Entwicklung und Validierung der in vivo zeitharmonischen Ultraschall-Elastografie des menschlichen Gehirns für die klinische Anwendung

Motivation: In neurology, the determination of intracranial pressure (ICP) is of central importance for the diagnosis of brain damage. However, reliable ICP measurements are realized by invasive techniques such as lumbar puncture or surgically implanted pressure probes. Cerebral stiffness (CS) measured by elastography could be a parameter sensitive to ICP variations. However, CS is currently measured exclusively by magnetic resonance elastography, which is associated with long examinations and limited availability. Time harmonic shear wave excitation used in magnetic resonance elastography combined with transcranial ultrasound (cerebral THE) can provide reproducible and stable elastograms over a large field-of-view in real-time. Initial applications of cerebral THE in healthy volunteers during performance of the Valsalva maneuver demonstrated sensitivity of CS to blood flow and pressure changes in the brain. The goal of this PhD project was to optimize and validate cerebral THE that I previously developed to quantify CS, identify it as a marker of cerebral perfusion, and provide initial evidence for the potential clinical application of the method as a noninvasive technique for estimating ICP. Methods: To this end, I conducted two studies in healthy volunteers aimed at artificial manipulation of cerebral blood flow: (i) I investigated the effect of hypercapnia during breathing of carbon dioxide-enriched gas and (ii) the effect of dehydration and oral rehydration on CS measured by cerebral THE. Finally, I applied cerebral THE in a pilot clinical study in patients with idiopathic intracranial hypertension (IIH) who underwent lumbar puncture (LP) along with invasive quantification of cerebrospinal fluid (CSF) opening pressure and, if necessary, CSF drainage. Results: Hypercapnia increased CS by 6 ± 4% above baseline. In contrast, dehydration of healthy volunteers resulted in a decrease in CS of 4 ± 2%, whereas CS returned to baseline after oral rehydration. In patients with IIH, CS was 16 ± 5% higher than in healthy volunteers and correlated positively with CSF opening pressure (r = 0:69, p < 0:001). Approximately 30 min after LP, patients’ CS values were within the range of CS values in healthy volunteers. Conclusion: Cerebral THE proved to be a reproducible, stable imaging technique for real-time determination of CS. This project demonstrated that changes in CS are closely associated with changes in cerebral perfusion and ICP. These results suggest that cerebral THE may be a promising noninvasive diagnostic tool for determining ICP in routine clinical practice.Motivation: In der Neurologie ist die Bestimmung des intrakraniellen Drucks (ICP) von zentraler Bedeutung für die Diagnose von Hirnschäden. Zuverlässige ICP-Messungen werden jedoch durch invasive Techniken wie die Lumbalpunktion oder chirurgisch implantierte Drucksonden realisiert. Die mittels Elastografie gemessene zerebrale Steifigkeit (CS) könnte ein Parameter sein, der empfindlich auf ICP-Schwankungen reagiert. Allerdings wird die CS derzeit ausschließlich mit der Magnetresonanz-Elastografie gemessen, die mit langen Untersuchungen und begrenzter Verfügbarkeit verbunden ist. Zeitharmonische Scherwellenanregung, wie sie in der Magnetresonanz-Elastografie verwendet wird, kombiniert mit transkraniellem Ultraschall (zerebrale THE) kann reproduzierbare, stabile Elastogramme über ein großes Sichtfeld in Echtzeit liefern. Erste Anwendungen der zerebralen THE bei gesunden Probanden während der Durchführung des Valsalva-Manövers zeigten, dass die CS empfindlich auf Blutflussund Druckänderungen im Gehirn reagiert. Ziel dieses Promotionsprojekts war die Optimierung und Validierung der zerebralen THE, welche ich zuvor entwickelt habe, um CS zu quantifizieren, als Marker für zerebrale Perfusion zu identifizieren und erste Beweise für die potenzielle klinische Anwendung der Methode als nichtinvasive Technik zur Abschätzung des ICP zu liefern. Methoden: Zu diesem Zweck führte ich zwei Studien an gesunden Probanden durch, welche die künstliche Manipulation des zerebralen Blutflusses zum Ziel hatten: (i) Ich untersuchte die Auswirkung von Hyperkapnie während der Atmung von mit Kohlendioxid angereichertem Gas und (ii) die Auswirkung von Dehydrierung und oraler Rehydrierung auf die durch zerebrale THE gemessene CS. Schließlich habe ich die zerebrale THE in einer klinischen Pilotstudie bei Patienten mit idiopathischer intrakranieller Hypertension (IIH) angewandt, bei denen eine Lumbalpunktion (LP) zusammen mit einer invasiven Quantifizierung des Liquoröffnungsdrucks und, falls erforderlich, einer Liquordrainage durchgeführt wurde. Ergebnisse: Hyperkapnie erhöhte den CS um 6 4% über den Ausgangswert. Im Gegensatz dazu führte die Dehydratation gesunder Probanden zu einem Rückgang des CS um 4 2%, während der CS nach oraler Rehydrierung wieder den Ausgangswert erreichte. Bei Patienten mit IIH war die CS um 16 5% höher als bei gesunden Probanden und korrelierte positiv mit dem Liquoröffnungsdruck (r = 0:69, p < 0:001). Etwa 30 Minuten nach der LP lagen die CS Werte der Patienten im Bereich der CS Werte gesunder Probanden. Schlussfolgerung: Die zerebrale THE erwies sich als reproduzierbares, stabiles bildgebendes Verfahren zur Echtzeit-Bestimmung der CS. Dieses Projekt zeigte, dass Änderungen des CS eng mit Änderungen der zerebralen Perfusion und des ICP verbunden sind. Diese Ergebnisse deuten darauf hin, dass die zerebrale THE ein vielversprechendes nichtinvasives Diagnoseinstrument zur Bestimmung des ICP in der klinischen Routinepraxis sein könnte

