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Background Several pre-participation screening algorithms (PPSAs) have been proposed to assess sports eligibility in different populations. They are usually based on self-administered questionnaires, without further medical assessment if no risk factors are documented. The Med-Ex “Formula Benessere” worksite program includes a complete cardiovascular (CV) screening for all participants. The purpose of this study was to assess PPSAs accuracy in detecting medical and/or CV abnormalities in the general population, comparing the results with the date derived from Med-Ex program. Methods The Med-Ex medical evaluation, consisting of medical history, physical examination (including body composition), resting electrocardiogram (ECG) and exercise stress test in 464 male subjects (38.4 aged) was analyzed and matched to several PPSAs – Physical Activity Readiness Questionnaire (PAR-Q) (2002–2020), American Heart Association (AHA)/American College of Sport Medicine (ACSM) (1998–2009–2014–2015), European Association of Cardiovascular Prevention and Rehabilitation (EACPR) (2011) – retrospectively simulated. Results Five-hundred and 67 abnormalities were detected though Med-Ex medical evaluation, and one-fourth (24%) would have been undetected applying PPSA alone. In particular 28% of high blood pressure, 21% of impaired fasting glycaemia, 21% of high Body Mass Index (BMI) values and 19% of ECG abnormalities would have been missed, on average, by all PPSAs. Conclusions The simulation analysis model performed in this study allowed to highlight the limits of PPSAs in granting sport eligibility, compared to a medical-guided CV screening. These findings emphasize the importance of a more balanced approach to pre-participation screening that includes a thorough evaluation of the cost/benefit ratio.

Thyroid cancer is the most common type of endocrine cancer, and its prevalence continue to rise. Non-metastatic thyroid cancer patients are successfully treated. However, looking for new therapeutic strategies is of great importance for metastatic thyroid cancers that still lead to death. With respect to this, the tumor microenvironment (TME), which plays a key role in tumor progression, should be considered as a new promising therapeutic target to hamper thyroid cancer progression. Indeed, thyroid tumors consist of cancer cells and a heterogeneous and ever-changing niche, represented by the TME, which contributes to establishing most of the features of cancer cells. The TME consists of extracellular matrix (ECM) molecules, soluble factors, metabolites, blood and lymphatic tumor vessels and several stromal cell types that, by interacting with each other and with tumor cells, affect TME remodeling, cancer growth and progression. Among the thyroid TME components, cancer-associated fibroblasts (CAFs) have gained more attention in the last years. Indeed, recent important evidence showed that thyroid CAFs strongly sustain thyroid cancer growth and progression by producing soluble factors and ECM proteins, which, in turn, deeply affect thyroid cancer cell behavior and aggressiveness. Hence, in this article, we describe the thyroid TME, focusing on the desmoplastic stromal reaction, which is a powerful indicator of thyroid cancer progression and an invasive growth pattern. In addition, we discuss the origins and features of the thyroid CAFs, their influence on thyroid cancer growth and progression, their role in remodeling the ECM and their immune-modulating functions. We finally debate therapeutic perspectives targeting CAFs.

Cutaneous melanoma (CM) tissue represents a network constituted by cancer cells and tumor microenvironment (TME). A key feature of CM is the high structural and cellular plasticity of TME, allowing its evolution with disease and adaptation to cancer cell and environmental alterations. In particular, during melanoma development and progression each component of TME by interacting with each other and with cancer cells is subjected to dramatic structural and cellular modifications. These alterations affect extracellular matrix (ECM) remodelling, phenotypic profile of stromal cells, cancer growth and therapeutic response. The stromal fibroblast populations of the TME include normal fibroblasts and melanoma-associated fibroblasts (MAFs) that are highly abundant and flexible cell types interacting with melanoma and stromal cells and differently influencing CM outcomes. The shift from the normal microenvironment to TME and from normal fibroblasts to MAFs deeply sustains CM growth. Hence, in this article we review the features of the normal microenvironment and TME and describe the phenotypic plasticity of normal dermal fibroblasts and MAFs, highlighting their roles in normal skin homeostasis and TME regulation. Moreover, we discuss the influence of MAFs and their secretory profiles on TME remodelling, melanoma progression, targeted therapy resistance and immunosurveillance, highlighting the cellular interactions, the signalling pathways and molecules involved in these processes.

The generation of pluripotent stem cells from adult somatic cells by cell reprogramming has put a whole new perspective on stem cell biology and stem cell-based regenerative medicine. Cell reprogramming acts through the introduction of key genes that regulate and maintain the pluripotent cell state. In this chapter, we describe the optimized protocol for the efficient isolation of fibroblasts from a skin punch biopsy and the subsequent easy and effective generation of integration-free induced pluripotent stem cell (iPSC) colonies forcing the expression of specific factors by non-modified RNAs.

The generation of pluripotent stem cells from adult somatic cells by cell reprogramming has put a whole new perspective on stem cell biology and stem cell-based regenerative medicine. Cell reprogramming acts through the introduction of key genes that regulate and maintain the pluripotent cell state. In this chapter, we describe the optimized protocol for the efficient isolation of fibroblasts from a skin punch biopsy and the subsequent easy and effective generation of integration-free induced pluripotent stem cell (iPSC) colonies forcing the expression of specific factors by non-modified RNAs. © 2021, Springer Science+Business Media, LLC.

