Development Of Pathology Research Articles

Effects of serotonergic manipulation in the brainstem and hypothalamus of overnourished rats during lactation

Previous studies suggest that being overweight and obese is capable of altering brain function during critical periods due to the higher plasticity of the brain during development. However, more literature still needs to be on the immediate effect of overnutrition and serotonin modulation in brain areas. In this study, we aimed to evaluate the effects of serotoninergic manipulation on oxidative stress and pro-inflammatory markers in the brainstem and hypothalamus of overnourished rats. The fluoxetine treatment was performed from postnatal day 3 (PND3) to postnatal day 21 (PND21) to evaluate mitochondrial function, oxidative balance, and the mRNA levels of monoaminergic molecules such as serotonin and dopamine, pro-inflammatory cytokines, and BDNF. Neonatal overnutrition induces molecular and biochemical changes in the brainstem and hypothalamus. These nutritional disturbances during lactation dysregulate energy balance and cellular redox, inducing oxidative stress. Conversely, modulation of the serotoninergic system through pharmacological use of fluoxetine was able to reverse the deleterious effects of overnutrition during lactation, enabling better brain development and delaying the development of pathologies and oxidative stress.

Forgotten Children

Fetal and infant skeletal remains in anatomical collections remain an underutilized yet important source of information on the interactions of gender, class, and religion during pregnancy and infant loss. The W. D. Trotter Anatomy Museum at the University of Otago in New Zealand, founded in the nineteenth century, houses more than 2,000 models and anatomical “specimens,” with many of the skeletal remains lacking provenience information. This research aimed to both provide an inventory of the fetal and infant skeletal remains and identify through archival analysis the social context of those whose remains were obtained by the museum. An osteological analysis was carried out to assess the minimum number of individuals (MNI), estimate age-at-death, and provide evidencefor pathology and dissection. The results are interpreted in the context of the archival accession data and historical information at the time to provide the social context in which these people lived and therefore information that may be related to their deaths and acquisition of their bodies by the university. Most of the infants within the collection died around the time of full-termbirth, and some individuals have evidence for developmental pathology, birth trauma, and/or postmortem dissection. The historical context and legislation around body donation suggests structural inequality played a role in the acquisition of these infants’ remains from mothers (primarily poor and/or unmarried women) and that these remains went through a process of commodification and objectification and were retained as developmental teaching “specimens” and examples of pathology.

Spatial differences in gene expression across the dorsal raphe nucleus in a model of early Alzheimer's disease.

Persons with Alzheimer's disease (AD) present with changes in mood, sleep, and arousal that may precede the clinical manifestation of cognitive decline. These early symptoms can be driven by changes in the serotonergic (5-HT) nuclei of the brainstem, particularly the dorsal raphe nucleus (DRN). It is unclear why all 5-HT neurons do not simultaneously develop AD pathology that progresses at the same rate. We sought to identify any underlying genetic components associated with susceptibility or resistance of 5-HT neurons to AD pathology. The Visium Spatial Gene Expression platform was used to identify transcriptomic changes across the DRN in a preclinical model of early AD, human tau-overexpressing mice (htau mice). We further used RNAscope and immunohistochemical assessment to validate findings of primary interest. We find that the DRN of htau mice differentially expresses AD-related genes, including those related to kinase binding, ion channel activity, ligand-receptor interactions, and regulation of serine/threonine kinases. We further find that computational sub-clustering of the DRN is consistent with previous circuitry-driven characterizations, allowing for spatial bounding of distinct subregions within the DRN. Of these, we find the dorsolateral DRN is preferentially impacted by 5-HT neuron loss and development of tau pathology, which coincides with increased expression of the long noncoding RNA Map2k3os. Map2k3os may serve regulatory roles relevant for tau phosphorylation and warrants further investigation to characterize its interactions. Overall, this report demonstrates the power of large-scale spatial transcriptomics technologies, while underscoring the need for convergent-data validation to overcome their limitations.

