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- Remote Sensing (7)
- SMN (7)
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- algorithm (7)
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- antigen (7)
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- cGMP (7)
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- immunofluorescence (7)
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- immunoprecipitation (7)
- latency (7)
- locomotion (7)
- luciferase (7)
- macrophage (7)
- mapping (7)
- mechanotransduction (7)
- meta-analysis (7)
- methylation (7)
- microenvironment (7)
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The Josephson effect describes the generic appearance of a supercurrent in a weak link between two superconductors. Its exact physical nature deeply influences the properties of the supercurrent. In recent years, considerable efforts have focused on the coupling of superconductors to the surface states of a three-dimensional topological insulator. In such a material, an unconventional induced p-wave superconductivity should occur, with a doublet of topologically protected gapless Andreev bound states, whose energies vary 4π-periodically with the superconducting phase difference across the junction. In this article, we report the observation of an anomalous response to rf irradiation in a Josephson junction made of a HgTe weak link. The response is understood as due to a 4π-periodic contribution to the supercurrent, and its amplitude is compatible with the expected contribution of a gapless Andreev doublet. Our work opens the way to more elaborate experiments to investigate the induced superconductivity in a three-dimensional insulator.
lt is known that 5-azacytidine (5-AC) induces tumors in several organs of rats and mice. The mechanisms of these effects are still poorly understood although it is known that 5-AC can be incorporated into DNA. Furthermore, it can inhibit DNA methylation. The known data on its clastogenic andjor gene mutation-inducing potential are still controversial. Therefore, we have investigated the kinds of genotoxic effects caused by 5-AC in Syrian hamster embryo (SHE) fibroblasts. Three different endp6ints (micronucleus formation, unscheduled DNA synthesis (UDS) and cell transforrnation) were assayed under similar conditions of metabolism and dose at target in this cell system. 5-AC induces morphological transformation of SHE cells, but not UDS. Therefore, 5-AC does not seem to cause repairable DNA lesions. Furthermore, our studies revealed that 5-AC is a potent inducer of mkronuclei in the SHE system. Immunocytochemical analysis revealed that a certain percentage of these contain kinetochores indicating that 5-AC may induce both clastogenic events and numerical chromosome changes.
The purpose of this study was to evaluate whether spatial hippocampus-dependent learning is affected by the serotonergic system and stress. Therefore, 5-HTT knockout (-/-), heterozygous (+/-) and wildtype (+/+) mice were subjected to the Barnes maze (BM) and the Morris water maze (WM), the latter being discussed as more aversive. Additionally, immediate early gene (IEG) expression, hippocampal adult neurogenesis (aN), and blood plasma corticosterone were analyzed.
While the performance of 5-HTT-/- mice in the BM was undistinguishable from both other genotypes, they performed worse in the WM. However, in the course of the repeated WM trials 5-HTT-/- mice advanced to wildtype level. The experience of a single trial of either the WM or the BM resulted in increased plasma corticosterone levels in all genotypes. After several trials 5-HTT-/- mice exhibited higher corticosterone concentrations compared with both other genotypes in both tests. Corticosterone levels were highest in 5-HTT-/- mice tested in the WM indicating greater aversiveness of the WM and a greater stress sensitivity of 5-HTT deficient mice.
Quantitative immunohistochemistry in the hippocampus revealed increased cell counts positive for the IEG products cFos and Arc as well as for proliferation marker Ki67 and immature neuron marker NeuroD in 5-HTT-/- mice compared to 5-HTT+/+ mice, irrespective of the test. Most differences were found in the suprapyramidal blade of the dentate gyrus of the septal hippocampus. Ki67-immunohistochemistry revealed a genotype x environment interaction with 5-HTT genotype differences in naïve controls and WM experience exclusively yielding more Ki67-positive cells in 5-HTT+/+ mice. Moreover, in 5-HTT-/- mice we demonstrate that learning performance correlates with the extent of aN.
Overall, higher baseline IEG expression and increased an in the hippocampus of 5-HTT-/- mice together with increased stress sensitivity may constitute the neurobiological correlate of raised alertness, possibly impeding optimal learning performance in the more stressful WM.
Anxiety disorders and depression are common comorbidities in cardiac patients. Mice lacking the serotonin transporter (5-HTT) exhibit increased anxiety-like behavior. However, the role of 5-HTT deficiency on cardiac aging, and on healing and remodeling processes after myocardial infarction (MI), remains unclear. Cardiological evaluation of experimentally naïve male mice revealed a mild cardiac dysfunction in ≥4-month-old 5-HTT knockout (−/−) animals. Following induction of chronic cardiac dysfunction (CCD) by MI vs. sham operation 5-HTT−/− mice with infarct sizes >30% experienced 100% mortality, while 50% of 5-HTT+/− and 37% of 5-HTT+/+ animals with large MI survived the 8-week observation period. Surviving (sham and MI < 30%) 5-HTT−/− mutants displayed reduced exploratory activity and increased anxiety-like behavior in different approach-avoidance tasks. However, CCD failed to provoke a depressive-like behavioral response in either 5-Htt genotype. Mechanistic analyses were performed on mice 3 days post-MI. Electrocardiography, histology and FACS of inflammatory cells revealed no abnormalities. However, gene expression of inflammation-related cytokines (TGF-β, TNF-α, IL-6) and MMP-2, a protein involved in the breakdown of extracellular matrix, was significantly increased in 5-HTT−/− mice after MI. This study shows that 5-HTT deficiency leads to age-dependent cardiac dysfunction and disrupted early healing after MI probably due to alterations of inflammatory processes in mice.
Selective serotonin reuptake inhibitors are among the most prescribed antidepressants. Fluoxetine is the lead molecule which exerts its therapeutic effects, at least in part, by promoting neuroplasticity through increased brain-derived neurotrophic factor (BDNF)/tropomyosin-related receptor kinase B (TrkB) signalling. It is unclear however, to which extent the neuroplastic effects of fluoxetine are solely mediated by the inhibition of the serotonin transporter (5-HTT). To answer this question, the effects of fluoxetine on neuroplasticity were analysed in both wild type (WT) and 5-Htt knock-out (KO) mice. Using Western blotting and RT-qPCR approaches, we showed that fluoxetine 10 µM activated BDNF/TrkB signalling pathways in both CD1 and C57BL/6J mouse primary cortical neurons. Interestingly, effects on BDNF signalling were observed in primary cortical neurons from both 5-Htt WT and KO mice. In addition, a 3-week in vivo fluoxetine treatment (15 mg/kg/d; i.p.) increased the expression of plasticity genes in brains of both 5-Htt WT and KO mice, and tended to equally enhance hippocampal cell proliferation in both genotypes, without reaching significance. Our results further suggest that fluoxetine-induced neuroplasticity does not solely depend on 5-HTT blockade, but might rely, at least in part, on 5-HTT-independent direct activation of TrkB.
Background: The role of serotonin (5-hydroxytrptamine, 5-HT) in the modulation of pain has been widely studied. Previous work led to the hypothesis that 5-hydroxyindolacetic acid (5-HIAA), a main metabolite of serotonin, might by itself influence pain thresholds. Results: In the present study, we investigated the role of 5-HIAA in inflammatory pain induced by intraplantar injection of complete Freund’s adjuvant (CFA) into the hind paw of mice. Wild-type mice were compared to mice deficient of the 5-HT transporter (5-HTT-/- mice) using behavioral tests for hyperalgesia and high-performance liquid chromatography (HPLC) to determine tissue levels of 5-HIAA. Wild-type mice reproducibly developed thermal hyperalgesia and paw edema for 5 days after CFA injection. 5-HTT-/- mice treated with CFA had reduced thermal hyperalgesia on day 1 after CFA injection and normal responses to heat hereafter. The 5-HIAA levels in spinal cord and sciatic nerve as measured with HPLC were lower in 5-HTT-/- mice than in wild-type mice after CFA injection. Pretreatment of wild-type mice with intraperitoneal injection of para-chlorophenylalanine (p-CPA), a serotonin synthesis inhibitor, resulted in depletion of the 5-HIAA content in spinal cord and sciatic nerve and decrease in thermal hyperalgesia in CFA injected mice. The application of exogenous 5-HIAA resulted in potentiation of thermal hyperalgesia induced by CFA in 5-HTT-/- mice and in wild-type mice pretreated with p- CPA, but not in wild-type mice without p-CPA pretreatment. Further, methysergide, a broad-spectrum serotonin receptor antagonist, had no effect on 5-HIAA-induced potentiation of thermal hyperalgesia in CFA-treated wildtype mice. Conclusion: Taken together, the present results suggest that 5-HIAA plays an important role in modulating peripheral thermal hyperalgesia in CFA induced inflammation, probably via a non-serotonin receptor mechanism.
