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Introduction: It has been hypothesized that leptin's modifications in Eating Disorders (EDs) aren't just a consequence of malnutrition, but they also represent a maintenance factor for pathological reward-related eating behaviours, such as dietary restraint and compensatory physical exercise, due to the role of leptin in the regulation of dopaminergic tone in brain reward circuits. Objectives: To evaluate the role of leptin in maintaining dietary restriction and compensatory exercise in subjects with Anorexia Nervosa (AN) and Bulimia Nervosa (BN). Methods: 56 patients were enrolled, 39 with AN and 17 with BN; 43 healthy controls (HCs) were also recruited. Blood leptin levels were measured and psychometric tests for the evaluation of general psychopathology (SCL-90-R) and of EDs specific pathological behaviours and psychopathology (EDE-Q) were administered. Results: Lower leptin levels were found in subjects withANand BN as compared with HCs. Leptin was negatively associated with the frequency of compensatory exercise, even when adjusting for age and BMI, but only in subjects with EDs. The moderated mediation models confirmed the role of leptin as a complete mediator for the relationship between low BMI and increased dietary restriction and compensatory exercise, but only for high levels of EDs specific psychopathology. Conclusions: These results suggest that for subjects with severe EDs psychopathology a low BMI maintains dietary restraint and compensatory exercise through a decrease in leptin levels, probably because of the role of leptin in reward circuits and due to the presence in EDs patients of an aberrant food-related reward that makes losing weight instead of feeding the desirable aim. [ABSTRACT FROM AUTHOR]

Growth differentiation factor 15 (GDF15) is a TGF-β superfamily member involved in diverse physiological and pathological processes. It is expressed in various tissues and its circulating levels rise during exercise, aging, pregnancy, and conditions such as cancer, cardiovascular disease, and infections. The biological activities of GDF15, including anorexia and cachexia, are primarily mediated through the GFRAL receptor, localized in the brainstem and functioning via RET co-receptor recruitment. This signaling is crucial for energy homeostasis and nausea induction. Recent studies suggest a broader GFRAL distribution, potentially explaining GDF15's distinct roles. These findings sparked interest in leveraging GDF15-GFRAL pathways for therapeutic development. Two primary strategies include GDF15 analogues as GFRAL agonists for obesity treatment and GDF15-derived peptides as antagonists to counteract cancer-induced cachexia and related disorders. This review highlights advancements in understanding GDF15-GFRAL signaling and its implications, summarizing bioactive GDF15-derived molecules, their pharmacological applications, and offering insights into novel treatment avenues for GDF15-associated conditions., (© 2025 The Authors. ChemMedChem published by Wiley-VCH GmbH.)

Background: Growth Differentiation Factor 15 (GDF15) has been described as influencing skeletal physiology. Nevertheless, no systematic appraisal of the effect of GDF15 on skeletal muscle tissues has been developed to the present day., Objective: The aim of the present work was to review the evidence on the topic., Methods: In this preregistered systematic review (https://osf.io/wa8xr), articles were retrieved from MEDLINE/PubMed, EMBASE, and WebOfScience. Inclusion criteria comprised studies on humans or animal models, assessment of peripheral or local tissue GDF15 concentrations, as well as the direct expression of GDF15 in skeletal muscle, and direct or indirect correlates of GDF15 with physical activity/ sarcopenia/trophism/ function., Results: A total of 646 studies were retrieved, and 144 finally included. Molecular inducers or inhibitors of GDF15 in skeletal muscle tissues were described. GDF15 was reported to promote skeletal muscle health, metabolic homeostasis, and overall physical conditioning. In pathology, GDF15 seems to be correlated to the degree of muscle impairment and mitochondrial stress. GDF15 has also been described as having the potential to stratify patients based on clinical prognosis and functional outcome., Conclusion: A hormetic hypothesis for GDF15 on skeletal muscle was proposed. In fact, GDF15 exhibited beneficial effects when expressed at high levels facing acute stressors (i.e., "myoprotection"). Conversely, GDF15 exhibited maladaptive effects, such as chronic low-grade inflammation, when chronically expressed in pathological processes (e.g., obesity, aging). GDF15 may be a potential molecular target for disease-modifying interventions. The current review underscores the need for further research on GDF15 to elucidate its therapeutic potential across different pathological states. The study protocol, registered before data collection and analysis, can be retrieved at https://osf.io/wa8xr. It should be noted that the study deviated from the protocol after peer review, including other electronic databases beyond MEDLINE/PubMed alone., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Holotomography (HT) is a cutting-edge fast live-cell quantitative label-free imaging technique. Based on the principle of quantitative phase imaging, it combines holography and tomography to record a three-dimensional map of the refractive index, used as intrinsic optical and quantitative imaging contrast parameter of biological samples, at a sub-micrometer spatial resolution. In this study HT has been employed for the first time to analyze the changes of fibroblasts differentiating towards myofibroblasts - recognized as the main cell player of fibrosis - when cultured in vitro with the pro-fibrotic factor, namely transforming growth factor-β1. In parallel, F-actin, vinculin, α-smooth muscle actin, phospho-myosin light chain 2, type-1 collagen, peroxisome proliferator-activated receptor-gamma coactivator-1α expression and mitochondria were evaluated by confocal laser scanning microscopy. Plasmamembrane passive properties and transient receptor potential canonical channels' currents were also recorded by whole-cell patch-clamp. The fluorescence images and electrophysiological results have been compared to the data obtained by HT and their congruence has been discussed. HT turned out to be a valid approach to morphologically distinguish fibroblasts from well differentiated myofibroblasts while obtaining objective measures concerning volume, surface area, projection area, surface index and dry mass (i.e., the mass of the non-aqueous content inside the cell including proteins and subcellular organelles) of the entire cell, nuclei and nucleoli with the major advantage to monitor outer and inner features in living cells in a non-invasive, rapid and label-free approach. HT might open up new research opportunities in the field of fibrotic diseases. RESEARCH HIGHLIGHTS: Holotomography (HT) is a label-free laser interferometric imaging technology exploiting the intrinsic optical property of cells namely refractive index (RI) to enable a direct imaging and analysis of whole cells or intracellular organelles. HT turned out a valid approach to distinguish morphological features of living unlabeled fibroblasts from differentiated myofibroblasts. HT provided quantitative information concerning volume, surface area, projection area, surface index and dry mass of the entire fibroblasts/myofibroblasts, nuclei and nucleoli., (© 2024 Wiley Periodicals LLC.)

