Quantitative imaging of intermediate filaments (IF) during the advanced phase of the assembly process is technically difficult, since the structures are several µm long and therefore they exceed the ...field of view of many electron (EM) or atomic force microscopy (AFM) techniques. Thereby quantitative studies become extremely laborious and time-consuming. To overcome these difficulties, we prepared fluorescently labeled vimentin for visualization by total internal reflection fluorescence microscopy (TIRFM). In order to investigate if the labeling influences the assembly properties of the protein, we first determined the association state of unlabeled vimentin mixed with increasing amounts of labeled vimentin under low ionic conditions by analytical ultracentrifugation. We found that bona fide tetrameric complexes were formed even when half of the vimentin was labeled. Moreover, we demonstrate by quantitative atomic force microscopy and electron microscopy that the morphology and the assembly properties of filaments were not affected when the fraction of labeled vimentin was below 10%. Using fast frame rates we observed the rapid deposition of fluorescently labeled IFs on glass supports by TIRFM in real time. By tracing their contours, we have calculated the persistence length of long immobilized vimentin IFs to 1 µm, a value that is identical to those determined for shorter unlabeled vimentin. These results indicate that the structural properties of the filaments were not affected significantly by the dye. Furthermore, in order to analyze the late elongation phase, we mixed long filaments containing either Alexa 488- or Alexa 647-labeled vimentin. The 'patchy' structure of the filaments obtained unambiguously showed the elongation of long IFs through direct end-to-end annealing of individual filaments.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The influence of cholesterol's alkyl side chain on membrane properties was studied using a series of synthetic cholesterol derivatives without a side chain or with a branched side chain consisting of ...5 to 14 carbon atoms. Cholesterol's side chain is crucial for all membrane properties investigated and therefore essential for the membrane properties of eukaryotic cells.
Lipid analogues carrying three nitrilotriacetic acid (tris‐NTA) head groups were developed for the selective targeting of His‐tagged proteins into liquid ordered (lo) or liquid disordered (ld) lipid ...phases. Strong partitioning into the lo phase of His‐tagged proteins bound to tris‐NTA conjugated to saturated alkyl chains (tris‐NTA DODA) was achieved, while tris‐NTA conjugated to an unsaturated alkyl chain (tris‐NTA SOA) predominantly resided in the ld phase. Interestingly, His‐tag‐mediated lipid crosslinking turned out to be required for efficient targeting into the lo phase by tris‐NTA DODA. Robust partitioning into lo phases was confirmed by using viral lipid mixtures and giant plasma membrane vesicles. Moreover, efficient protein targeting into lo and ld domains within the plasma membrane of living cells was demonstrated by single‐molecule tracking, thus establishing a highly generic approach for exploring lipid microdomains in situ.
Rapid rafting: Lipid analogues with multivalent chelator head groups allow the tethering of proteins to membrane liquid‐ordered and liquid‐disordered lipid phases. His‐tagged proteins can thus be stably and specifically targeted into lipid microdomains within the plasma membrane of living cells.
The reaction between BF3⋅OEt2 and one of two guanidines, 1,8‐bis(tetramethylguanidinyl)naphthalene (btmgn) and 1,2,4,5‐tetrakis(tetramethylguanidinyl)naphthalene (ttmgn), yields the salts ...(btmgn)(BF2)BF4 and (ttmgn)(BF2)2(BF4)2. NMR spectroscopic data show that the boron atoms in the cation and anion exchange in the case of (ttmgn)(BF2)2(BF4)2, but not in the case of (btmgn)(BF2)BF4. The rate constant for this exchange was estimated to be 4 s−1 at 80 °C for solutions in CH3CN. These salts were subsequently used for the reduction of dihalides Br2 or I2 to give polyhalide salts. We report the synthesis and first complete characterization (including structural analysis) of salts that contain pentabromide monoanions. In these salts, the Br5− anions interact to give dimeric units or polymeric chains. Our results are compared to previous quantum chemical calculations on the gas‐phase structure of the Br5− anion. The possible pathways that lead to the polyhalides are evaluated. In the case of (ttmgn)(BF2)2(BF4)2, reduction is accompanied by ttmgn oxidation, whereas in the case of (btmgn)(BF2)BF4 reduction is initiated by aromatic substitution.
