Left ventricular (LV) hypertrophy is characterized by cardiomyocyte hypertrophy and interstitial fibrosis ultimately leading to increased myocardial stiffness and reduced contractility. There is ...substantial evidence that the altered expression of matrix metalloproteinases (MMP) and Tenascin-C (TN-C) are associated with the progression of adverse LV remodeling. However, the role of TN-C in the development of LV hypertrophy because of chronic pressure overload as well as the regulatory role of TN-C on MMPs remains unknown.
In a knockout mouse model of TN-C, we investigated the effect of 10 weeks of pressure overload using transverse aortic constriction (TAC). Cardiac function was determined by magnetic resonance imaging. The expression of MMP-2 and MMP-9, CD147 as well as myocardial fibrosis were assessed by immunohistochemistry. The expression of TN-C was assessed by RT-qPCR and ELISA. TN-C knockout mice showed marked reduction in fibrosis (P < 0.001) and individual cardiomyocytes size (P < 0.01), in expression of MMP-2 (P < 0.05) and MMP-9 (P < 0.001) as well as preserved cardiac function (P < 0.01) in comparison with wild-type mice after 10 weeks of TAC. In addition, CD147 expression was markedly increased under pressure overload (P < 0.01), irrespectively of genotype. TN-C significantly increased the expression of the markers of hypertrophy such as ANP and BNP as well as MMP-2 in H9c2 cells (P < 0.05, respectively).
Our results are pointed toward a novel signaling mechanism that contributes to LV remodeling via MMPs upregulation, cardiomyocyte hypertrophy as well as myocardial fibrosis by TN-C under chronic pressure overload.
L-carnitine is an indispensable metabolite facilitating the transport of fatty acids into the mitochondrial matrix and has been previously postulated to exert a nutrigenomic effect. However, the ...underlying molecular mechanisms remain mostly unclear. We hypothesized that L-carnitine interacts with nuclear receptors involved in metabolic regulation, thereby modulating downstream targets of cellular metabolism. Therefore, we investigated the effect of L-carnitine supplementation on protein activity, mRNA expression, and binding affinities of nuclear receptors as well as mRNA expression of downstream targets in skeletal muscle cells, hepatocytes, and differentiated adipocytes. L-carnitine supplementation to hepatocytes increased the protein activity of multiple nuclear receptors (RAR, RXR, VDR, PPAR, HNF4, ER, LXR). Diverging effects on the mRNA expression of PPAR-α, PPAR-δ, PPAR-γ, RAR-β, LXR-α, and RXR-α were observed in adipocytes, hepatocytes, and skeletal muscle cells. mRNA levels of PPAR-α, a key regulator of lipolysis and β-oxidation, were significantly upregulated, emphasizing a role of L-carnitine as a promoter of lipid catabolism. L-carnitine administration to hepatocytes modulated the transcription of key nuclear receptor target genes, including ALDH1A1, a promoter of adipogenesis, and OGT, a contributor to insulin resistance. Electrophoretic mobility shift assays proved L-carnitine to increase binding affinities of nuclear receptors to their promoter target sequences, suggesting a molecular mechanism for the observed transcriptional modulation. Overall, these findings indicate that L-carnitine modulates the activity and expression of nuclear receptors, thereby promoting lipolytic gene expression and decreasing transcription of target genes linked to adipogenesis and insulin resistance.
Chemokines are important signaling molecules in the immune and nervous system. Using a fluorescence reporter mouse model, we demonstrate that the chemokine CCL17, a ligand of the chemokine receptor ...CCR4, is produced in the murine brain, particularly in a subset of hippocampal CA1 neurons. We found that basal expression of Ccl17 in hippocampal neurons was strongly enhanced by peripheral challenge with lipopolysaccharide (LPS). LPS‐mediated induction of Ccl17 in the hippocampus was dependent on local tumor necrosis factor (TNF) signaling, whereas upregulation of Ccl22 required granulocyte‐macrophage colony‐stimulating factor (GM‐CSF). CCL17 deficiency resulted in a diminished microglia density under homeostatic and inflammatory conditions. Further, microglia from naïve Ccl17‐deficient mice possessed a reduced cellular volume and a more polarized process tree as assessed by computer‐assisted imaging analysis. Regarding the overall branching, cell surface area, and total tree length, the morphology of microglia from naïve Ccl17‐deficient mice resembled that of microglia from wild‐type mice after LPS stimulation. In line, electrophysiological recordings indicated that CCL17 downmodulates basal synaptic transmission at CA3–CA1 Schaffer collaterals in acute slices from naïve but not LPS‐treated animals. Taken together, our data identify CCL17 as a homeostatic and inducible neuromodulatory chemokine affecting the presence and morphology of microglia and synaptic transmission in the hippocampus.
