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During critical prenatal periods nutrient supply programs central control of energy balance, feed intake and body weight. In mammals as well in birds, prenatal glucose oversupply, for instance, may increases the offspring risk for metabolic disorders and related diseases throughout life. We investigated the long-lasting influence of temporary inovo hyperglycemia in chickens on neuronal mechanisms of the nucleus infundibuli hypothalami (NI), the 'critical' centre regulation metabolism, feed intake and body weight as well as on related peripheral parameters. Experiments were carried out in Lohmann White Leghorn chickens starting in embryos in three experimental groups: one untreated control group, one in-ovo injection control (daily single injection of 0.5 ml NaCl-solution) and one group prenatally experienced with temporary hyperglycemia by daily single injection of 0.5 ml glucose solution (30 mmol/l) from days 14 to 17 of incubation. After three weeks post-hatching in the untreated control group for the first time the avian neuronal glucose and leptin sensitivity was investigated using extracellular recordings in brain slices. Human recombinant leptin was also tested in combination with GABAB-receptor agonist and antagonist. The longlasting effect of temporary in-ovo hyperglycemia was examined on neuronal glucose sensitivity, the glucose transporter GLUT1 and 3, the leptin and insulin receptor expression as well as on the total body fat and blood glucose level. Additional, in male and female chickens the body weight development was recorded in all experimental groups during the total 3-weeks growing period. Results can be summarized as follows (RANCOURT et al., 2015; TZSCHENTKE et al., 2015; BOGATYREV et al., 2017): 1) NI-neurons show mammalian-like responsiveness after acute glucose, leptin and GABA application. GABAB-mechanisms involved in GABA release play a likely important role in the leptin mediated effects on NI neurons via functional leptin receptors. 4) Temporary late prenatal hyperglycemia induces lasting hypothalamic 'glucose-resistance' via strongly decreased neuronal glucose sensitivity as well glucose transporter, insulin- and leptin receptor expression. 3) After 3 weeks post-hatching the strong changes in the neuronal mechanisms were not accompanied by changes in related peripheral metabolic parameters as well in the body weight in male and female chickens. In conclusion, although in 3 weeks old chickens no changes in peripheral metabolic parameters and body weight development could be observed, just temporary hyperglycemia during the second half of incubation, may leading to persistent malprogramming in central control of energy balance, feed intake and body weight as predisposition for the development of metabolic disorders. It shows a risk of in-ovo feeding. Further, the chicken embryo provides a valuable new model for investigating early central nervous origins of 'diabesity' and related diseases. [ABSTRACT FROM AUTHOR]

Objective: The European Health Data Space (EHDS) shapes the digital transformation of healthcare in Europe. The EHDS regulation will also accelerate the use of health data for research, innovation, policy-making, and regulatory activities for secondary use of data (known as EHDS2). The Integration of heterogeneous Data and Evidence towards Regulatory and HTA Acceptance (IDERHA) project builds one of the first pan-European health data spaces in alignment with the EHDS2 requirements, addressing lung cancer as a pilot., Methods: In this study, we conducted a comprehensive review of the EHDS regulation, technical requirements for EHDS2, and related projects. We also explored the results of the Joint Action Towards the European Health Data Space (TEHDAS) to identify the framework of IDERHA's alignment with EHDS2. We also conducted an internal webinar and an external workshop with EHDS experts to share expertise on the EHDS requirements and challenges., Results: We identified the lessons learned from the existing projects and the minimum-set of requirements for aligning IDERHA infrastructure with EHDS2, including user journey, concepts, terminologies, and standards. The IDERHA framework (i.e., platform architecture, standardization approaches, documentation, etc.) is being developed accordingly., Discussion: The IDERHA's alignment plan with EHDS2 necessitates the implementation of three categories of standardization for: data discoverability: Data Catalog Vocabulary (DCAT-AP), enabling semantics interoperability: Observational Medical Outcomes Partnership (OMOP), and health data exchange (DICOM and FHIR). The main challenge is that some standards are still being refined, e.g., the extension of the DCAT-AP (HealthDCAT-AP). Additionally, extensions to the Observational Health Data Sciences and Informatics (OHDSI) OMOP Common Data Model (CDM) to represent the patient-generated health data are still needed. Finally, proper mapping between standards (FHIR/OMOP) is a prerequisite for proper data exchange., Conclusions: The IDERHA's plan and our collaboration with other EHDS initiatives/projects are critical in advancing the implementation of EHDS2., Competing Interests: No competing interests were disclosed., (Copyright: © 2024 Hussein R et al.)

