Evaluating the impact of distance learning on gender-affirming healthcare competence: knowledge acquisition and satisfaction among healthcare professionals in Italy.

Background: Transgender and gender diverse (TGD) individuals face significant healthcare barriers, with one of the most critical being the inadequate knowledge and skills of healthcare professionals (HCPs) in TGD health. To address this issue, we undertook a project to develop a distance learning course for all healthcare professions, encompassing a comprehensive range of topics related to TGD health issues. Objectives: This study aimed to evaluate the impact of a course on gender-affirming healthcare competence, with a focus on knowledge acquisition and satisfaction levels. The hypothesis was that participating in the course would enhance the participants' knowledge on the covered topics. Methods: A distance learning course, designed for all Continuing Medical Education professions, was conducted between March and September 2023. The course was structured according to the Problem-Based Learning methodology. We implemented a pre-test vs. post-test study design to evaluate the enhancement of knowledge, based on a set of Multiple Choice Questions (MCQs), and investigated users' satisfaction through the administration of a semi-structured questionnaire. We examined the pre- and post-course proportions of correct responses to questions, along with the mean score difference, categorized by learners' sex, age, and geographical area. Eventually, a Satisfaction Training Index was created. Results: The maximum capacity was reached, with 29,998 out of 30,000 available spots filled. Of those enrolled, 18,282 HCPs successfully completed the training. Post-test results revealed an increase in correct answers across all MCQs, with overall mean score rising from 48.8 to 68.0 (p < 0.001). Stratified analysis indicated improvements across all participant categories. A higher average increase among female (19.87) compared to male enrollees (17.06) was detected (p < 0.001). Both "over 55" and "46-55" age groups showed the greatest score increases compared to "35-46" and "under 35" groups, despite no significant differences in pre-test scores. Course satisfaction was high, averaging 4.38 out of 5. Top-rated aspects included "learning new concepts" (4.49), "accessibility" (4.46), and "platform functionality" (4.46). Conclusion: Our research hypothesis was confirmed by the significant increase in knowledge going from pre-test to post-test and by the high level of user satisfaction. The obtained results serve as a foundation for planning additional professional education in TGD health.

Back to the Future: Is GnRHa Treatment in Transgender and Gender Diverse Adolescents Only an Extended Evaluation Phase?

CONTEXT: The role of body modifications induced by gonadal suppression in transgender and gender diverse adolescents on psychological functioning has not yet been evaluated. OBJECTIVE: The main aim of the present study was to explore several hormone, physical and psychological functioning changes during gonadotropin-releasing hormone analog (GnRHa) treatment in transgender and gender diverse adolescents (TGDAs). The potential relationship between the physical and hormone effects of GnRHa and psychological well-being, along with its magnitude, was assessed for the first time. METHODS: This prospective multidisciplinary study included 36 TGDA (22 assigned female at birth, and 14 assigned male at birth) who received psychological assessment followed by triptorelin prescription after referring to the Florence Gender Clinic. This study consisted of 3 time points: first referral (T0), psychological assessment (T1); and treatment with intramuscular injections of triptorelin for 3 up to 12 months (T2). Psychometric questionnaires were administered at each time point, and clinical and biochemical evaluations were performed at T1 and T2. RESULTS: The following results were found: (1) GnRHa showed efficacy in inhibiting puberty progression in TGDAs; (2) an increase in psychopathology was observed before starting GnRHa (T1) compared with baseline levels; (3) during GnRHa treatment (T2), a significant improvement in psychological functioning, as well as decrease in suicidality, body uneasiness, depression, and anxiety levels were observed; (4) hormone and physical changes (in terms of gonadotropin and sex steroid levels, height and body mass index percentiles, waist-hip ratio, and acne severity) observed during triptorelin treatment significantly correlated with a reduction in suicidal ideation, anxiety, and body image concerns. CONCLUSION: Psychological improvement in TGDA on GnRHa seems to be related to the objective body changes induced by a GnRHa. Therefore, the rationale for treatment with a GnRHa may not only be considered an extension of the evaluation phase, but also the start of a medical (even if reversible) gender-affirming path, especially in TGDAs whose puberty has already progressed.

Sex and gender differences in migraines: a narrative review.