In Italy, recent legislation (Law No. 10/2020) has tuned regulations concerning the donation of one's postmortem body and tissues for study, training, and scientific research purposes. This study discusses several specific issues to optimise the applicability and effectiveness of such an important, novel regulatory setting. Some of these unsolved issues may involve the grantees of teaching and training activities, the role of academic anatomical institutes, the role of family members in the donation process, the universal time limit indicated for any donation, the handling of corpses, and the limited body donation and its subordination to the donation of organs and tissues. Critical issues arise concerning the learners, the type of training and teaching activities that can be planned, the position of academic anatomy institutes, the role of family members in the donation process, the time frame of the donation process, the eligibility of partial donation, or the simultaneous donation of organs and tissues to patients awaiting transplantation. In particular, a universal time limit for donations (i.e., one year) makes it impossible to plan the long-term use of specific body parts, which could be effectively preserved for the advanced teaching and training of medical students and surgeons. The abovementioned conditions lead to the limited use of corpses, thus resulting in the inefficiency of the whole system of body donation. Overall, the donors' scope for the donation of their body could be best honoured by a more flexible and tuneable approach that can be used on a case-by-case basis. Furthermore, it is deemed necessary to closely monitor the events scheduled for corpses in public nonacademic institutions or private enterprises. This paper presents useful insights from Italian anatomists with the hope of providing inspiration for drafting the regulations. In conclusion, this paper focuses on the critical issues derived from the recently introduced Italian law on the donation and use of the body after death and provides suggestions to lawmakers for future implementations.

Neuromotor training can improve motor performance in athletes and patients. However, few data are available about their effect on reaction time (RT). We investigated the influence of video observation/motor imagery (VO/MI) on simple RT to visual and auditory stimuli. The experimental group comprised 21 cadets who performed VO/MI training over 4 weeks. Nineteen cadets completed a sham intervention as control. The main outcome measure was RT to auditory and visual stimuli for the upper and lower limbs. The RT to auditory stimuli improved significantly post-intervention in both groups (control vs. experimental mean change for upper limbs: &minus, 40 ms vs. &minus, 40 ms, p = 0.0008, for lower limbs: &minus, 50 ms vs. &minus, 30 ms, p = 0.0174). A trend towards reduced RT to visual stimuli was observed (for upper limbs: &minus, 30 ms vs. &minus, 20 ms, p = 0.0876, 20 ms, p = 0.0675). The interaction term was not significant. Only the specific VO/MI training produced a linear correlation between the improvement in the RT to auditory and visual stimuli for the upper (r = 0.703) and lower limbs (r = 0.473). In conclusion, VO/MI training does not improve RT when compared to control, but it may be useful in individuals who need to simultaneously develop a fast response to different types of stimuli.

Cutaneous melanoma (CM) is a highly aggressive and drug resistant solid tumor, showing an impressive metabolic plasticity modulated by oncogenic activation. In particular, melanoma cells can generate adenosine triphosphate (ATP) during cancer progression by both cytosolic and mitochondrial compartments, although CM energetic request mostly relies on glycolysis. The upregulation of glycolysis is associated with constitutive activation of BRAF/MAPK signaling sustained by BRAFV600E kinase mutant. In this scenario, the growth and progression of CM are strongly affected by melanoma metabolic changes and interplay with tumor microenvironment (TME) that sustain tumor development and immune escape. Furthermore, CM metabolic plasticity can induce a metabolic adaptive response to BRAF/MEK inhibitors (BRAFi/MEKi), associated with the shift from glycolysis toward oxidative phosphorylation (OXPHOS). Therefore, in this review article we survey the metabolic alterations and plasticity of CM, its crosstalk with TME that regulates melanoma progression, drug resistance and immunosurveillance. Finally, we describe hallmarks of melanoma therapeutic strategies targeting the shift from glycolysis toward OXPHOS.

Induced pluripotent stem cells (iPSCs) could be considered, to date, a promising source of pluripotent cells for the management of currently untreatable diseases, for the reconstitution and regeneration of injured tissues and for the development of new drugs. Despite all the advantages related to the use of iPSCs, such as the low risk of rejection, the lessened ethical issues, and the possibility to obtain them from both young and old patients without any difference in their reprogramming potential, problems to overcome are still numerous. In fact, cell reprogramming conducted with viral and integrating viruses can cause infections and the introduction of required genes can induce a genomic instability of the recipient cell, impairing their use in clinic. In particular, there are many concerns about the use of c-Myc gene, well-known from several studies for its mutation-inducing activity. Fibroblasts have emerged as the suitable cell population for cellular reprogramming as they are easy to isolate and culture and are harvested by a minimally invasive skin punch biopsy. The protocol described here provides a detailed step-by-step description of the whole procedure, from sample processing to obtain cell cultures, choice of reagents and supplies, cleaning and preparation, to cell reprogramming by the means of a commercial non-modified RNAs (NM-RNAs)-based reprogramming kit. The chosen reprogramming kit allows an effective reprogramming of human dermal fibroblast to iPSCs and small colonies can be seen as early as 24 h after the first transfection, even with modifications with the respect to the standard datasheet. The reprogramming procedure used in this protocol offers the advantage of a safe reprogramming, without the risk of infections caused by viral vector-based methods, reduces the cellular defense mechanisms, and allows the generation of xeno-free iPSCs, all critical features that are mandatory for further clinical applications.