The Annexin A1 Protein Mimetic Peptide Ac2-26 prevents cellular senescence of CHON-001 chondrocytes against tumor necrosis factor-α via the Nrf2/NF-κB pathway.

Osteoarthritis (OA) is a degenerative joint disorder characterized by progressive cartilage degradation. Excessive oxidative stress (OS), inflammatory responses, extracellular matrix breakdown, and cellular senescence of chondrocytes play crucial roles in the pathological development of OA. Currently, curing OA remains a significant challenge. In this study, we aimed to elucidate the protective effects of Annexin A1 protein Mimetic Peptide (Ac2-26) against tumor necrosis factor-α (TNF-α)-induced damage in CHON-001 chondrocytes by assessing cellular senescence, OS, and the expression levels of matrix metalloproteinase-13 (MMP-13) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4. Our results show that Ac2-26 mitigated the reduction of telomerase activity and the exacerbation of cellular senescence induced by TNF-α in CHON-001 chondrocytes. Treatment with TNF-α led to decreased expression of the human telomerase reverse transcriptase gene and increased expression of the telomeric repeat-binding factor 2 gene, which were reversed by Ac2-26 treatment. The TNF-α-induced increases in the gene and protein expressions of p53 and p16 were restored by Ac2-26 in a dose-dependent manner. Additionally, we found that TNF-α caused elevations in the mRNA and protein levels of MMP-13 and ADAMTS-4, which were reduced by Ac2-26 in a dose-dependent fashion. Furthermore, TNF-α triggered the activation of nuclear factor κ-B (NF-κB) by increasing the levels of phosphorylated NF-κB p65 and the luciferase activity of NF-κB. Notably, Ac2-26 alleviated OS by reducing mitochondrial reactive oxygen species levels and promoting the activation of NF-E2-related factor 2 (Nrf2) in TNF-α-challenged CHON-001 chondrocytes. Silencing Nrf2 abolished the Ac2-26-induced activation of NF-κB and cellular senescence in CHON-001 chondrocytes. Collectively, these findings offer new insights into the potential therapeutic use of Ac2-26 for treating OA.

MRI-based Whole-Brain Elastography and Volumetric Measurements to Predict Brain Age

Abstract Brain age, as a correlate of an individual’s chronological age obtained from structural and functional neuroimaging data, enables assessing developmental or neurodegenerative pathology relative to the overall population. Accurately inferring brain age from brain magnetic resonance imaging (MRI) data requires imaging methods sensitive to tissue health and sophisticated statistical models to identify the underlying age-related brain changes. Magnetic resonance elastography (MRE) is a specialized MRI technique which has emerged as a reliable, non-invasive method to measure the brain’s mechanical properties, such as the viscoelastic shear stiffness and damping ratio. These mechanical properties have been shown to change across the life span, reflect neurodegenerative diseases, and are associated with individual differences in cognitive function. Here, we aim to develop a machine learning framework to accurately predict a healthy individual’s chronological age from maps of brain mechanical properties. This framework can later be applied to understand neurostructural deviations from normal in individuals with neurodevelopmental or neurodegenerative conditions. Using 3D convolutional networks as deep learning models and more traditional statistical models, we relate chronological age as a function of multiple modalities of whole-brain measurements: stiffness, damping ratio, and volume. Evaluations on held-out subjects show that combining stiffness and volume in a multimodal approach achieves the most accurate predictions. Interpretation of the different models highlight important regions that are distinct between the modalities. The results demonstrate the complementary value of MRE measurements in brain age models, which, in future studies, could improve model sensitivity to brain integrity differences in individuals with neuropathology.