Main Points
CCL17 is constitutively expressed in pyramidal CA1 neurons and is further induced under inflammatory conditions.
CCL17‐deficiency causes alterations in microglia morphology and basal synaptic transmission at CA3‐CA1 Schaffer collaterals.
Bone healing involves a variety of different cell types and biological processes. Although certain key molecules have been identified, the molecular interactions of the healing progress are not ...completely understood. Moreover, a clinical routine for predicting the quality of bone healing after a fracture in an early phase is missing. This is mainly due to a lack of techniques to comprehensively screen for cytokines, growth factors and metabolites at their local site of action. Since all soluble molecules of interest are present in the fracture hematoma, its in-depth assessment could reveal potential markers for the monitoring of bone healing. Here, we describe an approach for sampling and quantification of cytokines and metabolites by using microdialysis, combined with solid phase extractions of proteins from wound fluids. By using a control group with an isolated soft tissue wound, we could reveal several bone defect-specific molecular features. In bone defect dialysates the neutrophil chemoattractants CXCL1, CXCL2 and CXCL3 were quantified with either a higher or earlier response compared to dialysate from soft tissue wound. Moreover, by analyzing downstream adaptions of the cells on protein level and focusing on early immune response, several proteins involved in the immune cell migration and activity could be identified to be specific for the bone defect group, e.g. immune modulators, proteases and their corresponding inhibitors. Additionally, the metabolite screening revealed different profiles between the bone defect group and the control group. In summary, we identified potential biomarkers to indicate imbalanced healing progress on all levels of analysis.
MotiQ is an open-source software for automated quantification of microglial motility and morphology. MotiQ can be applied to in vivo, ex vivo, and in vitro data from confocal, epifluorescence, and ...two-photon microscopy. MotiQ is not limited to microglia—it can also be applied to other cell types.
Microglia are the primary resident innate immune cells of the CNS. They possess branched, motile cell processes that are important for their cellular functions. To study the pathways that control microglial morphology and motility under physiological and disease conditions, it is necessary to quantify microglial morphology and motility precisely and reliably. Several image analysis approaches are available for the quantification of microglial morphology and motility. However, they are either not automated, not freely accessible, and/or limited in the number of morphology and motility parameters that can be assessed. Thus, we have developed MotiQ, an open-source, freely accessible software for automated quantification of microglial motility and morphology. MotiQ allows quantification of a diverse set of cellular motility and morphology parameters, including the parameters that have become the gold standard in the microglia field. We demonstrate that MotiQ can be applied to in vivo, ex vivo , and in vitro data from confocal, epifluorescence, or two-photon microscopy, and we compare its results to other analysis approaches. We suggest MotiQ as a versatile and customizable tool to study microglia.
Accurate determination of the size distribution of nanoparticle ensembles remains a challenge in nanotechnology‐related applications due to the limitations of established methods. Here, a ...microstructured fiber‐assisted nanoparticle tracking analysis (FaNTA) realization is introduced that breaks existing limitations through the recording of exceptionally long trajectories of rapidly diffusing polydisperse nanoparticles, resulting in excellent sizing precision and unprecedented separation capabilities of bimodal nanoparticle mixtures. An effective‐single‐mode antiresonant‐element fiber allows to efficiently confine nanoparticles in a light‐guiding microchannel and individually track them over more than 1000 frames, while aberration‐free imaging is experimentally confirmed by cross‐correlation analysis. Unique features of the approach are (i) the highly precise determination of the size distribution of monodisperse nanoparticle ensembles (only 7% coefficient of variation) and (ii) the accurate characterization of individual components in a bimodal mixture with very close mean diameters, both experimentally demonstrated for polymer nanospheres. The outstanding performance of the FaNTA realization can be quantified by introducing a new model for the bimodal separation index. Since FaNTA is applicable to all types of nano‐objects down to sub‐20 nm diameters, the method will improve the precision standard of mono‐ and polydisperse nanoparticle samples such as nano‐plastics or extracellular vesicles.