Decellularized extracellular matrix is one of the most promising biological scaffold supporting in vitro tissue growth and in vivo tissue regeneration in both preclinical research and clinical practice. In case of thick tissues or even organs, conventional static decellularization methods based on chemical or enzymatic treatments are not effective in removing the native cellular material without affecting the extracellular matrix. To overcome this limitation, dynamic decellularization methods, mostly based on perfusion and agitation, have been proposed. In this study, we developed a low-cost scalable 3D-printed sample-holder for agitation-based decellularization purposes, designed for treating multiple specimens simultaneously and for improving efficiency, homogeneity and reproducibility of the decellularization treatment with respect to conventional agitation-based approaches. In detail, the proposed sample-holder is able to house up to four specimens and, immersed in the decellularizing solution within a beaker placed on a magnetic stirrer, to expose them to convective flow, enhancing the solution transport through the specimens while protecting them. Computational fluid dynamics analyses were performed to investigate the fluid phenomena establishing within the beaker and to support the sample-holder design. Exploratory biological tests performed on human skin specimens demonstrated that the sample-holder reduces process duration and increases treatment homogeneity and reproducibility.

Introduction: Age, gender, body mass index percentiles and the adherence to Mediterranean diet were investigated as potential predictor factors in this assessment. Aim: To assess the parental perception of children's weight status. Methods: This cross-sectional observational study was carried out during a corporate wellness program (Ferrari Formula Benessere) implemented by Ferrari S.p.A. and managed by Med-Ex s.r.l. The children’s real weight status was assessed through Body Mass Index percentiles (ArthroPlus software—WHO). Results: 328 children (66.4%) were normal-weight, 10 were underweight (2%), 66 were overweight (13.4%) and 90 were obese (18.2%). 289 children (59%) were classified correctly by parents, while 205 children (41%) were not. 64 of 66 overweight children and 90 of 90 children with obesity have been completely underestimated (53 of 90 children with obesity were judged normal weight). The parents’ probability to estimate children’s weight status correctly decreased increasing body mass index percentiles paradoxically [OR = 0.96 (0.95–0.97)] and was lower in boys [OR = 0.65 (0.44–0.98)]. Although not statistically significant, children with higher adherence to Mediterranean diet seems to have higher probability to be correctly estimated [low adherence: reference, medium adherence: OR = 1.06 (0.61–1.85), high adherence: OR = 1.48 (0.81–2.75)]. Conclusions: A high percentage of children is overweight or obese and almost half of parents classified their weight status incorrectly. Increasing children’s body mass index percentiles decreases the probability to be correctly classified.

The complex and highly organized environment in which cells reside consists primarily of the extracellular matrix (ECM) that delivers biological signals and physical stimuli to resident cells. In the native myocardium, the ECM contributes to both heart compliance and cardiomyocyte maturation and function. Thus, myocardium regeneration cannot be accomplished if cardiac ECM is not restored. We hypothesize that decellularized human skin might make an easily accessible and viable alternate biological scaffold for cardiac tissue engineering (CTE). To test our hypothesis, we decellularized specimens of both human skin and human myocardium and analyzed and compared their composition by histological methods and quantitative assays. Decellularized dermal matrix was then cut into 600-μm-thick sections and either tested by uniaxial tensile stretching to characterize its mechanical behavior or used as three-dimensional scaffold to assess its capability to support regeneration by resident cardiac progenitor cells (hCPCs) in vitro. Histological and quantitative analyses of the dermal matrix provided evidence of both effective decellularization with preserved tissue architecture and retention of ECM proteins and growth factors typical of cardiac matrix. Further, the elastic modulus of the dermal matrix resulted comparable with that reported in literature for the human myocardium and, when tested in vitro, dermal matrix resulted a comfortable and protective substrate promoting and supporting hCPC engraftment, survival and cardiomyogenic potential. Our study provides compelling evidence that dermal matrix holds promise as a fully autologous and cost-effective biological scaffold for CTE.

Background. Recent findings indicate that the connective tissue of tendons hosts cells that can self-renew and are multipotent. Even if these cells seemingly fail to accomplish tendon regeneration in pathological conditions, their identification and characterization represents a milestone in the research and development of new biological therapies for tendinopathies. Methods. We isolated the adult human tendon-derived stem cells (hTDSCs) from the fragments of patellar or calcaneal tendons and characterized these cells in vitro by immunochemistry and histochemistry. Subsequently, the MTT test and Trypan Blue were used for the evaluation of cytotoxicity of the supplements/drugs commonly used for the treatment of musculoskeletal disorders: Curcumin, Hyaluronic Acid, Palmi-toylethanolamide, Diclofenac sodium, Triamcinolone acetonide and Thiocolchicoside. Results. Cells obtained by outgrowth expressed mesenchymal markers, were clono-genic and differentiated into chondroblasts, osteoblasts and adipocytes. High concen-trations of the anti-inflammatory glucocorticoid Triamcinolone and the analge-sic fatty-acid amide Palmitoylethanolamide significantly reduced cell viability. Only curcumin had a positive effect on cell survival, both in the normal and oxidative stress conditions. Conclusions. Adult human tendons posses stem cells. The influence of several drugs or supplements used for the treatment of musculoskeletal disorders should be taken into consideration in order to take the full advantage of the healing properties of stem cells within tendons.

Cancer associated fibroblasts (CAFs) are the main stromal cell type of solid tumour microenvironment and undergo an activation process associated with secretion of growth factors, cytokines, and paracrine interactions. One of the important features of solid tumours is the metabolic reprogramming that leads to changes of bioenergetics and biosynthesis in both tumour cells and CAFs. In particular, CAFs follow the evolution of tumour disease and acquire a catabolic phenotype: in tumour tissues, cancer cells and tumour microenvironment form a network where the crosstalk between cancer cells and CAFs is associated with cell metabolic reprogramming that contributes to CAFs activation, cancer growth, and progression and evasion from cancer therapies. In this regard, the study of CAFs metabolic reprogramming could contribute to better understand their activation process, the interaction between stroma, and cancer cells and could offer innovative tools for the development of new therapeutic strategies able to eradicate the protumorigenic activity of CAFs. Therefore, this review focuses on CAFs metabolic reprogramming associated with both differentiation process and cancer and stromal cells crosstalk. Finally, therapeutic responses and potential anticancer strategies targeting CAFs metabolic reprogramming are reviewed.