Hypoxia-inducible factor (HIF)-1α represents an oxygen-sensitive subunit of HIF transcriptional factor, which is usually degraded in normoxia and stabilized in hypoxia to regulate several target gene expressions. Nevertheless, in the skeletal muscle satellite stem cells (SCs), an oxygen level-independent regulation of HIF-1α has been observed. Although HIF-1α has been highlighted as a SC function regulator, its spatio-temporal expression and role during myogenic progression remain controversial. Herein, using biomolecular, biochemical, morphological and electrophysiological analyses, we analyzed HIF-1α expression, localization and role in differentiating murine C2C12 myoblasts and SCs under normoxia. In addition, we evaluated the role of matrix metalloproteinase (MMP)-9 as an HIF-1α effector, considering that MMP-9 is involved in myogenesis and is an HIF-1α target in different cell types. HIF-1α expression increased after 24/48 h of differentiating culture and tended to decline after 72 h/5 days. Committed and proliferating mononuclear myoblasts exhibited nuclear HIF-1α expression. Differently, the more differentiated elongated and parallel-aligned cells, which are likely ready to fuse with each other, show a mainly cytoplasmic localization of the factor. Multinucleated myotubes displayed both nuclear and cytoplasmic HIF-1α expression. The MMP-9 and MyoD (myogenic activation marker) expression synchronized with that of HIF-1α, increasing after 24 h of differentiation. By means of silencing HIF-1α and MMP-9 by short-interfering RNA and MMP-9 pharmacological inhibition, this study unraveled MMP-9's role as an HIF-1α downstream effector and the fact that the HIF-1α/MMP-9 axis is essential in morpho-functional cell myogenic commitment.

Mechanomimetic materials are particularly attractive for modeling in vitro fibroblast to myofibroblast (Myof) transition, a key process in the physiological repair of damaged tissue, and recognized as the core cellular mechanism of pathological fibrosis in different organs. In vivo, mechanical stimuli from the extracellular matrix (ECM) are crucial, together with cell-cell contacts and the pro-fibrotic transforming growth factor (TGF)-β1, in promoting fibroblast differentiation. Here, we explore the impact of hydrogels made by polyacrylamide with different composition on fibroblast behavior. By appropriate modulation of the hydrogel composition (e.g. adjusting the crosslinker content), we produce and fully characterize three kinds of scaffolds with different Young modulus (E). We observe that soft hydrogels (E < 1 kPa) induced fibroblast differentiation better than stiffer ones, also in the absence of TGF-β1. This study provides a readily accessible biomaterial platform to promote Myof generation. The easy approach used and the commercial availability of the monomers make these hydrogels suitable to a wide range of biomedical applications combined with high reproducibility and simple preparation protocols., Competing Interests: Declaration of competing interest The authors declare no competing interest., (Copyright © 2023. Published by Elsevier B.V.)

Adipokines are peptide hormones produced by the adipose tissue involved in several biological functions. Among adipokines, adiponectin (ADPN) has antidiabetic and anti-inflammatory properties. It can also modulate food intake at central and peripheral levels, acting on hypothalamus and facilitating gastric relaxation. ADPN exerts its action interacting with two distinct membrane receptors and triggering some well-defined signaling cascades. The ceramidase activity of ADPN receptor has been reported in many tissues: it converts ceramide into sphingosine. In turn, sphingosine kinase (SK) phosphorylates it into sphingosine-1 phosphate (S1P), a crucial mediator of many cellular processes including contractility. Using a multidisciplinary approach that combined biochemical, electrophysiological and morphological investigations, we explored for the first time the possible role of S1P metabolism in mediating ADPN effects on the murine gastric fundus muscle layer. By using a specific pharmacological inhibitor of SK2, we showed that ADPN affects smooth muscle cell membrane properties and contractile machinery via SK2 activation in gastric fundus, adding a piece of knowledge to the action mechanisms of this hormone. These findings help to identify ADPN and its receptors as new therapeutic targets or as possible prognostic markers for diseases with altered energy balance and for pathologies with fat mass content alterations.

Pheochromocytomas/paragangliomas (PPGLs) are neuroendocrine tumours, mostly resulting from mutations in predisposing genes. Mutations of succinate dehydrogenase (SDH) subunit B (SDHB) are associated with high probability of metastatic disease. Since bioelectrical properties and signalling in cancer are an emerging field, we investigated the metabolic, functional and electrophysiological characteristics in human succinate dehydrogenase subunit B (SDHB)-deficient pheochromocytoma cells. These cells exhibited reduced SDH function with elevated succinate-to-fumarate ratio and reduced intracellular ATP levels. The analysis of membrane passive properties revealed a more hyperpolarized membrane potential and a lower cell capacitance of SDHB-deficient cells compared to the parental ones. These bioelectrical changes were associated with reduced proliferation and adhesion capacity of SDHB-deficient cells. Only in SDHB-deficient cells, we also observed an increased amplitude of potassium currents suggesting an activation of ATP-sensitive potassium channels (KATP). Indeed, exposure of the SDHB-deficient cells to glibenclamide, a specific KATP inhibitor, or to ATP caused normalization of potassium current features and altered proliferation and adhesion. In this work, we show for the first time that reduced intracellular ATP levels in SDHB-deficient chromaffin cells impaired cell bioelectrical properties, which, in turn, are associated with an increased cell aggressiveness. Moreover, we first ever demonstrated that glibenclamide not only reduced the outward potassium currents in SDHB-deficient cells but increased their growth capacity, reduced their ability to migrate and shifted their phenotype towards one more similar to that of parental one.