That's a wrap: After wrapping it in a cationic boron complex, the organic reducing agent 1,2,4,5‐tetrakis(tetramethylguanidinyl)naphthalene (ttmgn) was used for the synthesis of polybromide and polyiodide monoanions (see scheme).
RATIONALE:Cell-based therapies are a promising option in patients with chronic postinfarction heart failure (ischemic cardiomyopathy ICM). However, the responses after intracoronary infusion of ...autologous bone marrow–derived mononuclear cells (BMCs) are heterogeneous, which may be related to impaired cell retention in patients with ICM. Ischemic injury is associated with upregulation of prototypical chemoattractant cytokines mediating retention and homing of circulating cells. The development of ultrasensitive tests to measure high-sensitive troponin T (hs-TnT) serum levels revealed the presence of ongoing minute myocardial injury even in patients with stable ICM.
OBJECTIVE:To test the hypothesis that serum levels of hs-TnT correlate with cell retention and determine the response to intracoronary BMC application in patients with ICM.
METHODS AND RESULTS:About 157 patients with stable ICM and no substantial impairment of kidney function received intracoronary BMC administration. Immediately prior to cell application, hs-TnT levels to measure myocardial injury and NT-proBNP levels as marker of left ventricular wall stress were determined. Patients with elevated hs-TnT were older and had more severe heart failure. Importantly, only patients with elevated baseline hs-TnT≥15.19 pg/mL (upper tertile) demonstrated a significant (P=0.04) reduction in NT-proBNP serum levels (−250 −1465; 33 pg/mL; relative reduction −24%) 4 months after BMC administration, whereas NT-proBNP levels remained unchanged in patients in the 2 lower hs-TnT tertiles. The absolute decrease in NT-proBNP at 4 months was inversely correlated with baseline hs-TnT (r=−0.27, P=0.001). Finally, retention of intracoronarily infused, Indium-labeled cells within the heart was closely associated with hs-TnT levels in patients with chronic ischemic heart failure (P=0.0008, n=10, triple measurements).
CONCLUSIONS:The extent of ongoing myocardial injury as measured by serum levels of hs-TnT predicts the reduction of NT-proBNP serum levels at 4 months after intracoronary BMC administration in patients with ICM, suggesting that the beneficial effects of BMC application on LV remodeling and wall stress are confined to patients with ongoing minute myocardial injury.
CLINICAL TRIAL REGISTRATION:URLwww.clinicaltrials.gov. Unique identifierNCT00962364.
Abstract
Using digital twins for decision making is a very promising concept which combines simulation models with corresponding experimental sensor data in order to support maintenance decisions or ...to investigate the reliability. The quality of the prognosis strongly depends on both the data quality and the quality of the digital twin. The latter comprises both the modeling assumptions as well as the correct parameters of these models. This article discusses the challenges when applying this concept to real measurement data for a demonstrator bridge in the lab, including the data management, the iterative development of the simulation model as well as the identification/updating procedure using Bayesian inference with a potentially large number of parameters. The investigated scenarios include both the iterative identification of the structural model parameters as well as scenarios related to a damage identification. In addition, the article aims at providing all models and data in a reproducible way such that other researcher can use this setup to validate their methodologies.
