It is well established that there is a link between inflammation and depression, with several studies reporting increased circulating levels of the pro-inflammatory cytokine, interleukin-6 (IL6), in ...depressed individuals. Peripheral epigenetic marks, including DNA methylation, hold promise as biomarkers for a range of complex conditions, with potential to inform diagnosis and tailor interventions. The aim of this study was to determine whether individuals with depression display differential methylation of the IL6 gene promoter compared to individuals without depression.
The ESPRIT study of later life neuropsychiatric disorders used a random sampling framework to select non-institutionalised participants aged ≥65 years and over living in the Montpellier region of France. Major depressive disorder (MDD) was assessed using the Mini International Neuropsychiatric Interview (MINI) according to DSM-IV criteria. High levels of depressive symptoms were defined as a score of ≥16 on the Centre for Epidemiologic Studies Depression Scale (CES-D). IL6 promoter DNA methylation was measured on a sub-sample of 380 participants who provided buccal samples.
Individuals with depression (current MDD or high depressive symptoms) had lower IL6 methylation levels at one of the four sites investigated, however the effect size was small (∆ 2.4%, SE 0.009, p = 0.006). Interestingly, antidepressant use was independently associated with higher IL-6 methylation at the same site (∆ 4.6%, SE 0.019, p = 0.015). In multivariate linear regression analyses adjusting for covariates, including sex and smoking status, these associations remained. There was no effect modification when considering IL6 genotype.
This study presents evidence that IL6 methylation may be a marker of depression status in older individuals, however further work is now needed to replicate these findings and to assess the association with inflammatory status of individuals.
Immune-mediated induction of cytidine deaminase APOBEC3B (A3B) expression leads to HBV covalently closed circular DNA (cccDNA) decay. Here, we aimed to decipher the signalling pathway(s) and ...regulatory mechanism(s) involved in A3B induction and related HBV control.
Differentiated HepaRG cells (dHepaRG) knocked-down for NF-κB signalling components, transfected with siRNA or micro RNAs (miRNA), and primary human hepatocytes ± HBV or HBVΔX or HBV-RFP, were treated with lymphotoxin beta receptor (LTβR)-agonist (BS1). The biological outcomes were analysed by reverse transcriptase-qPCR, immunoblotting, luciferase activity, chromatin immune precipitation, electrophoretic mobility-shift assay, targeted-bisulfite-, miRNA-, RNA-, genome-sequencing, and mass-spectrometry.
We found that canonical and non-canonical NF-κB signalling pathways are mandatory for A3B induction and anti-HBV effects. The degree of immune-mediated A3B production is independent of A3B promoter demethylation but is controlled post-transcriptionally by the miRNA 138-5p expression (hsa-miR-138-5p), promoting A3B mRNA decay. Hsa-miR-138-5p over-expression reduced A3B levels and its antiviral effects. Of note, established infection inhibited BS1-induced A3B expression through epigenetic modulation of A3B promoter. Twelve days of treatment with a LTβR-specific agonist BS1 is sufficient to reduce the cccDNA pool by 80% without inducing significant damages to a subset of cancer-related host genes. Interestingly, the A3B-mediated effect on HBV is independent of the transcriptional activity of cccDNA as well as on rcDNA synthesis.
Altogether, A3B represents the only described enzyme to target both transcriptionally active and inactive cccDNA. Thus, inhibiting hsa-miR-138-5p expression should be considered in the combinatorial design of new therapies against HBV, especially in the context of immune-mediated A3B induction.
Immune-mediated induction of cytidine deaminase APOBEC3B is transcriptionally regulated by NF-κB signalling and post-transcriptionally downregulated by hsa-miR-138-5p expression, leading to cccDNA decay. Timely controlled APOBEC3B-mediated cccDNA decay occurs independently of cccDNA transcriptional activity and without damage to a subset of cancer-related genes. Thus, APOBEC3B-mediated cccDNA decay could offer an efficient therapeutic alternative to target hepatitis B virus chronic infection.