Whole Slide Imaging, Artificial Intelligence, and Machine Learning in Pediatric and Perinatal Pathology: Current Status and Future Directions

The integration of artificial intelligence (AI) into healthcare is becoming increasingly mainstream. Leveraging digital technologies, such as AI and deep learning, impacts researchers, clinicians, and industry due to promising performance and clinical potential. Digital pathology is now a proven technology, enabling generation of high-resolution digital images from glass slides (whole slide images; WSI). WSIs facilitates AI-based image analysis to aid pathologists in diagnostic tasks, improve workflow efficiency, and address workforce shortages. Example applications include tumor segmentation, disease classification, detection, quantitation and grading, rare object identification, and outcome prediction. While advancements have occurred, integration of WSI-AI into clinical laboratories faces challenges, including concerns regarding evidence quality, regulatory adaptations, clinical evaluation, and safety considerations. In pediatric and developmental histopathology, adoption of AI could improve diagnostic efficiency, automate routine tasks, and address specific diagnostic challenges unique to the specialty, such as standardizing placental pathology and developmental autopsy findings, as well as mitigating staffing shortages in the subspeciality. Additionally, AI-based tools have potential to mitigate medicolegal implications by enhancing reproducibility and objectivity in diagnostic evaluations. An overview of recent developments and challenges in applying AI to pediatric and developmental pathology, focusing on machine learning methods applied to WSIs of pediatric pathology specimens is presented.

Level of tryptophan signaling molecules in children with different dynamics of obesity development

The high incidence of obesity in children is an extremely pressing problem in the modern health care system of many countries. There is an increase in the incidence of obesity in children in Russia, which is dangerous not only due to the development of somatic concomitant pathologies, but also to a violation of the social adaptation of children. Without a doubt, the search for early biochemical indicators of stabilization and progress of obesity in older age is very important for the formation of a risk group and timely prevention of the development of obesity and its complications. Our study showed that when assessing the various dynamics of obesity development, it is necessary to take into account the gender of the child, which is ignored in many cases. A decrease in serum serotonin concentrations may be considered as an indicator associated with the stabilization or progression of obesity in girls. Whereas in boys prone to stabilization and/or progression of obesity, a reduced serum level of indole-3-acrylate may act as a predictor marker.

Modeling of Healing of Intra-Articular Fractures of Long Tubular Bones in Rats

Background: We present a new original method of modeling an intra-articular fracture of the long tubular bone in laboratory animals.Objective: To create a new experimental model of an intra-articular fracture of the long tubular bone and to approve it for comprehensive evaluation of morphofunctional parameters of cartilage and bone healing.Materials and methods: The experiment was performed on 30 white sexually mature Wistar rats. Modeling involved layer-by-layer dissection of tissues along the anteromedial surface and formation of an incomplete fracture of the femoral medial condyle using a sharp L-shaped working end of an 0.2 × 0.2 сm instrument perpendicular to the medial condyle with a slight single muscle force. The rats were euthanized in groups of 10 animals on days 7, 14, and 30. We performed knee radiography and determined blood osteocalcin and BMP-7 levels. The Mankin score was used in the histological evaluation of the articular cartilage changes. The groups were compared using the Kruskal-Wallis test. P < .05 was considered statistically significant.Results: We noted statistically significant differences (P < .0001) when comparing the groups using the Mankin score. The post hoc test revealed statistically significant differences in the Mankin scores between days 7 and 14 (P = .014) and between days 7 and 30 (P < .0001). Group comparisons showed no statistically significant differences in blood osteocalcin levels (P = .518). However, there were statistically significant differences (P = .0028) between the groups in blood BMP-7 levels. The post hoc test revealed statistically significant differences in blood BMP-7 levels between days 7 and 14 (P = .048) and between days 14 and 30 (P = .0195).Conclusions: The developed experimental model of intra-articular fractures of long tubular bones allows for a comprehensive assessment of reparative processes of cartilage and bone tissue. The model is close to the real mechanism of pathology development.

Drosophila glial system: an approach towards understanding molecular complexity of neurodegenerative diseases.

Glia is pivotal in regulating neuronal stem cell proliferation, functioning, and nervous system homeostasis, significantly influencing neuronal health and disorders. Dysfunction in glial activity is a key factor in the development and progression of brain pathology. However, a deeper understanding of the intricate nature of glial cells and their diverse role in neurological disorders is still required. To this end, we conducted data mining to retrieve literature from PubMed and Google Scholar using the keywords: glia, Drosophila, neurodegeneration, and mammals. The retrieved literature was manually screened and used to comprehensively understand and present the different glial types in Drosophila, i.e., perineurial, subperineurial, cortex, astrocyte-like and ensheathing glia, their relevance with mammalian counterparts, mainly microglia and astrocytes, and their potential to reveal complex neuron-glial molecular networks in managing neurodegenerative processes.