Early healing after myocardial infarction (MI) is characterized by a strong inflammatory reaction. Most leukotrienes are pro-inflammatory and are therefore potential mediators of healing and remodeling after myocardial ischemia. The enzyme 5-lipoxygenase (5-LOX) has a key role in the transformation of arachidonic acid in leukotrienes. Thus, we tested the effect of 5-LOX on healing after MI. After chronic coronary artery ligation, early mortality was significantly increased in 5-LOX\(^{−/−}\) when compared to matching wildtype (WT) mice due to left ventricular rupture. This effect could be reproduced in mice treated with the 5-LOX inhibitor Zileuton. A perfusion mismatch due to the vasoactive potential of leukotrienes is not responsible for left ventricular rupture since local blood flow assessed by magnetic resonance perfusion measurements was not different. However, after MI, there was an accentuation of the inflammatory reaction with an increase of pro-inflammatory macrophages. Yet, mortality was not changed in chimeric mice (WT vs. 5-LOX\(^{−/−}\) bone marrow in 5-LOX\(^{−/−}\) animals), indicating that an altered function of 5-LOX\(^{−/−}\) inflammatory cells is not responsible for the phenotype. Collagen production and accumulation of fibroblasts were significantly reduced in 5-LOX\(^{−/−}\) mice in vivo after MI. This might be due to an impaired migration of 5-LOX\(^{−/−}\) fibroblasts, as shown in vitro to serum. In conclusion, a lack or inhibition of 5-LOX increases mortality after MI because of healing defects. This is not mediated by a change in local blood flow, but through an altered inflammation and/or fibroblast function.
Two 5-methylcytosine (5-MeC)-rich heterochromatic regions were demonstrated in metaphase chromosomes of the Indian muntjac by indirect immunofluorescence using a monoclonal anti-5-MeC antibody. The metaphases were obtained from diploid and triploid cell lines. A major region is located in the ‘neck' of the 3;X fusion chromosome and can be detected after denaturation of the chromosomal DNA with UV-light irradiation for 1 h. It is located exactly at the border of the X chromosome and the translocated autosome 3. A minor region is found in the centromeric region of the free autosome 3 after denaturing the chromosomal DNA for 3 h or longer. The structure and possible function of the major hypermethylated region as barrier against spreading of the X-inactivation process into the autosome 3 is discussed.
Background: Ga-[1,4,7,10-tetraazacyclododecane-N,N0,N00,N000-tetraacetic acid]-d-Phe1,Tyr3-octreotate (DOTATATE) positron emission tomography (PET) is commonly used for the visualization of somatostatin receptor (SSTR)-positive neuroendocrine tumors. SSTR is also known to be expressed on macrophages, which play a major role in inflammatory processes in the walls of coronary arteries and large vessels. Therefore, imaging SSTR expression has the potential to visualize vulnerable plaques. We assessed 68Ga-DOTATATE accumulation in large vessels in comparison to 18F-2-fluorodeoxyglucose (FDG) uptake, calcified plaques (CPs), and cardiovascular risk factors. Methods: Sixteen consecutive patients with neuroendocrine tumors or thyroid cancer underwent both 68Ga-DOTATATE and 18F-FDG PET/CT for staging or restaging purposes. Detailed clinical data, including common cardiovascular risk factors, were recorded. For a separate assessment, they were divided into a high-risk and a low-risk group. In each patient, we calculated the maximum target-to-background ratio (TBR) of eight arterial segments. The correlation of the TBRmean of both tracers with risk factors including plaque burden was assessed. Results: The mean TBR of 68Ga-DOTATATE in all large arteries correlated significantly with the presence of CPs (r = 0.52; p < 0.05), hypertension (r = 0.60; p < 0.05), age (r = 0.56; p < 0.05), and uptake of 18F-FDG (r = 0.64; p < 0.01). There was one significant correlation between 18F-FDG uptake and hypertension (0.58; p < 0.05). Out of the 37 sites with the highest focal 68Ga-DOTATATE uptake, 16 (43.2%) also had focal 18F-FDG uptake. Of 39 sites with the highest 18F-FDG uptake, only 11 (28.2%) had a colocalized 68Ga-DOTATATE accumulation. Conclusions: In this series of cancer patients, we found a stronger association of increased 68Ga-DOTATATE uptake with known risk factors of cardiovascular disease as compared to 18F-FDG, suggesting a potential role for plaque imaging in large arteries. Strikingly, we found that focal uptake of 68Ga-DOTATATE and 18F-FDG does not colocalize in a significant number of lesions.
The SEED project is developing ePortfolios for teaching at the Julius-Maximilians-Universität Würzburg. ePortfolios make it possible to further develop teaching and learning in a competence-oriented manner and to strengthen the individual reflection aspect of academic studies to support training professional skills. A total of seven use cases were developed. They provide examples of how ePortfolios can be used in university teaching. Four exam variants help to illustrate both subject-specific and reflective components when examining using ePortfolios.
Vitamin B6 deficiency has been linked to cognitive impairment in human brain disorders for decades. Still, the molecular mechanisms linking vitamin B6 to these pathologies remain poorly understood, and whether vitamin B6 supplementation improves cognition is unclear as well. Pyridoxal phosphatase (PDXP), an enzyme that controls levels of pyridoxal 5’-phosphate (PLP), the co-enzymatically active form of vitamin B6, may represent an alternative therapeutic entry point into vitamin B6-associated pathologies. However, pharmacological PDXP inhibitors to test this concept are lacking. We now identify a PDXP and age-dependent decline of PLP levels in the murine hippocampus that provides a rationale for the development of PDXP inhibitors. Using a combination of small molecule screening, protein crystallography and biolayer interferometry, we discover and analyze 7,8-dihydroxyflavone (7,8-DHF) as a direct and potent PDXP inhibitor. 7,8-DHF binds and reversibly inhibits PDXP with low micromolar affinity and sub-micromolar potency. In mouse hippocampal neurons, 7,8-DHF increases PLP in a PDXP-dependent manner. These findings validate PDXP as a druggable target. Of note, 7,8-DHF is a well-studied molecule in brain disorder models, although its mechanism of action is actively debated. Our discovery of 7,8-DHF as a PDXP inhibitor offers novel mechanistic insights into the controversy surrounding 7,8-DHF-mediated effects in the brain.
Alzheimer's disease (AD) is a multifactorial disease and the most common form of dementia. There are no treatments to cure, prevent or slow down the progression of the disease. Natural products hold considerable interest for the development of preventive neuroprotectants to treat neurodegenerative disorders like AD, due to their low toxicity and general beneficial effects on human health with their anti-inflammatory and antioxidant features. In this work we describe regioselective synthesis of 7-O-ester hybrids of the flavonoid taxifolin with the phenolic acids cinnamic and ferulic acid, namely 7-O-cinnamoyltaxifolin and 7-O-feruloyltaxifolin. The compounds show pronounced overadditive neuroprotective effects against oxytosis, ferroptosis and ATP depletion in the murine hippocampal neuron HT22 cell model. Furthermore, 7-O-cinnamoyltaxifolin and 7-O-feruloyltaxifolin reduced LPS-induced neuroinflammation in BV-2 microglia cells as assessed by effects on the levels of NO, IL6 and TNFα. In all in vitro assays the 7-O-esters of taxifolin and ferulic or cinnamic acid showed strong overadditive activity, significantly exceeding the effects of the individual components and the equimolar mixtures thereof, which were almost inactive in all of the assays at the tested concentrations. In vivo studies confirmed this overadditive effect. Treatment of an AD mouse model based on the injection of oligomerized Aβ\(_{25-35}\) peptide into the brain to cause neurotoxicity and subsequently memory deficits with 7-O-cinnamoyltaxifolin or 7-O-feruloyltaxifolin resulted in improved performance in an assay for short-term memory as compared to vehicle and mice treated with the respective equimolar mixtures. These results highlight the benefits of natural product hybrids as a novel compound class with potential use for drug discovery in neurodegenerative diseases due to their pharmacological profile that is distinct from the individual natural components.