The neuroendocrine control of reproduction is strictly coordinated at the central level by the pulsatile release of gonadotropin-releasing hormone (GnRH) by the hypothalamic GnRH neurons. Alterations of the GnRH-network, especially during development, lead to long-term reproductive and systemic consequences, also causing infertility. Recent evidence shows that benzo[a]pyrene (BaP), a diffuse pollutant that can play a role as an endocrine disruptor, affects gonadal function and gamete maturation, whereas data demonstrating its impact at hypothalamic level are very scarce. This study investigated the effects of BaP (10 μM) in a primary cell culture isolated from the human fetal hypothalamus (hfHypo) and exhibiting a clear GnRH neuron phenotype. BaP significantly decreased gene and protein expression of both GnRH and kisspeptin receptor (KISS1R), the master regulator of GnRH neuron function. Moreover, BaP exposure increased phospho-ERK1/2 signaling, a well-known mechanism associated with KISS1R activation. Interestingly, BaP altered the electrophysiological membrane properties leading to a significant depolarizing effect and it also significantly increased GnRH release, with both effects being not affected by kisspeptin addition. In conclusion, our findings demonstrate that BaP may alter GnRH neuron phenotype and function, mainly interfering with KISS1R signaling and GnRH secretion and therefore with crucial mechanisms implicated in the central neuroendocrine control of reproduction., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Adiponectin (ADPN), a hormone produced by adipose tissue, facilitates gastric relaxation and can be a satiety signal in the network connecting peripheral organs and the central nervous system for feeding behavior control. Here, we performed preclinical research by morpho-functional analyses on murine gastric fundus smooth muscle to add insights into the molecular mechanisms underpinning ADPN action. Moreover, we conducted a clinical study to evaluate the potential use of ADPN as a biomarker for eating disorders (ED) based on the demonstrated gastric alterations and hormone level fluctuations that are often associated with ED. The clinical study recruited patients with ED and healthy controls who underwent blood draws for ADPN dosage and psychopathology evaluation tests. The findings of this basic research support the ADPN relaxant action, as indicated by the smooth muscle cell membrane pro-relaxant effects, with mild modifications of contractile apparatus and slight inhibitory effects on gap junctions. All of these actions engaged the ADPN/nitric oxide/guanylate cyclase pathway. The clinical data failed to unravel a correlation between ADPN levels and the considered ED, thus negating the potential use of ADPN as a valid biomarker for ED management for the moment. Nevertheless, this adipokine can modulate physiological eating behavior, and its effects deserve further investigation.

Cachexia is a systemic disease associated with several pathologies, including cancer, that leads to excessive weight loss due to enhanced protein degradation. Previously, we showed that cachectic features in myotubes are provoked by a metabolic shift toward lactic fermentation. Our previous results led us to hyphotesise that increasing pyruvate concentration could impede the metabolic modifications responsible for induction of cachexia in myotubes. Here, we demonstrated that the addition of sodium pyruvate in conditioned media from CT26 colon cancer cells (CM CT26) prevents the onset of either phenotypic and metabolic cachectic features. Myotubes treated with CM CT26 containing sodium pyruvate show a phenotype similar to the healthy counterpart and display lactate production, oxygen consumption, and pyruvate dehydrogenase activity as control myotubes. The use of the Mitochondrial Pyruvate Carrier inhibitor UK5099, highlights the importance of mitochondrial pyruvate amount in the prevention of cachexia. Indeed, UK5099-treated myotubes show cachectic features as those observed in myotubes treated with CM CT26. Finally, we found that sodium pyruvate is able to decrease STAT3 phosphorylation level, a signaling pathway involved in the induction of cachexia in myotubes. Collectively, our results show that cachexia in myotubes could be prevented by the utilization of sodium pyruvate which impedes the metabolic modifications responsible for the acquisition of the cachectic features., (© 2022 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Resistin, among its several actions, has been reported to exert central anorexigenic effects in rodents. Some adipokines which centrally modulate food intake have also been reported to affect the activity of gastric smooth muscle, whose motor responses represent a source of peripheral signals implicated in the control of the hunger-satiety cycle through the gut-brain axis. On this basis, in the present experiments, we investigated whether resistin too could affect the mechanical responses in the mouse longitudinal gastric fundal strips. Electrical field stimulation (EFS) elicited tetrodotoxin- and atropine-sensitive contractile responses. Resistin reduced the amplitude of the EFS-induced contractile responses. This effect was no longer detected in the presence of L-NNA, a nitric oxide (NO) synthesis inhibitor. Resistin did not influence the direct muscular response to methacholine. In the presence of carbachol and guanethidine, EFS elicited inhibitory responses whose amplitude was increased by resistin. L-NNA abolished the inhibitory responses evoked by EFS, indicating their nitrergic nature. In the presence of L-NNA, resistin did not have any effect on the EFS-evoked inhibitory responses. Western blot and immunofluorescence analysis revealed a significant increase in neuronal nitric oxide synthase (nNOS) expression in neurons of the myenteric plexus following resistin exposure. In conclusion, the present results offer the first evidence that resistin acts on the gastric fundus, likely through a modulatory action on the nitrergic neurotransmission., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Idrizaj, Garella, Nistri, Squecco and Baccari.)