INTRODUCTION: Gender medicine is a new medical approach aimed at the study of the differences between women and men in terms of prevention, diagnosis, and the outcome of all diseases. Migraines are among these. They represent the most common neurological illness; they are most prevalent in adults between 20 and 50 years of age and are three to four times more frequent in woman than in men. Affecting people in working age, migraines are a problem that strongly impacts the psychophysical health and productivity of workers, regardless of the specific job task they have. METHODS: A narrative review was performed, searching for the most relevant articles describing gender differences in people suffering from migraines, and particularly in workers. RESULTS: Migraine global prevalence is 20.7% in women and 9.7% in men whereas prevalence in Italy is 32.9% for women and only 13.0% for men. This difference is partly explained by hormonal differences, as well as by differences in brain structure, genetic polymorphisms and neuronal pathways. Sex differences may also play a role in the progression from episodic to chronic migraine. In workers, migraines are mostly associated with strenuous physical work in men, whilst migraines triggered by night shifts, lack of sleep, or irregular sleep patterns are more common in women. CONCLUSIONS: To this day, the reasons of sex/gender disparity for migraine are still obscure. However, migraines, chronic migraine in particular, have a negative impact on the lives of all individuals affected by this disease, but particularly in women in which family cares and working activity are often superimposed. Migraine prevention strategies should be planned in workers through the occupational health physician.

The Natural Estrogen Receptor Beta Agonist Silibinin as a Promising Therapeutic Tool in Diffuse Large B-cell Lymphoma.

BACKGROUND/AIM: About 40% of patients with diffuse large cell lymphoma (DLBCL) still have a poor prognosis. Additionally, DLBCL patients treated with doxorubicin are at risk of cardiac failure. Growing evidence suggests an antitumor and cardioprotective activity exerted by estrogen via its binding to estrogen receptor (ER) ß. The aim of this study was to evaluate the anticancer activity of the phytoestrogen silibinin, an ERß selective agonist, on DLBCL growth, and its potential cardioprotective effect. MATERIALS AND METHODS: DLBCL cell lines SUDHL-8, SUDHL-6, and RIVA were used. The anti-tumor activity of silibinin was also evaluated in vivo in NOD/SCID/IL2Rg-/- (NSG) xenografted mice. AC16 human ventricular cardiomyocytes were used to investigate the cardioprotective effects of silibinin. RESULTS: In vitro silibinin induced apoptosis and autophagy, and blocked tumor cell proliferation, also protecting AC16 cardiomyocytes from doxorubicin-induced toxicity. In vivo silibinin induced cell death and autophagy, and reduced tumor volume. CONCLUSION: Silibinin represents a promising therapeutic tool.

On the role of sphingolipids in cell survival and death.

Sphingolipids, universal components of biological membranes of all eukaryotic organisms, from yeasts to mammals, in addition of playing a structural role, also play an important part of signal transduction pathways. They participate or, also, ignite several fundamental subcellular signaling processes but, more in general, they directly contribute to key biological activities such as cell motility, growth, senescence, differentiation as well as cell fate, i.e., survival or death. The sphingolipid metabolic pathway displays an intricate network of reactions that result in the formation of multiple sphingolipids, including ceramide, and sphingosine-1-phosphate. Different sphingolipids, that have key roles in determining cell fate, can induce opposite effects: as a general rule, sphingosine-1-phosphate promotes cell survival and differentiation, whereas ceramide is known to induce apoptosis. Furthermore, together with cholesterol, sphingolipids also represent the basic lipid component of lipid rafts, cholesterol- and sphingolipid-enriched membrane microdomains directly involved in cell death and survival processes. In this review, we briefly describe the characteristics of sphingolipids and lipid membrane microdomains. In particular, we will consider the involvement of various sphingolipids per se and of lipid rafts in apoptotic pathway, both intrinsic and extrinsic, in nonapoptotic cell death, in autophagy, and in cell differentiation. In addition, their roles in the most common physiological and pathological contexts either as pathogenetic elements or as biomarkers of diseases will be considered. We would also hint how the manipulation of sphingolipid metabolism could represent a potential therapeutic target to be investigated and functionally validated especially for those diseases for which therapeutic options are limited or ineffective.

Educational impact of hand motion analysis in the evaluation of FAST examination skills.

