Modeling hypothalamic pathophysiology in vitro for metabolic, circadian, and sleep disorders.

The hypothalamus, a small and intricate brain structure, orchestrates numerous neuroendocrine functions through specialized neurons and nuclei. Disruption of this complex circuitry can result in various diseases, including metabolic, circadian, and sleep disorders. Advances in in vitro models and their integration with new technologies have significantly benefited research on hypothalamic function and pathophysiology. We explore existing in vitro hypothalamic models and address their challenges and limitations as well as translational findings. We also highlight how collaborative efforts among multidisciplinary teams are essential to develop relevant and translational experimental models capable of replicating intricate neural circuits and neuroendocrine pathways, thereby advancing our understanding of therapeutic targets and drug discovery in hypothalamus-related disorders.

Orogenital and anal infection by Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, and other sexually transmitted infections in men who have sex with men in Lisbon.

BACKGROUND: Men who have sex with men (MSM) are at risk for sexually transmitted infections (STIs), but more data on extragenital carriage are needed. AIM: We assessed the genital and extragenital prevalence of bacterial and other STIs in MSM in a Lisbon sexual health clinic. METHODS: We screened oral, anal, and urine samples of MSM visiting the GAT-CheckpointLX clinic June 2017-December 2021 for Chlamydia trachomatis (including lymphogranuloma venereum, LGV), Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis, Mycoplasma hominis, Ureaplasma urealyticum, and U. parvum. Ano-oro-genital lesions were tested for LGV, Treponema pallidum, and Herpes Simplex Virus. Blood was tested for HIV and T. pallidum antibodies. RESULTS: N. gonorrhoeae was found in 16.6% of the MSM followed by C. trachomatis (13.2%), M. genitalium (10.3%) and T. vaginalis (0.2%). The most frequent occurrence was anorectal (C. trachomatis, M. genitalium) and oral (N. gonorrhoeae). We found high carriage of U. urealyticum (36.1%) and M. hominis (22.1%). LGV was detected in 21.8% of chlamydia-positive anorectal swabs. Syphilis was detected in 22.6% of tested MSM, while 13.8% had HIV. Gonorrhoea and chlamydia were significantly more prevalent in MSM with concomitant HIV or syphilis. CONCLUSION: The substantial extragenital prevalence of bacterial STIs in MSM, and HIV and syphilis coinfections, suggest screening has value in identifying hidden carriage and in contributing for providing better care.

Impaired neuron differentiation in GBA-associated Parkinson's disease is linked to cell cycle defects in organoids.

The mechanisms underlying Parkinson's disease (PD) etiology are only partially understood despite intensive research conducted in the field. Recent evidence suggests that early neurodevelopmental defects might play a role in cellular susceptibility to neurodegeneration. To study the early developmental contribution of GBA mutations in PD we used patient-derived iPSCs carrying a heterozygous N370S mutation in the GBA gene. Patient-specific midbrain organoids displayed GBA-PD relevant phenotypes such as reduction of GCase activity, autophagy impairment, and mitochondrial dysfunction. Genome-scale metabolic (GEM) modeling predicted changes in lipid metabolism which were validated with lipidomics analysis, showing significant differences in the lipidome of GBA-PD. In addition, patient-specific midbrain organoids exhibited a decrease in the number and complexity of dopaminergic neurons. This was accompanied by an increase in the neural progenitor population showing signs of oxidative stress-induced damage and premature cellular senescence. These results provide insights into how GBA mutations may lead to neurodevelopmental defects thereby predisposing to PD pathology.

Extra-Neural Metastases of Glioblastoma: A Case Series.

Glioblastoma (GBM) is the most common brain tumor and has a median survival of less than two years. The current standard of care consists of surgery followed by concomitant radio and chemotherapy with temozolomide. Although it is an aggressive tumor, distant metastases are extremely rare. The dissemination mechanisms are not fully understood and currently there is no standard of care for its treatment. In this study, the authors present a comprehensive analysis of all the existing cases of extra-neural metastases of GBM in a tertiary care center over the last five years, along with the procedures carried out in each case.