Cerebral Ultrasound Time-Harmonic Elastography Reveals Softening of the Human Brain Due to Dehydration

Hydration influences blood volume, blood viscosity, and water content in soft tissues - variables that determine the biophysical properties of biological tissues including their stiffness. In the brain, the relationship between hydration and stiffness is largely unknown despite the increasing importance of stiffness as a quantitative imaging marker. In this study, we investigated cerebral stiffness (CS) in 12 healthy volunteers using ultrasound time-harmonic elastography (THE) in different hydration states: (i) during normal hydration, (ii) after overnight fasting, and (iii) within 1 h of drinking 12 ml of water per kg body weight. In addition, we correlated shear wave speed (SWS) with urine osmolality and hematocrit. SWS at normal hydration was 1.64 ± 0.02 m/s and decreased to 1.57 ± 0.04 m/s (p < 0.001) after overnight fasting. SWS increased again to 1.63 ± 0.01 m/s within 30 min of water drinking, returning to values measured during normal hydration (p = 0.85). Urine osmolality at normal hydration (324 ± 148 mOsm/kg) increased to 784 ± 107 mOsm/kg (p < 0.001) after fasting and returned to normal (288 ± 128 mOsm/kg, p = 0.83) after water drinking. SWS and urine osmolality correlated linearly (r = -0.68, p < 0.001), while SWS and hematocrit did not correlate (p = 0.31). Our results suggest that mild dehydration in the range of diurnal fluctuations is associated with significant softening of brain tissue, possibly due to reduced cerebral perfusion. To ensure consistency of results, it is important that cerebral elastography with a standardized protocol is performed during normal hydration

Cerebral tomoelastography based on multifrequency MR elastography in two and three dimensions

Magnetic resonance elastography (MRE) generates quantitative maps of the mechanical properties of biological soft tissues. However, published values obtained by brain MRE vary largely and lack detail resolution, due to either true biological effects or technical challenges. We here introduce cerebral tomoelastography in two and three dimensions for improved data consistency and detail resolution while considering aging, brain parenchymal fraction (BPF), systolic blood pressure, and body-mass-index. Multifrequency MRE with 2D- and 3D-tomoelastography postprocessing was applied to the brains of 31 volunteers (age range: 22-61 years) for analyzing the coefficient of variation (CV) and effects of biological factors. Eleven volunteers were rescanned after one day and one year to determine intraclass correlation coefficient (ICC) and identify possible long-term changes. White matter shear-wave-speed (SWS) was slightly higher in 2D-MRE (1.28±0.02m/s) than 3D-MRE (1.22±0.05m/s, p<0.0001), with less variation after one day in 2D (0.33±0.32%) than in 3D (0.96±0.66%, p=0.004), which was also reflected in a slightly lower CV and higher ICC in 2D (1.84%, 0.97 [0.88-0.99]) than in 3D (3.89%, 0.95 [0.76-0.99]). Remarkably, 3D-MRE was sensitive to a decrease in white matter SWS within only one year, whereas no change in white matter volume was observed during this follow-up period. Across volunteers, stiffness correlated with age and BPF, but not with blood pressure and body-mass-index. Cerebral tomoelastography provides high-resolution viscoelasticity maps with excellent consistency. Brain MRE in 2D shows less variation across volunteers in shorter scan times than 3D-MRE, while 3D-MRE appears to be more sensitive to subtle biological effects such as aging