Background: Adiponectin (Adn), released by adipocytes and other cell types such as skeletal muscle, has insulin-sensitizing and anti-inflammatory properties. Sphingosine 1-phosphate (S1P) is reported to act as effector of diverse biological actions of Adn in different tissues. S1P is a bioactive sphingolipid synthesized by the phosphorylation of sphingosine catalyzed by sphingosine kinase (SK) 1 and 2. Consolidated findings support the key role of S1P in the biology of skeletal muscle., Methods and Results: Here we provide experimental evidence that S1P signalling is modulated by globular Adn treatment being able to increase the phosphorylation of SK1/2 as well as the mRNA expression levels of S1P 4 in C2C12 myotubes. These findings were confirmed by LC-MS/MS that showed an increase of S1P levels after Adn treatment. Notably, the involvement of S1P axis in Adn action was highlighted since, when SK1 and 2 were inhibited by PF543 and ABC294640 inhibitors, respectively, not only the electrophysiological changes but also the increase of oxygen consumption and of aminoacid levels induced by the hormone, were significantly inhibited., Conclusion: Altogether, these findings show that S1P biosynthesis is necessary for the electrophysiological properties and oxidative metabolism of Adn in skeletal muscle cells.

New Findings: What is the central question of this study? It is a challenge to discover effective therapies for fibrosis. Increasing evidence supports the antifibrotic potential of platelet-rich plasma (PRP) as a source of bioactive molecules, such as vascular endothelial growth factor (VEGF)-A. However, the effects and mechanisms of action of PRP need to be clarified. What is the main finding and its importance? This report clarifies the mechanisms mediating the antifibrotic action of PRP, strengthening the role of VEGF-A/VEGF receptor, and identifies gap junction currents and connexin 43 as novel targets of this pathway in the fibroblast-to-myofibroblast transition induced by the transforming growth factor-β1., Abstract: Despite increasing experimental evidence, the antifibrotic potential of platelet-rich plasma (PRP) remains controversial, and its mechanisms of action are not fully clarified. This short report extends our previous research on the capability of PRP to prevent the in vitro differentiation of fibroblasts toward myofibroblasts, the key effectors of fibrosis, induced by the profibrotic agent transforming growth factor-β1 (TGF-β1). In particular, we focused on the involvement of signalling mediated by vascular endothelial growth factor (VEGF)-A/VEGF receptor (VEGFR) in the PRP-induced fibroblast response, highlighting gap junction features. Electrophysiological and morphological analyses revealed that PRP hindered morphofunctional differentiation of both murine NIH/3T3 and human primary adult skin fibroblasts toward myofibroblasts as judged by the analysis of membrane phenomena, α-smooth muscle actin and vinculin expression and cell morphology. Neutralization of VEGF-A by blocking antibodies or pharmacological inhibition of VEGFR by KRN633 in TGF-β1-treated fibroblasts prevented the PRP-promoted effects, such as the reduction of voltage-dependent transjunctional currents in cell pairs and a decreased expression of connexin 43, the typical connexin isoform forming voltage-dependent connexons. The role of VEGF-A in inhibiting these events was confirmed by treating TGF-β1-stimulated fibroblasts with soluble VEGF-A. The results obtained when cells were differentiated using KRN633 alone suggest an antagonistic cross-talk between TGF-β1 and VEGFR. In conclusion, this study identifies, for the first time, gap junction currents as crucial targets in the VEGF-A/VEGFR-mediated antifibrotic pathway and provides new insights into mechanisms behind the action of PRP in preventing differentiation of fibroblasts to myofibroblasts., (© 2021 The Authors. Experimental Physiology © 2021 The Physiological Society.)

Development of biological tissues in vitro is not a trivial task and requires the correct maturation of the selected cell line. To this aim, many attempts were done mainly by mimicking the biological environment using micro/nanopatterned or stimulated scaffolds. However, the obtainment of functional tissues in vitro is still far from being achieved. In contrast with the standard methods, we here present an easy approach for the maturation of myotubes toward the reproduction of muscular tissue. By using liquid crystalline networks with different stiffness and molecular alignment, we demonstrate how the material itself can give favorable interactions with myoblasts helping a correct differentiation. Electrophysiological studies demonstrate that myotubes obtained on these polymers have more adult-like morphology and better functional features with respect to those cultured on standard supports. The study opens to a platform for the differentiation of other cell lines in a simple and scalable way., Competing Interests: The authors declare no competing interests., (© 2021 The Author(s).)

The widespread environmental pollutant 2,2',4,4',5,5'-hexachlorobiphenyl (PCB153) is a non-dioxin-like toxicant. It is a potential carcinogen compound able to induce gap junction (GJ) intercellular communication impairment, probably the first non-genomic event leading to tumor promotion. Although PCBs have been known for many years, the molecular mode of PCB153 action is still unclear. Recent studies from our research group have shown that the toxicant elicits a transient modulation of connexin (Cx) 43-formed GJs in hepatic stem-like WB-F344 cells involving sphingosine 1-phosphate (S1P) path. Taking into account that other strictly related bioactive sphingolipids, such as ceramide (Cer), may have different effects from S1P, here we aim to clarify the signaling paths engaged by PCB153 in the control of GJs, focusing primarily on the role of Cer. Accordingly, we have achieved a combined biomolecular and electrophysiological analysis of GJs in cultured WB-F344 cells treated with PCB153 at different time points. We have found that the toxicant elicited a time-dependent regulation of GJs formed by different Cx isoforms, through a transient modulation of Cer/Cer kinase (CerK) axis and, in turn, of protein phosphatase 2A (PP2A). Our new findings demonstrate the existence of a specific molecular mechanism downstream to Cer, which distinctly affects the voltage-dependent and -independent GJs in liver stem-like cells, and open new opportunities for the identification of additional potential targets of these environmental toxicants.