Accurate size characterization of nanoparticle ensembles in solution is realized by tracking their individual motion inside an optical fiber‐integrated microchannel. As a prerequisite, the fiber design provides effective single‐mode light guidance and high imaging quality. Bimodal mixtures of polystyrene nanospheres with mean hydrodynamic diameters as close as 110 and 132 nm are resolved.
Expert committees of the German medical associations provide a free and out-of-court evaluation of putative cases of medical malpractice. They prepare reports that contain valuable information on ...process steps that precede the actual treatment error. The aim of the present study was to identify and systematically categorize individual process steps in the expert reports and thus to lay the foundations for the understanding of malpractice evaluation processes.
In this study, ten randomly selected and anonymized expert reports of the Expert Committee for Questions of Medical Liability of the District Medical Association of South Württemberg with identified GP treatment errors were evaluated, using the method of qualitative content analysis. In an iterative process, central elements of expert reports were classified into a deductively and inductively built category system.
Six main categories with associated subcategories were identified: 1) structural aspects of the report, 2) doctor-patient communication, 3) medical course, 4) patient's experience, 5) action by the GP team, and 6) coordinative role in the health care system. The category system showed sufficient reliability with repeated use.
This study offers an opportunity to learn from errors. The proposed system allows to structure the complexity of expert reports on GP malpractice and may thus serve as a tool in various contexts. In particular, it facilitates the preparation and comparative analysis of reports in a structured way. It could also be used in health care research as well as in education and training.
Introduction
Salmonella spp. are a recognized and global cause of serious health issues from gastroenteritis to invasive disease. The mouse model of human typhoid fever, which uses Salmonella ...enterica serovar Typhimurium (STM) in susceptible mouse strains, has revealed that the bacteria gain access to extraintestinal tissues from the gastrointestinal tract to cause severe systemic disease. Previous analysis of the immune responses against Salmonella spp. revealed the crucial role played by dendritic cells (DCs) in carrying STM from the intestinal mucosa to the mesenteric lymph nodes (mLNs), a key site for antigen presentation and T cell activation. In this study, we investigated the influence of chemokine CCL17 on the dissemination of STM.
Methods
WT, CCL17/EGFP reporter, or CCL17‐deficient mice were infected orally with STM (SL1344) or mCherry‐expressing STM for 1–3 days. Colocalization of STM with CCL17‐expressing DCs in Peyer's patches (PP) and mLN was analyzed by fluorescence microscopy. In addition, DCs and myeloid cell populations from naïve and Salmonella‐infected mice were analyzed by flow cytometry. Bacterial load was determined in PP, mLN, spleen, and liver 1 and 3 days after infection.
Results
Histological analysis revealed that CCL17‐expressing cells are located in close proximity to STM in the dome area of PP. We show that, in mLN, STM were preferentially located within CCL17+ rather than CCL17− DCs, besides other mononuclear phagocytes, and identified the CD103+ CD11b− DC subset as the main STM‐carrying DC population in the intestine. STM infection triggered upregulation of CCL17 expression in specific intestinal DC subsets in a tissue‐specific manner. The dissemination of STM from the gut to the mLN, however, was only moderately influenced by the presence of CCL17.
Conclusion
CCL17‐expressing DCs were preferentially infected by Salmonella in the intestine in comparison to other DC. Nevertheless, the production of CCL17 was not essential for the early dissemination of Salmonella from the gut to systemic organs.
Oral infection with Salmonella enhances the frequency of dendritic cells expressing the pro‐inflammatory chemokine CCL17, promoting dendritic cell–T cell interaction. In Peyer's patches, CCL17‐expressing cells colocalize with Salmonella and Salmonella‐containing cells in mesenteric lymph nodes are preferentially CCL17‐positive. While CCL17‐expressing cells participate in the early dissemination of Salmonella, CCL17 itself appears to play a redundant role in this process.
