Unraveling the molecular architecture of autoimmune thyroid diseases at spatial resolution.

Autoimmune thyroid diseases (AITD) such as Graves' disease (GD) or Hashimoto's thyroiditis (HT) are organ-specific diseases that involve complex interactions between distinct components of thyroid tissue. Here, we use spatial transcriptomics to explore the molecular architecture, heterogeneity and location of different cells present in the thyroid tissue, including thyroid follicular cells (TFCs), stromal cells such as fibroblasts, endothelial cells, and thyroid infiltrating lymphocytes. We identify damaged antigen-presenting TFCs with upregulated CD74 and MIF expression in thyroid samples from AITD patients. Furthermore, we discern two main fibroblast subpopulations in the connective tissue including ADIRF+ myofibroblasts, mainly enriched in GD, and inflammatory fibroblasts, enriched in HT patients. We also demonstrate an increase of fenestrated PLVAP+ vessels in AITD, especially in GD. Our data unveil stromal and thyroid epithelial cell subpopulations that could play a role in the pathogenesis of AITD.

Enzymatic Degradation Behavior of Self-Degradable Lipase-Embedded Aliphatic and Aromatic Polyesters and Their Blends.

Over the past decade, the preparation of novel materials by enzyme-embedding into biopolyesters has been proposed as a straightforward method to produce self-degrading polymers. This paper reports the preparation and enzymatic degradation of extruded self-degradable films of three different biopolyesters: poly(lactic acid) (PLA), poly(butylene adipate-co-terephthalate) (PBAT), and poly(butylene succinate) (PBS), as well as three binary/ternary blends. Candida antarctica lipase B (CalB) has been employed for the enzyme-embedding procedure, and to the best of our knowledge, the use of this approach in biopolyester blends has not been reported before. The three homopolymers exhibited differentiated degradation and suggested a preferential attack of CalB on PBS films over PBAT and PLA. Moreover, the self-degradable films obtained from the blends showed slow degradation, probably due to the higher content in PLA and PBAT. These observations pave the way for exploring enzymes capable of degrading all blend components or an enzymatic mixture for blend degradation.

Analysis of T follicular and T peripheral helper lymphocytes in autoimmune thyroid disease.

PURPOSE: Peripheral helper T (Tph) cells have an important role in the induction of humoral immune responses and autoantibody production. Accordingly, it is feasible that this lymphocyte subset has a relevant role in the pathogenesis of autoimmune thyroid diseases (AITD). In this study we aim to analyze the levels and function of Tph cells in blood samples from patients with AITD. METHODS: We performed an observational study with cases and controls. Blood samples were obtained from nineteen patients with Hashimoto's thyroiditis (HT), twenty-four with Graves' disease (GD), and fifteen healthy controls. In addition, the levels of follicular T helper (Tfh) cells and Tph cells, the release of interleukin-21 (IL-21) by these lymphocytes and the number of plasmablasts were analyzed by multi-parametric flow cytometry analyses. RESULTS: Increased percentages of Tfh and Tph lymphocytes were detected in patients with HT and GD. Furthermore, an enhanced synthesis of the cytokine IL-21 by these cells was observed. Accordingly, we detected significant higher percentages of plasmablasts in patients with GD, and these values tended to be also higher in HT patients. Moreover, significant positive associations were observed between the levels of Tfh or Tph and the number of plasmablast or anti-TSHR Ab titers in patients with AITD. CONCLUSION: Our data suggest that Tph lymphocytes may have a relevant role in the pathogenesis of AITD.

Integrative clinical, hormonal, and molecular data associate with invasiveness in acromegaly: REMAH study.