Objective: The recent conceptualization of ghrelin as a stress hormone suggested that its chronic alterations may have a role in maintaining overeating behaviors in subjects with eating disorders (EDs) reporting childhood traumatic experiences. The aim of this study was to investigate the alterations of ghrelin levels in patients with EDs, their associations with early trauma, binge and emotional eating, and possible moderation/mediation models., Method: Sixty-four patients with EDs and 42 healthy controls (HCs) had their plasma ghrelin levels measured and completed questionnaires evaluating general and ED-specific psychopathology, emotional eating, and childhood traumatic experiences., Results: Participants with anorexia nervosa had higher ghrelin levels than HCs in body mass index (BMI)-adjusted comparisons. Moreover, patients reporting a history of childhood trauma had higher ghrelin levels. Childhood sexual abuse (CSA), BMI, and self-induced vomiting were independent predictors of ghrelin levels. Moderation analyses showed that ghrelin levels were associated with binge and emotional eating only for higher levels of childhood trauma. Elevated ghrelin was a significant mediator for the association of CSA with binge eating., Conclusions: These results support the hypothesis that chronic alterations in ghrelin levels following childhood traumatic experiences could represent a neurobiological maintaining factor of pathological overeating behaviors in EDs., (© 2021 The Authors. European Eating Disorders Review published by Eating Disorders Association and John Wiley & Sons Ltd.)

Bone marrow-mesenchymal stem/stromal cells (MSCs) may offer promise for skeletal muscle repair/regeneration. Growing evidence suggests that the mechanisms underpinning the beneficial effects of such cells in muscle tissue reside in their ability to secrete bioactive molecules (secretome) with multiple actions. Hence, we examined the effects of MSC secretome as conditioned medium (MSC-CM) on ex vivo murine extensor digitorum longus muscle injured by forced eccentric contraction (EC). By combining morphological (light and confocal laser scanning microscopies) and electrophysiological analyses we demonstrated the capability of MSC-CM to attenuate EC-induced tissue structural damages and sarcolemnic functional properties' modifications. MSC-CM was effective in protecting myofibers from apoptosis, as suggested by a reduced expression of pro-apoptotic markers, cytochrome c and activated caspase-3, along with an increase in the expression of pro-survival AKT factor. Notably, MSC-CM also reduced the EC-induced tissue redistribution and extension of telocytes/CD34 + stromal cells, distinctive cells proposed to play a "nursing" role for the muscle resident myogenic satellite cells (SCs), regarded as the main players of regeneration. Moreover, it affected SC functionality likely contributing to replenishment of the SC reservoir. This study provides the necessary groundwork for further investigation of the effects of MSC secretome in the setting of skeletal muscle injury and regenerative medicine.

Skeletal muscle atrophy is characterized by a decrease in muscle mass causing reduced agility, increased fatigability and higher risk of bone fractures. Inflammatory cytokines, such as tumor necrosis factor-alpha (TNFα), are strong inducers of skeletal muscle atrophy. The bioactive sphingolipid sphingosine 1-phoshate (S1P) plays an important role in skeletal muscle biology. S1P, generated by the phosphorylation of sphingosine catalyzed by sphingosine kinase (SK1/2), exerts most of its actions through its specific receptors, S1P 1-5 . Here, we provide experimental evidence that TNFα induces atrophy and autophagy in skeletal muscle C2C12 myotubes, modulating the expression of specific markers and both active and passive membrane electrophysiological properties. NMR-metabolomics provided a clear picture of the deep remodelling of skeletal muscle fibre metabolism induced by TNFα challenge. The cytokine is responsible for the modulation of S1P signalling axis, upregulating mRNA levels of S1P 2 and S1P 3 and downregulating those of SK2. TNFα increases the phosphorylated form of SK1, readout of its activation. Interestingly, pharmacological inhibition of SK1 and specific antagonism of S1P 3 prevented the increase in autophagy markers and the changes in the electrophysiological properties of C2C12 myotubes without affecting metabolic remodelling induced by the cytokine, highlighting the involvement of S1P signalling axis on TNFα-induced atrophy in skeletal muscle.

Some adipokines, such as adiponectin (ADPN), other than being implicated in the central regulation of feeding behavior, may influence gastric motor responses, which are a source of peripheral signals that also influence food intake. The present study aims to elucidate the signaling pathways through which ADPN exerts its actions in the mouse gastric fundus. To this purpose, we used a multidisciplinary approach. The mechanical results showed that ADPN caused a decay of the strip basal tension, which was abolished by the nitric oxide (NO) synthesis inhibitor, L-N G -nitro arginine (L-NNA). The electrophysiological experiments confirmed that all ADPN effects were abolished by L-NNA, except for the reduction of Ca 2+ current, which was instead prevented by the inhibitor of AMP-activated protein kinase (AMPK), dorsomorphin. The activation of the AMPK signaling by ADPN was confirmed by immunofluorescence analysis, which also revealed the ADPN R1 receptor (AdipoR1) expression in glial cells of the myenteric plexus. In conclusion, our results indicate that ADPN exerts an inhibitory action on the gastric smooth muscle by acting on AdipoR1 and involving the AMPK signaling pathway at the peripheral level. These findings provide novel bases for considering AMPK as a possible pharmacologic target for the potential treatment of obesity and eating disorders.

