Continual proteomic divergence of HepG2 cells as a consequence of long-term spheroid culture

Three-dimensional models are considered a powerful tool for improving the concordance between in vitro and in vivo phenotypes. However, the duration of spheroid culture may infuence the degree of correlation between these counterparts. When using immortalised cell lines as model systems, the assumption for consistency and reproducibility is often made without adequate characterization or validation. It is therefore essential to defne the biology of each spheroid model by investigating proteomic dynamics, which may be altered relative to culture duration. As an example, we assessed the infuence of culture duration on the relative proteome abundance of HepG2 cells cultured as spheroids, which are routinely used to model aspects of the liver. Quantitative proteomic profling of whole cell lysates labelled with tandem-mass tags was conducted using liquid chromatographytandem mass spectrometry (LC–MS/MS). In excess of 4800 proteins were confdently identifed, which were shared across three consecutive time points over 28 days. The HepG2 spheroid proteome was divergent from the monolayer proteome after 14 days in culture and continued to change over the successive culture time points. Proteins representing the recognised core hepatic proteome, cell junction, extracellular matrix, and cell adhesion proteins were found to be continually modulated.The National Research Foundation (NRF) Tuthuka PhD Track Funding Scheme, the NRF Innovation Masters and Doctoral Scholarship Program, University of Pretoria Doctoral Bursary as well as the NRF National Equipment Program.http://www.nature.com/srep/index.htmlpm2022PharmacologyPhysiolog

Social media and COVID-19 – perceptions and public deceptions regarding colchicine, hydroxychloroquine and ivermectin and resultant lessons for future pandemics

Background: The capacity for social media to influence the consumption of re-purposed medicines to manage COVID-19 despite limited safety and efficacy data at the start of the pandemic is cause for concern. Objective: To ascertain links between social media reports and utilization for three re-purposed medicines (hydroxychloroquine (HCQ), ivermectin and colchicine) to direct future activities. Methods: A combined retrospective analysis of social media posts for these re-purposed medicines was performed in South Africa between January and June 2021 together with utilization and clinical trials data. Utilisation data from IQVIA from three different platforms included private and public markets. Clinical trials data was obtained from various databases. Chloroquine data was analysed in South Africa (HCQ was not available). Results: 77257 posts were collected across key social media platforms during the study period of which 6884 were relevant. Ivermectin had the highest number of posts (55%) followed by HCQ (44%), with limited posts for colchicine (1%). The spike in ivermectin utilisation was closely correlated with social media posts. Similarly, with chloroquine social media interest was enhanced by comments from local politicians. Sentiment analysis showed that the posts regarding the effectiveness of particularly ivermectin and HCQ were positive. Of concern is that the origin of the majority of reporters in social media (85%) was unidentifiable. Conclusion: This is the first study of its kind in South Africa providing evidence that social media is a driver of re-purposed medicine use. Healthcare professionals have a key role to provide evidence-based advice especially with unidentifiable posts

Social media and COVID-19 – perceptions and public deceptions of ivermectin, colchicine and hydroxychloroquine : lessons for future pandemics

The capacity for social media to influence the utilization of re-purposed medicines to manage COVID-19, despite limited availability of safety and efficacy data, is a cause for concern within health care systems. This study sought to ascertain links between social media reports and utili-zation for three re-purposed medicines: hydroxychloroquine (HCQ), ivermectin and colchicine. A combined retrospective analysis of social media posts for these three re-purposed medicines was undertaken, along with utilization and clinical trials data, in South Africa, between January 2020 and June 2021. In total, 77,257 posts were collected across key social media platforms, of which 6884 were relevant. Ivermectin had the highest number of posts (55%) followed by HCQ (44%). The spike in ivermectin use was closely correlated to social media posts. Similarly, regarding chlo-roquine (as HCQ is not available in South Africa), social media interest was enhanced by local politicians. Sentiment analysis revealed that posts regarding the effectiveness of these repurposed medicines were positive. This was different for colchicine, which contributed only a small number of mentions (1%). Of concern is that the majority of reporters in social media (85%) were uniden-tifiable. This study provides evidence of social media as a driver of re-purposed medicines. Healthcare professionals have a key role in providing evidence-based advice especially with unidentifiable post

Characterization and reproducibility of HepG2 hanging drop spheroids toxicology in vitro