Abstract 4137500: The Histone Methyltransferase MLL1 Regulates Notch Signaling and T Cell Phenotype During Pathologic Abdominal Aortic Aneurysm Development

Objective: Abdominal aortic aneurysms (AAA) are characterized by pathological vascular remodeling and an imbalance between anti-inflammatory regulatory T H cells (Tregs) and pro-inflammatory T H 17 cells within the aortic wall. The mechanisms regulating CD4 + T H cell differentiation during AAA development remain unknown. Recently, the histone methyltransferase, mixed lineage leukemia 1 (MLL1), has been shown to influence T H cell phenotype by directly altering key transcription factors for T H 17 (RORγ) and Treg (FOXp3) differentiation in varying disease states. As such the objective of this study was to evaluate the role of MLL1 in promoting T H 17 activity within AAA development and to determine key T H 17 cell signaling pathways that are altered Methods: Single-cell sequencing was conducted on human AAAs and control tissue samples. For our murine model, C57BL/6 mice were injected with an AAV encoding a PCSK9 gain-of-function mutation and fed a saturated fat diet followed by either AngII infusion to induce AAAs (1 µg/kg/min) or saline. Mice with T-cell specific deletion of Notch1 signaling (Notch1 f/f CD4 cre+ ) or MLL1 ( Mll1 f/f CD4 cre+ ) were subjected to the AngII-induced AAA model and AAA diameters quantified. CD4+ T-cells were isolated by MACs and gene expression analyzed by qPCR as well as T-cell phenotype by flow cytometry. ChIP was used to evaluate histone 3 lysine trimethylation (H3K4me3). Results: Single-cell RNA sequencing of human aortic tissue revealed that the epigenetic enzyme MLL1 and Notch1 signaling were profoundly upregulated in human AAA CD4+ cells compared to non-aneurysmal control samples. Congruently, CD4 + T H cells isolated from the murine AngII-induced AAA model displayed a predominance of T H 17 phenotype and elevated expression of MLL1. Histone methylation was evaluated by ChIP in AngII-induced AAA CD4+ cells and demonstrated increased levels of the MLL1-mediated activation histone methylation mark, H3K4me3, on the Notch 1 promoter. Mice deficient in CD4 + T H cell Notch signaling ( Notch1 f/f CD4 cre+ ) or MLL1 ( Mll1 f/f CD4 cre+ ) subjected to the AngII-induced murine model displayed significant reduction AAA development (n=10/group, p<0.05) with decreased CD4+ T H 17 cell activity and reciprocal increase in Treg expression. Conclusions: MLL1, a histone methyltransferase, regulates Notch1 signaling and T-cell differentiation in AAA development, causing pathologic inflammation. This pathway may serve as a potential therapeutic target to prevent aortic dilation.

Fungal Apoptosis-Related Proteins.

Programmed cell death (PCD) plays a crucial role in the development and homeostasis maintenance of multicellular organisms. Apoptosis is a form of PCD that prevents pathological development by eliminating damaged or useless cells. Despite the complexity of fungal apoptosis mechanisms being similar to those of plants and metazoans, fungal apoptosis lacks the core regulatory elements of animal apoptosis. Apoptosis-like PCD in fungi can be triggered by a variety of internal and external factors, participating in biological processes such as growth, development, and stress response. Although the core regulatory elements are not fully understood, apoptosis-inducing factor and metacaspase have been found to be involved. This article summarizes various proteins closely related to fungal apoptosis, such as apoptosis-inducing factor, metacaspase, and inhibitors of apoptosis proteins, as well as their structures and functions. This research provides new strategies and ideas for the development of natural drugs targeting fungal apoptosis and the control of fungal diseases.