It has been hypothesized that leptin level alterations in Eating Disorders (EDs) represent a maintaining factor for pathological reward-related ED behaviors, given leptin role in the dopaminergic reward systems. The aim of the present study was to evaluate the role of leptin in EDs as a mediator for the relationship between Body Mass Index (BMI) and several pathological behaviors, such as dietary restraint, compensatory exercise, vomiting, binge eating and emotional eating. Sixty-two patients with EDs and 41 healthy controls (HC) had their blood drawn and completed psychometric tests for the evaluation of general psychopathology, ED psychopathology and emotional eating. Moderated linear regression models showed that, in the presence of high levels of ED psychopathology, leptin levels were negatively associated with dietary restraint and compensatory exercise, and positively with emotional eating and binge eating. Finally, leptin showed an indirect effect on the association between BMI and all these reward-related behaviors. These results suggest that a variation of BMI maintains these pathological ED behaviors through a variation in leptin levels. Considering the role of leptin in reward circuits, the results seem to confirm an aberrant food-related reward mechanism in ED patients., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Glucagon-like peptide-2 (GLP-2) has been reported to indirectly relax gastric smooth muscle. In the present study we investigated, through a combined mechanical and immunohistochemical approach, whether GLP-2 interferes with the electrical field stimulation (EFS)-induced vipergic relaxant responses and the mechanism through which it occurs. For functional experiments, strips from the mouse gastric fundus were mounted in organ baths for isometric recording of the mechanical activity. Vasoactive intestinal peptide (VIP) immunoreactivity in GLP-2 exposed specimens was also evaluated by immunohistochemistry. In carbachol pre-contracted strips, GLP-2 (20 nM) evoked a tetrodotoxin (TTX)-sensitive relaxation, similar in shape to the TTX-insensitive of 100 nM VIP. In the presence of GLP-2, VIP had no longer effects and no more response to GLP-2 was observed following VIP receptor saturation. EFS (4-16 Hz) induced a fast relaxant response followed, at the higher stimulation frequencies (≥ 8 Hz), by a slow one. This latter was abolished either by GLP-2 or VIP receptor saturation as well as by the VIP receptor antagonist, VIP 6-28 (10 μM). A decrease of VIP-immunoreactive nerve structures in the GLP-2 exposed specimens was observed. These results suggest that, in the mouse gastric fundus, GLP-2 influences the EFS-induced slow relaxant response by promoting neuronal VIP release., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The regulation of food intake is a complex mechanism, and the hypothalamus is the main central structure implicated. In particular, the arcuate nucleus appears to be the most critical area in the integration of multiple peripheral signals. Among these signals, those originating from the white adipose tissue and the gastrointestinal tract are known to be involved in the regulation of food intake. The present paper focuses on adiponectin, an adipokine secreted by white adipose tissue, which is reported to have a role in the control of feeding by acting centrally. The recent observation that adiponectin is also able to influence gastric motility raises the question of whether this action represents an additional peripheral mechanism that concurs with the central effects of the hormone on food intake. This possibility, which represents an emerging aspect correlating the central and peripheral effects of adiponectin in the hunger-satiety cycle, is discussed in the present paper., Competing Interests: Conflict-of-interest statement: No conflicts of interest, financial or otherwise, are declared by the authors., (©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.)

Skeletal muscle repair/regeneration may benefit by Platelet-Rich Plasma (PRP) treatment owing to PRP pro-myogenic and anti-fibrotic effects. However, PRP anti-fibrotic action remains controversial. Here, we extended our previous researches on the inhibitory effects of PRP on in vitro transforming growth factor (TGF)-β1-induced differentiation of fibroblasts into myofibroblasts, the effector cells of fibrosis, focusing on gap junction (GJ) intercellular communication. The myofibroblastic phenotype was evaluated by cell shape analysis, confocal fluorescence microscopy and Western blotting analyses of α-smooth muscle actin and type-1 collagen expression, and electrophysiological recordings of resting membrane potential, resistance, and capacitance. PRP negatively regulated myofibroblast differentiation by modifying all the assessed parameters. Notably, myofibroblast pairs showed an increase of voltage-dependent GJ functionality paralleled by connexin (Cx) 43 expression increase. TGF-β1-treated cells, when exposed to a GJ blocker, or silenced for Cx43 expression, failed to differentiate towards myofibroblasts. Although a minority, myofibroblast pairs also showed not-voltage-dependent GJ currents and coherently Cx26 expression. PRP abolished the TGF-β1-induced voltage-dependent GJ current appearance while preventing Cx43 increase and promoting Cx26 expression. This study adds insights into molecular and functional mechanisms regulating fibroblast-myofibroblast transition and supports the anti-fibrotic potential of PRP, demonstrating the ability of this product to hamper myofibroblast generation targeting GJs., Competing Interests: The authors declare no conflict of interest.

It has been well established, particularly in animal models, that oestrogens exert neuroprotective effects in brain areas linked to cognitive processes. A key protective role could reside in the capacity of oestrogen to modulate the inflammatory response. However, the direct neuroprotective actions of oestrogens on neurones are complex and remain to be fully clarified. In the present study, we took advantage of a previously characterised primary culture of human cholinergic neurones (hfNBM) from the foetal nucleus basalis of Meynert, which is known to regulate hippocampal and neocortical learning and memory circuits, aiming to investigate the direct effects of oestrogens under inflammatory conditions. Exposure of cells to tumour necrosis factor (TNF)α (10 ng mL -1 ) determined the activation of an inflammatory response, as demonstrated by nuclear factor-kappa B p65 nuclear translocation and cyclooxygenase-2 mRNA expression. These effects were inhibited by treatment with either 17β-oestradiol (E 2 ) (10 nmol L -1 ) or G1 (100 nmol L -1 ), the selective agonist of the G protein-coupled oestrogen receptor (GPER1). Interestingly, the GPER1 antagonist G15 abolished the effects of E 2 in TNFα-treated cells, whereas the ERα/ERβ inhibitor tamoxifen did not. Electrophysiological measurements in hfNBMs revealed a depolarising effect caused by E 2 that was specifically blocked by tamoxifen and not by G15. Conversely, G1 specifically hyperpolarised the cell membrane and also increased both inward and outward currents elicited by a depolarising stimulus, suggesting a modulatory action on hfNBM excitability by GPER1 activation. Interestingly, pretreating cells with TNFα completely blocked the effects of G1 on membrane properties and also significantly reduced GPER1 mRNA expression. In addition, we found a peculiar subcellular localisation of GPER1 to focal adhesion sites that implicates new possible mechanisms of action of GPER1 in the neuronal perception of mechanical stimuli. The results obtained in the present study indicate a modulatory functional role of GPER1 with respect to mediating the oestrogen neuroprotective effect against inflammation in brain cholinergic neurones and, accordingly, may help to identify protective strategies for preventing cognitive impairments., (© 2020 British Society for Neuroendocrinology.)

