Austrian Drug Screening Institute (Austria)

Abstract Smoking is one of the single most important preventable risk factors for cancer and other adverse health outcomes 1,2 . Smoking cessation represents a key public health intervention with the potential to reduce its negative health outcomes 2–4 . While epidemiological, cross-sectional, and individual longitudinal ‘omic’ or biomarker studies have evaluated the impact of smoking cessation, no study to date has systematically profiled molecular and clinical changes in several organ systems or tissues longitudinally over the course of smoking cessation that could allow for more detailed assessment of response biomarkers and the identification of interindividual differences in the recovery of physiological functions. Here, we report the first human longitudinal multi-omic study of smoking cessation, evaluating 2,501 unique single or composite features from 1,094 longitudinal samples. Our comprehensive analysis, leveraging over half a million longitudinal data points, revealed a profound effect of smoking cessation on epigenetic biomarkers and microbiome features across multiple organ systems within 6 months of smoking cessation, alongside shifts in the immune and blood oxygenation system. Moreover, our multi-omic analysis provided unprecedented granularity that allows for identification of new cross-ome associations for mechanistic discovery. We anticipate that data and an interactive app from the Tyrol Lifestyle Atlas ( eutops.github.io/lifestyle-atlas ), comprising the current study and a parallel study arm evaluating the impact of diet on biomarkers of health and disease, will provide the basis for future discovery, biomarker benchmarking in their responsiveness to health-promoting interventions, and study of individualised response group, representing a major advance for personalised health monitoring using biomarkers.

To explore whether the reported lower pathogenicity in infected individuals of variant of concern (VoC) Omicron and its current subvariants compared to VoC Delta may be related to fundamental differences in the initial virus-tissue interaction, we assessed their ability to penetrate, replicate and cause damage in a human 3D respiratory model. For this, we used TEER measurements, real-time PCR, LDH, cytokine and complex confocal imaging analyses. We observed that Delta readily penetrated deep into the respiratory epithelium and this was associated with major tissue destruction, high LDH activity, high viral loads and pronounced innate immune activation as observed by intrinsic C3 activation and IL-6 release at infection sites. In contrast, Omicron subvariants BA.5, BQ.1.1 and BF7 remained superficially in the mucosal layer resulting merely in outward-directed destruction of cells, maintenance of epithelial integrity, minimal LDH activity and low basolateral release of virus at infection sites, as well as significantly smaller areas of complement activation and lower IL-6 secretion. Interestingly, also within Omicron subvariants differences were observed with newer Omicron subvariants BQ.1.1 and BF.7 illustrating significantly reduced viral loads, IL-6 release and LDH activity compared to BA.5. Our data indicate that earliest interaction events after SARS-CoV-2 transmission may have a role in shaping disease severity.

Delta-like homolog 1 (Dlk1), an inhibitor of adipogenesis, controls the cell fate of adipocyte progenitors. Experimental data presented here identify two independent regulatory mechanisms, transcriptional and translational, by which Ifrd1 (TIS7) and its orthologue Ifrd2 (SKMc15) regulate Dlk1 levels. Mice deficient in both Ifrd1 and Ifrd2 (dKO) had severely reduced adipose tissue and were resistant to high fat diet-induced obesity. Wnt signaling, a negative regulator of adipocyte differentiation was significantly up regulated in dKO mice. Elevated levels of the Wnt/β-catenin target protein Dlk1 inhibited the expression of adipogenesis regulators Pparg and Cebpa, and fatty acid transporter Cd36. Although both, Ifrd1 and Ifrd2, contributed to this phenotype, they utilized two different mechanisms. Ifrd1 acted by controlling Wnt signaling and thereby transcriptional regulation of Dlk1. On the other hand, distinctive experimental evidence showed that Ifrd2 acts as a general translational inhibitor significantly affecting Dlk1 protein levels. Novel mechanisms of Dlk1 regulation in adipocyte differentiation involving Ifrd1 and Ifrd2 are based on experimental data presented here.

In this study, different injection solutions containing opioid and nonopioid compounds used for patient‐controlled analgesia in hospice and palliative care were evaluated in terms of analyte stability. Investigated injection solutions contained different combinations of morphine, hydromorphone, metamizole and esketamine. For the practical implementation, samples from infusion pumps were daily drawn over a period of 7 days at 22 and 37°C. Quantitative measurements were performed on a high‐performance liquid chromatography system with ultraviolet detection applying a validated analytical method. All compounds apart from morphine showed no evident changes in concentration. However, a significant loss of morphine was observed for injection mixtures containing both morphine and metamizole at 37°C. After 7 days, only 72% of the initially measured morphine concentration was measured in the binary and 77% in the ternary mixture. Furthermore, an additional compound was detected that could represent the morphine‐metamizole‐adduct, “metamorphine”. Based on these results, a significantly reduced morphine concentration must be expected after only 3 days if an injection solution mixture containing both morphine and metamizole is administered to a patient at 37°C. Since the analgesic effects of morphine–metamizole adducts have not yet been thoroughly investigated, further clinical studies are necessary before accurate conclusions can be drawn in this regard.