Innovative three-dimensional models for understanding mechanisms underlying lung diseases: powerful tools for translational research.

Chronic lung diseases result from alteration and/or destruction of lung tissue, inevitably causing decreased breathing capacity and quality of life for patients. While animal models have paved the way for our understanding of pathobiology and the development of therapeutic strategies for disease management, their translational capacity is limited. There is, therefore, a well-recognised need for innovative in vitro models to reflect chronic lung diseases, which will facilitate mechanism investigation and the advancement of new treatment strategies. In the last decades, lungs have been modelled in healthy and diseased conditions using precision-cut lung slices, organoids, extracellular matrix-derived hydrogels and lung-on-chip systems. These three-dimensional models together provide a wide spectrum of applicability and mimicry of the lung microenvironment. While each system has its own limitations, their advantages over traditional two-dimensional culture systems, or even over animal models, increases the value of in vitro models. Generating new and advanced models with increased translational capacity will not only benefit our understanding of the pathobiology of lung diseases but should also shorten the timelines required for discovery and generation of new therapeutics. This article summarises and provides an outline of the European Respiratory Society research seminar "Innovative 3D models for understanding mechanisms underlying lung diseases: powerful tools for translational research", held in Lisbon, Portugal, in April 2022. Current in vitro models developed for recapitulating healthy and diseased lungs are outlined and discussed with respect to the challenges associated with them, efforts to develop best practices for model generation, characterisation and utilisation of models and state-of-the-art translational potential.

RUFY3 regulates endolysosomes perinuclear positioning, antigen presentation and migration in activated phagocytes.

Endo-lysosomes transport along microtubules and clustering in the perinuclear area are two necessary steps for microbes to activate specialized phagocyte functions. We report that RUN and FYVE domain-containing protein 3 (RUFY3) exists as two alternative isoforms distinguishable by the presence of a C-terminal FYVE domain and by their affinity for phosphatidylinositol 3-phosphate on endosomal membranes. The FYVE domain-bearing isoform (iRUFY3) is preferentially expressed in primary immune cells and up-regulated upon activation by microbes and Interferons. iRUFY3 is necessary for ARL8b + /LAMP1+ endo-lysosomes positioning in the pericentriolar organelles cloud of LPS-activated macrophages. We show that iRUFY3 controls macrophages migration, MHC II presentation and responses to Interferon-γ, while being important for intracellular Salmonella replication. Specific inactivation of rufy3 in phagocytes leads to aggravated pathologies in mouse upon LPS injection or bacterial pneumonia. This study highlights the role of iRUFY3 in controlling endo-lysosomal dynamics, which contributes to phagocyte activation and immune response regulation.

Alterations of bovine nucleus pulposus cells with aging.

Aging is one of the major etiological factors driving intervertebral disc (IVD) degeneration, the main cause of low back pain. The nucleus pulposus (NP) includes a heterogeneous cell population, which is still poorly characterized. Here, we aimed to uncover main alterations in NP cells with aging. For that, bovine coccygeal discs from young (12 months) and old (10-16 years old) animals were dissected and primary NP cells were isolated. Gene expression and proteomics of fresh NP cells were performed. NP cells were labelled with propidium iodide and analysed by flow cytometry for the expression of CD29, CD44, CD45, CD146, GD2, Tie2, CD34 and Stro-1. Morphological cell features were also dissected by imaging flow cytometry. Elder NP cells (up-regulated bIL-6 and bMMP1 gene expression) presented lower percentages of CD29+, CD44+, CD45+ and Tie2+ cells compared with young NP cells (upregulated bIL-8, bCOL2A1 and bACAN gene expression), while GD2, CD146, Stro-1 and CD34 expression were maintained with age. NP cellulome showed an upregulation of proteins related to endoplasmic reticulum (ER) and melanosome independently of age, whereas proteins upregulated in elder NP cells were also associated with glycosylation and disulfide bonds. Flow cytometry analysis of NP cells disclosed the existence of 4 subpopulations with distinct auto-fluorescence and size with different dynamics along aging. Regarding cell morphology, aging increases NP cell area, diameter and vesicles. These results contribute to a better understanding of NP cells aging and highlighting potential anti-aging targets that can help to mitigate age-related disc disease.