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•Impairment of NF-κB signalling prevents APOBEC3B induction and cccDNA decay.•APOBEC3B is post-transcriptionally regulated by the hsa-miR-138-5p.•Over-expression of the hsa-miR-138-5p inhibits APOBEC3B expression and cccDNA decay.•A3B timely induces cccDNA decay without damage to cancer-related genes.•APOBEC3B-mediated cccDNA decay is independent of cccDNA transcriptional activity.
Hepatic innate immune control of viral infections has largely been attributed to Kupffer cells, the liver-resident macrophages. However, hepatocytes, the parenchymal cells of the liver, also possess ...potent immunological functions in addition to their known metabolic functions. Owing to their abundance in the liver and known immunological functions, we aimed to investigate the direct antiviral mechanisms employed by hepatocytes.
Using lymphocytic choriomeningitis virus (LCMV) as a model of liver infection, we first assessed the role of myeloid cells by depletion prior to infection. We investigated the role of hepatocyte-intrinsic innate immune signaling by infecting mice lacking canonical NF-κB signaling (IkkβΔHep) specifically in hepatocytes. In addition, mice lacking hepatocyte-specific interferon-α/β signaling-(IfnarΔHep), or interferon-α/β signaling in myeloid cells-(IfnarΔMyel) were infected.
Here, we demonstrate that LCMV activates NF-κB signaling in hepatocytes. LCMV-triggered NF-κB activation in hepatocytes did not depend on Kupffer cells or TNFR1 signaling but rather on Toll-like receptor signaling. LCMV-infected IkkβΔHep livers displayed strongly elevated viral titers due to LCMV accumulation within hepatocytes, reduced interferon-stimulated gene (ISG) expression, delayed intrahepatic immune cell influx and delayed intrahepatic LCMV-specific CD8+ T cell responses. Notably, viral clearance and ISG expression were also reduced in LCMV-infected primary hepatocytes lacking IKKβ, demonstrating a hepatocyte-intrinsic effect. Similar to livers of IkkβΔHep mice, enhanced hepatocytic LCMV accumulation was observed in livers of IfnarΔHep mice, whereas IfnarΔMyel mice were able to control LCMV infection. Hepatocytic NF-κB signaling was also required for efficient ISG induction in HDV-infected dHepaRG cells and interferon-α/β-mediated inhibition of HBV replication in vitro.
Together, these data show that hepatocyte-intrinsic NF-κB is a vital amplifier of interferon-α/β signaling, which is pivotal for strong early ISG responses, immune cell infiltration and hepatic viral clearance.
Innate immune cells have been ascribed a primary role in controlling viral clearance upon hepatic infections. We identified a novel dual role for NF-κB signaling in infected hepatocytes which was crucial for maximizing interferon responses and initiating adaptive immunity, thereby efficiently controlling hepatic virus replication.
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•LCMV infection activates NF-κB signaling in hepatocytes.•Macrophages, TNFR1 signaling do not induce LCMV-driven hepatocyte NF-κB-activation.•IkkβΔHep mice display increased viral infection/replication and lower ISG induction.•IfnarΔHep mice recapitulate aberrant virus replication as observed in IkkβΔHep mice.•NF-κB signaling is required for efficient ISG induction in HBV-/HDV-infected HepaRG.
In vitro models of the human blood-brain barrier (BBB) are highly desirable for drug development. This study aims to analyze a set of ten different BBB culture models based on primary cells, human ...induced pluripotent stem cells (hiPSCs), and multipotent fetal neural stem cells (fNSCs). We systematically investigated the impact of astrocytes, pericytes, and NSCs on hiPSC-derived BBB endothelial cell function and gene expression. The quadruple culture models, based on these four cell types, achieved BBB characteristics including transendothelial electrical resistance (TEER) up to 2,500 Ω cm2 and distinct upregulation of typical BBB genes. A complex in vivo-like tight junction (TJ) network was detected by freeze-fracture and transmission electron microscopy. Treatment with claudin-specific TJ modulators caused TEER decrease, confirming the relevant role of claudin subtypes for paracellular tightness. Drug permeability tests with reference substances were performed and confirmed the suitability of the models for drug transport studies.