Although telocytes (TCs) have been proposed to play a "nursing" role in resident satellite cell (SC)-mediated skeletal muscle regeneration, currently there is no evidence of TC-SC morpho-functional interaction following tissue injury. Hence, we explored the presence of TCs and their relationship with SCs in an ex vivo model of eccentric contraction (EC)-induced muscle damage. EC-injured muscles showed structural/ultrastructural alterations and changes in electrophysiological sarcolemnic properties. TCs were identified in control and EC-injured muscles by either confocal immunofluorescence (i.e. CD34 + CD31 - TCs) or transmission electron microscopy (TEM). In EC-injured muscles, an extended interstitial network of CD34 + TCs/telopodes was detected around activated SCs displaying Pax7 + and MyoD + nuclei. TEM revealed that TCs invaded the SC niche passing with their telopodes through a fragmented basal lamina and contacting the underlying activated SCs. TC-SC interaction after injury was confirmed in vitro by culturing single endomysial sheath-covered myofibers and sprouting TCs and SCs. EC-damaged muscle-derived TCs showed increased expression of the recognized pro-myogenic vascular endothelial growth factor-A, and SCs from the same samples exhibited increased MyoD expression and greater tendency to fuse into myotubes. Here, we provide the essential groundwork for further investigation of TC-SC interactions in the setting of skeletal muscle injury and regenerative medicine.

Some adipokines known to regulate food intake at a central level can also affect gastrointestinal motor responses. These are recognized to be peripheral signals able to influence feeding behavior as well. In this view, it has been recently observed that adiponectin (ADPN), which seems to have a role in sending satiety signals at the central nervous system level, actually affects the mechanical responses in gastric strips from mice. However, at present, there are no data in the literature about the electrophysiological effects of ADPN on gastric smooth muscle. To this aim, we achieved experiments on smooth muscle cells (SMCs) of gastric fundus to find out a possible action on SMC excitability and on membrane phenomena leading to the mechanical response. Experiments were made inserting a microelectrode in a single cell of a muscle strip of the gastric fundus excised from adult female mice. We found that ADPN was able to hyperpolarize the resting membrane potential, to enhance the delayed rectifier K + currents and to reduce the voltage-dependent Ca 2+ currents. Our overall results suggest an inhibitory action of ADPN on gastric SMC excitation-contraction coupling. In conclusion, the depressant action of ADPN on the gastric SMC excitability, here reported for the first time, together with its well-known involvement in metabolism, might lead us to consider a possible contribution of ADPN also as a peripheral signal in the hunger-satiety cycle and thus in feeding behavior.

Hearing loss is among the most prevalent sensory impairments in humans. Cochlear implantable devices represent the current therapies for hearing loss but have various shortcomings. ERM (ezrin- radixin -moesin) are a family of adaptor proteins that link plasma membrane with actin cytoskeleton, playing a crucial role in cell morphology and in the formation of membrane protrusions. Recently, bioactive sphingolipids have emerged as regulators of ERM proteins. Sphingosine 1-phosphate (S1P) is a pleiotropic sphingolipid which regulates fundamental cellular functions such as proliferation, survival, migration as well as processes such as development and inflammation mainly via ligation to its specific receptors S1PR (S1P 1-5 ). Experimental findings demonstrate a key role for S1P signaling axis in the maintenance of auditory function. Preservation of cellular junctions is a fundamental function both for S1P and ERM proteins, crucial for the maintenance of cochlear integrity. In the present work, S1P was found to activate ERM in a S1P 2 -dependent manner in murine auditory epithelial progenitors US/VOT-E36. S1P-induced ERM activation potently contributed to actin cytoskeletal remodeling and to the appearance of ionic currents and membrane passive properties changes typical of more differentiated cells. Moreover, PKC and Akt activation was found to mediate S1P-induced ERM phosphorylation. The obtained findings contribute to demonstrate the role of S1P signaling pathway in inner ear biology and to disclose potential innovative therapeutical approaches in the field of hearing loss prevention and treatment., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The present review focuses on adipocytes-released peptides known to be involved in the control of gastrointestinal motility, acting both centrally and peripherally. Thus, four peptides have been taken into account: leptin, adiponectin, nesfatin-1, and apelin. The discussion of the related physiological or pathophysiological roles, based on the most recent findings, is intended to underlie the close interactions among adipose tissue, central nervous system, and gastrointestinal tract. The better understanding of this complex network, as gastrointestinal motor responses represent peripheral signals involved in the regulation of food intake through the gut-brain axis, may also furnish a cue for the development of either novel therapeutic approaches in the treatment of obesity and eating disorders or potential diagnostic tools., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Aim: To investigate whether the adipocytes derived hormone adiponectin (ADPN) affects the mechanical responses in strips from the mouse gastric fundus., Methods: For functional experiments, gastric strips from the fundal region were cut in the direction of the longitudinal muscle layer and placed in organ baths containing Krebs-Henseleit solution. Mechanical responses were recorded via force-displacement transducers, which were coupled to a polygraph for continuous recording of isometric tension. Electrical field stimulation (EFS) was applied via two platinum wire rings through which the preparation was threaded. The effects of ADPN were investigated on the neurally-induced contractile and relaxant responses elicited by EFS. The expression of ADPN receptors, Adipo-R1 and Adipo-R2, was also evaluated by touchdown-PCR analysis., Results: In the functional experiments, EFS (4-16 Hz) elicited tetrodotoxin (TTX)-sensitive contractile responses. Addition of ADPN to the bath medium caused a reduction in amplitude of the neurally-induced contractile responses ( P < 0.05). The effects of ADPN were no longer observed in the presence of the nitric oxide (NO) synthesis inhibitor L-N G -nitro arginine (L-NNA) ( P > 0.05). The direct smooth muscle response to methacholine was not influenced by ADPN ( P > 0.05). In carbachol precontracted strips and in the presence of guanethidine, EFS induced relaxant responses. Addition of ADPN to the bath medium, other than causing a slight and progressive decay of the basal tension, increased the amplitude of the neurally-induced relaxant responses ( P < 0.05). Touchdown-PCR analysis revealed the expression of both Adipo-R1 and Adipo-R2 in the gastric fundus., Conclusion: The results indicate for the first time that ADPN is able to influence the mechanical responses in strips from the mouse gastric fundus., Competing Interests: Conflict-of-interest statement: No conflicts of interest, financial or otherwise, are declared by the authors.