Impact of Donor Age on Long-Term Outcomes in Simultaneous Pancreas-Kidney Transplantation.

BACKGROUND: Simultaneous pancreas-kidney transplantation (SPKT) is the treatment of choice for type 1 diabetes patients with end-stage renal disease. Donor characteristics are determinants of graft and patient survival. We aimed to study the impact of donor age on outcomes in SPKT. METHODS: We retrospectively studied 254 patients submitted to SPKT between 2000 and 2021. Patients were classified as "younger donor" (donor age <40 years) and "older donor" (donor age ≥40 years). RESULTS: Fifty-three patients received grafts from older donors. Pancreas graft survival rates at 1, 5, 10, and 15 years were 89%, 83%, 77%, and 73% in the younger donor group, respectively, and 77%, 73%, 67%, and 62% in the older donor group, respectively (P = .052). Older donors and previous major adverse cardiovascular events (MACEs) were associated with pancreas graft failure at 15 years. Kidney transplant survival (1, 5, 10, and 15 years) was lower in the older donor cohort (94%, 92%, 69%, 60% vs 97%, 94%, 89%, and 84%, respectively; P = .004). Older donor, recipient age, and previous MACE predicted kidney graft failure at 15 years. Patient survival rates at 1, 5, 10, and 15 years were 98%, 95%, 91%, and 81% in the younger donor group, respectively, versus 92%, 90%, 84%, and 72% in the older donor group, respectively (P = .127). CONCLUSIONS: The kidney graft survival rate was lower in the older donor group, whereas pancreas graft survival and patient survival did not differ significantly. Multivariate analysis showed that a donor age of ≥40 years was an independent predictor of pancreas and kidney graft failure at 15 years in SPKT patients.

Corrigendum: Ionic-liquid-based approaches to improve biopharmaceuticals downstream processing and formulation.

[This corrects the article DOI: 10.3389/fbioe.2023.1037436.].

Cerebral Air Embolism: A Case of a Rare Transthoracic Needle Biopsy Complication.

Transthoracic needle biopsy (TNB) is a fundamental procedure in the diagnosis of a wide spectrum of thoracic diseases replacing more invasive surgical procedures. The procedure may be performed with computed tomography (CT) or ultrasound imaging guidance, with CT being the more commonly utilized. Although less invasive than surgery, there is still a complication risk associated with this procedure. These can be local such as pneumothorax, parenchymal hemorrhage, tumor seeding, and hemoptysis, or systemic such as air embolism. The authors report a case of cerebral circulation air embolism as a complication of TNB in a 54-year-old male with suspected lung tumor followed by a brief review of the current literature.

Synthesis, In Vitro Biological Evaluation of Antiproliferative and Neuroprotective Effects and In Silico Studies of Novel 16E-Arylidene-5α,6α-epoxyepiandrosterone Derivatives.