The properties of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) as an antagonist ligand for A\(_1\) adenosirre receptors were examined and conipared with other radioligands for this receptor. DPCPX competitively antagonized both the inhibition of adenylate cyclase activity via A\(_1\) adenosirre receptors and the stimulationvia A\(_2\) adenosirre receptors. The K\(_i\)-values of this antagonism were 0.45 nM at the A\(_1\) receptor of rat fat cells, and 330 nM at the A\(_2\) receptor of human platelets, giving a more than 700-fold A\(_1\)-selectivity. A similar A\(_1\)-selectivity was determined in radioligand binding studies. Even at high concentrations, DPCPX did not significantly inhibit the soluble cAMPphosphodiesterase activity of human platelets. [\(^3\)H]DPCPX (105 Ci/mmol) bound in a saturable manner with high affinity to A\(_1\) receptors in membranes of bovine brain and heart, and rat brain and fat cells (K\(_D\) -values 50-190 pM). Its nonspecific binding was about 1% of total at K\(_D\) , except in bovine myocardial membranes (about 10%). Binding studies with bovine myocardial membranes allowed the analysis of both the high and low agonist affinity states of this receptor in a tissue with low receptor density. The binding properties of [\(^3\)H]DPCPX appear superior to those of other agonist and antagonist radioligands for the A\(_1\) receptor.
High programmed cell death 1 ligand 1 (PD-L1) protein expression and copy number alterations (CNAs) of the corresponding genomic locus 9p24.1 in Hodgkin- and Reed–Sternberg cells (HRSC) have been shown to be associated with favourable response to anti-PD-1 checkpoint inhibition in relapsed/refractory (r/r) classical Hodgkin lymphoma (cHL). In the present study, we investigated baseline 9p24.1 status as well as PD-L1 and major histocompatibility complex (MHC) class I and II protein expression in 82 biopsies from patients with early stage unfavourable cHL treated with anti-PD-1-based first-line treatment in the German Hodgkin Study Group (GHSG) NIVAHL trial (ClinicalTrials.gov Identifier: NCT03004833). All evaluated specimens showed 9p24.1 CNA in HRSC to some extent, but with high intratumoral heterogeneity and an overall smaller range of alterations than reported in advanced-stage or r/r cHL. All but two cases (97%) showed PD-L1 expression by the tumour cells in variable amounts. While MHC-I was rarely expressed in >50% of HRSC, MHC-II expression in >50% of HRSC was found more frequently. No obvious impact of 9p24.1 CNA or PD-L1 and MHC-I/II expression on early response to the highly effective anti-PD-1-based NIVAHL first-line treatment was observed. Further studies evaluating an expanded panel of potential biomarkers are needed to optimally stratify anti-PD-1 first-line cHL treatment.
France, Italy, and Spain are three Romance-speaking countries which – at least in Europe – have been affected to a very high degree by the consequences of the Corona pandemic. This paper examines discursive strategies on social media (Twitter and Facebook) by the three heads of government/state of the aforementioned countries – namely Emmanuel Macron (France), Giuseppe Conte (Italy), and Pedro Sánchez (Spain)- from a corpuslinguistic point of view. For this purpose, a corpus was created which contains all Twitter and Facebook messages posted by these heads of government/state from the beginning of February until the end of April 2020. By applying corpus-linguistic methods we find that all three politicians consciously use social media to sensitize, inform, and – in view of a dramatic pandemic situation – unite their respective populations behind them.
Stem cell therapy holds great promise for tissue regeneration and cancer treatment, although its efficacy is still inconclusive and requires further understanding and optimization of the procedures. Non-invasive cell tracking can provide an important opportunity to monitor in vivo cell distribution in living subjects. Here, using a combination of positron emission tomography (PET) and in vitro 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) direct cell labelling, the feasibility of engrafted stem cell monitoring was tested in multiple animal species. Human mesenchymal stem cells (MSCs) were incubated with phosphate-buffered saline containing [18F]FDG for in vitro cell radiolabelling. The pre-labelled MSCs were administrated via peripheral vein in a mouse (n=1), rats (n=4), rabbits (n=4) and non-human primates (n=3), via carotid artery in rats (n=4) and non-human primates (n=3), and via intra-myocardial injection in rats (n=5). PET imaging was started 10 min after cell administration using a dedicated small animal PET system for a mouse and rats. A clinical PET system was used for the imaging of rabbits and non-human primates. After MSC administration via peripheral vein, PET imaging revealed intense radiotracer signal from the lung in all tested animal species including mouse, rat, rabbit, and non-human primate, suggesting administrated MSCs were trapped in the lung tissue. Furthermore, the distribution of the PET signal significantly differed based on the route of cell administration. Administration via carotid artery showed the highest activity in the head, and intra-myocardial injection increased signal from the heart. In vitro [18F]FDG MSC pre-labelling for PET imaging is feasible and allows non-invasive visualization of initial cell distribution after different routes of cell administration in multiple animal models. Those results highlight the potential use of that imaging approach for the understanding and optimization of stem cell therapy in translational research.
We have recently demonstrated CXCR4 overexpression in vestibular schwannomas (VS). This study investigated the feasibility of CXCR4-directed positron emission tomography/computed tomography (PET/CT) imaging of VS using the radiolabeled chemokine ligand [\(^{68}\)Ga]Pentixafor.
Methods: 4 patients with 6 primarily diagnosed or pre-treated/observed VS were enrolled. All subjects underwent [\(^{68}\)Ga]Pentixafor PET/CT prior to surgical resection. Images were analyzed visually and semi-quantitatively for CXCR4 expression including calculation of tumor-to-background ratios (TBR). Immunohistochemistry served as standard of reference in three patients.
Results: [\(^{68}\)Ga]Pentixafor PET/CT was visually positive in all cases. SUV\(_{mean}\) and SUV\(_{max}\) were 3.0 ± 0.3 and 3.8 ± 0.4 and TBR\(_{mean}\) and TBR\(_{max}\) were 4.0 ± 1.4 and 5.0 ± 1.7, respectively. Histological analysis confirmed CXCR4 expression in tumors.
Conclusion: Non-invasive imaging of CXCR4 expression using [\(^{68}\)Ga]Pentixafor PET/CT of VS is feasible and could prove useful for in vivo assessment of CXCR4 expression.
Chemokine (C-X-C motif) receptor 4 (CXCR4) is a key factor for tumor growth and metastasis in several types of human cancer including multiple myeloma (MM). Proof-of-concept of CXCR4-directed radionuclide therapy in MM has recently been reported. This study assessed the diagnostic performance of the CXCR4-directed radiotracer [\(^{68}\)Ga]Pentixafor in MM and a potential role for stratifying patients to CXCR4-directed therapies.
Thirty-five patients with MM underwent [\(^{68}\)Ga]Pentixafor-PET/CT for evaluation of eligibility for endoradiotherapy. In 19/35 cases, [\(^{18}\)F]FDG-PET/CT for correlation was available. Scans were compared on a patient and on a lesion basis. Tracer uptake was correlated with standard clinical parameters of disease activity.
[\(^{68}\)Ga]Pentixafor-PET detected CXCR4-positive disease in 23/35 subjects (66%). CXCR4-positivity at PET was independent from myeloma subtypes, cytogenetics or any serological parameters and turned out as a negative prognostic factor. In the 19 patients in whom a comparison to [\(^{18}\)F]FDG was available, [\(^{68}\)Ga]Pentixafor-PET detected more lesions in 4/19 (21%) subjects, [\(^{18}\)F]FDG proved superior in 7/19 (37%). In the remaining 8/19 (42%) patients, both tracers detected an equal number of lesions. [\(^{18}\)F]FDG-PET positivity correlated with [\(^{68}\)Ga]Pentixafor-PET positivity (p=0.018).
[\(^{68}\)Ga]Pentixafor-PET provides further evidence that CXCR4 expression frequently occurs in advanced multiple myeloma, representing a negative prognostic factor and a potential target for myeloma specific treatment. However, selecting patients for CXCR4 directed therapies and prognostic stratification seem to be more relevant clinical applications for this novel imaging modality, rather than diagnostic imaging of myeloma.
We present herein an in‐depth study of complexes in which a molecule containing a boron‐boron triple bond is bound to tellurate cations. The analysis allows the description of these salts as true π complexes between the B−B triple bond and the tellurium center. These complexes thus extend the well‐known Dewar‐Chatt‐Duncanson model of bonding to compounds made up solely of p block elements. Structural, spectroscopic and computational evidence is offered to argue that a set of recently reported heterocycles consisting of phenyltellurium cations complexed to diborynes bear all the hallmarks of \(\pi\)‐complexes in the \(\pi\)‐complex/metallacycle continuum envisioned by Joseph Chatt. Described as such, these compounds are unique in representing the extreme of a metal‐free continuum with conventional unsaturated three‐membered rings (cyclopropenes, azirenes, borirenes) occupying the opposite end.
\(^{11}\)C-methionine-PET in multiple myeloma: a combined study from two different institutions
(2017)
\(^{11}\)C-methionine (MET) has recently emerged as an accurate marker of tumor burden and disease activity in patients with multiple myeloma (MM). This dual-center study aimed at further corroboration of the superiority of MET as positron emission tomography (PET) tracer for staging and re-staging MM, as compared to \(^{18}\)F-2`-deoxy-2`-fluoro-D-glucose (FDG).