Standard monitoring of heart rate, blood pressure and arterial oxygen saturation during endoscopy is recommended by current guidelines on procedural sedation. A number of studies indicated a ...reduction of hypoxic (art. oxygenation < 90% for > 15 s) and severe hypoxic events (art. oxygenation < 85%) by additional use of capnography. Therefore, U.S. and the European guidelines comment that additional capnography monitoring can be considered in long or deep sedation. Integrated Pulmonary Index® (IPI) is an algorithm-based monitoring parameter that combines oxygenation measured by pulse oximetry (art. oxygenation, heart rate) and ventilation measured by capnography (respiratory rate, apnea > 10 s, partial pressure of end-tidal carbon dioxide PetCO
2
). The aim of this paper was to analyze the value of IPI as parameter to monitor the respiratory status in patients receiving propofol sedation during PEG-procedure. Patients reporting for PEG-placement under sedation were randomized 1:1 in either standard monitoring group (SM) or capnography monitoring group including IPI (IM). Heart rate, blood pressure and arterial oxygen saturation were monitored in SM. In IM additional monitoring was performed measuring PetCO
2
, respiratory rate and IPI. Capnography and IPI values were recorded for all patients but were only visible to the endoscopic team for the IM-group. IPI values range between 1 and 10 (10 = normal; 8–9 = within normal range; 7 = close to normal range, requires attention; 5–6 = requires attention and may require intervention; 3–4 = requires intervention; 1–2 requires immediate intervention). Results on capnography versus standard monitoring of the same study population was published previously. A total of 147 patients (74 in SM and 73 in IM) were included in the present study. Hypoxic events occurred in 62 patients (42%) and severe hypoxic events in 44 patients (29%), respectively. Baseline characteristics were equally distributed in both groups. IPI = 1, IPI < 7 as well as the parameters PetCO
2
= 0 mmHg and apnea > 10 s had a high sensitivity for hypoxic and severe hypoxic events, respectively (IPI = 1: 81%/81% hypoxic/severe hypoxic event, IPI < 7: 82%/88%, PetCO
2
: 69%/68%, apnea > 10 s: 84%/84%). All four parameters had a low specificity for both hypoxic and severe hypoxic events (IPI = 1: 13%/12%, IPI < 7: 7%/7%, PetCO
2
: 29%/27%, apnea > 10 s: 7%/7%). In multivariate analysis, only SM and PetCO
2
= 0 mmHg were independent risk factors for hypoxia. IPI (IPI = 1 and IPI < 7) as well as the individual parameters PetCO
2
= 0 mmHg and apnea > 10 s allow a fast and convenient conclusion on patients’ respiratory status in a morbid patient population. Sensitivity is good for most parameters, but specificity is poor. In conclusion, IPI can be a useful metric to assess respiratory status during propofol-sedation in PEG-placement. However, IPI was not superior to PetCO
2
and apnea > 10 s.
In response to a wide-spread decline in forest vitality associated with acid rain in the 1980s, liming of soils has been implemented in many federal states in Germany to buffer further acid ...deposition and improve availability of nutrients such as calcium and magnesium. As a consequence, it may also increase vitality and depth of fine-root systems and hence improve the drought tolerance of species such as Norway spruce
(L.) Karst., which occurs mostly on acidic forest soils. However, the influence of repeated liming on drought tolerance of trees has never been studied. Here we compared the resistance, recovery and resilience of radial growth in
in relation to drought in limed and control stands and assessed how the dosage and interval between lime application and drought year influences the radial growth response of
. We analyzed radial growth in 198
trees of six experimental sites in south-west Germany. An analysis of the radial increment over the last 30 years allowed the analysis of drought events shortly after the first liming (short-term effect) as well as posterior drought events (mid- to long-term effects). Generalized linear models were developed to assess the influence of drought intensity, site and period since first liming on the drought tolerance of Norway spruce. Regardless of drought intensity, there was no general increase in drought resistance of Norway spruce in response to liming. However, drought resistance of radial growth improved on a loamy site that was additionally treated with wood ash 30 years after the first lime application. Furthermore, recovery and resilience of radial growth after severe drought events were generally better in spruce trees of limed treatments. This indicates a shorter stress period in spruce trees growing on limed soil, which may reduce their susceptibility to secondary, drought-related pests and pathogens.
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