PURPOSE: Increasing pressure pushes towards the objective competence assessment of clinical operators. Hand motion analysis (HMA) was introduced to measure surgical and clinical procedures; its recent application to FAST examinations leaves unsolved issues. This study aimed at determining optimal HMA parameters to discriminate between operators' skill levels, and which FAST tasks are experience-dependent. METHODS: Ten experienced (EG) and 13 beginner (BG) sonographers performed a FAST examination on one female and one male model. A motion capture system returned the duration, working volume, number of movements (absolute and time normalized), and hand path length (absolute and time normalized) of each view. RESULTS: BG took more time in completing specific views, with a higher working volume (p = 0.003) and longer hands path (p < 0.001). The number of movements was lower in the EG (p < 0.001) and differed between views (p = 0.014). No significant Group/Model differences were found for the normalized number of movements. The LUQ view required a higher number of movements (p < 0.001). CONCLUSIONS: HMA identified kinematic parameters discriminating between proficiency level and critical subtasks in the FAST examination. These findings could be the base for a focused HMA-based evaluation of performances following a proctored training period. There is room to incorporate HMA into simulation metrics and evidence-based credentialing standards for clinical ultrasound applications.

Trehalose administration in C57BL/6N old mice affects healthspan improving motor learning and brain anti-oxidant defences in a sex-dependent fashion: a pilot study.

Aim of this study was to characterize the effects of oral trehalose administration (2%w/v) on healthspan in old mice. Trehalose was administered in drinking water for 1 month to male and female C57BL/6N mice aged 25-months. After behavioral phenotyping (grip strength, beam walking and rotarod tests), autophagy (LC3-II/actin) and oxidative stress were tested in the cerebral cortex and gastrocnemius muscle. The latter parameter was indirectly assessed by evaluating carbonyl groups added to proteins as a result of oxidative reactions, in addition to central levels of NRF2 protein, a transcription factor that regulates the expression of antioxidant enzymes. In comparison with sex-matched controls, trehalose-treated males performed better in motor planning and coordination tasks. This behavioral phenotype was associated with an activation of the ubiquitin-proteasome system, autophagy and antioxidant defences in cerebral cortex. Independently from trehalose administration, females were characterized by better motor performance and showed higher levels of ubiquitinated proteins and NRF2 in cerebral cortex, suggesting an up-regulation of basal antioxidant defences. In conclusion, trehalose was effective in counteracting some aspects of age-related decay, with specific effects in male and female subjects.

Not only FAST The MUSEC® experience in training surgeons.

For a long time surgeons have been discussing the need to improve their skills in the use of ultrasound (US). However in the recent years it has become evident the importancxe for general aklnd trauma surgeons treating critically-ill patients to learn basic and advanced US. The two last editions (9th and 10th) of the ATLS manual have officially included FAST and e-FAST in the primary assessment of trauma patients, making this tool an essential skill for surgeons. In the acute care setting FAST, e-FAST and other applications have gained a pivotal, evidence-based role in this fields. Nevertheless, surgeons are rarely performing US exams by themselves, losing a major decision-making tool. The Modular Ultrasound ESTES Course (MUSEC®) was developed to provide both fundamental and advanced US training for surgeons in trauma and acute care settings. We are strongly convinced, in the light of the results from both the surveys carried out and the customer satisfaction tests administered to all the participants in the MUSEC courses, that US courses such as these should be part of the general surgery residency programs. KEY WORDS: e-FAST, MUSEC Ultrasound in Emergency Department, Ultrasound Training Trauma Patients.

Inflammatory cytokines associated with cancer growth induce mitochondria and cytoskeleton alterations in cardiomyocytes.