78 patients with a history of solitary plasmacytoma (n=4), smoldering MM (SMM, n=5), and symptomatic MM (n=69) underwent both MET- and FDG-PET/computed tomography (CT) at the University Centers of Würzburg, Germany and Navarra, Spain. Scans were compared on a patient and on a lesion basis. Inter-reader agreement was also evaluated. In 2 patients, tumor biopsies for verification of discordant imaging results were available.
MET-PET detected focal lesions (FL) in 59/78 subjects (75.6%), whereas FDG-PET/CT showed lesions in only 47 patients (60.3%; p<0.01), accordingly disease activity would have been missed in 12 patients. Directed biopsies of discordant results confirmed MET-PET/CT results in both cases.
MET depicted more FL in 44 patients (56.4%; p<0.01), whereas in two patients (2/78), FDG proved superior. In the remainder (41.0%, 32/78), both tracers yielded comparable results. Inter-reader agreement for MET was higher than for FDG (κ = 0.82 vs κ = 0.72).
This study demonstrates higher sensitivity of MET in comparison to standard FDG to detect intra- and extramedullary MM including histologic evidence of FDG-negative, viable disease exclusively detectable by MET-PET/CT. MET holds the potential to replace FDG as functional imaging standard for staging and re-staging of MM.
Multiple myeloma (MM) remains an essentially incurable hematologic malignancy. However, new treatment modalities and novel drugs have been introduced and thus additional tools for therapy monitoring are increasingly needed. Therefore, we evaluated the radiotracers \(^{11}\)C-Methionine (paraprotein-biosynthesis) and \(^{18}\)F-FDG (glucose-utilization) for monitoring response to anti-myeloma-therapy and outcome prediction. Influence of proteasome-inhibition on radiotracer-uptake of different MM cell-lines and patient-derived CD138\(^{+}\) plasma cells was analyzed and related to tumor-biology. Mice xenotransplanted with MM. 1S tumors underwent MET- and FDG-\(\mu\)PET. Tumor-to-background ratios before and after 24 h, 8 and 15 days treatment with bortezomib were correlated to survival. Treatment reduced both MET and FDG uptake; changes in tracer-retention correlated with a switch from high to low CD138-expression. In xenotransplanted mice, MET-uptake significantly decreased by 30-79% as early as 24 h after bortezomib injection. No significant differences were detected thus early with FDG. This finding was confirmed in patient-derived MM cells. Importantly, early reduction of MET-but not FDG-uptake correlated with improved survival and reduced tumor burden in mice. Our results suggest that MET is superior to FDG in very early assessment of response to anti-myeloma-therapy. Early changes in MET-uptake have predictive potential regarding response and survival. MET-PET holds promise to individualize therapies in MM in future.
Background
\(^{177}\)Lu is used in peptide receptor radionuclide therapies for the treatment of neuroendocrine tumors. Based on the recent literature, SST2 antagonists are superior to agonists in tumor uptake. The compound OPS201 is the novel somatostatin antagonist showing the highest SST2 affinity. The aim of this study was to measure the in vivo biodistribution and dosimetry of \(^{177}\)Lu-OPS201 in five anesthetized Danish Landrace pigs as an appropriate substitute for humans to quantitatively assess the absorbed doses for future clinical applications.
Results
\(^{177}\)Lu-OPS201 was obtained with a specific activity ranging from 10 to 17 MBq/μg. Prior to administration, the radiochemical purity was measured as s > 99.7 % in all cases. After injection, fast clearance of the compound from the blood stream was observed. Less than 5 % of the injected activity was presented in blood 10 min after injection. A series of SPECT/CT and whole-body scans conducted until 10 days after intravenous injection showed uptake mostly in the liver, spine, and kidneys. There was no visible uptake in the spleen. Blood samples were taken to determine the time-activity curve in the blood. Time-activity curves and time-integrated activity coefficients were calculated for the organs showing visible uptake. Based on these data, the absorbed organ dose coefficients for a 70-kg patient were calculated with OLINDA/EXM. For humans after an injection of 5 GBq \(^{177}\)Lu-OPS201, the highest predicted absorbed doses are obtained for the kidneys (13.7 Gy), the osteogenic cells (3.9 Gy), the urinary bladder wall (1.8 Gy), and the liver (1.0 Gy). No metabolites of 177Lu-OPS201 were found by radio HPLC analysis. None of the absorbed doses calculated will exceed organ toxicity levels.
Conclusions
The \(^{177}\)Lu-OPS201 was well tolerated and caused no abnormal physiological or behavioral signs. In vivo distributions and absorbed doses of pigs are comparable to those observed in other publications. According to the biodistribution data in pigs, presented in this work, the expected radiation exposure in humans will be within the acceptable range.
\(^{11}\)C-methionine (\(^{11}\)C-MET) is a new positron emission tomography (PET) tracer for the assessment of disease activity in multiple myeloma (MM) patients, with preliminary data suggesting higher sensitivity and specificity than \(^{18}\)F-fluorodeoxyglucose (\(^{18}\)F-FDG). However, the value of tumor burden biomarkers has yet to be investigated. Our goals were to corroborate the superiority of \(^{11}\)C-MET for MM staging and to compare its suitability for the assessment of metabolic tumor burden biomarkers in comparison to \(^{18}\)F-FDG. Twenty-two patients with newly diagnosed, treatment-naïve symptomatic MM who had undergone \(^{11}\)C-MET and \(^{18}\)F-FDG PET/CT were evaluated. Standardized uptake values (SUV) were determined and compared with total metabolic tumor volume (TMTV) for both tracers: total lesion glycolysis (TLG) and total lesion \(^{11}\)C-MET uptake (TLMU). PET-derived values were compared to Revised International Staging System (R-ISS), cytogenetic, and serologic MM markers such as M component, beta 2 microglobulin (B2M), serum free light chains (FLC), albumin, and lactate dehydrogenase (LDH). In 11 patients (50%), \(^{11}\)C-MET detected more focal lesions (FL) than FDG (p < 0.01). SUVmax, SUVmean, SUVpeak, TMTV, and TLMU were also significantly higher in \(^{11}\)C-MET than in \(^{18}\)F-FDG (p < 0.05, respectively). \(^{11}\)C-MET PET biomarkers had a better correlation with tumor burden (bone marrow plasma cell infiltration, M component; p < 0.05 versus p = n.s. respectively). This pilot study suggests that \(^{11}\)C-MET PET/CT is a more sensitive marker for the assessment of myeloma tumor burden than \(^{18}\)F-FDG. Its implications for prognosis evaluation need further investigation.
Prostate-specific membrane antigen (PSMA)-targeted PET imaging for prostate cancer with \(^{68}\)Ga-labeled compounds has rapidly become adopted as part of routine clinical care in many parts of the world. However, recent years have witnessed the start of a shift from \(^{68}\)Ga- to \(^{18}\)F-labeled PSMA-targeted compounds. The latter imaging agents have several key advantages, which may lay the groundwork for an even more widespread adoption into the clinic. First, facilitated delivery from distant suppliers expands the availability of PET radiopharmaceuticals in smaller hospitals operating a PET center but lacking the patient volume to justify an onsite \(^{68}\)Ge/\(^{68}\)Ga generator. Thus, such an approach meets the increasing demand for PSMA-targeted PET imaging in areas with lower population density and may even lead to cost-savings compared to in-house production. Moreover, \(^{18}\)F-labeled radiotracers have a higher positron yield and lower positron energy, which in turn decreases image noise, improves contrast resolution, and maximizes the likelihood of detecting subtle lesions. In addition, the longer half-life of 110 min allows for improved delayed imaging protocols and flexibility in study design, which may further increase diagnostic accuracy. Moreover, such compounds can be distributed to sites which are not allowed to produce radiotracers on-site due to regulatory issues or to centers without access to a cyclotron. In light of these advantageous characteristics, \(^{18}\)F-labeled PSMA-targeted PET radiotracers may play an important role in both optimizing this transformative imaging modality and making it widely available. We have aimed to provide a concise overview of emerging \(^{18}\)F-labeled PSMA-targeted radiotracers undergoing active clinical development. Given the wide array of available radiotracers, comparative studies are needed to firmly establish the role of the available \(^{18}\)F-labeled compounds in the field of molecular PCa imaging, preferably in different clinical scenarios.
Background
The emergence of antibiotic resistant bacteria in recent decades has highlighted the importance of developing new drugs to treat infections. However, in addition to the design of new drugs, the development of accurate preclinical testing methods is essential. In vivo imaging technologies such as bioluminescence imaging (BLI) or magnetic resonance imaging (MRI) are promising approaches. In a previous study, we showed the effectiveness of \(^{19}\)F MRI using perfluorocarbon (PFC) emulsions for detecting the site of Staphylococcus aureus infection. In the present follow-up study, we investigated the use of this method for in vivo visualization of the effects of antibiotic therapy.