INTRODUCTION: Growth hormone (GH)-secreting pituitary tumors (GHomas) are the most common acromegaly cause. At diagnosis, most of them are macroadenomas, and up to 56% display cavernous sinus invasion. Biomarker assessment associated with tumor growth and invasion is important to optimize their management. OBJECTIVES: The study aims to identify clinical/hormonal/molecular biomarkers associated with tumor size and invasiveness in GHomas and to analyze the influence of pre-treatment with somatostatin analogs (SSAs) or dopamine agonists (DAs) in key molecular biomarker expression. METHODS: Clinical/analytical/radiological variables were evaluated in 192 patients from the REMAH study (ambispective multicenter post-surgery study of the Spanish Society of Endocrinology and Nutrition). The expression of somatostatin/ghrelin/dopamine system components and key pituitary/proliferation markers was evaluated in GHomas after the first surgery. Univariate/multivariate regression studies were performed to identify association between variables. RESULTS: Eighty percent of patients harbor macroadenomas (63.8% with extrasellar growth). Associations between larger and more invasive GHomas with younger age, visual abnormalities, higher IGF1 levels, extrasellar/suprasellar growth, and/or cavernous sinus invasion were found. Higher GH1 and lower PRL/POMC/CGA/AVPR1B/DRD2T/DRD2L expression levels (P < .05) were associated with tumor invasiveness. Least Absolute Shrinkage and Selection Operator's penalized regression identified combinations of clinical and molecular features with areas under the curve between 0.67 and 0.82. Pre-operative therapy with DA or SSAs did not alter the expression of any of the markers analyzed except for DRD1/AVPR1B (up-regulated with DA) and FSHB/CRHR1 (down-regulated with SSAs). CONCLUSIONS: A specific combination of clinical/analytical/molecular variables was found to be associated with tumor invasiveness and growth capacity in GHomas. Pre-treatment with first-line drugs for acromegaly did not significantly modify the expression of the most relevant biomarkers in our association model. These findings provide valuable insights for risk stratification and personalized management of GHomas.

Predictors of biochemical response to somatostatin receptor ligands in acromegaly.

Although predictors of response to first-generation somatostatin receptor ligands (fg-SRLs), and to a lesser extent to pasireotide, have been studied in acromegaly for many years, their use is still not recommended in clinical guidelines. Is there insufficient evidence to use them? Numerous biomarkers including various clinical, functional, radiological and molecular markers have been identified. The first ones are applicable pre-surgery, while the molecular predictors are utilized for patients not cured after surgery. In this regard, factors predicting a good response to fg-SRLs are specifically: low basal GH, a low GH nadir in the acute octreotide test, T2 MRI hypointensity, a densely granulated pattern, high immunohistochemistry staining for somatostatin receptor 2 (SSTR2), and E-cadherin. However, there is still a lack of consensus regarding which of these biomarkers is more useful or how to integrate them into clinical practice. With classical statistical methods, it is complex to define reliable and generalizable cut-off values for a single biomarker. The potential solution to the limitations of traditional methods involves combining systems biology with artificial intelligence, which is currently providing answers to such long-standing questions that may eventually be finally included into the clinical guidelines and make personalized medicine a reality. The aim of this review is to describe the current knowledge of the main fg-SRLs and pasireotide response predictors, discuss their current usefulness, and point to future directions in the research of this field.

Salt-induced Fmoc-tripeptide supramolecular hydrogels: a combined experimental and computational study of the self-assembly.

Delving into the mechanism behind the molecular interactions at the atomic level of short-sequence peptides plays a key role in the development of nanomaterials with specific structure-property-function relationships from a bottom-up perspective. Due to their poor water solubility, the self-assembly of Fmoc-bearing peptides is usually induced by dissolution in an organic solvent, followed by a dilution step in water, pH changes, and/or a heating-cooling process. Herein, we report a straightforward methodology for the gelation of Fmoc-FFpY (F: phenylalanine; Y: tyrosine; and p: PO42-), a negatively charged tripeptide, in NaCl solution. The electrostatic interactions between Fmoc-FFpY and Na+ ions give rise to different nanofibrillar hydrogels with rheological properties and nanofiber sizes modulated by the NaCl concentration in pure aqueous media. Initiated by the electrostatic interactions between the peptide phosphate groups and the Na+ ions, the peptide self-assembly is stabilized thanks to hydrogen bonds between the peptide backbones and the π-π stacking of aromatic Fmoc and phenyl units. The hydrogels showed self-healing and thermo-responsive properties for potential biomedical applications. Molecular dynamics simulations from systems devoid of prior training not only confirm the aggregation of peptides at a critical salt concentration and the different interactions involved, but also corroborate the secondary structure of the hydrogels at the microsecond timescale. It is worth highlighting the remarkable achievement of reproducing the morphological behavior of the hydrogels using atomistic simulations. To our knowledge, this study is the first to report such a correspondence.