Artificial Gravity Attenuates the Transcriptomic Response to Spaceflight in the Optic Nerve and Retina.

The development of eye pathology is a serious concern for astronauts who spend time in deep space. Microgravity is a major component of the spaceflight environment which could have adverse effects on ocular health. The use of centrifugation to exert forces that partially or fully mimic Earth-level gravity in space is a possible countermeasure to mitigate the effects of microgravity on the eye. Therefore, we subjected mice on the International Space Station (ISS) to microgravity (0 G) or artificial gravity by centrifugation at 0.33 G, 0.67 G, and 1 G, and then performed RNA sequencing (RNA-seq) on optic nerve and retinal tissue after returning them to Earth alive. We find that the microgravity environment induces transcriptomic changes in the optic nerve and retina consistent with an increased oxidative stress load, inflammation, apoptosis, and lipid metabolic stress. We also find that adding artificial gravity on board the ISS attenuates the transcriptomic response to microgravity in a dose-dependent manner. Such attenuation may effectively protect from and mitigate spaceflight-induced detrimental effects on ocular tissue.

Serum IgG Antibody against Helicobacter pylori and the CagA-VacA s1 Genotype as a Predictor of Gastric Cancer Development: A Case-control Study

Helicobacter pylori (H. pylori)-induced inflammation increases the risk of developing various upper gastrointestinal conditions which may progress to gastric cancer (CA). Early prediction and detection of infection are crucial for reducing cancer-induced mortality rates. The present case-control study aimed to investigate the combination of serum and molecular markers and H. pylori-associated gastro-duodenal conditions as risk factors for predicting CA development in dyspeptic patients. Consecutive gastric biopsies and blood samples were collected from 100 adult dyspeptic patients. Serum IgG antibody levels against H. pylori were determined, and receiver operating characteristic (ROC) analysis was performed. The expression of the virulence genes cagA and vacA was evaluated by polymerase chain reaction (PCR). A significant association was reported between the disease condition and the status of several risk factors, such as family history, serum IgG antibody concentration, and the virulence genes cagA and vacA. Among the 71 H. pylori-positive patients, 35.2% (25/71) had CA. Both cagA and vacA genes were found in 46 out of 71 (64.7%) patients, and 92% (23/25) of CA patients carried the cagA+vacA s1 gene. ROC analysis of the serum IgG concentrations revealed AUC values of 0.81 and 0.78 for differentiating patients with non-ulcer dyspepsia from those with ulceration/inflammation and CA, respectively. The concordance between the IgG-positive and PCR-positive patients was 84% (k value=0.41). Patients who had a family history of CA with an increased serum IgG concentration and the presence of H. pylori cagA-vacA s1 genotypes may be considered strong predictors of future development of gastric pathologies, including CA.

Hippocampus under pressure: molecular mechanisms of cognitive impairment in shr rats

In clinical studies and in animal experiments, data have been obtained indicating the association of chronic hypertension with the development of cognitive impairment. The review examines structural and biochemical changes in the hippocampus of SHR rats with genetic hypertension, which are used as a model of essential hypertension, as well as vascular dementia. The dysfunction of the hypothalamic-pituitary-adrenocortical system, observed in SHR rats at an early age, may, along with the development of hypertension, be a key factor in the damage to the hippocampus at the structural and molecular levels. Global changes at the body level (hypertension, neurohumoral dysfunction) are associated with the development of vascular pathology and destruction of the blood-brain barrier. Changes in multiple biochemical glucocorticoid-dependent processes in the hippocampus (dysfunction of steroid hormone receptors, disorders of neurotransmitter systems, BDNF deficiency, oxidative stress, neuroinflammation) are accompanied by structural changes including cellular processes of neuroinflammation (microgliosis, astrogliosis), disorders of neurogenesis in the subgranular neurogenic niche, neurodegenerative processes at the level of synapses, axons and dendrites up to neuronal cell death. The consequence of this is dysfunction of the hippocampus, a key structure of the limbic system necessary for the realization of cognitive functions. Summarizing of the available results at various levels, from the level of the organism and the structure of the brain (hippocampus) to the molecular one, allows us to confirm the translational validity of SHR rats for modeling the mechanisms of vascular dementia.