Samples for histological analyses are often formalin-fixed paraffin-embedded (FFPE) and slide-mounted, which complicates RNA extraction for many downstream molecular applications. Furthermore, when the region of interest is extremely small due to isolation with laser microdissection (LMD), extracting RNA of adequate quality and quantity is difficult. We describe an optimized protocol for maximizing RNA output from FFPE tissue devised to identify and analyze gene expression of human maternal uterine scar tissue obtained from uterotomy scars resulting from prior cesarean deliveries. Gomori trichrome staining allowed for region identification for LMD. Successful RNA isolation, reverse transcription and, importantly, quantitative real-time PCR (qRT-PCR) were performed. This report provides an optimized step-by-step protocol yielding sufficient RNA for qRT-PCR analyses from challenging tissue/LMD-FFPE samples.

Nitrogen mustard (NM) is a cytotoxic vesicant known to cause acute lung injury which progresses to fibrosis. Herein, we developed a murine model of NM-induced pulmonary toxicity with the goal of assessing inflammatory mechanisms of injury. C57BL/6J mice were euthanized 1-28 d following intratracheal exposure to NM (0.08 mg/kg) or PBS control. NM caused progressive alveolar epithelial thickening, perivascular inflammation, bronchiolar epithelial hyperplasia, interstitial fibroplasia and fibrosis, peaking 14 d post exposure. Enlarged foamy macrophages were also observed in the lung 14 d post NM, along with increased numbers of microparticles in bronchoalveolar lavage fluid (BAL). Following NM exposure, rapid and prolonged increases in BAL cells, protein, total phospholipids and surfactant protein (SP)-D were also detected. Flow cytometric analysis showed that CD11b + Ly6G - F4/80 + Ly6C hi proinflammatory macrophages accumulated in the lung after NM, peaking at 3 d. This was associated with macrophage expression of HMGB1 and TNFα in histologic sections. CD11b + Ly6G - F4/80 + Ly6C lo anti-inflammatory/pro-fibrotic macrophages also increased in the lung after NM peaking at 14 d, a time coordinate with increases in TGFβ expression and fibrosis. NM exposure also resulted in alterations in pulmonary mechanics including increases in tissue elastance and decreases in compliance and static compliance, most prominently at 14 d. These findings demonstrate that NM induces structural and inflammatory changes in the lung that correlate with aberrations in pulmonary function. This mouse model will be useful for mechanistic studies of mustard lung injury and for assessing potential countermeasures., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Inc.)

Study Question: Do fertility treatments, including ovulation induction (OI), alter epigenetic mechanisms such as DNA methylation at imprinted loci?, Summary Answer: We observed small but statistically significant differences in certain imprinting control regions (ICRs) based on the method of conception, however, these small changes in methylation did not correlate to the overall transcriptional levels of the genes adjacent to the ICRs (such as KCNQ1 and SNRPN)., What Is Known and What This Paper Adds: Assisted reproductive technology (ART) has been associated with an increase in the risk of rare childhood disorders caused by loss of imprinting (LOI). This study provides novel epigenetic analyses on infants conceived by OI and examines how methylation levels correlate with gene expression., Design: Data and biospecimens used in this study were from 147 participants of the Epigenetic Birth Cohort comprising 1941 mother-child dyads recruited between June 2007 and June 2009 at the Department of Obstetrics, Gynecology and Reproductive Biology at Brigham and Women's Hospital (BWH) in Boston, MA, USA. Wilcoxon rank-sum tests were used to examine the differences in median percent methylation at each differentially methylated region (DMR) between the spontaneous conception control group and the fertility treatment groups (OI and IVF)., Participants and Setting: For each woman who reported IVF we selected a woman who conceived spontaneously matched on age (± 2 years). To increase efficiency, we matched the same controls from the spontaneously conceived group to participants who reported OI. If an appropriate control was not identified that had been previously matched to an IVF participant, a new control was selected. The final analytic sample consisted of 61 spontaneous, 59 IVF and 27 OI conceptions., Main Results and the Role of Chance: No functionally relevant differences in methylation levels were observed across five (out of six) imprinted DMRs in either the placenta or cord blood of infants conceived with OI or IVF compared with infants conceived spontaneously. While KCNQ1, SNRPN and H19 DMRs demonstrated small but statistically significant differences in methylation based on the method of conception, expression levels of the genes related to these control regions only correlated with the methylation levels of H19., Bias, Confounding and Other Reasons for Caution: Limitations of our study include the limited sample size, lack of information on OI medication used and culture medium for the IVF procedures and underlying reasons for infertility among OI and IVF patients. We did not perform allele-specific expression analyses and therefore cannot make any inferences about LOI., Generalizability to Other Populations: These results are likely to be generalizable to non-Hispanic white individuals in populations with similar ART and fertility treatments.