Steroids constitute an important class of pharmacologically active molecules, playing key roles in human physiology. Within this group, 16E-arylideneandrostane derivatives have been reported as potent anti-cancer agents for the treatment of leukemia, breast and prostate cancers, and brain tumors. Additionally, 5α,6α-epoxycholesterol is an oxysterol with several biological activities, including regulation of cell proliferation and cholesterol homeostasis. Interestingly, pregnenolone derivatives combining these two modifications were described as potential neuroprotective agents. In this research, novel 16E-arylidene-5α,6α-epoxyepiandrosterone derivatives were synthesized from dehydroepiandrosterone by aldol condensation with different aldehydes followed by a diastereoselective 5α,6α-epoxidation. Their cytotoxicity was evaluated on tumoral and non-tumoral cell lines by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Furthermore, the assessment of the neuroprotective activity of these derivatives was performed in a dopaminergic neuronal cell line (N27), at basal conditions, and in the presence of the neurotoxin 6-hydroxydopamine (6-OHDA). Interestingly, some of these steroids had selective cytotoxic effects in tumoral cell lines, with an IC50 of 3.47 µM for the 2,3-dichlorophenyl derivative in the breast cancer cell line (MCF-7). The effects of this functionalized epoxide on cell proliferation (Ki67 staining), cell necrosis (propidium iodide staining), as well as the analysis of the nuclear area and near neighbor distance in MCF-7 cells, were analyzed. From this set of biological studies, strong evidence of the activation of apoptosis was found. In contrast, no significant neuroprotection against 6-OHDA-induced neurotoxicity was observed for the less cytotoxic steroids in N27 cells. Lastly, molecular docking simulations were achieved to verify the potential affinity of these compounds against important targets of steroidal drugs (androgen receptor, estrogen receptor α, and 5α-reductase type 2, 17α-hydroxylase-17,20-lyase and aromatase enzymes). This in silico study predicted a strong affinity between most novel steroidal derivatives and 5α-reductase and 17α-hydroxylase-17,20-lyase enzymes.

Understanding neuron-glia crosstalk and biological clocks in insomnia.

According to the World Health Organization, about one-third of the population experiences insomnia symptoms, and about 10-15% suffer from chronic insomnia, the most common sleep disorder. Sleeping difficulties associated with insomnia are often linked to chronic sleep deprivation, which has a negative health impact partly due to disruption in the internal synchronisation of biological clocks. These are regulated by clock genes and modulate most biological processes. Most studies addressing circadian rhythm regulation have focused on the role of neurons, yet glial cells also impact circadian rhythms and sleep regulation. Chronic insomnia and sleep loss have been associated with glial cell activation, exacerbated neuroinflammation, oxidative stress, altered neuronal metabolism and synaptic plasticity, accelerated age-related processes and decreased lifespan. It is, therefore, essential to highlight the importance of glia-neuron interplay on sleep/circadian regulation and overall healthy brain function. Hence, in this review, we aim to address the main neurobiological mechanisms involved in neuron-glia crosstalk, with an emphasis on microglia and astrocytes, in both healthy sleep, chronic sleep deprivation and chronic insomnia.

Glycemic Control in Type 1 Diabetes Mellitus and COVID-19: What We Learned From the Lockdown Experience.

INTRODUCTION: Confinement measures that were imposed during the COVID-19 pandemic drastically changed the routines of the population. Some studies on the impact of confinement on glycemic control suggest a reduction of 0.1 to 0.5% in glycated hemoglobin. The objective of this study was to evaluate the impact of the COVID-19 pandemic lockdown on glycemic control in adult patients with type 1 diabetes mellitus. METHODS: An observational retrospective cohort study of patients with type 1 diabetes mellitus followed in a Diabetes Unit was performed. The study compared the metabolic control of these patients before (between January 1st and March 18th, 2020) and after (between May 3rd and July 31st, 2020) the lockdown. RESULTS: The study included 102 patients with type 1 diabetes mellitus (51% females), with a median age of 36 years (interquartile range 18.75, (24.25-43)) and a median duration of diabetes of 15 years (interquartile range 13, (8-21)). After lockdown, a significant decrease of 0.28±0.71% in glycated hemoglobin was observed (7.88±1.33% vs 7.59±1.23%, p=<0.001). In patients using continuous glucose monitoring a significant improvement in time in range was also noted (47.25±17.33% vs 49.97±18.61%, p=0.008). CONCLUSIONS: This study demonstrated an improvement in glycemic control after the lockdown. This might be explained by the positive impact of stable schedules, healthy meals and greater availability to make therapeutic adjustments to glycemic control. The fact that diabetes was considered a risk factor for the development of severe COVID-19 disease might also influence patients to increase their efforts to optimize their glycemic control.