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•Establishment of a standardized human BBB co-culture model based on hiPSCs and fNSCs•Reflection of physiological BBB integrity and expression of relevant transporters/TJs•Confirmation of TJ network functionality by claudin-specific TJ modulators•Validation of physiological transcellular model tightness by permeability studies
In this article, Metzger and colleagues present the establishment of physiologically relevant human blood-brain barrier quadruple culture models based on induced pluripotent and multipotent stem cells. The novel model can be used as a powerful tool in pharmaceutical drug research and ADMET studies, which may allow a more reliable translation of promising drug candidates into clinical application.
Pancreatic cystic lesions (PCL) are increasingly diagnosed. Endoscopic ultrasound fine-needle aspiration (EUS-FNA) cytology is often used for diagnostic confirmation but can be inconclusive. In this ...study, the role of molecular analyses in the pre-operative diagnostics of PCL is evaluated. Targeted Next Generation Sequencing (NGS) applied on cytology smears was retrospectively evaluated in a cohort of 37 resected PCL. Usefulness of NGS on fresh cyst fluids was tested in a prospective cohort of patients with newly diagnosed PCL (n = 71). In the retrospective cohort, cytology plus NGS displayed higher sensitivity (94.1% vs. 87.1%) and specificity (100% vs. 50%) than cytology alone for the detection of mucinous neoplasms. In the prospective cohort, sensitivity and specificity of conventional cytology alone were 54.2% and 100% for the detection of mucinous neoplasia and 50.0% and 100% for the detection of high-grade dysplasia, respectively. Adding NGS, all lesions which underwent histopathologic verification (12/71, 17%) could be classified without false positive or false negative results regarding the detection of mucinous neoplasm so far. NGS analysis of cfDNA in PCL fluids is feasible and can increase diagnostic accuracy in the detection of mucinous neoplasms compared to cytology alone. However, algorithms for the detection of high-risk lesions need further improvement.
Many pathogenic bacteria produce pore-forming toxins to attack and kill human cells. We have determined the 4.5 Å structure of the ~2.2 MDa pore complex of pneumolysin, the main virulence factor of
, ...by cryoEM. The pneumolysin pore is a 400 Å ring of 42 membrane-inserted monomers. Domain 3 of the soluble toxin refolds into two ~85 Å β-hairpins that traverse the lipid bilayer and assemble into a 168-strand β-barrel. The pore complex is stabilized by salt bridges between β-hairpins of adjacent subunits and an internal α-barrel. The apolar outer barrel surface with large sidechains is immersed in the lipid bilayer, while the inner barrel surface is highly charged. Comparison of the cryoEM pore complex to the prepore structure obtained by electron cryo-tomography and the x-ray structure of the soluble form reveals the detailed mechanisms by which the toxin monomers insert into the lipid bilayer to perforate the target membrane.
To fulfill its role in protein biogenesis, the endoplasmic reticulum (ER) depends on the Hsp70-type molecular chaperone BiP, which requires a constant ATP supply. However, the carrier that catalyzes ...ATP uptake into the ER was unknown. Here, we report that our screen of gene expression datasets for member(s) of the family of solute carriers that are co-expressed with BiP and are ER membrane proteins identifies SLC35B1 as a potential candidate. Heterologous expression of SLC35B1 in E. coli reveals that SLC35B1 is highly specific for ATP and ADP and acts in antiport mode. Moreover, depletion of SLC35B1 from HeLa cells reduces ER ATP levels and, as a consequence, BiP activity. Thus, human SLC35B1 may provide ATP to the ER and was named AXER (ATP/ADP exchanger in the ER membrane). Furthermore, we propose an ER to cytosol low energy response regulatory axis (termed lowER) that appears as central for maintaining ER ATP supply.
When barriers ignore the “rule-of-five” Krämer, Stefanie D.; Aschmann, Hélène E.; Hatibovic, Maja ...