Functional losses in ground spider communities due to habitat structure degradation under tropical land-use change

Deforestation and land‐use change in tropical regions result in habitat loss and extinction of species that are unable to adapt to the conditions in agricultural landscapes. If the associated loss of functional diversity is not compensated by species colonizing the converted habitats, extinctions might be followed by a reduction or loss of ecosystem functions including biological control. To date, little is known about how land‐use change in the tropics alters the functional diversity of invertebrate predators and which key environmental factors may mitigate the decline in functional diversity and predation in litter and soil communities. We applied litter sieving and heat extraction to study ground spider communities and assessed structural characteristics of vegetation and parameters of litter in rainforest and agricultural land‐use systems (jungle rubber, rubber, and oil palm monocultures) in a Southeast Asian hotspot of rainforest conversion: Sumatra, Indonesia. We found that (1) spider density, species richness, functional diversity, and community predation (energy flux to spiders) were reduced by 57–98% from rainforest to oil palm monoculture; (2) jungle rubber and rubber monoculture sustained relatively high diversity and predation in ground spiders, but small cryptic spider species strongly declined; (3) high species turnover compensated losses of some functional trait combinations, but did not compensate for the overall loss of functional diversity and predation per unit area; (4) spider diversity was related to habitat structure such as amount of litter, understory density, and understory height, while spider predation was better explained by plant diversity. Management practices that increase habitat‐structural complexity and plant diversity such as mulching, reduced weeding, and intercropping monocultures with other plants may contribute to maintaining functional diversity of and predation services provided by ground invertebrate communities in plantations

Stiffness pulsation of the human brain detected by non-invasive time-harmonic elastography

Introduction: Cerebral pulsation is a vital aspect of cerebral hemodynamics. Changes in arterial pressure in response to cardiac pulsation cause cerebral pulsation, which is related to cerebrovascular compliance and cerebral blood perfusion. Cerebrovascular compliance and blood perfusion influence the mechanical properties of the brain, causing pulsation-induced changes in cerebral stiffness. However, there is currently no imaging technique available that can directly quantify the pulsation of brain stiffness in real time.Methods: Therefore, we developed non-invasive ultrasound time-harmonic elastography (THE) technique for the real-time detection of brain stiffness pulsation. We used state-of-the-art plane-wave imaging for interleaved acquisitions of shear waves at a frequency of 60 Hz to measure stiffness and color flow imaging to measure cerebral blood flow within the middle cerebral artery. In the second experiment, we used cost-effective lineby-line B-mode imaging to measure the same mechanical parameters without flow imaging to facilitate future translation to the clinic.Results: In 10 healthy volunteers, stiffness increased during the passage of the arterial pulse wave from 4.8% ± 1.8% in the temporal parenchyma to 11% ± 5% in the basal cisterns and 13% ± 9% in the brain stem. Brain stiffness peaked in synchrony with cerebral blood flow at approximately 180 ± 30 ms after the cardiac R-wave. Line-by-line THE provided the same stiffness values with similar time resolution as high-end plane-wave THE, demonstrating the robustness of brain stiffness pulsation as an imaging marker.Discussion: Overall, this study sets the background and provides reference values for time-resolved THE in the human brain as a cost-efficient and easy-touse mechanical biomarker associated with cerebrovascular compliance