Cardiac dysfunction is often observed in patients with cancer also representing a serious problem limiting chemotherapeutic intervention and even patient survival. In view of the recently established role of the immune system in the control of cancer growth, the present work has been undertaken to investigate the effects of a panel of the most important inflammatory cytokines on the integrity and function of mitochondria, as well as of the cytoskeleton, two key elements in the functioning of cardiomyocytes. Either mitochondria features or actomyosin cytoskeleton organization of in vitro-cultured cardiomyocytes treated with different inflammatory cytokines were analyzed. In addition, to investigate the interplay between tumor growth and cardiac function in an in vivo system, immunocompetent female mice were inoculated with cancer cells and treated with the chemotherapeutic drug doxorubicin at a dosing schedule able to suppress tumor growth without inducing cardiac alterations. Analyses carried out in cardiomyocytes treated with the inflammatory cytokines, such as tumor necrosis factor α (TNF-α), interferon γ (IFN-γ), interleukin 6 (IL-6), IL-8, and IL-1ß revealed severe phenotypic changes, for example, of contractile cytoskeletal elements, mitochondrial membrane potential, mitochondrial reactive oxygen species production and mitochondria network organization. Accordingly, in immunocompetent mice, the tumor growth was accompanied by increased levels of the inflammatory cytokines TNF-α, IFN-γ, IL-6, and IL-8, either in serum or in the heart tissue, together with a significant reduction of ventricular systolic function. The alterations of mitochondria and of microfilament system of cardiomyocytes, due to the systemic inflammation associated with cancer growth, could be responsible for remote cardiac injury and impairment of systolic function observed in vivo.

Inhibition of autophagy increases susceptibility of glioblastoma stem cells to temozolomide by igniting ferroptosis.

The role of autophagy in cancer onset and progression appears still controversial. On one hand, autophagy allows cancer cell to survive in unfavorable environmental conditions, on the other hand, once internal energy resources are exhausted, it leads to cell death. In addition, autophagy interpheres with cell cycle progression, de facto exerting a cytostatic activity. Hence, it represents an important target for anticancer therapy. For example, temozolomide (TMZ), of use for glioblastoma (GBM) treatment, appears as capable of inducing autophagy partially inhibiting cancer cell proliferation. However, GBM, a very aggressive brain tumor with poor prognosis even after surgery and radio-chemotherapy, invariably recurs and leads to patient death. Since cancer stem cells have been hypothesized to play a role in refractory/relapsing cancers, in the present work we investigated if autophagy could represent a constitutive cytoprotection mechanism for glioblastoma stem-like cells (GSCs) and if the modulation of autophagic process could affect GBM growth and survival. Thus, in the present study we first evaluated the relevance of autophagy in GBM tumor specimens, then its occurrence in GSCs and, finally, if modulation of autophagy could influence GSC response to TMZ. Our results suggested that, in vitro, the impairing autophagic process with quinacrine, a compound able to cross the blood-brain barrier, increased GSC susceptibility to TMZ. Death of GSCs was apparently due to the iron dependent form of programmed cell death characterized by the accumulation of lipid peroxides called ferroptosis. These results underscore the relevance of the modulation of autophagy in the GSC survival and death and suggest that triggering of ferroptosis in GSCs could represent a novel and important target for the management of glioblastoma.

Recruitment of mitofusin 2 into "lipid rafts" drives mitochondria fusion induced by Mdivi-1.

The regulation of the mitochondrial dynamics and the balance between fusion and fission processes are crucial for the health and fate of the cell. Mitochondrial fusion and fission machinery is controlled by key proteins such as mitofusins, OPA-1 and several further molecules. In the present work we investigated the implication of lipid rafts in mitochondrial fusion induced by Mdivi-1. Our results underscore the possible implication of lipid "rafts" in mitochondrial morphogenetic changes and their homeostasis.

Preclinical models in the study of sex differences.

The biology of sex differences deals with the study of the disparities between females and males and the related biological mechanisms. Gender medicine focuses on the impact of gender and sex on human physiology, pathophysiology and clinical features of diseases that are common to women and men. The term gender refers to a complex interrelation and integration of sex-as a biological and functional determinant-and psychological and cultural behaviours (due to ethnical, social or religious background). The attention to the impact of gender differences on the pathophysiology and, therefore, on the clinical management of the most common diseases, such as cardiovascular diseases (CVD), neurodegenerative disorders, immune and autoimmune diseases as well as several tumours, is in fact often neglected. Hence, studies covering different fields of investigation and including sex differences in the pathogenesis, in diagnostic and prognostic criteria as well as in response to therapy appear mandatory. However, prerequisites for this development are preclinical studies, including in vitro and in vivo approaches. They represent the first step in the development of a drug or in the comprehension of the pathogenetic mechanisms of diseases, in turn a necessary step for the development of new or more appropriate therapeutic strategies. However, sex differences are still poorly considered and the great majority of preclinical studies do not take into account the relevance of such disparities. In this review, we describe the state of the art of these studies and provide some paradigmatic examples of key fields of investigation, such as oncology, neurology and CVD, where preclinical models should be improved.