Primary Cilia as a Tumor Marker in Pituitary Neuroendocrine Tumors.

Pituitary neuroendocrine tumors (PitNETs) account for approximately 15% of all intracranial neoplasms. Although they usually appear to be benign, some tumors display worse behavior, displaying rapid growth, invasion, refractoriness to treatment, and recurrence. Increasing evidence supports the role of primary cilia (PC) in regulating cancer development. Here, we showed that PC are significantly increased in PitNETs and are associated with increased tumor invasion and recurrence. Serial electron micrographs of PITNETs demonstrated different ciliation phenotypes (dot-like versus normal-like cilia) that represented PC at different stages of ciliogenesis. Molecular findings demonstrated that 123 ciliary-associated genes (eg, doublecortin domain containing protein 2, Sintaxin-3, and centriolar coiled-coil protein 110) were dysregulated in PitNETs, representing the upregulation of markers at different stages of intracellular ciliogenesis. Our results demonstrate, for the first time, that ciliogenesis is increased in PitNETs, suggesting that this process might be used as a potential target for therapy in the future.

Neonatal point-of-care lung ultrasound: what should be known and done out of the NICU?

Lung ultrasound is rapidly becoming a useful tool in the care of neonates: its ease of use, reproducibility, low cost, and negligible side effects make it a very suitable tool for the respiratory care of all neonates. This technique has been extensively studied by different approaches in neonatal intensive care unit (NICU), both for diagnostic and prognostic aims and to guide respiratory treatments. However, many neonates are being born in level I/II hospitals without NICU facilities so all pediatricians, not just neonatal intensivists, should be aware of its potential. This is made possible by the increasing access to ultrasound machines in a modern hospital setting. In this review, we describe the ultrasonographic characteristics of the normal neonatal lung. We also discuss the ultrasound features of main neonatal respiratory diseases: transient tachypnea of the neonate (TTN), respiratory distress syndrome (RDS), meconium aspiration syndrome (MAS), pneumothorax (PNX), pleural effusion (PE), or pneumonia. Finally, we mention two functional approaches to lung ultrasound: 1. The use of lung ultrasound in level I delivery centers as a mean to assess the severity of neonatal respiratory distress and request a transport to a higher degree structure in a timely fashion. 2. The prognostic accuracy of lung ultrasound for early and targeted surfactant replacement. CONCLUSION:  LU is still a useful tool in level I/II neonatal units, both for diagnostic and functional issues. WHAT IS KNOWN: • Neonatal lung ultrasound has been recently introduced in the usual care in many Neonatal Intensive Care Units. WHAT IS NEW: • It also has many advantages in level I/II neonatal units, both for neonatologist or even pediatricians that treat neonates in those sites.

Separate gut-brain circuits for fat and sugar reinforcement combine to promote overeating.

Food is a powerful natural reinforcer that guides feeding decisions. The vagus nerve conveys internal sensory information from the gut to the brain about nutritional value; however, the cellular and molecular basis of macronutrient-specific reward circuits is poorly understood. Here, we monitor in vivo calcium dynamics to provide direct evidence of independent vagal sensing pathways for the detection of dietary fats and sugars. Using activity-dependent genetic capture of vagal neurons activated in response to gut infusions of nutrients, we demonstrate the existence of separate gut-brain circuits for fat and sugar sensing that are necessary and sufficient for nutrient-specific reinforcement. Even when controlling for calories, combined activation of fat and sugar circuits increases nigrostriatal dopamine release and overeating compared with fat or sugar alone. This work provides new insights into the complex sensory circuitry that mediates motivated behavior and suggests that a subconscious internal drive to consume obesogenic diets (e.g., those high in both fat and sugar) may impede conscious dieting efforts.