Human striatal precursor cells (HSPs) isolated from ganglionic eminence may differentiate in electrophysiologically functional excitable neuron-like cells and a number of endogenous molecules such as hormones, neurotransmitters or growth factors can actually regulate neuronal growing and differentiation. The purpose of this research was to assess, by electrophysiological and immunocytochemical analysis, if the type of culture medium could specifically impact on the neuronal differentiation potential of HSPs. Accordingly, HSPs were maintained in different inductive media such as cortical and spinal cord conditioned media, and we estimated the possible changes in the main ion currents, excitability and expression of neuronal markers indicative of neuronal differentiation. Our results have shown that 36 h exposure to each of the conditioned media, with their blend of autocrine and paracrine growth factors, was able to modify significantly the electrophysiological membrane properties and the functional expression of inward ionic currents in selected neuronal HSPs. Moreover, although both types of conditioned media determined neuronal maturation (increased neuritogenesis and increased expression of neuronal and striatal markers), each of them leads to the occurrence of different functional features. Particularly, the spinal medium caused a stronger depolarization of the membrane potential and significantly increased the amplitude of Na + current as well as L- and N- type Ca 2+ currents, definitely modifying their kinetics. In contrast, the cortical medium mainly caused a significant and more marked increase of the membrane conductance and time constant values. These results strongly support the plasticity of our cellular model that, although already committed towards a specific phenotype, it can be differently affected by the conditioned media, thereby resulting functionally modifiable according to environmental cues., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Aim: To investigate the signaling pathways involved in the relaxin (RLX) effects on ileal preparations from mice through mechanical and electrophysiological experiments., Methods: For mechanical experiments, ileal preparations from female mice were mounted in organ baths containing Krebs-Henseleit solution. The mechanical activity was recorded via force-displacement transducers, which were coupled to a polygraph for continuous recording of isometric tension. Electrophysiological measurements were performed in current- and voltage-clamp conditions by a microelectrode inserted in a single smooth muscle cell (SMC) of the ileal longitudinal layer. Both the membrane passive properties and inward voltage-dependent L-type Ca 2+ currents were recorded using suitable solutions and voltage stimulation protocols., Results: Mechanical experiments showed that RLX induced a decay of the basal tension and a reduction in amplitude of the spontaneous contractions. The effects of RLX were partially reduced by 1H-[1,2,4]oxadiazolo[4,3- a ]-quinoxalin-1-one (ODQ) or 9-cyclopentyladenine mesylate (9CPA), inhibitors of guanylate cyclase (GC) and adenylate cyclase (AC), respectively, and were abolished in the concomitant presence of both drugs. Electrophysiological experiments demonstrated that RLX directly influenced the biophysical properties of ileal SMCs, decreasing the membrane conductance, hyperpolarizing the resting membrane potential, reducing the L-type calcium current amplitude and affecting its kinetics. The voltage dependence of the current activation and inactivation time constant was significantly speeded by RLX. Each electrophysiological effect of RLX was reduced by ODQ or 9CPA, and abolished in the concomitant presence of both drugs as observed in mechanical experiments., Conclusion: Our new findings demonstrate that RLX influences ileal muscle through a dual mechanism involving both GC and AC., Competing Interests: Conflict-of-interest statement: No conflicts of interest, financial or otherwise, are declared by the authors.

Aim: To investigate whether glucagon-like peptide-2 (GLP-2) influences the neurally-induced responses in gastric strips from mice, since no data are available., Methods: For functional experiments, gastric fundal strips were mounted in organ baths containing Krebs-Henseleit solution. Mechanical responses were recorded via force-displacement transducers, which were coupled to a polygraph for continuous recording of isometric tension. Electrical field stimulation (EFS) was applied via two platinum wire rings through which the preparation was threaded. The effects of GLP-2 (2 and 20 nmol/L) were evaluated on the neurally-induced contractile and relaxant responses elicited by EFS. Neuronal nitric oxide synthase (nNOS) enzyme was evaluated by immunohistochemistry., Results: In the functional experiments, electrical field stimulation (EFS, 4-16 Hz) induced tetrodotoxin (TTX)-sensitive contractile responses, which were reduced in amplitude by GLP-2 ( P < 0.05). In the presence of the nitric oxide (NO) synthesis inhibitor L-NNA, GLP-2 no longer influenced the neurally-evoked contractile responses ( P > 0.05). The direct smooth muscle response to methacholine was not influenced by GLP-2 ( P > 0.05). In the presence of guanethidine and carbachol, the addition of GLP-2 to the bath medium evoked TTX-sensitive relaxant responses that were unaffected by L-NNA ( P > 0.05). EFS induced a fast NO-mediated relaxation, whose amplitude was enhanced in the presence of the hormone ( P < 0.05). Immunohistochemical experiments showed a significant increase ( P < 0.05) in nNOS immunoreactivity in the nerve structures after GLP-2 exposure., Conclusion: The results demonstrate that in gastric fundal strips, GLP-2 influences the amplitude of neurally-induced responses through the modulation of the nitrergic neurotransmission and increases nNOS expression., Competing Interests: Conflict-of-interest statement: No conflicts of interest, financial or otherwise, are declared by the authors.