Childhood sexual abuse as a predictor of Complex Posttraumatic Stress Disorder: A meta-analysis

Sexual abuse, especially when it occurs during childhood, is an experience that causes deep and lasting harm. Currently, its study as a risk factor for the development of trauma-related pathologies is of great relevance. In 2018, Complex Post-Traumatic Stress Disorder (CPTSD) was officially recognized as a distinct syndrome in the 11th Revision of the International Classification of Diseases (ICD-11), with the aim of distinguishing it from neurotic disorders secondary to stressful situations, somatoform disorders, and those specifically associated with stress. The inclusion of CPTSD in the ICD-11 marked the culmination of two decades of research dedicated to understanding its symptoms, treatments, and risk factors. This article aims to conduct a meta-analysis that explores the relationship between sexual abuse and the development of CPTSD. Fifteen studies were selected for analysis, and the results revealed several key risk factors associated with the development of CPTSD, with the primary one being childhood sexual abuse (k = 15; OR = 3.007).

Targeting Endogenous Tau in Seeded Tauopathy Models Inhibits Tau Spread.

The transmission of tau pathology has been proposed as one of the major mechanisms for the spatiotemporal spreading of tau pathology in neurodegenerative diseases. Over the last decade, studies have demonstrated that targeting total or pathological tau using tau antibodies can mitigate the development of tau pathology in tauopathy or Alzheimer's disease (AD) mouse models, and multiple tau immunotherapy agents have progressed to clinical trials. Tau antibodies are believed to inhibit the internalization of pathologic seeds and/or block seed elongation after seed internalization. To further address the mechanism of tau antibody inhibition of pathological spread, we conducted immunotherapy studies in mouse primary neurons and wild-type mice (females) seeded with AD patient-derived tau to induce the formation and spreading of tau pathology. Notably, we evaluated the effect of a mouse tau-specific antibody (mTau8) which does not interact with AD-tau seeds in these models. Our results show that mTau8 crosses the blood-brain barrier at levels similar to other antibodies and effectively decreases AD-tau-seeded tau pathology in vitro and in vivo. Importantly, our data suggest that mTau8 binds to endogenous intraneuronal mouse tau, thereby inhibiting the elongation of internalized tau seeds. These findings provide valuable insights into the possible mechanism underlying antibody-based therapies for treating tauopathies.Significance Statement The transmission of tau pathology plays key role in the pathoclinical progression of tauopathy. Studies have shown that tau antibody treatment can mitigate tau pathology in transgenic and spreading models of tauopathy. To explore the mechanisms involved in this procedure, we conducted immunotherapy studies on human tau seeds induced tau spreading models using a mouse tau-specific antibody (mTau8), which does not interact with human-tau seeds. Our findings in the study enhance our understanding of antibody-based therapies for tauopathies.

Reference TSH intervals in children and adolescents

The determination of thyroid-stimulating hormone (TSH) is a reliable test in the diagnosis of thyroid dysfunction. When interpreting the results of quantitative studies, clinicians and laboratory specialists most often resort to assessing the compliance of the result with reference intervals (RI). At the same time, each laboratory must determine them for the analytes being measured itself, based on the characteristics of the examined contingent, the characteristics of the analytical systems used, achieving the necessary diagnostic effectiveness of the methodology, and its own practical and economic capabilities. A retrospective analysis of TSH results obtained from patients under the age of 19 years was performed for 54970 patients without hypothyroidism, thyrotoxicosis and pituitary disorders. For this work, reagent kits “Roche Cobas”, “Mindray” and “Beckman Coulter” (ICLA) were used. As a result, RI was established for the study of TSH for three sets of reagents with appropriate equipment and for four age groups: 1 day – 6 months, 6 months – 7 years, 7–14 years, 14–19 years. This makes it possible to use RI as a screening for possible developmental pathology in childhood and to determine the limits of clinical decisions.