Metabolic Reprogramming of Breast Tumor-Educated Macrophages Revealed by NMR Metabolomics.

The metabolic crosstalk between tumor cells and tumor-associated macrophages (TAMs) has emerged as a critical contributor to tumor development and progression. In breast cancer (BC), the abundance of immune-suppressive TAMs positively correlates with poor prognosis. However, little is known about how TAMs reprogram their metabolism in the BC microenvironment. In this work, we have assessed the metabolic and phenotypic impact of incubating THP-1-derived macrophages in conditioned media (CM) from two BC cell lines cultured in normoxia/hypoxia: MDA-MB-231 cells (highly metastatic, triple-negative BC), and MCF-7 cells (less aggressive, luminal BC). The resulting tumor-educated macrophages (TEM) displayed prominent differences in their metabolic activity and composition, compared to control cells (M0), as assessed by exo- and endometabolomics. In particular, TEM turned to the utilization of extracellular pyruvate, alanine, and branched chain keto acids (BCKA), while exhibiting alterations in metabolites associated with several intracellular pathways, including polyamines catabolism (MDA-TEM), collagen degradation (mainly MCF-TEM), adenosine accumulation (mainly MDA-TEM) and lipid metabolism. Interestingly, following a second-stage incubation in fresh RPMI medium, TEM still displayed several metabolic differences compared to M0, indicating persistent reprogramming. Overall, this work provided new insights into the metabolic plasticity of TEM, revealing potentially important nutritional exchanges and immunoregulatory metabolites in the BC TME.

Ionic-liquid-based approaches to improve biopharmaceuticals downstream processing and formulation.

The emergence of biopharmaceuticals, including proteins, nucleic acids, peptides, and vaccines, revolutionized the medical field, contributing to significant advances in the prophylaxis and treatment of chronic and life-threatening diseases. However, biopharmaceuticals manufacturing involves a set of complex upstream and downstream processes, which considerably impact their cost. In particular, despite the efforts made in the last decades to improve the existing technologies, downstream processing still accounts for more than 80% of the total biopharmaceutical production cost. On the other hand, the formulation of biological products must ensure they maintain their therapeutic performance and long-term stability, while preserving their physical and chemical structure. Ionic-liquid (IL)-based approaches arose as a promise alternative, showing the potential to be used in downstream processing to provide increased purity and recovery yield, as well as excipients for the development of stable biopharmaceutical formulations. This manuscript reviews the most important progress achieved in both fields. The work developed is critically discussed and complemented with a SWOT analysis.

IgA Vasculitis with scrotal involvement - a rare presentation in adults.

IgA vasculitis (IgAV) is a small-vessel vasculitis common in children but rare in adults. It is usually an auto-limited disease in children but has a more severe course and worse prognosis in adults. The classical manifestations are non-thrombocytopenic purpura, arthralgias, gastrointestinal involvement and renal involvement. Herein we report a case of a 39-year-old man with a rash of the lower limbs associated with testicular and lower abdominal pain. The initial study revealed increased inflammatory biomarkers and enlarged left testis with bilateral ischemic areas on doppler ultrasound. A cutaneous biopsy later revealed leukocytoclastic vasculitis, confirming the diagnosis of IgAV with scrotal involvement. The patient started prednisolone, with improvement in the first week and sustained remission after two years of follow-up. This case report describes an adult with IgAV and scrotal involvement, which is rarely reported in adults and appears to be different from the one in children. The prevalence of scrotal involvement is presumably underestimated. In all men with IgAV, a scrotal examination should be performed and ultrasonography accordingly since it affects the treatment and follow-up. Recommendations for IgAV diagnosis and treatment in adults are still lacking and more research is needed.