Background: New findings on adipose tissue physiology and obesity-associated inflammation status suggest that modification of the adipokine level can be relevant for the long-term prevention of obesity-associated chronic disease. Objectives: The scope of the present study was to investigate the effectiveness of physical exercise in reducing the systemic inflammation related to obesity in children. Methods: We conducted a systematic review with meta-analysis of controlled randomized trials, identified through electronic database search, which investigated the effect of physical exercise, without concomitant dietary intervention, on adiponectin, leptin, and/or other inflammatory markers in children up to age 18 years with a body mass index greater than the 95th percentile for age and sex. Results: Seven trials were included in the meta-analysis, with a total of 250 participants. Compared with the control group without any lifestyle modification, the physical exercise resulted in a reduction in leptin [standardized mean difference (SMD) −1.13; 95% confidence interval (95%CI): −1.89 to −0.37; I2 = 79.9%] and interleukin-6 (SMD −0.84; 95%CI: −1.45 to −0.23, I2 = 0.9%) and an increase in adiponectin plasma concentration (SMD 0.69; 95%CI: 0.02–1.35; I2 = 74.3%). Conclusions: These results indicate that physical exercise improved the inflammatory state in children with obesity. It is unclear whether this effect can reduce the risk of cardiovascular and metabolic disease in adulthood. Clinical trials with a uniform intervention protocol and outcome measurements are required to put our knowledge on adipose tissue biology into a clinical perspective.

Official tests are used to assess the fitness status of soccer referees, and their results correlate with match performance. However, FIFA-approved tests expose the referees to high physical demands and are difficult to implement during the sportive year. The aim of our study was to evaluate the correlation between the 6 × 40-m sprint and Yo-Yo Intermittent Recovery Level 1 (IR1) official tests and other field-based tests that require no or little equipment, are not time-consuming, and impose low physical demands. All tests were performed by male referees from the Regional Section of the Italian Referee Association (n = 30). We observed a strong correlation between 6 × 40-m sprint and Illinois agility tests (r = 0.63, p = 0.001) and a moderate correlation between Yo-Yo IR1 and hand-grip strength in the dominant (r = 0.45, p = 0.014) and non-dominant hand (r = 0.41, p = 0.031). Interestingly, only a moderate correlation (r = −0.42, p = 0.025) was observed between the FIFA official tests, 6 × 40-m sprint and Yo-Yo IR1. These results suggest that Illinois agility and hand-grip tests could represent simple and low-physical-impact tools for repeated assessment and monitoring of referee fitness throughout the sportive season.

Cardiac tissue engineering by means of synthetic or natural scaffolds combined with stem/progenitor cells is emerging as the response to the unsatisfactory outcome of approaches based solely on the injection of cells. Parenchymal and supporting cells are surrounded, in vivo, by a specialized and tissue-specific microenvironment, consisting mainly of extracellular matrix (ECM) and soluble factors incorporated in the ECM. Since the naturally occurring ECM is the ideal platform for ensuring cell engraftment, survival, proliferation, and differentiation, the acellular native ECM appears by far the most promising and appealing substrate among all biomaterials tested so far. To obtain intact scaffold of human native cardiac ECM while preserving its composition, we compared the decellularized ECM (d-ECM) produced through five different protocols of decellularization (named Pr1, Pr2, Pr3, Pr4, and Pr5) in terms of efficiency of decellularization, composition, and three-dimensional architecture of d-ECM scaffolds and of their suitability for cell repopulation. The decellularization procedures proved substantially different. Specifically, only three, of the five protocols tested, proved effective in producing thoroughly acellular d-ECM. In addition, the d-ECM delivered differed in architecture and composition and, more importantly, in its ability to support engraftment, survival, and differentiation of cardiac primitive cells in vitro.

Breast cancers are very heterogeneous tissues with several cell types and metabolic pathways together sustaining the initiation and progression of disease and contributing to evasion from cancer therapies. Furthermore, breast cancer cells have an impressive metabolic plasticity that is regulated by the heterogeneous tumour microenvironment through bidirectional interactions. The structure and accessibility of nutrients within this unstable microenvironment influence the metabolism of cancer cells that shift between glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) to produce adenosine triphosphate (ATP). In this scenario, the mitochondrial energetic pathways of cancer cells can be reprogrammed to modulate breast cancer’s progression and aggressiveness. Moreover, mitochondrial alterations can lead to crosstalk between the mitochondria and the nucleus, and subsequently affect cancer tissue properties. This article reviewed the metabolic plasticity of breast cancer cells, focussing mainly on breast cancer mitochondrial metabolic reprogramming and the mitochondrial alterations influencing nuclear pathways. Finally, the therapeutic strategies targeting molecules and pathways regulating cancer mitochondrial alterations are highlighted.