Methods/Principal findings
Mice were infected with S. aureus Xen29 and treated with 0.9% NaCl solution, vancomycin or linezolid. Mock treatment led to the highest bioluminescence values during infection followed by vancomycin treatment. Counting the number of colony-forming units (cfu) at 7 days post-infection (p.i.) showed the highest bacterial burden for the mock group and the lowest for the linezolid group. Administration of PFCs at day 2 p.i. led to the accumulation of \(^{19}\)F at the rim of the abscess in all mice (in the shape of a hollow sphere), and antibiotic treatment decreased the \(^{19}\)F signal intensity and volume. Linezolid showed the strongest effect. The BLI, cfu, and MRI results were comparable.
Conclusions
\(^{19}\)F-MRI with PFCs is an effective non-invasive method for assessing the effects of antibiotic therapy in vivo. This method does not depend on pathogen specific markers and can therefore be used to estimate the efficacy of antibacterial therapy against a broad range of clinically relevant pathogens, and to localize sites of infection.
Chemokine receptor-4 (CXCR4) has been reported to be overexpressed in glioblastoma (GBM) and to be associated with poor survival. This study investigated the feasibility of non-invasive CXCR4-directed imaging with positron emission tomography/computed tomography (PET/CT) using the radiolabelled chemokine receptor ligand \(^{68}\)Ga-Pentixafor.
15 patients with clinical suspicion on primary or recurrent glioblastoma (13 primary, 2 recurrent tumors) underwent \(^{68}\)Ga-Pentixafor-PET/CT for assessment of CXCR4 expression prior to surgery. O-(2-\(^{18}\)F-fluoroethyl)-L-tyrosine (\(^{18}\)F-FET) PET/CT images were available in 11/15 cases and were compared visually and semi-quantitatively (SUV\(_{max}\), SUV\(_{mean}\)). Tumor-to-background ratios (TBR) were calculated for both PET probes. \(^{68}\)Ga-Pentixafor-PET/CT results were also compared to histological CXCR4 expression on neuronavigated surgical samples.
\(^{68}\)Ga-Pentixafor-PET/CT was visually positive in 13/15 cases with SUV\(_{mean}\) and SUV\(_{max}\) of 3.0±1.5 and 3.9±2.0 respectively. Respective values for \(^{18}\)F-FET were 4.4±2.0 (SUV\(_{mean}\)) and 5.3±2.3 (SUV\(_{max}\)). TBR for SUV\(_{mean}\) and SUV\(_{max}\) were higher for \(^{68}\)Ga-Pentixafor than for \(^{18}\)F-FET (SUV\(_{mean}\) 154.0±90.7 vs. 4.1±1.3; SUV\(_{max}\) 70.3±44.0 and 3.8±1.2, p<0.01), respectively. Histological analysis confirmed CXCR4 expression in tumor areas with high \(^{68}\)Ga-Pentixafor uptake; regions of the same tumor without apparent \(^{68}\)Ga-Pentixafor uptake showed no or low receptor expression.
In this pilot study, \(^{68}\)Ga-Pentixafor retention has been observed in the vast majority of glioblastoma lesions and served as readout for non-invasive determination of CXCR4 expression. Given the paramount importance of the CXCR4/SDF-1 axis in tumor biology, \(^{68}\)Ga-Pentixafor-PET/CT might prove a useful tool for sensitive, non-invasive in-vivo quantification of CXCR4 as well as selection of patients who might benefit from CXCR4-directed therapy.
A cyclic (alkyl)(amino)carbene (CAAC) has been shown to react with a covalent azide similar to the Staudinger reaction. The reaction of \(^{Me}\)CAAC with trimethylsilyl azide afforded the N‐silylated 2‐iminopyrrolidine (\(^{Me}\)CAAC=NSiMe\(_{3}\)), which was fully characterized. This compound undergoes hydrolysis to afford the 2‐iminopyrrolidine and trimethylsiloxane which co‐crystallize as a hydrogen‐bonded adduct. The N‐silylated 2‐iminopyrrolidine was used to transfer the novel pyrrolidine‐2‐iminato ligand onto both main‐group and transition‐metal centers. The reaction of the tetrabromodiborane bis(dimethyl sulfide) adduct with two equivalents of \(^{Me}\)CAAC=NSiMe\(_{3}\) afforded the disubstituted diborane. The reaction of \(^{Me}\)CAAC=NSiMe\(_{3}\) with TiCl\(_{4}\) and CpTiCl\(_{3}\) afforded \(^{Me}\)CAAC=NTiCl\(_{3}\) and \(^{Me}\)CAAC=NTiCl\(_{2}\)Cp, respectively.
Context: Patients with primary adrenal insufficiency (PAI) or congenital adrenal hyperplasia (CAH) are at a high risk of adrenal crisis (AC). Glucocorticoid sensitivity is at least partially genetically determined by polymorphisms of the glucocorticoid receptor (GR).
Objectives: To determine if a number of intercurrent illnesses and AC are associated with the GR gene polymorphism \(Bcl\)I in patients with PAI and CAH.
Design and patients: This prospective, longitudinal study over 37.7 ± 10.1 months included 47 PAI and 25 CAH patients. During the study period, intercurrent illness episodes and AC were documented.
Results: The study period covered 223 patient years in which 21 AC occurred (9.4 AC/100 pat years). There were no significant differences between \(Bcl\)I polymorphisms (CC (n=29), CG (n=34) and GG (n=9)) regarding BMI, hydrocortisone equivalent daily dose and blood pressure. We did not find a difference in the number of intercurrent illnesses/patient year among \(Bcl\)I polymorphisms (CC (1.5±1.4/pat year), CG (1.2±1.2/pat year) and GG (1.6±2.2/pat year)). The occurrence of AC was not significantly different among the homozygous (GG) genotype (32.5 AC/100 pat years), the CC genotype (6.7 AC/100 pat years) and the CG genotype (4.9 AC/100 pat years). Concomitant hypothyroidism was the highest in the GG genotype group (5/9), compared to others (CC (11/29) and CG (11/34)).
Conclusions: Although sample sizes were relatively small and results should be interpreted with caution, this study suggests that the GR gene polymorphism \(Bcl\)I may not be associated with the frequencies of intercurrent illnesses and AC.
Life-threatening systemic infections often occur due to the translocation of pathogens across the gut barrier and into the bloodstream. While the microbial and host mechanisms permitting bacterial gut translocation are well characterized, these mechanisms are still unclear for fungal pathogens such as Candida albicans, a leading cause of nosocomial fungal bloodstream infections. In this study, we dissected the cellular mechanisms of translocation of C. albicans across intestinal epithelia in vitro and identified fungal genes associated with this process. We show that fungal translocation is a dynamic process initiated by invasion and followed by cellular damage and loss of epithelial integrity. A screen of >2,000 C. albicans deletion mutants identified genes required for cellular damage of and translocation across enterocytes. Correlation analysis suggests that hypha formation, barrier damage above a minimum threshold level, and a decreased epithelial integrity are required for efficient fungal translocation. Translocation occurs predominantly via a transcellular route, which is associated with fungus-induced necrotic epithelial damage, but not apoptotic cell death. The cytolytic peptide toxin of C. albicans, candidalysin, was found to be essential for damage of enterocytes and was a key factor in subsequent fungal translocation, suggesting that transcellular translocation of C. albicans through intestinal layers is mediated by candidalysin. However, fungal invasion and low-level translocation can also occur via non-transcellular routes in a candidalysin-independent manner. This is the first study showing translocation of a human-pathogenic fungus across the intestinal barrier being mediated by a peptide toxin. IMPORTANCE Candida albicans, usually a harmless fungus colonizing human mucosae, can cause lethal bloodstream infections when it manages to translocate across the intestinal epithelium. This can result from antibiotic treatment, immune dysfunction, or intestinal damage (e.g., during surgery). However, fungal processes may also contribute. In this study, we investigated the translocation process of C. albicans using in vitro cell culture models. Translocation occurs as a stepwise process starting with invasion, followed by epithelial damage and loss of epithelial integrity. The ability to secrete candidalysin, a peptide toxin deriving from the hyphal protein Ece1, is key: C. albicans hyphae, secreting candidalysin, take advantage of a necrotic weakened epithelium to translocate through the intestinal layer.