Evidence for the involvement of lipid rafts localized at the ER-mitochondria associated membranes in autophagosome formation.

Mitochondria-associated membranes (MAMs) are subdomains of the endoplasmic reticulum (ER) that interact with mitochondria. This membrane scrambling between ER and mitochondria appears to play a critical role in the earliest steps of autophagy. Recently, lipid microdomains, i.e. lipid rafts, have been identified as further actors of the autophagic process. In the present work, a series of biochemical and molecular analyses has been carried out in human fibroblasts with the specific aim of characterizing lipid rafts in MAMs and to decipher their possible implication in the autophagosome formation. In fact, the presence of lipid microdomains in MAMs has been detected and, in these structures, a molecular interaction of the ganglioside GD3, a paradigmatic "brick" of lipid rafts, with core-initiator proteins of autophagy, such as AMBRA1 and WIPI1, was revealed. This association seems thus to take place in the early phases of autophagic process in which MAMs have been hypothesized to play a key role. The functional activity of GD3 was suggested by the experiments carried out by knocking down ST8SIA1 gene expression, i.e., the synthase that leads to the ganglioside formation. This experimental condition results in fact in the impairment of the ER-mitochondria crosstalk and the subsequent hindering of autophagosome nucleation. We thus hypothesize that MAM raft-like microdomains could be pivotal in the initial organelle scrambling activity that finally leads to the formation of autophagosome.

AMBRA1 and SQSTM1 expression pattern in prostate cancer.

Prostate cancer is among the most commonly diagnosed male diseases and a leading cause of cancer mortality in men. There is emerging evidence that autophagy plays an important role in malignant cell survival and offers protection from the anti-cancer drugs in prostate cancer cells. AMBRA1 and the autophagic protein sequestosome-1 (SQSTM1; p62) expression were evaluated by immunohistochemistry and western blot on tissue samples from both benign and malignant prostatic lesions. The data reported in this pilot study demonstrated an increased expression of AMBRA1 and SQSTM1, which were also associated with an accumulation of LC3II in prostate cancer but not in benign lesion. In the present study we found that: (i) at variance with benign lesion, prostate cancer cells underwent SQSTM1 accumulation, i.e., clearly displayed a defective autophagic process but, also, (ii) prostate cancer accumulated AMBRA1 and (iii) this increase positively correlated with the Gleason score. These results underscore a possible implication of autophagy in prostate cancer phenotype and of AMBRA1 as possible cancer progression biomarker in this malignancy.

Autophagic flux and autophagosome morphogenesis require the participation of sphingolipids.

Apoptosis and autophagy are two evolutionary conserved processes that exert a critical role in the maintenance of tissue homeostasis. While apoptosis is a tightly regulated cell program implicated in the removal of damaged or unwanted cells, autophagy is a cellular catabolic pathway that is involved in the lysosomal degradation and recycling of proteins and organelles, and is thereby considered an important cytoprotection mechanism. Sphingolipids (SLs), which are ubiquitous membrane lipids in eukaryotes, participate in the generation of various membrane structures, including lipid rafts and caveolae, and contribute to a number of cellular functions such as cell proliferation, apoptosis and, as suggested more recently, autophagy. For instance, SLs are hypothesized to be involved in several intracellular processes, including organelle membrane scrambling, whilst at the plasma membrane lipid rafts, acting as catalytic domains, strongly contribute to the ignition of critical signaling pathways determining cell fate. In particular, by targeting several shared regulators, ceramide, sphingosine-1-phosphate, dihydroceramide, sphingomyelin and gangliosides seem able to differentially regulate the autophagic pathway and/or contribute to the autophagosome formation. This review illustrates recent studies on this matter, particularly lipid rafts, briefly underscoring the possible implication of SLs and their alterations in the autophagy disturbances and in the pathogenesis of some human diseases.

Mineralocorticoid receptor antagonism induces browning of white adipose tissue through impairment of autophagy and prevents adipocyte dysfunction in high-fat-diet-fed mice.