This work is the first to describe the use of Direct Analysis in Real Time Mass Spectrometry (DART-MS) for the rapid quantification of cannabidiol (CBD) in CBD oils. For this study, self-prepared samples spiked with CBD in hemp seed oil as well as commercial CBD oils from the Austrian market with different CBD contents were analyzed. CBD concentrations were between 5 and 30% (m/m) for the spiked samples as well as between 5 and 15% (m/m) for the real samples. The performance of quantification by means of DART-MS was assessed against a validated liquid chromatography-mass spectrometry (LC-MS) method. The correlation of the quantification results of both methods was high with a correlation factor greater than 0.98 and a maximum bias of 9.8%. Furthermore, the relative standard deviation values of the DART-MS measurments were below the tolerable limit of 12%. These results demonstrate that quantification of CBD by DART-MS is reliable and hence suitable as a rapid and cost-effective alternative method for quality control of CBD content in CBD oils.

Electronic cigarettes (e-cigarettes) have gained increasing popularity in recent years, mostly because they are supposed to be less harmful than regular cigarettes. Therefore, it is highly imperative to investigate possible noxious effects to protect the consumers. E-liquids consist of propylene glycol, glycerol, aroma compounds and sweeteners. One of these sweeteners is a chlorinated version of sucrose, namely sucralose. The aim of this work was to investigate degradation products of sucralose in the presence of propylene glycol and glycerol at different temperatures of commercially available e-cigarettes. Chemical analysis and biological tests were simultaneously performed on e-liquid aerosol condensates. The results of the chemical analysis, which was executed by employing GC‐MS/GC-FID, demonstrated high amounts of various chloropropanols. The most abundant one is extremely toxic, namely 3-chloropropane-1,2-diol, which can be detected at concentrations ranging up to 10,000 mg/kg. Furthermore, a cytotoxicity investigation of the condensates was performed on HUVEC/Tert2 cells in which metabolic activity was determined by means of resazurin assay. The cellular metabolic activity significantly decreased by treatment with e-liquid aerosol condensate. Due to the results of this study, we advise against the use of sucralose as sweetener in e-liquids.

BackgroundExcessive inflammation triggered by a hitherto undescribed mechanism is a hallmark of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections and is associated with enhanced pathogenicity and mortality.ObjectiveComplement hyperactivation promotes lung injury and was observed in patients suffering from Middle East respiratory syndrome-related coronavirus, SARS-CoV-1, and SARS-CoV-2 infections. Therefore, we investigated the very first interactions of primary human airway epithelial cells on exposure to SARS-CoV-2 in terms of complement component 3 (C3)-mediated effects.MethodsFor this, we used highly differentiated primary human 3-dimensional tissue models infected with SARS-CoV-2 patient isolates. On infection, viral load, viral infectivity, intracellular complement activation, inflammatory mechanisms, and tissue destruction were analyzed by real-time RT-PCR, high content screening, plaque assays, luminex analyses, and transepithelial electrical resistance measurements.ResultsHere, we show that primary normal human bronchial and small airway epithelial cells respond to SARS-CoV-2 infection by an inflated local C3 mobilization. SARS-CoV-2 infection resulted in exaggerated intracellular complement activation and destruction of the epithelial integrity in monolayer cultures of primary human airway cells and highly differentiated, pseudostratified, mucus-producing, ciliated respiratory tissue models. SARS-CoV-2–infected 3-dimensional cultures secreted significantly higher levels of C3a and the proinflammatory cytokines IL-6, monocyte chemoattractant protein 1, IL-1α, and RANTES.ConclusionsCrucially, we illustrate here for the first time that targeting the anaphylotoxin receptors C3a receptor and C5a receptor in nonimmune respiratory cells can prevent intrinsic lung inflammation and tissue damage. This opens up the exciting possibility in the treatment of COVID-19.

How cells adjust nutrient transport across their membranes is incompletely understood. Previously, we have shown that S. cerevisiae broadly re-configures the nutrient transporters at the plasma membrane in response to amino acid availability, through endocytosis of sugar- and amino acid transporters (AATs) (Müller et al., 2015). A genome-wide screen now revealed that the selective endocytosis of four AATs during starvation required the α-arrestin family protein Art2/Ecm21, an adaptor for the ubiquitin ligase Rsp5, and its induction through the general amino acid control pathway. Art2 uses a basic patch to recognize C-terminal acidic sorting motifs in AATs and thereby instructs Rsp5 to ubiquitinate proximal lysine residues. When amino acids are in excess, Rsp5 instead uses TORC1-activated Art1 to detect N-terminal acidic sorting motifs within the same AATs, which initiates exclusive substrate-induced endocytosis. Thus, amino acid excess or starvation activate complementary α-arrestin-Rsp5-complexes to control selective endocytosis and adapt nutrient acquisition.