Melanoma is one of the most aggressive solid tumors and includes a stromal microenvironment that regulates cancer growth and progression. The components of stromal microenvironment such as fibroblasts, fibroblast aggregates and cancer-associated fibroblasts (CAFs) can differently influence the melanoma growth during its distinct stages. In this work, we have developed and studied a stromal microenvironment model, represented by fibroblasts, proto-myofibroblasts, myofibroblasts and aggregates of inactivated myofibroblasts, such as spheroids. In particular, we have generated proto-myofibroblasts from primary cutaneous myofibroblasts. The phenotype of proto-myofibroblasts is characterized by a dramatic reduction of &alpha, smooth muscle actin (&alpha, SMA) and cyclooxygenase-2 (COX-2) protein levels, as well as an enhancement of cell viability and migratory capability compared with myofibroblasts. Furthermore, proto-myofibroblasts display the mesenchymal marker vimentin and less developed stress fibers, with respect to myofibroblasts. The analysis of crosstalk between the stromal microenvironment and A375 or A2058 melanoma cells has shown that the conditioned medium of proto-myofibroblasts is cytotoxic, mainly for A2058 cells, and dramatically reduces the migratory capability of both cell lines compared with the melanoma-control conditioned medium. An array analysis of proto-myofibroblast and melanoma cell-conditioned media suggests that lower levels of some cytokines and growth factors in the conditioned medium of proto-myofibroblasts could be associated with their anti-tumor activity. Conversely, the conditioned media of melanoma cells do not influence the cell viability, outgrowth, and migration of proto-myofibroblasts from spheroids. Interestingly, the conditioned medium of proto-myofibroblasts does not alter the cell viability of both BJ-5ta fibroblast cells and myofibroblasts. Hence, proto-myofibroblasts could be useful in the study of new therapeutic strategies targeting melanoma.

Cutaneous melanoma (CM) represents one of the most metastasizing and drug resistant solid tumors. CM is characterized by a remarkable metabolic plasticity and an important connection between oncogenic activation and energetic metabolism. In fact, melanoma cells can use both cytosolic and mitochondrial compartments to produce adenosine triphosphate (ATP) during cancer progression. However, the CM energetic demand mainly depends on glycolysis, whose upregulation is strictly linked to constitutive activation of BRAF/MAPK pathway affected by BRAFV600E kinase mutant. Furthermore, the impressive metabolic plasticity of melanoma allows the development of resistance mechanisms to BRAF/MEK inhibitors (BRAFi/MEKi) and the adaptation to microenvironmental changes. The metabolic interaction between melanoma cells and tumor microenvironment affects the immune response and CM growth. In this review article, we describe the regulation of melanoma metabolic alterations and the metabolic interactions between cancer cells and microenvironment that influence melanoma progression and immune response. Finally, we summarize the hallmarks of melanoma therapies and we report BRAF/MEK pathway targeted therapy and mechanisms of metabolic resistance.

Induced pluripotent stem cells (iPSCs) are adult somatic cells genetically reprogrammed to an embryonic stem cell‐like state. Notwithstanding their autologous origin and their potential to differentiate towards cells of all three germ layers, iPSC reprogramming is still affected by low efficiency. As dermal fibroblast is the most used human cell for reprogramming, we hypothesize that the variability in reprogramming is, at least partially, because of the skin fibroblasts used. Human dermal fibroblasts harvested from five different anatomical sites (neck, breast, arm, abdomen and thigh) were cultured and their morphology, proliferation, apoptotic rate, ability to migrate, expression of mesenchymal or epithelial markers, differentiation potential and production of growth factors were evaluated in vitro. Additionally, gene expression analysis was performed by real‐time PCR including genes typically expressed by mesenchymal cells. Finally, fibroblasts isolated from different anatomic sites were reprogrammed to iPSCs by integration‐free method. Intriguingly, while the morphology of fibroblasts derived from different anatomic sites differed only slightly, other features, known to affect cell reprogramming, varied greatly and in accordance with anatomic site of origin. Accordingly, difference also emerged in fibroblasts readiness to respond to reprogramming and ability to form colonies. Therefore, as fibroblasts derived from different anatomic sites preserve positional memory, it is of great importance to accurately evaluate and select dermal fibroblast population prior to induce reprogramming.

Aims The aim of our study was to assess the clinical significance of the exercise stress testing endpoints, namely 85% of maximal theoretical heart rate (MTHR), metabolic equivalent of task, and rating of perceived exertion (RPE), and their relation to electrocardiographic (ECG) changes in a healthy adult population. Methods A cross-sectional study was conducted on 408 males and 52 females (mean age 39.4 ± 8.6 years) who performed the maximal cycle ergometer exercise stress test until volitional exhaustion, reporting the RPE score at 85% of MTHR and at peak exercise. Metabolic equivalents of task were indirectly calculated from the maximum workload and compared with the predicted values. Sitting torso-lead ECG and blood pressure were recorded at rest, during exercise and during recovery. Results Of 460 participants, 73% exceeded 85% of MTHR. The RPE score represented the overall most significant endpoint of exercise stress testing, with the median value of 17 at peak exercise. ECG events were detected in 23/124 (18.5%) who reached ≤ 85% of MTHR and in 61/336 (18.2%) who achieved >85% of MTHR ( p = 0.92). In the latter group, 54% of ECG changes occurred at 85% of MTHR ( p = 0.51). If the exercise stress testing had been interrupted at ≤ 85% of MTHR, almost half of the ECG events would have remained undetected and 35% of the cardiovascular abnormalities observed at the diagnostic follow-up would have remained undiagnosed. Conclusion Terminating exercise stress testing before volitional exhaustion and an RPE score of 17 limits the test accuracy and reduces the possibility to detect cardiovascular abnormalities in apparently healthy adult populations.