Hepatotoxicity remains a major challenge in drug development despite preclinical toxicity screening using hepatocytes of human origin. To overcome some limitations of reproducing the hepatic phenotype, more structurally and functionally authentic cultures in vitro can be introduced by growing cells in 3D spheroid cultures. Characterisation and reproducibility of HepG2 spheroid cultures using a high-throughput hanging drop technique was performed and features contributing to potential phenotypic variation highlighted. Cultured HepG2 cells were seeded into Perfecta 3D® 96-well hanging drop plates and assessed over time for morphology, viability, cell cycle distribution, protein content and protein-mass profiles. Divergent aspects which were assessed included cell stocks, seeding density, volume of culture medium and use of extracellular matrix additives. Hanging drops are advantageous due to no complex culture matrix being present, enabling background free extractions for downstream experimentation. Varying characteristics were observed across cell stocks and batches, seeding density, culture medium volume and extracellular matrix when using immortalized HepG2 cells. These factors contribute to wide-ranging cellular responses and highlights concerns with respect to generating a reproducible phenotype in HepG2 hanging drop spheroids.A National Research Foundation of South Africa Thuthuka grant (Grant No. 87880).http://www.elsevier.com/locate/tiv2019-08-01hj2018Pharmacolog

Survey of neonatal respiratory care and surfactant administration in very preterm infants in the Italian neonatal network

Introduction: Variation of respiratory care is described between centers around the world.The Italian Neonatal Network (INN), as a national group of the Vermont-Oxford Network (VON) allows to perform a wide analysis of respiratory care in very low birth weight infants. Methods:We analyzed the dataset of infants enrolled in the INN in 2009 and 2010 and, for surfactant administration only, from 2006 to 2010 from 83 participating centers. All definitions are those of the (VON). A questionnaire analysis was also performed with a questionnaire on centers practices. Results: We report data for 8297 infants. Data on ventilator practices and outcomes are outlined. Variation for both practices and outcome is found. Trend in surfactant administration is also analyzed. Conclusions. The great variation across hospitals in all the surveyed techniques points to the possibility of implementing potentially better practices with the aim of reducing unwanted variation. These data also show the power of large neonatal networks in identifying areas for potential improvement. \ua9 Mattioli 1885

Survey of neonatal respiratory care and surfactant administration in very preterm infants in the Italian neonatal network

Introduction: Variation of respiratory care is described between centers around the world.The Italian Neonatal Network (INN), as a national group of the Vermont-Oxford Network (VON) allows to perform a wide analysis of respiratory care in very low birth weight infants. Methods:We analyzed the dataset of infants enrolled in the INN in 2009 and 2010 and, for surfactant administration only, from 2006 to 2010 from 83 participating centers. All definitions are those of the (VON). A questionnaire analysis was also performed with a questionnaire on centers practices. Results: We report data for 8297 infants. Data on ventilator practices and outcomes are outlined. Variation for both practices and outcome is found. Trend in surfactant administration is also analyzed. Conclusions. The great variation across hospitals in all the surveyed techniques points to the possibility of implementing potentially better practices with the aim of reducing unwanted variation. These data also show the power of large neonatal networks in identifying areas for potential improvement. © Mattioli 1885

Association of maternal hypertension and chorioamnionitis with preterm outcomes

OBJECTIVES: We compared the relative effect of hypertensive disorders of pregnancy and chorioamnionitis on adverse neonatal outcomes in very preterm neonates, and studied whether gestational age (GA) modulates these effects. METHODS: A cohort of neonates 23 to 30 weeks' GA, born in 2008 to 2011 in 82 hospitals adhering to the Italian Neonatal Network, was analyzed. Infants born from mothers who had hypertensive disorders (N = 2096) were compared with those born after chorioamnionitis (N = 1510). Statistical analysis employed logistic models, adjusting for GA, hospital, and potential confounders. RESULTS: Overall mortality was higher after hypertension than after chorioamnionitis (odds ratio [OR], 1.39; 95% confidence interval [CI], 1.08-1.80), but this relationship changed across GA weeks; the OR for hypertension was highest at low GA, whereas from 28 weeks' GA onward, mortality was higher for chorioamnionitis. For other outcomes, the relative risks were constant across GA; infants born after hypertension had an increased risk for bronchopulmonary dysplasia (OR, 2.20; 95% CI, 1.68-2.88) and severe retinopathy of prematurity (OR, 1.48; 95% CI, 1.02-2.15), whereas there was a lower risk for early-onset sepsis (OR, 0.25; 95% CI, 0.19-0.34), severe intraventricular hemorrhage (OR, 0.65; 95% CI, 0.48-0.88), periventricular leukomalacia (OR, 0.70; 95% CI, 0.48-1.01), and surgical necrotizing enterocolitis or gastrointestinal perforation (OR, 0.47; 95% CI, 0.31-0.72). CONCLUSIONS: Mortality and other adverse outcomes in very preterm infants depend on antecedents of preterm birth. Hypertension and chorioamnionitis are associated with different patterns of outcomes; for mortality, the effect changes across GA weeks. Copyright \uc2\ua9 2014 by the American Academy of Pediatrics