Effects of High Altitude and Diet on Gastric Disease Severity in Helicobacter pylori Infection in Peru.

Helicobacter pylori is a bacterium that infects approximately half of the world's population, being more prevalent in low- and middle-income countries. H. pylori can cause gastritis, peptic ulcer disease, mucosa-associated lymphoid tissue lymphoma, and gastric cancer, which is among the five most frequent cancers worldwide. Other factors such as a diet low in vegetables and high in processed red meat have been associated with gastric cancer. Here, we studied the effects of high altitude and diet on gastric disease severity in H. pylori infection in a multicenter cross-sectional study in Peru (N = 343). We recruited people from villages with distinct eating habits (high meat consumption, mixed, and limited meat consumption diet) in the Andes (Puno), and compared them to people living at sea level with a mixed diet (Lima). H. pylori infection prevalence was higher at high altitude than at sea level. High altitude, diet, and age showed a significant correlation with the severity of gastric disease, whereas H. pylori infection and sex did not. However, high altitude was not found to be a risk factor for intestinal metaplasia, while diet and age were. At high altitude, a meat-rich diet was associated with a higher incidence of metaplasia compared to a plant-based diet. This study provides a comparison of communities living at high altitude with spontaneously different diets, showing that high processed red meat consumption is a risk factor for gastric disease. Further studies are needed to explain this phenomenon and its impact on the development and progression of gastric pathologies.

Advanced soft tissue visualization in conjunction with bone structures using contrast-enhanced micro-CT.

Micro-computed tomography (CT) analysis of soft tissues alongside bone remains challenging due to significant differences in X-ray absorption, preventing spatial inspection of bone remodeling including the cellular intricacies of mineralized tissues in developmental biology and pathology. The goal was to develop a protocol for contrast-enhanced micro-CT imaging that effectively visualizes soft tissues and cells in conjunction with bone while minimizing bone attenuation by decalcification. Murine femur samples were decalcified in ethylenediaminetetraacetic acid and treated with three different contrast agents: (i)iodine in ethanol, (ii)phosphotungstic acid in water, and (iii)Lugol's iodine. Micro-CT scans were performed in the laboratory setup SkyScan 1172 and at the synchrotron radiation for medical physics beamline in synchrotron radiation facility Elettra. Soft and hard tissue contrast-to-noise ratio (CNR) and contrast efficiency after decalcification were measured. In laboratory micro-CT, Lugol's iodine demonstrated a threefold higher CNR in the bone marrow, representing the soft tissue portion, compared with the bone. Contrast efficiencies, measured in synchrotron micro-CT, were consistent with these findings. Higher resolutions and the specificity of Lugol's iodine to cellular structures enabled detailed visualization of bone-forming cells in the epiphyseal plate. The combination of decalcification and the utilization of the contrast agent Lugol's iodine facilitated an enhanced soft tissue visualization in conjunction with bone.

FGF8-mediated gene regulation affects regional identity in human cerebral organoids.

The morphogen FGF8 establishes graded positional cues imparting regional cellular responses via modulation of early target genes. The roles of FGF signaling and its effector genes remain poorly characterized in human experimental models mimicking early fetal telencephalic development. We used hiPSC-derived cerebral organoids as an in vitro platform to investigate the effect of FGF8 signaling on neural identity and differentiation. We found that FGF8 treatment increases cellular heterogeneity, leading to distinct telencephalic and mesencephalic-like domains that co-develop in multi-regional organoids. Within telencephalic domains, FGF8 affects the anteroposterior and dorsoventral identity of neural progenitors and the balance between GABAergic and glutamatergic neurons, thus impacting spontaneous neuronal network activity. Moreover, FGF8 efficiently modulates key regulators responsible for several human neurodevelopmental disorders. Overall, our results show that FGF8 signaling is directly involved in both regional patterning and cellular diversity in human cerebral organoids and in modulating genes associated with normal and pathological neural development.