External Validation of a Multivariate Model for Targeted Surfactant Replacement.

INTRODUCTION: Early targeted surfactant therapy for preterm infants is recommended but the best criteria to personalize treatment are unclear. We validate a previously published multivariate prognostic model based on gestational age (GA), lung ultrasound score (LUS), and oxygen saturation to inspire oxygen fraction ratio (SatO2/FiO2) using an independent data set. METHODS: Pragmatic, observational study in 10 Italian and Spanish NICUs, including preterm babies (250 and 336 weeks divided into 3 GA intervals) with clinical signs of respiratory distress syndrome and stabilized on CPAP. LUS and SatO2/FiO2 were collected soon after stabilization. Their prognostic accuracy was evaluated on the subsequent surfactant administration by a rigorously masked physician. RESULTS: One hundred seventy-five infants were included in the study. Surfactant was given to 74% infants born at 25-27 weeks, 38.5% at 28-30 weeks, and 26.5% at 31-33 weeks. The calibration curve comparing the validation and the development populations showed significant overlap with an intercept = 0.08, 95% CI (-0.34; 0.5) and a slope = 1.53, 95% CI (1.07-1.98). The validation cohort had a high predictive accuracy. Its ROC curve showed an AUC = 0.95, 95% CI (0.91-0.99) with sensitivity = 0.93, 95% CI (0.83-0.98), specificity = 0.81, 95% CI (0.73-0.88), PPV = 0.76, 95% CI (0.65-0.84), NPV = 0.95, 95% CI (0.88-0.98). LUS ≥9 demonstrated the highest sensitivity (0.91, 95% CI [0.82-0.97]) and specificity = 0.81, 95% CI (0.72-0.88) as individual predictor. LUS and SatO2/FiO2 prognostic performances varied with GA. CONCLUSION: We validated a prognostic model based on LUS and Sat/FiO2 to facilitate early, customized surfactant administration that may improve respiratory management of preterm neonates.

Lung UltrasouNd Guided surfactant therapy in preterm infants: an international multicenter randomized control trial (LUNG study).

BACKGROUND: The management of respiratory distress syndrome (RDS) in premature newborns is based on different types of non-invasive respiratory support and on surfactant replacement therapy (SRT) to avoid mechanical ventilation as it may eventually result in lung damage. European guidelines currently recommend SRT only when the fraction of inspired oxygen (FiO2) exceeds 0.30. The literature describes that early SRT decreases the risk of bronchopulmonary dysplasia (BPD) and mortality. Lung ultrasound score (LUS) in preterm infants affected by RDS has proven to be able to predict the need for SRT and different single-center studies have shown that LUS may increase the proportion of infants that received early SRT. Therefore, the aim of this study is to determine if the use of LUS as a decision tool for SRT in preterm infants affected by RDS allows for the reduction of the incidence of BPD or death in the study group. METHODS/DESIGN: In this study, 668 spontaneously-breathing preterm infants, born at 25+0 to 29+6 weeks' gestation, in nasal continuous positive airway pressure (nCPAP) will be randomized to receive SRT only when the FiO2 cut-off exceeds 0.3 (control group) or if the LUS score is higher than 8 or the FiO2 requirements exceed 0.3 (study group) (334 infants per arm). The primary outcome will be the difference in proportion of infants with BPD or death in the study group managed compared to the control group. DISCUSSION: Based on previous published studies, it seems that LUS may decrease the time to administer surfactant therapy. It is known that early surfactant administration decreases BPD and mortality. Therefore, there is rationale for hypothesizing a reduction in BPD or death in the group of patients in which the decision to administer exogenous surfactant is based on lung ultrasound scores. TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT05198375 . Registered on 20 January 2022.