Advanced drug delivery reviews,
06/2016, Letnik:
101
Journal Article
Recenzirano
Why are a few drugs with properties beyond the rule of 5 (bRo5) absorbed across the intestinal mucosa while most other bRo5 compounds are not? Are such exceptional bRo5 compounds exclusively taken up ...by carrier-mediated transport or are they able to permeate the lipid bilayer (passive lipoidal diffusion)? Our experimental data with liposomes indicate that tetracycline, which violates one rule of the Ro5, and rifampicin, violating three of the rules, significantly permeate a phospholipid bilayer with kinetics similar to labetalol and metoprolol, respectively. Published data from experimental work and molecular dynamics simulations suggest that the formation of intramolecular H-bonds and the possibility to adopt an elongated shape besides the presence of a significant fraction of net neutral species facilitate lipid bilayer permeation. As an alternative to lipid bilayer permeation, carrier proteins can be targeted to improve absorption, with the potential drawbacks of drug–drug interactions and non-linear pharmacokinetics.
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Objective We aimed to prospectively derive and validate a novel 0-/1-hour algorithm using high-sensitivity cardiac troponin I (hs-cTnI) for the early “rule-out” and “rule-in” of acute myocardial ...infarction (AMI). Methods In a prospective multicenter diagnostic study, we enrolled 1,500 patients presenting with suspected AMI to the emergency department. The final diagnosis was centrally adjudicated by 2 independent cardiologists blinded to hs-cTnI concentrations. The hs-cTnI (Siemens Vista) 0-/1-hour algorithm incorporated measurements performed at baseline and absolute changes within 1 hour, was derived in the first 750 patients (derivation cohort), and then validated in the second 750 (validation cohort). Results Overall, AMI was the final diagnosis in 16% of patients. Applying the hs-cTnI 0-/1-hour algorithm developed in the derivation cohort to the validation cohort, 57% of patients could be classified as “rule-out”; 10%, as “rule-in”; and 33%, as “observe.” In the validation cohort, the sensitivity and the negative predictive value for AMI in the “rule-out” zone were 100% (95% CI 96%-100%) and 100% (95% CI 99%-100%), respectively. The specificity and the positive predictive value (PPV) for AMI in the “rule-in” zone were 96% (95% CI 94%-97%) and 70% (95% CI 60%-79%), respectively. Negative predictive value and positive predictive value of the 0-/1-hour algorithm were higher compared to the standard of care combining hs-cTnI with the electrocardiogram (both P < .001). Conclusion The hs-cTnI 0-/1-hour algorithm performs very well for early rule-out as well as rule-in of AMI. The clinical implications are that used in conjunction with all other clinical information, the 0-/1-hour algorithm will be a safe and effective approach to substantially reduce time to diagnosis.
Obesity is a worldwide problem with different treatment options. Bariatric surgery is an effective treatment for severe obesity; however, it leads to drastic changes (e.g., changes in everyday life ...and eating behavior) for patients, which may lead to information needs. Our aim was to identify the information needs of patients undergoing bariatric surgery and to explore the information provision within the healthcare process of bariatric surgery in Germany.
We conducted a qualitative study (n = 14 single, semi-structured telephone interviews) between April 2018 and April 2019. The interview guide was designed prior to the interviews and consisted of four main sections (demographic information, pre-/postoperative healthcare provision, information needs). The interviews were transcribed verbatim and analyzed using qualitative content analysis with MAXQDA software.
There were unmet information needs with two factors (time: pre/postoperative and categories of information: general/specific) to be considered. Due to the patients' description of information, we categorized information into general (different surgical procedures, general nutritional information) and specific (occurring simultaneously with a problem) information. Most patients felt well informed concerning general information. However, it was pointed out that it was not possible to provide complete information preoperatively, as the need for information only arises when there are postoperative (specific) problems. In addition, there seems to be a high demand for specific postoperative information regarding nutrition and nutrition-related problems. However, patients stated that postoperative nutritional counseling is not reimbursed by health insurance funds. The information conveyed in support groups and the exchange of experiences are highly valued by patients. However, some patients describe the information provided within the support groups as unfiltered, frightening or exaggerated.
Overall, there were unmet information needs. Reimbursement by health insurance funds could increase the use of postoperative nutritional counseling and thus serve existing information needs. Support groups enable an exchange of experiences and therefore offer low-barrier access to information. Cooperation between support groups and healthcare professionals in information provision could be an approach to improving existing information needs or to avoiding the development of information gaps. Furthermore, the development and implementation of a digital solution for (postoperative) information dissemination could be helpful.
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