Molecular Mechanisms Behind the Role of Plasmacytoid Dendritic Cells in Systemic Sclerosis.

Systemic sclerosis (SSc) is a debilitating autoimmune disease that affects multiple systems. It is characterized by immunological deregulation, functional and structural abnormalities of small blood vessels, and fibrosis of the skin, and, in some cases, internal organs. Fibrosis has a devastating impact on a patient's life and lung fibrosis is associated with high morbimortality. Several immune populations contribute to the progression of SSc, and plasmacytoid dendritic cells (pDCs) have been identified as crucial mediators of fibrosis. Research on murine models of lung and skin fibrosis has shown that pDCs are essential in the development of fibrosis, and that removing pDCs improves fibrosis. pDCs are a subset of dendritic cells (DCs) that are specialized in anti-viral responses and are also involved in autoimmune diseases, such as SSc, systemic lupus erythematosus (SLE) and psoriasis, mostly due to their capacity to produce type I interferon (IFN). A type I IFN signature and high levels of CXCL4, both derived from pDCs, have been associated with poor prognosis in patients with SSc and are correlated with fibrosis. This review will examine the recent research on the molecular mechanisms through which pDCs impact SSc.

Responsiveness to pulmonary rehabilitation in COPD is associated with changes in microbiota.

BACKGROUND: Pulmonary Rehabilitation (PR) is one of the most cost-effective therapies for chronic obstructive pulmonary disease (COPD) management. There are, however, people who do not respond to PR and reasons for non-response are mostly unknown. PR is likely to change the airway microbiota and this could play a role in its responsiveness. In this study we have explored the association between PR effectiveness and specific alterations in oral microbiota and inflammation. METHODS: A prospective longitudinal study was conducted. Data on exercise capacity, dyspnoea, impact of disease and 418 saliva samples were collected from 76 patients, half of whom participated in a 12-weeks PR programme. Responders and non-responders to PR (dyspnoea, exercise-capacity and impact of disease) were defined based on minimal clinically important differences. RESULTS: Changes in microbiota, including Prevotella melaninogenica and Streptococcus were observed upon PR. Prevotella, previously found to be depleted in severe COPD, increased during the first month of PR in responders. This increase was negatively correlated with Streptococcus and Lautropia, known to be enriched in severe cases of COPD. Simultaneously, an anti-inflammatory commensal of the respiratory tract, Rothia, correlated strongly and negatively with several pro-inflammatory markers, whose levels were generally boosted by PR. Conversely, in non-responders, the observed decline in Prevotella correlated negatively with Streptococcus and Lautropia whose fluctuations co-occurred with several pro-inflammatory markers. CONCLUSIONS: PR is associated with changes in oral microbiota. Specifically, PR increases salivary Prevotella melaninogenica and avoids the decline in Rothia and the increase in Streptococcus and Lautropia in responders, which may contribute to the benefits of PR.

The impact of insomnia on frailty and the hallmarks of aging.

Throughout the course of life, there are age-related changes in sleep. Despite these normal changes, there is a high percentage of older adults that report sleep dissatisfaction with a high pervasiveness of chronic insomnia, the most common sleep disorder worldwide, with its prevalence being expected to continuously increase due to the growing rates of aging and obesity. This can have different adverse health outcomes, especially by promoting both physical and cognitive decline, which ultimately may aggravate frailty in older adults. Moreover, age-related frailty and sleep dysfunction may have a common mechanism related to the hallmarks of cellular aging. Cellular aging was categorized into nine hallmarks, such as DNA damage, telomere attrition and epigenetic changes. In the context of geriatric and chronic insomnia research, this review aims at discussing the current evidence from both animal models and human cohorts addressing the link between chronic insomnia, the hallmarks of aging and their impact on frailty. Moreover, the most recent research about the putative effect of insomnia therapeutic approaches on hallmarks of aging will be also highlighted.