the recognition and translocation of synthesized mitochondrial precursor proteins, and its protein kinase CK2-dependent phosphorylation is mandatory for TOMM complex biogenesis and proper mitochondrial protein import. In mammals, the biological function of protein kinase CSNK2/CK2 remains vastly elusive and it is unknown whether CSNK2-dependent phosphorylation of TOMM protein subunits has a similar role as that in yeast. To address this issue, we used a skeletal muscle-specific Csnk2b/Ck2β-conditional knockout (cKO) mouse model. Phenotypically, these skeletal muscle Csnk2b cKO mice showed reduced muscle strength and abnormal metabolic activity of mainly oxidative muscle fibers, which point towards mitochondrial dysfunction. Enzymatically, active muscle lysates from skeletal muscle Csnk2b cKO mice phosphorylate murine TOMM22, the mammalian ortholog of yeast Tom22, to a lower extent than lysates prepared from controls. Mechanistically, CSNK2-mediated phosphorylation of TOMM22 changes its binding affinity for mitochondrial precursor proteins. However, in contrast to yeast, mitochondrial protein import seems not to be affected in vitro using mitochondria isolated from muscles of skeletal muscle Csnk2b cKO mice. PINK1, a mitochondrial health sensor that undergoes constitutive import under physiological conditions, accumulates within skeletal muscle Csnk2b cKO fibers and labels abnormal mitochondria for removal by mitophagy as demonstrated by the appearance of mitochondria-containing autophagosomes through electron microscopy. Mitophagy can be normalized by either introduction of a phosphomimetic TOMM22 mutant in cultured myotubes, or by in vivo electroporation of phosphomimetic Tomm22 into muscles of mice. Importantly, transfection of the phosphomimetic Tomm22 mutant in muscle cells with ablated Csnk2b restored their oxygen consumption rate comparable to wild-type levels. In sum, our data show that mammalian CSNK2-dependent phosphorylation of TOMM22 is a critical switch for mitophagy and reveal CSNK2-dependent physiological implications on metabolism, muscle integrity and behavior.

The muscle-specific ubiquitin ligase atrogin-1 (MAFbx) has been identified as a critical regulator of pathologic and physiological cardiac hypertrophy; it regulates these processes by ubiquitinating transcription factors [nuclear factor of activated T-cells and forkhead box O (FoxO) 1/3]. However, the role of atrogin-1 in regulating transcription factors in aging has not previously been described. Atrogin-1 cardiomyocyte-specific transgenic (Tg(+)) adult mice (α-major histocompatibility complex promoter driven) have normal cardiac function and size. Herein, we demonstrate that 18-month-old atrogin-1 Tg(+) hearts exhibit significantly increased anterior wall thickness without functional impairment versus wild-type mice. Histologic analysis at 18 months revealed atrogin-1 Tg(+) mice had significantly less fibrosis and significantly greater nuclei and cardiomyocyte cross-sectional analysis. Furthermore, by real-time quantitative PCR, atrogin-1 Tg(+) had increased Col 6a4, 6a5, 6a6, matrix metalloproteinase 8 (Mmp8), and Mmp9 mRNA, suggesting a role for atrogin-1 in regulating collagen deposits and MMP-8 and MMP-9. Because atrogin-1 Tg(+) mice exhibited significantly less collagen deposition and protein levels, enhanced Mmp8 and Mmp9 mRNA may offer one mechanism by which collagen levels are kept in check in the aged atrogin-1 Tg(+) heart. In addition, atrogin-1 Tg(+) hearts showed enhanced FoxO1/3 activity. The present study shows a novel link between atrogin-1–mediated regulation of FoxO1/3 activity and reduced collagen deposition and fibrosis in the aged heart. Therefore, targeting FoxO1/3 activity via the muscle-specific atrogin-1 ubiquitin ligase may offer a muscle-specific method to modulate aging-related cardiac fibrosis.

Background There is no clear consensus as to the optimal method of entry in laparoscopic renal surgery and no reports have compared them in Urology. To analyze contemporary practice patterns in entry technique and port placement for laparoscopic kidney surgery. Methods We identified 60 high volume urological laparoscopic centers. A purpose-built questionnaire was sent to surgeons. The survey included 22 questions regarding access techniques and port configuration during laparoscopic kidney surgery. Data on were collected and retrospectively analyzed. Concordance among port configurations was assessed using Cohen's Kappa statistics. Results The survey was sent to 60 surgeons and completed by 32 of them. Surgical procedures included were laparoscopic radical nephrectomy (1177 LRN/year) and laparoscopic partial nephrectomy (1047 LPN/year). The transperitoneal route was preferred (85%). Hasson technique was used for the access in 55% of the cases. Patient lateral recumbent position is the most frequently used during the port placement (41%). Although there is a high variability in the port positioning among the surgeons, in more than 90% of cases it was found a specific concordance in triangulation of optics and operating trocars. There were no significant differences between port configuration in LRN and LPN. Limitations include retrospective design and limited sample. Conclusions A standard port configuration has not been previously reported in urological literature. Our study suggests that the transperitoneal approach, the Hasson technique and a specific triangulation of optics and operating trocars have a significant concordance in some high volume laparoscopic urologic centers.

The ability of whole body vibration (WBV) to increase energy expenditure (EE) has been investigated to some extent in the past using short-term single exercises or sets of single exercises. However, the current practice in WBV training for fitness is based on the execution of multiple exercises during a WBV training session for a period of at least 20 min; nevertheless, very limited and inconsistent data are available on EE during long term WBV training session. This crossover study was designed to demonstrate, in an adequately powered sample of participants, the ability of WBV to increase the metabolic cost of exercise vs. no vibration over the time span of a typical WBV session for fitness (20 min). Twenty-two physically active young males exercised on a vibration platform (three identical sets of six different exercises) using an accelerometer-verified vibration stimulus in both the WBV and no vibration condition. Oxygen consumption was measured with indirect calorimetry and expressed as area under the curve (O2(AUC)). Results showed that, in the overall 20-min training session, WBV increased both the O2(AUC) and the estimated EE vs. no vibration by about 22% and 20%, respectively (P

Aims Pompe disease is an autosomal recessive lysosomal storage disorder resulting from deficiency of acid α-glucosidase (GAA) enzyme. Histopathological hallmarks in skeletal muscle tissue are fiber vacuolization and autophagy. Since 2006, ERT is the only approved treatment with human recombinant GAA alglucosidase alfa. We designed a study to examine ERT-related skeletal muscle changes in 18 modestly to moderately affected LOPD patients along with the relationship between morphological/biochemical changes and clinical outcomes. Treatment duration was short-to-long term. Methods We examined muscle biopsies from 18 LOPD patients at both histopathological and biochemical level. All patients underwent two muscle biopsies, before and after ERT administration respectively. The study is partially retrospective because the first biopsies were taken before the study was designed whereas the second biopsy was always performed after at least six months of ERT administration. Results After ERT, 15 out of 18 patients showed improved 6MWT (p=0.0007) and most of them achieved respiratory stabilization. Pre-treatment muscle biopsies disclosed marked histopathological variability, ranging from an almost normal pattern to a severe vacuolar myopathy. After treatment, we detected morphological improvement in 15 patients and worsening in 3 patients. Post-ERT GAA enzymatic activity was mildly increased compared with pre-treatment levels in all patients. Protein levels of the mature enzyme increased in 14 of the 18 patients (mean increase = +35%; p