Chlamydia trachomatis (Ct) is an obligate intracellular human pathogen. It causes blinding trachoma and sexually transmitted disease such as chlamydia, pelvic inflammatory disease and lymphogranuloma venereum. Ct has a unique biphasic development cycle and replicates in an intracellular vacuole called inclusion. Normally it has two forms: the infectious form, elementary body (EB); and the non-infectious form, reticulate body (RB). Ct is not easily amenable to genetic manipulation. Hence, to understand the infection process, it is crucial to study how the metabolic activity of Ct exactly evolves in the host cell and what roles of EB and RB play differentially in Ct metabolism during infection. In addition, Ct was found regularly coinfected with other pathogens in patients who got sexually transmitted diseases (STDs). A lack of powerful methods to culture Ct outside of the host cell makes the detailed molecular mechanisms of coinfection difficult to study.
In this work, a genome-scale metabolic model with 321 metabolites and 277 reactions was first reconstructed by me to study Ct metabolic adaptation in the host cell during infection. This model was calculated to yield 84 extreme pathways, and metabolic flux strength was then modelled regarding 20hpi, 40hpi and later based on a published proteomics dataset. Activities of key enzymes involved in target pathways were further validated by RT-qPCR in both HeLa229 and HUVEC cell lines. This study suggests that Ct's major active pathways involve glycolysis, gluconeogenesis, glycerolphospholipid biosynthesis and pentose phosphate pathway, while Ct's incomplete tricarboxylic acid cycle and fatty acid biosynthesis are less active. EB is more activated in almost all these carbohydrate pathways than RB. Result suggests the survival of Ct generally requires a lot of acetyl-CoA from the host. Besides, both EB and RB can utilize folate biosynthesis to generate NAD(P)H but may use different pathways depending on the demands of ATP. When more ATP is available from both host cell and Ct itself, RB is more activated by utilizing energy providing chemicals generated by enzymes associated in the nucleic acid metabolism. The forming of folate also suggests large glutamate consumption, which is supposed to be converted from glutamine by the glutamine-fructose-6-phosphate transaminase (glmS) and CTP synthase (pyrG).
Then, RNA sequencing (RNA-seq) data analysis was performed by me in a coinfection study. Metatranscriptome from patient RNA-seq data provides a realistic overview. Thirteen patient samples were collected and sequenced by our collaborators. Six male samples were obtained by urethral swab, and seven female samples were collected by cervicovaginal lavage. All the samples were Neisseria gonorrhoeae (GC) positive, and half of them had coinfection with Ct. HISAT2 and Stringtie were used for transcriptomic mapping and assembly respectively, and differential expression analysis by DESeq2, Ballgown and Cuffdiff2 are parallelly processed for comparison. Although the measured transcripts were not sufficient to assemble Ct's transcriptome, the differential expression of genes in both the host and GC were analyzed by comparing Ct positive group (Ct+) against Ct-uninfected group. The results show that in the Ct+ group, the host MHC class II immune response was highly induced. Ct infection is associated with the regulation of DNA methylation, DNA double-strand damage and ubiquitination. The analysis also shows Ct infection enhances host fatty acid beta oxidation, thereby inducing mROS, and the host responds to reduce ceramide production and glycolysis. The coinfection upregulates GC's own ion transporters and amino acid uptake, while it downregulates GC's restriction and modification systems. Meanwhile, GC has the nitrosative and oxidative stress response and also increases the ability for ferric uptake especially in the Ct+ group compared to Ct-uninfected group.
In conclusion, methods in bioinformatics were used here in analyzing the metabolism of Ct itself, and the responses of the host and GC respectively in a coinfection study with and without Ct. These methods provide metabolic and metatranscriptomic details to study Ct metabolism during infection and Ct associated coinfection in the human microbiota.
Clostridioides difficile is a bacterial species well known for its ability to cause C. difficile
infection (also known as CDI). The investigation of the role of this species in the human
gut has been so far dominated by a disease-centred perspective, focused on studying
C. difficile in relation to its associated disease.
In this context, the first aim of this thesis was to combine publicly available
metagenomic data to analyse the microbial composition of stool samples from patients
diagnosed with CDI, with a particular focus on identifying a CDI-specific microbial
signature.
However, similarly to many other bacterial species inhabiting the human gut, C.
difficile association with disease is not valid in absolute terms, as C. difficile can be
found also among healthy subjects. Further aims of this thesis were to 1) identify
potential C. difficile reservoirs by screening a wide range of habitats, hosts, body sites
and age groups, and characterize the biotic context associated with C. difficile
presence, and 2) investigate C. difficile within-species diversity and its toxigenic
potential across different age groups.
The first part of the thesis starts with the description of the concepts and
definitions used to identify bacterial species and within-species diversity, and then
proceeds to provide an overview of the bacterial species at the centre of my
investigation, C. difficile. The first Chapter includes a detailed description of the
discovery, biology and physiology of this clinically relevant species, followed by an
overview of the diagnostic protocols used in the clinical setting to diagnose CDI.
The second part of the thesis describes the methodology used to investigate
the questions mentioned above, while the third part presents the results of such
investigative effort. I first show that C. difficile could be found in only a fraction of the
CDI samples and that simultaneous colonization of multiple enteropathogenic species
able to cause CDI-like clinical manifestations is more common than previously
thought, raising concerns about CDI overdiagnosis. I then show that the CDIassociated
gut microbiome is characterized by a specific microbial signature,
distinguishable from the community composition associated with non-CDI diarrhea.
Beyond the nosocomial and CDI context, I show that while rarely found in adults, C.
difficile is a common member of the infant gut microbiome, where its presence is
associated with multiple indicators typical of a desirable healthy microbiome
development.
In addition, I describe C. difficile extensive carriage among asymptomatic
subjects, of all age groups and a potentially novel clade of C. difficile identified
exclusively among infants.
Finally, I discuss the limitations, challenges and future perspectives of my
investigation.
The role of regulatory T cells (Tregs) in bacterial sepsis remains controversial because antibody-mediated depletion experiments gave conflicting results. We employed DEREG mice (DEpletion of REGulatory T cells) and a caecal ligation and puncture model to elucidate the role of \(CD4^+Foxp3^+\) Tregs in sepsis. In DEREG mice natural Tregs can be visualized easily and selectively depleted by diphtheria toxin because the animals express the diphtheria toxin receptor and enhanced green fluorescent protein as a fusion protein under the control of the foxp3 locus. We confirmed rapid Treg-activation and an increased ratio of Tregs to Teffs in sepsis. Nevertheless, 24 h after sepsis induction, Treg-depleted and control mice showed equally strong inflammation, immune cell immigration into the peritoneum and bacterial dissemination. During the first 36 h of disease survival was not influenced by Treg-depletion. Later, however, only Treg-competent animals recovered from the insult. We conclude that the suppressive capacity of Tregs is not sufficient to control overwhelming inflammation and early mortality, but is a prerequisite for the recovery from severe sepsis.
Highlights
• The GLA variant S126G is not associated with Fabry symptoms in the presented case
• S126G has no effect on α-GAL A activity or Gb3 levels in this patient
• S126G sensory neurons show no electrophysiological abnormalities
Abstract
Fabry disease (FD) is a life-limiting disorder characterized by intracellular globotriaosylceramide (Gb3) accumulations. The underlying α-galactosidase A (α-GAL A) deficiency is caused by variants in the gene GLA. Variants of unknown significance (VUS) are frequently found in GLA and challenge clinical management. Here, we investigated a 49-year old man with cryptogenic lacunar cerebral stroke and the chance finding of the VUS S126G, who was sent to our center for diagnosis and initiation of a costly and life-long FD-specific treatment. We combined clinical examination with in vitro investigations of dermal fibroblasts (HDF), induced pluripotent stem cells (iPSC), and iPSC-derived sensory neurons. We analyzed α-GAL A activity in iPSC, Gb3 accumulation in all three cell types, and action potential firing in sensory neurons. Neurological examination and small nerve fiber assessment was normal except for reduced distal skin innervation. S126G iPSC showed normal α-GAL A activity compared to controls and no Gb3 deposits were found in all three cell types. Baseline electrophysiological characteristics of S126G neurons showed no difference compared to healthy controls as investigated by patch-clamp recordings. We pioneer multi-level cellular characterization of the VUS S126G using three cell types derived from a patient and provide further evidence for the benign nature of S126G in GLA, which is of great importance in the management of such cases in clinical practice.
Plexus injury often occurs after motor vehicle accidents and results in lifelong disability with severe neuropathic pain. Surgical treatment can partially restore motor functions, but sensory loss and neuropathic pain persist. Regenerative medicine concepts, such as cell replacement therapies for restoring dorsal root ganglia (DRG) function, set high expectations. However, up to now, it is unclear which DRG cell types are affected by nerve injury and can be targeted in regenerative medicine approaches.