TRY plant trait database – enhanced coverage and open access

Plant traits - the morphological, anatomical, physiological, biochemical and phenological characteristics of plants - determine how plants respond to environmental factors, affect other trophic levels, and influence ecosystem properties and their benefits and detriments to people. Plant trait data thus represent the basis for a vast area of research spanning from evolutionary biology, community and functional ecology, to biodiversity conservation, ecosystem and landscape management, restoration, biogeography and earth system modelling. Since its foundation in 2007, the TRY database of plant traits has grown continuously. It now provides unprecedented data coverage under an open access data policy and is the main plant trait database used by the research community worldwide. Increasingly, the TRY database also supports new frontiers of trait‐based plant research, including the identification of data gaps and the subsequent mobilization or measurement of new data. To support this development, in this article we evaluate the extent of the trait data compiled in TRY and analyse emerging patterns of data coverage and representativeness. Best species coverage is achieved for categorical traits - almost complete coverage for ‘plant growth form’. However, most traits relevant for ecology and vegetation modelling are characterized by continuous intraspecific variation and trait–environmental relationships. These traits have to be measured on individual plants in their respective environment. Despite unprecedented data coverage, we observe a humbling lack of completeness and representativeness of these continuous traits in many aspects. We, therefore, conclude that reducing data gaps and biases in the TRY database remains a key challenge and requires a coordinated approach to data mobilization and trait measurements. This can only be achieved in collaboration with other initiatives

Recombinant plasmids capable to replication in B. subtilis and E. coli

The plasmid pBC16 (4.25 kbases), ongtnally isolated from Bacillus cereus, determines tetracycline resistance and can be transformed into competent cells of B. subtilis. A miniplasmid of pBCl6 (pBCI6-1), 2,7 kb) which has lost an EcoRI fragment of pBCI6 retains the replication functions and the tetracycline resistance. This plasmid which carries only one EcoRI site has been joined in vitro to pBS], a cryptic plasmid previously isolated from B. subtilis and shown to carry also a single EcoRI site (Bernhard et aI., 1978). The recombinant plasmid is unstable and dissociates into the plasmid pBSl61 (8.2 kb) and the smaller plasmid pBS162 (2. I kb). Plasmid pBS161 retains the tetracycline resistance. It possesses a single EcoRI site and 6 HindlII sites. The largest HindIII fragment of pBS161 carries the tetracycline resistance gene and the replication function. After circularization in vitro of this fragment a new plasmid, pBS161-l is generated, which can be used as a HindlII and EcoRI cloning vector in Bacillus suhtilis. Hybrid plasmids consisting of the E. coli plasmids pBR322, p WL 7 or pACl84 and different HindlII fragments of pBSI61 were constructed in vitro. Hybrids containing together with the E. coli plasmid the largest HindlII fragment of pBS161 can replicate in E. coli and B. sublilis. In E. coli only the replicon of the E. coli plasmid part is functioning whereas in B. suhtilis replication of the hybrid plasmid is under the control of the Bacillus replicon. The tetracycline resistance of the B. subtilis plasmid is expressed in E. coli, but several antibiotic resistances of the E. coli plasmids (ampicillin, kanamycin and chloramphenicol) are not expressed in B. suhtilis. The hybrid plasmids seem to be more unstable in B. subtilis than in E. coli

Specificity of <i>O</i>-demethylation in extracts of the homoacetogenic <i>Holophaga foetida</i> and demethylation kinetics measured by a coupled photometric assay

The kinetics and specificity of O-demethylation were studied in cell-free extracts of the strictly anaerobic, methanethiol- and dimethylsulfide-producing homoacetogen Holophaga foetida strain TMBS4 with methanethiol and tetrahydrofolate (H4folate) as methyl acceptors. Extracts of cells grown with 3,4,5-trimethoxybenzoate contained an enzyme system that demethylated various phenyl methyl ethers with at least one ortho-positioned hydroxyl or methoxyl group (the ortho system) and also contained a decarboxylase. Extracts of cells grown with 3,5-dihydroxyanisole contained an enzyme system with a novel specificity that demethylated only the metahydroxylated compounds 3,5-dihydroxyanisole and 3-hydroxyanisole (the meta system) and lacked a decarboxylase. H4folate-dependent demethylation produced CH3-H4folate. For a photometric in vitro assay of the meta system, the NADPH-consuming phloroglucinol reductase (PR) reaction was coupled to the phloroglucinol-yielding demethylation of 3,5-dihydroxyanisole. The kinetics of the indicator enzyme PR were studied. The cell extract had a high and stable specific PR activity. PR was inhibited by phloroglucinol (substrate inhibition) and the substrate analogue 3,5-dihydroxyanisole. Doubling the PR activity of the coupled enzyme assay by additions of a PR-enriched fraction had no effect, showing that the PR activity supplied by cell extract did not limit reaction rates. Demethylation activity of the meta system with either methyl acceptor increased with the square of the protein concentration. With H4folate, the in vivo activity could be attained. Kinetic parameters for the methyl acceptors were determined