The chemokine CCL17, mainly produced by dendritic cells (DCs) in the immune system, is involved in the pathogenesis of various inflammatory diseases. As a ligand of CCR4, CCL17 induces chemotaxis and ...facilitates T cell-DC interactions. We report the identification of two novel RNA aptamers, which were validated in vitro and in vivo for their capability to neutralize CCL17. Both aptamers efficiently inhibited the directed migration of the CCR4+ lymphoma line BW5147.3 toward CCL17 in a dose-dependent manner. To study the effect of these aptamers in vivo, we used a murine model of contact hypersensitivity. Systemic application of the aptamers significantly prevented ear swelling and T cell infiltration into the ears of sensitized mice after challenge with the contact sensitizer. The results of this proof-of-principle study establish aptamers as potent inhibitors of CCL17-mediated chemotaxis. Potentially, CCL17-specific aptamers may be used therapeutically in humans to treat or prevent allergic and inflammatory diseases.
Display omitted
The chemokine CCL17 promotes T cell attraction to peripheral tissues and thereby exacerbates inflammatory reactions. Using SELEX, Fülle et al. selected high-affinity aptamers, which neutralize CCL17 in murine contact hypersensitivity. Because of their small size and structural properties, such aptamers may represent new versatile drugs for allergic skin diseases.
Atezolizumab/bevacizumab (atezo/bev) and lenvatinib have demonstrated efficacy as first-line therapies for hepatocellular carcinoma (HCC). However, vascular endothelial growth factor (VEGF) ...inhibition with these therapies may be associated with the risk of bleeding and thromboembolic events. In this study, we evaluated the efficacy and safety with focus on the bleeding and thromboembolic events of atezo/bev vs. lenvatinib in a large, multicenter real-world population.
This study is based on HCC cohorts from seven centers in Germany and Austria. Incidences of bleeding or thromboembolic events and efficacy outcomes were assessed and compared.
In total, 464 patients treated with atezo/bev (n = 325) or lenvatinib (n = 139) were analyzed. Both groups were balanced with respect to demographics, presence of liver cirrhosis, and variceal status. Duration of therapy did not differ between groups. Within 3 months of therapy, bleeding episodes were described in 57 (18%) patients receiving atezo/bev compared with 15 (11%) patients receiving lenvatinib (p = 0.07). Variceal hemorrhage occurred in 11 (3%) patients treated with atezo/bev compared with 4 (3%) patients treated with lenvatinib (p = 0.99). Thromboembolic events were reported in 19 (6%) of patients in the atezo/bev cohort compared with 5 (4%) patients in the lenvatinib cohort (p = 0.37). In addition, incidence of overall bleeding, variceal hemorrhage, and thromboembolic events did not differ significantly in patients who received either atezo/bev or lenvantinib for 6 months.
Safety considerations related to bleeding and thromboembolic events may not be helpful in guiding clinical decision-making when choosing between atezo/bev and lenvatinib.
The inhibition of VEGF by current first-line therapies for HCC, such as atezolizumab/bevacizumab or lenvatinib, may be associated with the risk of bleeding and thromboembolic events. Studies comparing the incidence of these side effects between atezolizumab/bevacizumab and lenvatinib, which are preferred treatments over sorafenib for HCC, are needed. Differences in this side effect profile may influence the choice of first-line therapy by treating physicians. Because no significant differences were observed regarding bleeding or thromboembolic events between both therapies in the present study, we conclude that safety considerations related to these events may not be helpful in guiding clinical decision-making when choosing between atezolizumab/bevacizumab and lenvatinib.
Display omitted
•Evaluation of the efficacy and safety of atezolizumab/bevacizumab and lenvatinib in a large real-world population of 464 patients with HCC.•Compared with lenvatinib, atezolizumab/bevacizumab improves ORR.•No significant differences in bleeding or thromboembolic events were observed.•In atezolizumab/bevacizumab-treated patients, spleen size and history of variceal bleeding were associated with GI-bleeding risk, while anticoagulation was associated with non-GI bleeding risk.
PDF