This study followed the hypothesis that satellite glial cells (SGCs) might be a suitable endogenous cell source for regenerative medicine concepts in the DRG. SGCs originate from the same neural crest-derived cell lineage as sensory neurons, making them attractive for neural repair strategies in the peripheral nervous system. Our hypothesis was investigated on three levels of experimentation. First, we asked whether adult SGCs have the potential of sensory neuron precursors and can be reprogrammed into sensory neurons in vitro. We found that adult mouse DRG harbor SGC-like cells that can still dedifferentiate into progenitor-like cells. Surprisingly, expression of the early developmental transcription factors Neurog1 and Neurog2 was sufficient to induce neuronal and glial cell phenotypes. In the presence of nerve growth factor, induced neurons developed a nociceptor-like phenotype expressing functional nociceptor markers, such as the ion channels TrpA1, TrpV1 and NaV1.9. In a second set of experiments, we used a rat model for peripheral nerve injury to look for changes in the DRG cell composition. Using an unbiased deep learning-based approach for cell analysis, we found that cellular plasticity responses after nerve injury activate SGCs in the whole DRG. However, neither injury-induced neuronal death nor gliosis was observed. Finally, we asked whether a severe nerve injury changed the cell composition in the human DRG. For this, a cohort of 13 patients with brachial plexus injury was investigated. Surprisingly, in about half of all patients, the injury-affected DRG showed no characteristic DRG tissue. The complete entity of neurons, satellite cells, and axons was lost and fully replaced by mesodermal/connective tissue. In the other half of the patients, the basic cellular entity of the DRG was well preserved. Objective deep learning-based analysis of large-scale bioimages of the “intact” DRG showed no loss of neurons and no signs of gliosis.
This study suggests that concepts for regenerative medicine for restoring DRG function need at least two translational research directions: reafferentation of existing DRG units or full replacement of the entire multicellular DRG structure. For DRG replacement, SGCs of the adult DRG are an attractive endogenous cell source, as the multicellular DRG units could possibly be rebuilt by transdifferentiating neural crest-derived sensory progenitor cells into peripheral sensory neurons and glial cells using Neurog1 and Neurog2.
This thesis provides an edition and commentary of a manuscript discovered by Michael Stolberg in the archives of the central library in Zurich under the title “Mon aprendisage à l'Hôtel Dieu de Paris 1704.” (My apprenticeship at the Hôtel-Dieu de Paris 1704). The manuscript contains records of a midwifery student at the Hôtel-Dieu de Paris, an old hospital famous among others for its education in midwifery in the maternity ward. We read about managing different births, recipes for common remedies, direct questions answered by the maîtresse sage-femme, the leading midwife at the Hôtel-Dieu de Paris and more.
Although other accounts exist of the maternity ward at the Hôtel-Dieu de Paris, \(Mon\) \(Aprendisage\) is the first and only account from a midwife’s perspective that gives more than just instructions on obstetrical techniques. It takes us into the day-to-day experience of a woman as she progressed through her training at the Hôtel-Dieu.
Surgical implantation of a biomaterial triggers foreign-body-induced fibrous encapsulation. Two major mechanisms of this complex physiological process are (I) chemotaxis of fibroblasts from surrounding tissue to the implant region, followed by (II) tissue remodeling. As an alternative to animal studies, we here propose a process-aligned \({in}\) \({vitro}\) test platform to investigate the material dependency of fibroblast chemotaxis and tissue remodeling mediated by material-resident macrophages.
Embedded in a biomimetic three-dimensional collagen hydrogel, chemotaxis of fibroblasts in the direction of macrophage-material-conditioned cell culture supernatant was analyzed by live cell imaging. A combination of statistical analysis with a complementary parameterized random walk model allowed quantitative and qualitative characterization of the cellular walk process. We thereby identified an increasing macrophage-mediated chemotactic potential ranking of biomaterials from glass over polytetrafluorethylene to titanium. To address long-term effects of biomaterial-resident macrophages on fibroblasts in a three-dimensional microenvironment, we further studied tissue remodeling by applying macrophage-material-conditioned medium on fibrous \({in}\) \({vitro}\) tissue models. A high correlation of the \({in}\) \({vitro}\) tissue model to state of the art \({in}\) \({vivo}\) study data was found. Titanium exhibited a significantly lower tissue remodeling capacity compared to polytetrafluorethylene. With this approach, we identified a material dependency of both chemotaxis and tissue remodeling processes, strengthening knowledge on their specific contribution to the foreign body reaction.
Measurements of ZZ production in the l(+)l(-)l'(+)l'(-) channel in proton-proton collisions at 13 TeV center-of-mass energy at the Large Hadron Collider are presented. The data correspond to 36.1 fb(-1) of collisions collected by the ATLAS experiment in 2015 and 2016. Here l and l ' stand for electrons or muons. Integrated and differential ZZ -> l(+)l(-)l'(+)l'(-) cross sections with Z -> l(+)l(-) candidate masses in the range of 66 GeV to 116 GeV are measured in a fiducial phase space corresponding to the detector acceptance and corrected for detector effects. The differential cross sections are presented in bins of twenty observables, including several that describe the jet activity. The integrated cross section is also extrapolated to a total phase space and to all standard model decays of Z bosons with mass between 66 GeV and 116 GeV, resulting in a value of 17.3 +/- 0.9 [+/- 0.6(start) +/- 0.5 (syst) +/- 0.6 (lumi)] pb. The measurements are found to be in good agreement with the standard model. A search for neutral triple gauge couplings is performed using the transverse momentum distribution of the leading Z boson candidate. No evidence for such couplings is found and exclusion limits are set on their parameters.
Several hypotheses explain the prevalence of undifferentiated sex chromosomes in poikilothermic vertebrates. Turnovers change the master sex determination gene, the sex chromosome or the sex determination system (e.g. XY to WZ). Jumping master genes stay main triggers but translocate to other chromosomes. Occasional recombination (e.g. in sex-reversed females) prevents sex chromosome degeneration. Recent research has uncovered conserved heteromorphic or even homomorphic sex chromosomes in several clades of non-avian and non-mammalian vertebrates. Sex determination in sturgeons (Acipenseridae) has been a long-standing basic biological question, linked to economical demands by the caviar-producing aquaculture. Here, we report the discovery of a sex-specific sequence from sterlet (Acipenser ruthenus). Using chromosome-scale assemblies and pool-sequencing, we first identified an approximately 16 kb female-specific region. We developed a PCR-genotyping test, yielding female-specific products in six species, spanning the entire phylogeny with the most divergent extant lineages (A. sturio, A. oxyrinchus versus A. ruthenus, Huso huso), stemming from an ancient tetraploidization. Similar results were obtained in two octoploid species (A. gueldenstaedtii, A. baerii). Conservation of a female-specific sequence for a long period, representing 180 Myr of sturgeon evolution, and across at least one polyploidization event, raises many interesting biological questions. We discuss a conserved undifferentiated sex chromosome system with a ZZ/ZW-mode of sex determination and potential alternatives.
This article is part of the theme issue ‘Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)’.
The signals that coordinate and control movement in vertebrates are transmitted from motoneurons (MNs) to their target muscle cells at neuromuscular junctions (NMJs). Human NMJs display unique structural and physiological features, which make them vulnerable to pathological processes. NMJs are an early target in the pathology of motoneuron diseases (MND). Synaptic dysfunction and synapse elimination precede MN loss suggesting that the NMJ is the starting point of the pathophysiological cascade leading to MN death. Therefore, the study of human MNs in health and disease requires cell culture systems that enable the connection to their target muscle cells for NMJ formation. Here, we present a human neuromuscular co-culture system consisting of induced pluripotent stem cell (iPSC)-derived MNs and 3D skeletal muscle tissue derived from myoblasts. We used self-microfabricated silicone dishes combined with Velcro hooks to support the formation of 3D muscle tissue in a defined extracellular matrix, which enhances NMJ function and maturity. Using a combination of immunohistochemistry, calcium imaging, and pharmacological stimulations, we characterized and confirmed the function of the 3D muscle tissue and the 3D neuromuscular co-cultures. Finally, we applied this system as an in vitro model to study the pathophysiology of Amyotrophic Lateral Sclerosis (ALS) and found a decrease in neuromuscular coupling and muscle contraction in co-cultures with MNs harboring ALS-linked SOD1 mutation. In summary, the human 3D neuromuscular cell culture system presented here recapitulates aspects of human physiology in a controlled in vitro setting and is suitable for modeling of MND.