The mineralocorticoid receptor (MR) controls adipocyte function, but its role in the conversion of white adipose tissue (WAT) into thermogenic fat has not been elucidated. We investigated responses to the MR antagonists spironolactone (spiro; 20 mg/kg/d) and drospirenone (DRSP; 6 mg/kg/d) in C57BL/6 mice fed a high-fat (HF) diet for 90 d. DRSP and spiro curbed HF diet-induced impairment in glucose tolerance, and prevented body weight gain and white fat expansion. Notably, either MR antagonist induced up-regulation of brown adipocyte-specific transcripts and markedly increased protein levels of uncoupling protein 1 (UCP1) in visceral and inguinal fat depots when compared with the HF diet group. Positron emission tomography and magnetic resonance spectroscopy confirmed acquisition of brown fat features in WAT. Interestingly, MR antagonists markedly reduced the autophagic rate both in murine preadipocytes in vitro (10(-5) M) and in WAT depots in vivo, with a concomitant increase in UCP1 protein expression. Moreover, the autophagy repressor bafilomycin A1 (10(-8) M) mimicked the effect of MR antagonists, increasing UCP1 protein expression in primary preadipocytes. Hence, we showed that adipocyte MR regulates brown remodeling of WAT through a modulation of autophagy. These results provide a rationale for the use of MR antagonists to prevent the adverse metabolic consequences of adipocyte dysfunction.

Evidence for the involvement of GD3 ganglioside in autophagosome formation and maturation.

Sphingolipids are structural lipid components of cell membranes, including membrane of organelles, such as mitochondria or endoplasmic reticulum, playing a role in signal transduction as well as in the transport and intermixing of cell membranes. Sphingolipid microdomains, also called lipid rafts, participate in several metabolic and catabolic cell processes, including apoptosis. However, the defined role of lipid rafts in the autophagic flux is still unknown. In the present study we analyzed the role of gangliosides, a class of sphingolipids, in autolysosome morphogenesis in human and murine primary fibroblasts by means of biochemical and analytical cytology methods. Upon induction of autophagy, by using amino acid deprivation as well as tunicamycin, we found that GD3 ganglioside, considered as a paradigmatic raft constituent, actively contributed to the biogenesis and maturation of autophagic vacuoles. In particular, fluorescence resonance energy transfer (FRET) and coimmunoprecipitation analyses revealed that this ganglioside interacts with phosphatidylinositol 3-phosphate and can be detected in immature autophagosomes in association with LC3-II as well as in autolysosomes associated with LAMP1. Hence, it appears as a structural component of autophagic flux. Accordingly, we found that autophagy was significantly impaired by knocking down ST8SIA1/GD3 synthase (ST8 α-N-acetyl-neuraminide α-2,8-sialyltransferase 1) or by altering sphingolipid metabolism with fumonisin B1. Interestingly, exogenous administration of GD3 ganglioside was capable of reactivating the autophagic process inhibited by fumonisin B1. Altogether, these results suggest that gangliosides, via their molecular interaction with autophagy-associated molecules, could be recruited to autophagosome and contribute to morphogenic remodeling, e.g., to changes of membrane curvature and fluidity, finally leading to mature autolysosome formation.

Antioxidants counteract lipopolysaccharide-triggered alterations of human colonic smooth muscle cells.

Gut dysmotility develops in individuals during and after recovering from infective acute gastroenteritis and it is apparently due to a direct effect of circulating lipopolysaccharides (LPS). This is an endotoxin with a prooxidant activity derived from gram-negative bacteria. Due to the lack of human models available so far, the mechanisms underlying LPS-induced gut dysmotility are, however, poorly investigated. In the present work long-term effects of LPS and their reversibility have been assessed by means of different analytical cytology methods on pure primary cultures of human colonic smooth muscle cells. We found that LPS triggered the following alterations: (i) a redox imbalance with profound changes of contractile microfilament network, and (ii) the induction of cell cycle progression with dedifferentiation from a contractile to a synthetic phenotype. These alterations persisted also after LPS removal. Importantly, two unrelated antioxidants, alpha-tocopherol and N-acetylcysteine, were able to reverse the cytopathic effects of LPS and to restore normal muscle cell function. The present data indicate that LPS is capable of triggering a persistent and long-term response that could contribute to muscle dysfunction occurring after an infective and related inflammatory burst and suggest a reappraisal of antioxidants in the management of postinfective motor disorders of the gut.