The esophagus is a muscular tube linking the pharynx to the stomach. It runs through the thorax, crosses the diaphragm, and enters the stomach. The authors examined the relationship of the esophagus with nearby organs in the cervical, thoracic, diaphragmatic, and abdominal portions. In particular, lymphatic drainage and its relations with lymph nodes and with the thoracic duct were discussed in order to clarify the possible dissemination of metastatic cells. The importance of the innervation, in its sensory and motor components, was evaluated also for its role in pain from esophagus and in salivatory protective reflex.

The esophagus is a muscular tube that conveys the bolus from the oral pharynx to the stomach. The microscopic structure of the esophagus is here examined with particular attention to the differentiative and morphological aspects which can be involved in the genesis of esophageal pathologies. In particular, the authors examined the protective mechanisms which act in normal conditions to avoid tissue damages due to the microenvironment of the low esophagus district, as the pre-epithelial barrier and the composition of the intercellular junction in the epithelium.

The pulmonary artery autograft (PA) is the ideal substitute for aortic valve disease in children and young adult. However, it is harnessed by the issue of long-term dilation and regurgitation, often requiring surgery. PA implanted in aortic position during the growth phase in children undergoes a process of mechanical remodeling. We previously developed a semiresorbable armored prosthesis able to mechanically sustain the neoaorta preventing dilation and to gradually integrate with the PA wall inducing a progressive arterial-like tissue positive remodeling. We also described the mechanisms of growth, remodeling and stress shielding of the reinforced PA through a mathematical model. We sought to demonstrate the biological counterpart and the potential molecular mechanisms underlying this histological and mechanical remodeling. A specific mathematical model was developed to describe mechanical behavior of the PA. Mallory trichrome red staining and immunohistochemistry for MMP-9 were performed to elucidate extracellular matrix remodeling phenomena. Apoptosis and cell proliferation were determined by TUNEL assay and immunohistochemistry for Ki67, respectively. An histological remodeling phenomenon sustained by increased level of MMP-9, augmented cell proliferation and reduced apoptosis in the reinforced PA was demonstrated. The mathematical model predicted the biomechanical behavior subtended by the histological changes of the PA in these settings. Changes in metalloproteinases (MMP-9), cell proliferation and apoptosis are the main actors in the remodeling process occurring after transposition of the PA into systemic regimens. Use of semiresorbable reinforcements might induce a positive remodeling of the PA in the context of Ross operation. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2785-2793, 2016.

Cardiac adverse remodeling is characterized by biological changes that affect the composition and architecture of the extracellular matrix (ECM). The consequently disrupted signaling can interfere with the balance between cardiogenic and pro-fibrotic phenotype of resident cardiac stromal primitive cells (CPCs). The latter are important players in cardiac homeostasis and can be exploited as therapeutic cells in regenerative medicine. Our aim was to compare the effects of human decellularized native ECM from normal (dECM-NH) or failing hearts (dECM-PH) on human CPCs. CPCs were cultured on dECM sections and characterized for gene expression, immunofluorescence, and paracrine profiles. When cultured on dECM-NH, CPCs significantly upregulated cardiac commitment markers (CX43, NKX2.5), cardioprotective cytokines (bFGF, HGF), and the angiogenesis mediator, NO. When seeded on dECM-PH, instead, CPCs upregulated pro-remodeling cytokines (IGF-2, PDGF-AA, TGF-&beta, ) and the oxidative stress molecule H2O2. Interestingly, culture on dECM-PH was associated with impaired paracrine support to angiogenesis, and increased expression of the vascular endothelial growth factor (VEGF)-sequestering decoy isoform of the KDR/VEGFR2 receptor. Our results suggest that resident CPCs exposed to the pathological microenvironment of remodeling ECM partially lose their paracrine angiogenic properties and release more pro-fibrotic cytokines. These observations shed novel insights on the crosstalk between ECM and stromal CPCs, suggesting also a cautious use of non-healthy decellularized myocardium for cardiac tissue engineering approaches.

The stromal microenvironment regulates mammary gland development and tumorigenesis. In normal mammary glands, the stromal microenvironment encompasses the ducts and contains fibroblasts, the main regulators of branching morphogenesis. Understanding the way fibroblast signaling pathways regulate mammary gland development may offer insights into the mechanisms of breast cancer (BC) biology. In fact, the unregulated mammary fibroblast signaling pathways, associated with alterations in extracellular matrix (ECM) remodeling and branching morphogenesis, drive breast cancer microenvironment (BCM) remodeling and cancer growth. The BCM comprises a very heterogeneous tissue containing non-cancer stromal cells, namely, breast cancer-associated fibroblasts (BCAFs), which represent most of the tumor mass. Moreover, the different components of the BCM highly interact with cancer cells, thereby generating a tightly intertwined network. In particular, BC cells activate recruited normal fibroblasts in BCAFs, which, in turn, promote BCM remodeling and metastasis. Thus, comparing the roles of normal fibroblasts and BCAFs in the physiological and metastatic processes, could provide a deeper understanding of the signaling pathways regulating BC dissemination. Here, we review the latest literature describing the structure of the mammary gland and the BCM and summarize the influence of epithelial-mesenchymal transition (EpMT) and autophagy in BC dissemination. Finally, we discuss the roles of fibroblasts and BCAFs in mammary gland development and BCM remodeling, respectively.