Burns affect millions every year and a model to mimic the pathophysiology of such injuries in detail is required to better understand regeneration. The current gold standard for studying burn wounds are animal models, which are under criticism due to ethical considerations and a limited predictiveness. Here, we present a three-dimensional burn model, based on an open-source model, to monitor wound healing on the epidermal level. Skin equivalents were burned, using a preheated metal cylinder. The healing process was monitored regarding histomorphology, metabolic changes, inflammatory response and reepithelialization for 14 days. During this time, the wound size decreased from 25% to 5% of the model area and the inflammatory response (IL-1β, IL-6 and IL-8) showed a comparable course to wounding and healing in vivo. Additionally, the topical application of 5% dexpanthenol enhanced tissue morphology and the number of proliferative keratinocytes in the newly formed epidermis, but did not influence the overall reepithelialization rate. In summary, the model showed a comparable healing process to in vivo, and thus, offers the opportunity to better understand the physiology of thermal burn wound healing on the keratinocyte level.
A 3D printed model of the female pelvis for practical education of gynecological pelvic examination
(2022)
Background
Pelvic palpation is a core component of every Gynecologic examination. It requires vigorous training, which is difficult due to its intimate nature, leading to a need of simulation. Up until now, there are mainly models available for mere palpation which do not offer adequate visualization of the concerning anatomical structures. In this study we present a 3D printed model of the female pelvis. It can improve both the practical teaching of gynecological pelvic examination for health care professionals and the spatial understanding of the relevant anatomy.
Methods
We developed a virtual, simplified model showing selected parts of the female pelvis. 3D printing was used to create a physical model.
Results
The life-size 3D printed model has the ability of being physically assembled step by step by its users. Consequently, it improves teaching especially when combining it with commercial phantoms, which are built solely for palpation training. This is achieved by correlating haptic and visual sensations with the resulting feedback received.
Conclusion
The presented 3D printed model of the female pelvis can be of aid for visualizing and teaching pelvic anatomy and examination to medical staff. 3D printing provides the possibility of creating, multiplying, adapting and sharing such data worldwide with little investment of resources. Thus, an important contribution to the international medical community can be made for training this challenging examination.
To compare the effects of a 3‐week multimodal rehabilitation involving supervised high‐intensity interval training (HIIT) on female breast cancer survivors with respect to key variables of aerobic fitness, body composition, energy expenditure, cancer‐related fatigue, and quality of life to those of a standard multimodal rehabilitation program. A randomized controlled trial design was administered. Twenty‐eight women, who had been treated for cancer were randomly assigned to either a group performing exercise of low‐to‐moderate intensity (LMIE; n = 14) or a group performing high‐intensity interval training (HIIT; n = 14) as part of a 3‐week multimodal rehabilitation program. No adverse events related to the exercise were reported. Work economy improved following both HIIT and LMIE, with improved peak oxygen uptake following LMIE. HIIT reduced mean total body fat mass with no change in body mass, muscle or fat‐free mass (best P < 0.06). LMIE increased muscle and total fat‐free body mass. Total energy expenditure (P = 0.45) did not change between the groups, whereas both improved quality of life to a similar high extent and lessened cancer‐related fatigue. This randomized controlled study demonstrates that HIIT can be performed by female cancer survivors without adverse health effects. Here, HIIT and LMIE both improved work economy, quality of life and cancer‐related fatigue, body composition or energy expenditure. Since the outcomes were similar, but HIIT takes less time, this may be a time‐efficient strategy for improving certain aspects of the health of female cancer survivors.
The present study was designed to assess the psycho-physiological responses of physically untrained individuals to mobile-based multi-stimulating, circuit-like, multiple-joint conditioning (Circuit\(_{HIIT}\)) performed either once (1xCircuitHIIT) or twice (2xCircuit\(_{HIIT}\)) daily for 4 weeks. In this single-center, two-arm randomized, controlled study, 24 men and women (age: 25 ± 5 years) first received no training instructions for 4 weeks and then performed 4 weeks of either 1xCircuitHIIT or 2xCircuit\(_{HIIT}\) (5 men and 7 women in each group) daily. The 1xCircuitHIIT and 2xCircuit\(_{HIIT}\) participants carried out 90.7 and 85.7% of all planned training sessions, respectively, with average heart rates during the 6-min sessions of 74.3 and 70.8% of maximal heart rate. Body, fat and fat-free mass, and metabolic rate at rest did not differ between the groups or between time-points of measurement. Heart rate while running at 6 km⋅h\(^{-1}\) declined after the intervention in both groups. Submaximal and peak oxygen uptake, the respiratory exchange ratio and heart rate recovery were not altered by either intervention. The maximal numbers of push-ups, leg-levers, burpees, 45°-one-legged squats and 30-s skipping, as well as perception of general health improved in both groups. Our 1xCircuit\(_{HIIT}\) or 2xCircuit\(_{HIIT}\) interventions improved certain parameters of functional strength and certain dimensions of quality of life in young untrained individuals. However, they were not sufficient to enhance cardio-respiratory fitness, in particular peak oxygen uptake.
A cloned cytolytic determinant from the genome of Bacillus cereus GP-4 has been characterized at the molecular Ievel. Nucleotide sequence determination revealed the presence of two open reading frames. 8oth open reading frames were found by deletion and complementation analysis to be necessary for expression of the hemolytic phenotype by Bacillus subtilis and Escherichia coli hosts. The 5' open reading frame was found to be nearly identical to a recently reported phospholipase C gene derived from a mutant B. cereus strain which overexpresses the respective protein, and it conferred a lecithinase-positive phenotype to the B. subtilis host. The 3' open reading frame encoded a sphingomyelinase. The two tandemly encoded activities, phospholipase C and sphingomyelinase, constitute a biologically functional cytolytic determinant of B. cereus termed cereolysin AB.
Deafness, the most frequent sensory deficit in humans, is extremely heterogeneous with hundreds of genes involved. Clinical and genetic analyses of an extended consanguineous family with pre-lingual, moderate-to-profound autosomal recessive sensorineural hearing loss, allowed us to identify CLRN2, encoding a tetraspan protein, as a new deafness gene. Homozygosity mapping followed by exome sequencing identified a 14.96 Mb locus on chromosome 4p15.32p15.1 containing a likely pathogenic missense variant in CLRN2 (c.494C > A, NM_001079827.2) segregating with the disease. Using in vitro RNA splicing analysis, we show that the CLRN2 c.494C > A variant leads to two events: (1) the substitution of a highly conserved threonine (uncharged amino acid) to lysine (charged amino acid) at position 165, p.(Thr165Lys), and (2) aberrant splicing, with the retention of intron 2 resulting in a stop codon after 26 additional amino acids, p.(Gly146Lysfs*26). Expression studies and phenotyping of newly produced zebrafish and mouse models deficient for clarin 2 further confirm that clarin 2, expressed in the inner ear hair cells, is essential for normal organization and maintenance of the auditory hair bundles, and for hearing function. Together, our findings identify CLRN2 as a new deafness gene, which will impact future diagnosis and treatment for deaf patients.
Evolution has endowed the lung with exceptional design providing a large surface area for gas exchange area (ca. 100 m\(^{2}\)) in a relatively small tissue volume (ca. 6 L). This is possible due to a complex tissue architecture that has resulted in one of the most challenging organs to be recreated in the lab. The need for realistic and robust in vitro lung models becomes even more evident as causal therapies, especially for chronic respiratory diseases, are lacking. Here, we describe the Cyclic In VItro Cell-stretch (CIVIC) “breathing” lung bioreactor for pulmonary epithelial cells at the air-liquid interface (ALI) experiencing cyclic stretch while monitoring stretch-related parameters (amplitude, frequency, and membrane elastic modulus) under real-time conditions. The previously described biomimetic copolymeric BETA membrane (5 μm thick, bioactive, porous, and elastic) was attempted to be improved for even more biomimetic permeability, elasticity (elastic modulus and stretchability), and bioactivity by changing its chemical composition. This biphasic membrane supports both the initial formation of a tight monolayer of pulmonary epithelial cells (A549 and 16HBE14o\(^{-}\)) under submerged conditions and the subsequent cell-stretch experiments at the ALI without preconditioning of the membrane. The newly manufactured versions of the BETA membrane did not improve the characteristics of the previously determined optimum BETA membrane (9.35% PCL and 6.34% gelatin [w/v solvent]). Hence, the optimum BETA membrane was used to investigate quantitatively the role of physiologic cyclic mechanical stretch (10% linear stretch; 0.33 Hz: light exercise conditions) on size-dependent cellular uptake and transepithelial transport of nanoparticles (100 nm) and microparticles (1,000 nm) for alveolar epithelial cells (A549) under ALI conditions. Our results show that physiologic stretch enhances cellular uptake of 100 nm nanoparticles across the epithelial cell barrier, but the barrier becomes permeable for both nano- and micron-sized particles (100 and 1,000 nm). This suggests that currently used static in vitro assays may underestimate cellular uptake and transbarrier transport of nanoparticles in the lung.