Granular Disulfide-Crosslinked Hyaluronic Hydrogels: A Systematic Study of Reaction Conditions on Thiol Substitution and Injectability Parameters.

Granular polymer hydrogels based on dynamic covalent bonds are attracting a great deal of interest for the design of injectable biomaterials. Such materials generally exhibit shear-thinning behavior and properties of self-healing/recovery after the extrusion that can be modulated through the interactions between gel microparticles. Herein, bulk macro-hydrogels based on thiolated-hyaluronic acid were produced by disulphide bond formation using oxygen as oxidant at physiological conditions and gelation kinetics were monitored. Three different thiol substitution degrees (SD%: 65%, 30% and 10%) were selected for hydrogel formation and fully characterized as to their stability in physiological medium and morphology. Then, extrusion fragmentation technique was applied to obtain hyaluronic acid microgels with dynamic disulphide bonds that were subsequently sterilized by autoclaving. The resulting granular hyaluronic hydrogels were able to form stable filaments when extruded through a syringe. Rheological characterization and cytotoxicity tests allowed to assess the potential of these materials as injectable biomaterials. The application of extrusion fragmentation for the formation of granular hyaluronic hydrogels and the understanding of the relation between the autoclaving processes and the resulting particle size and rheological properties should expand the development of injectable materials for biomedical applications.

Evaluation of Epithelial-Mesenchymal Transition Markers in Autoimmune Thyroid Diseases.

A state of chronic inflammation is common in organs affected by autoimmune disorders, such as autoimmune thyroid diseases (AITD). Epithelial cells, such as thyroid follicular cells (TFCs), can experience a total or partial transition to a mesenchymal phenotype under these conditions. One of the major cytokines involved in this phenomenon is transforming growth factor beta (TGF-ß), which, at the initial stages of autoimmune disorders, plays an immunosuppressive role. However, at chronic stages, TGF- ß contributes to fibrosis and/or transition to mesenchymal phenotypes. The importance of primary cilia (PC) has grown in recent decades as they have been shown to play a key role in cell signaling and maintaining cell structure and function as mechanoreceptors. Deficiencies of PC can trigger epithelial-mesenchymal transition (EMT) and exacerbate autoimmune diseases. A set of EMT markers (E-cadherin, vimentin, α-SMA, and fibronectin) were evaluated in thyroid tissues from AITD patients and controls through RT-qPCR, immunohistochemistry (IHC), and western blot (WB). We established an in vitro TGF-ß-stimulation assay in a human thyroid cell line to assess EMT and PC disruption. EMT markers were evaluated in this model using RT-qPCR and WB, and PC was evaluated with a time-course immunofluorescence assay. We found an increased expression of the mesenchymal markers α-SMA and fibronectin in TFCs in the thyroid glands of AITD patients. Furthermore, E-cadherin expression was maintained in these patients compared to the controls. The TGF-ß-stimulation assay showed an increase in EMT markers, including vimentin, α-SMA, and fibronectin in thyroid cells, as well as a disruption of PC. The TFCs from the AITD patients experienced a partial transition to a mesenchymal phenotype, preserving epithelial characteristics associated with a disruption in PC, which might contribute to AITD pathogenesis.

MicroRNAs in autoimmune thyroid diseases and their role as biomarkers.

MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression at the posttranscriptional level. They are emerging as potential biomarkers and as therapeutic targets for several diseases including autoimmune thyroid diseases (AITD). They control a wide range of biological phenomena, including immune activation, apoptosis, differentiation and development, proliferation and metabolism. This function makes miRNAs attractive as disease biomarker candidates or even as therapeutic agents. Because of their stability and reproducibility circulating miRNAs have been an interesting area of research in many diseases, and studies describing their role in the immune response and in autoimmune diseases have progressively developed. The mechanisms underlying AITD remain elusive. AITD pathogenesis is characterized by a multifactorial interplay based on the synergy between susceptibility genes and environmental stimulation, together with epigenetic modulation. Understanding the regulatory role of miRNAs could lead to identify potential susceptibility pathways, diagnostic biomarkers and therapeutic targets for this disease. Herein we update our present knowledge on the role of microRNAs in AITD and discuss on their importance as possible diagnostic and prognostic biomarkers in the most prevalent AITDs: Hashimoto's thyroiditis (HT), Graves' disease (GD) and Graves' Ophthalmopathy (GO). This review provides an overview of the state of the art in the pathological roles of microRNAs as well as in possible novel miRNA-based therapeutic approaches in AITD.

Minor Changes in Daily Rhythms Induced by a Skeleton Photoperiod Are Associated with Increased Adiposity and Glucose Intolerance.

Eating during the rest phase is associated with metabolic syndrome, proposed to result from a conflict between food consumption and the energy-saving state imposed by the circadian system. However, in nocturnal rodents, eating during the rest phase (day-feeding, DF) also implies food intake during light exposure. To investigate whether light exposure contributes to DF-induced metabolic impairments, animals receive food during the subjective day without light. A skeleton photoperiod (SP) is used to entrain rats to a 12:12 cycle with two short light pulses framing the subjective day. DF-induced adiposity is prevented by SP, suggesting that the conflict between light and feeding stimulates fat accumulation. However, all animals under SP conditions develop glucose intolerance regardless of their feeding schedule. Moreover, animals under SP with ad libitum or night-feeding have increased adiposity. SP animals show a delayed onset of the daily rise in body temperature and energy expenditure and shorter duration of nighttime activity, which may contribute to the metabolic disturbances. These data emphasize that metabolic homeostasis can only be achieved when all daily cycling variables are synchronized. Even small shifts in the alignment of different metabolic rhythms, such as those induced by SP, may predispose individuals to metabolic disease.

Circadian modulation of microglial physiological processes and immune responses.

Microglia is considered the central nervous system (CNS) resident macrophages that establish an innate immune response against pathogens and toxins. However, the recent studies have shown that microglial gene and protein expression follows a circadian pattern; several immune activation markers and clock genes are expressed rhythmically without the need for an immune stimulus. Furthermore, microglia responds to an immune challenge with different magnitudes depending on the time of the day. This review examines the circadian control of microglia function and the possible physiological implications. For example, we discuss that synaptic prune is performed in the cortex at a certain moment of the day. We also consider the implications of daily microglial function for maintaining biological rhythms like general activity, body temperature, and food intake. We conclude that the developmental stage, brain region, and pathological state are not the only factors to consider for the evaluation of microglial functions; instead, emerging evidence indicates that circadian time as an essential aspect for a better understanding of the role of microglia in CNS physiology.

pH modulates antibacterial activity of hydroxybenzaldehyde derivatives immobilized in chitosan films via reversible Schiff bases and its application to preserve freshly-squeezed juice.

Antimicrobial food grade hydroxybenzaldehyde derivatives were immobilized on the surface of chitosan films by means of reversible Schiff bases. Spectroscopy and elemental analysis evidenced the different ability of the aldehydes to form Schiff bases with chitosan. Chitosan films modified with Schiff bases of aldehydes exerted antimicrobial properties against E. coli under mild acidic environments. The efficacy of the films lied on the reversibility of synthetized imine bonds and release of the aldehydes which was promoted in mildly acid aqueous solutions. Besides acidity, imine bond reversibility depended on the chemical structure of the aldehyde covalently bonded. Films carrying salicylaldehyde presented the highest in vitro antimicrobial performance and thus, they were chosen to evaluate their effectivity in inhibiting E. coli proliferation in freshly-squeezed carrot-orange juice. Films were successfully activated by the acid environment of the juice and reduced the population of the inoculated pathogen. Salicylaldehyde migrated to the juice did not exert toxic effects on Caco-2 cells.