Objectives: DNA methylation is known to play a critical role in regulating development of placental morphology and physiology. The methylation of genes mediated by glucocorticoid hormones may be particularly vulnerable to intrauterine stress in the placenta. However little is known about DNA methylation of stress-related genes within a healthy placenta, and particularly whether methylation occurs uniformly across different regions of the placenta, which is a critical question for researchers seeking to analyze methylation patterns. We examined DNA methylation across four regions of the placenta to evaluate methylation levels of stress-related genes within a healthy placenta, and to evaluate whether methylation patterns vary by sampling location., Study Design: We evaluated levels of DNA methylation of three stress-related genes: NR3C1, BDNF, and 11B-HSD2 and of the repetitive element, LINE-1, in four different sample locations of 20 healthy placentas., Main Outcome Measures: Pyrosequencing was used to quantify levels of methylation at CpG sites within the promoter regions of each of the three stress-related genes, and global methylation of LINE-1., Results: Very low levels of methylation were found across all three stress-related genes; no gene showed a median methylation level greater than 4.20% across placental regions. Variation in methylation between placental regions for stress-related genes and for LINE-1 was minimal., Conclusions: Our data suggest that these frequently studied stress-related genes have low levels of methylation in healthy placenta tissue. Minimal variation between sites suggests that sampling location does not affect DNA methylation analyses of these genes or of LINE-1 repetitive elements., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

To recapitulate the stochastic nature of human cancer development, we have devised a strategy for generating mouse tumor models that involves stepwise genetic manipulation of embryonic stem (ES) cells and chimera generation. Tumors in the chimeric animals develop from engineered cells in the context of normal tissue. Adenocarcinomas arising in an allelic series of lung cancer models containing HER2 (also known as ERBB2), KRAS or EGFR oncogenes exhibit features of advanced malignancies. Treatment of EGFR(L858R) and KRAS(G12V) chimeric models with an EGFR inhibitor resulted in near complete tumor regression and no response to the treatment, respectively, accurately reflecting previous clinical observations. Transcriptome and immunohistochemical analyses reveal that PI3K pathway activation is unique to ERBB family tumors whereas KRAS-driven tumors show activation of the JNK/SAP pathway, suggesting points of therapeutic intervention for this difficult-to-treat tumor category.

The beneficial effects of supplemental oxygen delivered to patients suffering from acute respiratory distress is offset by its reduction to genotoxic reactive oxygen species (ROS) that inhibit proliferation and kill pulmonary cells. Cells respond to oxygen-induced damage by expressing the tumor suppressor p53 and the cyclin-dependent kinase inhibitor p21(Cip1/WAF1/Sdi1) (p21), which limits proliferation by blocking entry into S phase. Since preventing DNA synthesis during genotoxic stress may enhance survival, the current study examines whether hyperoxia induces p21 through a p53-dependent pathway and whether p21 protects cells from the toxic effects of oxygen. HCT116 colon carcinoma cells and clonal lines lacking p53 or p21were used in this study because they allow direct cytotoxic comparisons between isogenic cells, without complications arising from unknown genetic differences between nonhomologous cell lines. Hyperoxia (95% O2, 5% CO2) increased p53 abundance, phosphorylation of p53 on serine 15, and p21 mRNA and protein in parental HCT116 cells that ceased proliferation. In contrast, p21 was not detected in either p53- or p21-deficient HCT116 cells, which exited the G1 compartment and were arrested in S and G2/M phases during hyperoxia. Trypan blue-dye exclusion revealed that induction of p21 markedly enhanced survival during exposure and colony survival assays showed that p21 enhanced the ability to resume proliferation during recovery in room air. The observation that p53-dependent induction of p21 prevents exit from G1 and promotes survival during hyperoxia is consistent with the importance of limiting DNA replication during genotoxic stress caused by oxygen exposure.