We aimed to establish the prevalence of the musculocutaneous nerve (MCN) variations and the probability of the variation being pure or mixed in the same plexus. We applied the principles of evidence-based anatomy to find, appraise, and synthesize data through a meta-analysis of anatomical studies. The variations were grouped based on the presence and location of the communicating branch with the median nerve and the origin of branches to anterior arm muscles. Forty-three cadaveric studies met the inclusion criteria, providing data from 4124 plexuses. The overall pooled prevalence of plexuses with MCN variations was 20%. Based on the classification applied in our study, the pooled prevalence of variations was 17% in region 1A, 20% in region 1B, 36% in region 2 and 49% in region 3. Importantly, 64.58% of variations in region 1A and 74.14% of variations in region 1B were mixed, that is, associated with a variation in another region. The odds of finding another variation in the presence of a variation in region 2 or 3 were equal 0.37 and 0.52, respectively, demonstrating a significantly lower probability of finding mixed variations involving these regions, when compared with region 1A. Variations of the MCN are most common in the part distal to the exit from within or beneath the coracobrachialis muscle. Proximal variations are more often associated with another variation located along the nerve. These findings can assist health care professionals in the treatment of brachial plexus lesions. Clin. Anat. 32:183-195, 2019. © 2018 Wiley Periodicals, Inc.

Breast cancer is the most common cancer in women, which incidence has increased in recent years. It is constituted by very heterogeneous tissue characterized by an abnormal microenvironment regulating tumor progression and providing evasion from cancer therapies. Breast cancer-associated fibroblasts (BCAFs) are the main cell type of breast cancer microenvironment and can represent up to 80% of the tumor mass. In particular, BCAFs induce cancer initiation, proliferation, invasion and metastasis by undergoing an activation process associated with the secretion of growth factors, cytokines, and paracrine interactions. Therapy resistance is the main cause of poor therapeutic results or even failure in breast cancer patients. Despite recent advances in breast cancer management, there is a need for new prognostic markers and novel agents for targeting key signalling pathways to either improve the efficacy of the current therapies, or reduce toxicity. In this view, BCAFs represent markers useful to clinical diagnosis, therapy, and prognosis of breast cancer. This review focuses on the role of BCAFs in cancer, and describes the processes of endocrine/chemotherapy resistance linked to BCAFs action. Moreover, it points to molecules and pathways regulating therapy resistance induced by BCAFs. Finally, potential therapeutic strategies targeting BCAFs and offering new tools in breast cancer therapy are highlighted.

The increasing availability of food supplements, aggressive media advertising, and common beliefs that these substances have only positive effects on health and sport performance indicate a need for continuous monitoring of this phenomenon. The aim of this study was to investigate the habits and beliefs related to diet supplementation among medical, health professional, and other university/high school students by means of a cross-sectional anonymous survey online. Among the respondents aware of supplements, 37.4% were taking or had taken them in the past. Food supplement use was more common among university students (in particular, those in health professional graduate courses) than high school students. Individual sport practice, rather than team sport, was associated with higher likelihood of food supplement use. Multivitamins were most commonly used, while weight-loss formulations were the least popular. Strikingly, filling nutrient gaps was statistically not considered the main reason for taking food supplements. Instead, they were used to enhance mental performance or enhance well-being. There was statistical evidence that students not enrolled in health or medical professional studies strongly agreed more often than medical students that taking food supplements prevents illness. These results indicate a striking difference between the evidence-based and personal reasons for food supplement use. Arguably, it calls for an improvement in education about diet supplementation and a change in attitude of health care providers to its implementation.

Objective Only a few anatomic studies of the entire course of the hypoglossal nerve (cranial nerve XII) have been reported. We analyzed all relationships of the 12th nerve with surrounding structures from the brainstem to the tongue through a microscopic perspective. A comprehensive anatomically and clinically oriented classification of its different segments is proposed. Methods Ten formalin-fixed adult human cadaveric heads (20 sides) were dissected with the aim to explore the entire course of cranial nerve XII via lateral suboccipital, far lateral partial, or total transcondylar routes. Different segments of the nerve were identified based on the hypoglossal course and its relationship with surrounding structures. Measurements of every portion of the nerve were taken in all specimens during dissection. Results The hypoglossal nerve was divided into 5 segments: cisternal, intracanalar, descending, horizontal, and ascending. Detailed and comprehensive examination of basic anatomic relationships through the view of different transcranial and endoscope-assisted approaches was performed. A new perspective of the hypoglossal canal is proposed, and the venous plexus surrounding the intracanalar segment of the nerve is described in detail. Conclusions Classification of 5 segments for the hypoglossal nerve seems anatomically valid, and it is surgically oriented with respect to all surgical approaches. Precise knowledge of the relationships with the surrounding structures may help to prevent some complications during surgery, and it is useful to explain, segment by segment, the pathogenic mechanisms for nerve injuries that are evidenced by lesions that exist along the entire intracranial and extracranial course.

Objective Different surgical approaches have been used over the years in order to access skull base. The endoscopic endonasal approach represents a direct and minimally invasive approach to the suprasellar, retrosellar, and retroclival space, with the advantage of avoid brain retraction and visualize safely and effectively the surgical target. The present contribution aims to provide anatomical details of the skull base as seen from below (i.e., via an endoscopic endonasal approach). Methods Five human cadaver heads were dissected. The anatomical neurovascular structures within the skull base were visualized and carefully described from an endoscopic endonasal view. The advantages and limitations of the endoscopic endonasal route were discussed as well. Results The entire skull base region, as seen from the endoscopic endonasal viewpoint, has been divided in 4 main regions: anterior skull base, middle skull base, posterior skull base and parasellar area. Conclusion The development of endoscopic techniques has opened different perspectives over the skull base surgery. Endonasal surgery provides access to a wide range of skull base lesions via a natural surgical corridor (i.e., the nasal cavities).