Previous studies have shown that hyperoxia inhibits proliferation and increases the expression of the tumor suppressor p53 and its downstream target, the cyclin-dependent kinase inhibitor p21(CIP1/WAF1), which inhibits proliferation in the G1 phase of the cell cycle. To determine whether growth arrest was mediated through activation of the p21-dependent G1 checkpoint, the kinetics of cell cycle movement during exposure to 95% O2 were assessed in the Mv1Lu and A549 pulmonary adenocarcinoma cell lines. Cell counts, 5-bromo-2'-deoxyuridine incorporation, and cell cycle analyses revealed that growth arrest of both cell lines occurred in S phase, with A549 cells also showing evidence of a G1 arrest. Hyperoxia increased p21 in A549 but not in Mv1Lu cells, consistent with the activation of the p21-dependent G1 checkpoint. The ability of p21 to exert the G1 arrest was confirmed by showing that hyperoxia inhibited proliferation of HCT 116 colon carcinoma cells predominantly in G1, whereas an isogenic line lacking p21 arrested in S phase. The cell cycle arrest in S phase appears to be a p21-independent process caused by a gradual reduction in the rate of DNA strand elongation. Our data reveal that hyperoxia inhibits proliferation in G1 and S phase and demonstrate that p53 and p21 retain their ability to affect G1 checkpoint control during exposure to elevated O2 levels.

The authors, using the narrative form, examine three epistemological structures that are found in nursing curricula. Those fundamentally different approaches of knowledge acquisition are portrayed and discussed by the director and three staff members from a fictitious nursing school. Although faculty members naturally tend to adhere to their own embedded view of the way one learns, the authors stress the need for judicious blend of the three epistemic orientations when developing nursing curricula.

High concentrations of O(2) inhibit epithelial cell proliferation that resumes on recovery in room air. To determine whether growth arrest is mediated by transforming growth factor-beta (TGF-beta), changes in cell proliferation during exposure to hyperoxia were assessed in the mink lung epithelial cell line Mv1Lu and the clonal variant R1B, which is deficient for the type I TGF-beta receptor. Mv1Lu cells treated with TGF-beta accumulated in the G(1) phase of the cell cycle as determined by propidium iodide staining, whereas proliferation of R1B cells was unaffected by TGF-beta. In contrast, hyperoxia inhibited proliferation of both cell lines within 24 h of exposure through an accumulation in the S phase. Mv1Lu cells treated with TGF-beta and exposed to hyperoxia accumulated in the G(1) phase, suggesting that TGF-beta can inhibit the S phase accumulation observed with hyperoxia alone. Cyclin A was detected in cultures exposed to room air or growth arrested by hyperoxia while decreasing in cells growth arrested in the G(1) phase by TGF-beta. Finally, hyperoxia failed to activate a TGF-beta-dependent transcriptional reporter in both Mv1Lu and R1B cells. These findings reveal that simple growth arrest by hyperoxia involves a defect in S phase progression that is independent of TGF-beta signaling.

In hospitals administrators work with two major groups of medical professionals: physicians and nurses. Together, these three groups of professionals are primarily responsible for the care of patients. The quality of the service they provide is dependent on the working relationships among these disciplines. Potentially, the more harmony among administrators, physicians, and nurses, the better care patients can receive. The research reported in this article addresses the issue of interdisciplinary misunderstanding among these groups of professionals. The research was premised on the assumption that hospital administrators, physicians, and nurses differ in the way they access knowledge. That is, administrators, physicians, and nurses access knowledge in dissimilar ways and, by doing so, set the stage for crises and conflicts. Using a standardized instrument to assess knowledge-accessing modes, data were collected from samples of health administration, medical, and nursing students. The data indicated that the three groups did differ in the way they accessed knowledge. The individual discipline profiles generated from the data also revealed a finding common to the three disciplines, a finding may help resolve a very real problem for hospital patients. Based on the research results, the article discusses ways that administrators may be able to prevent crises and conflicts and enhance harmonious relations among the three disciplines, and thereby improve patient care. Avenues for further research are also suggested.