Myofibroblasts are activated fibroblasts, involved in tissue repair and cancer, characterized by de novo expression of alpha smooth muscle actin (α-SMA), increased secretion of growth factors and immunoregulatory phenotype [1]. At the end of wound healing myofibroblasts undergo apoptotic cell death, whereas in vitro they are also subjected to a programmed necrosis-like cell death, termed nemosis, associated with cyclooxygenase-2 (COX-2) expression induction and inflammatory response [1,2]. Moreover, fibroblasts form clusters during wound healing, fibrotic states and tumorigenesis. In this investigation, we produced and analysed clusters such as spheroids from human primary cutaneous myofibroblasts to evaluate apoptotic or necrotic cell death, inflammation and activation markers during myofibroblasts clustering. The spheroids formation does not induce apoptosis, necrotic cell death and COX-2 protein induction. The significant decrease of α-SMA in protein extracts of spheroids, the anti- migratory effect of spheroid-conditioned medium on normal cell lines and the absence of proliferation marker Ki-67 after 72 h of three-dimensional culture indicated that myofibroblasts undergo a deactivation process within spheroids. The cells of spheroids, reverted to adhesion growth, preserve their proliferation capability and are able to reacquire a myofibroblastic phenotype. Furthermore, the spontaneous formation of clusters and spheroids on plastic and glass substrates suggests that aggregates formation could be a physiological feature of cutaneous myofibroblasts. This study represents an experimental model to analyse myofibroblasts deactivation and indicates that fibroblast clusters could be a cell reservoir regulating tissue turnover., Italian Journal of Anatomy and Embryology, Vol. 122, No. 1 (Supplement) 2017

Myofibroblasts are activated fibroblasts involved in tissue repair and cancer. They are characterized by de novo expression of α-smooth muscle actin (α-SMA), immunoregulatory phenotype and paracrine interaction with normal and tumorigenic cells leading to cell proliferation. At the end of wound-healing myofibroblasts undergo apoptotic cell death, whereas in vitro-activated fibroblasts are also subjected to a programmed necrosis-like cell death, termed nemosis, associated with cyclooxygenase-2 (COX-2) expression induction and inflammatory response. Furthermore, myofibroblasts form clusters during wound healing, fibrotic states and tumorigenesis. In this study, we generated and analysed clusters such as spheroids from human primary cutaneous myofibroblasts, which represent a part of stromal microenvironment better than established cell lines. Therefore, we evaluated apoptotic or necrotic cell death, inflammation and activation markers during myofibroblasts clustering. The spheroids formation did not trigger apoptosis, necrotic cell death and COX-2 protein induction. The significant decrease of α-SMA in protein extracts of spheroids, the cytostatic effect exerted by spheroids conditioned medium on both normal and cancer cell lines and the absence of proliferation marker Ki-67 after 72 h of three-dimensional culture indicated that myofibroblasts have undergone a deactivation process within spheroids. The cells of spheroids reverted to adhesion growth preserved their proliferation capability and can re-acquire a myofibroblastic phenotype. Moreover, the spontaneous formation of clusters on plastic and glass substrates suggests that aggregates formation could be a physiological feature of cutaneous myofibroblasts. This study represents an experimental model to analyse myofibroblasts deactivation and suggests that fibroblast clusters could be a cell reservoir regulating tissues turnover.

Introduction Lateral epicondylitis is a common painful elbow disorder. Several approaches to treatment have been proposed, with a local injection of corticosteroids being the most frequently used. Recent insights into the pathophysiology encouraged the introduction of autologous blood injections as an alternative treatment method. The aim of this meta-analysis is to summarize quantitatively the evidence regarding the efficacy of corticosteroids and autologous blood injections for treatment of pain in lateral epicondylitis. Evidence acquisition Studies were considered eligible based on the following inclusion criteria: adult human, diagnosis of lateral epicondylitis, randomized controlled trials comparing corticosteroids versus autologous blood injections, pain assessment. Exclusion criteria were previous surgery for lateral epicondylitis or for other elbow disorders, concurrent treatment with drugs or physiotherapy, diagnosis of musculoskeletal systemic disorder. A systematic search of literature was performed according to the PRISMA statement. Effect size of each included study was calculated and analyzed in a random-effects model. Evidence synthesis Four studies, enrolling total of 218 patients (139 females and 79 males), were included in quantitative analysis. At 2 weeks, there was a trend towards a reduction of VAS score in the corticosteroid group (WMD=2.12 [95% CI: 4.38 to 0.14], P=0.07). No significant differences were recorded in the medium-term (4-12 weeks; WMD=0.85 [95% CI: -0.44 to 2.15], P=0.19) and long-term (24 weeks; WMD=0.63 [95% CI: -2.40 to 3.66], P=0.68) follow-up. Conclusions Few high-quality trials compare the efficacy of corticosteroid and autologous blood injections in the control of pain related to lateral epicondylitis. Available data indicate that corticosteroids tend to reduce VAS score in short-term follow-up, although these data are not statistically significant. No differences were recorded in the medium and long term. Contrary to popular opinion among medical professionals, and despite pathophysiological cues, the currently available data offer no support for the effectiveness of autologous blood injections in medium- and long-term follow-up. Further studies are necessary to establish which treatment has more impact on pain in lateral epicondylitis. These data could be then used as a basis for practical guidelines and new protocols of treatment.