15 research outputs found
Interaction of the Membrane-bound GlnK-AmtB Complex with the Master Regulator of Nitrogen Metabolism TnrA inBacillus subtilis
No full textP-II proteins are widespread and highly conserved signal transduction proteins occurring in bacteria, Archaea, and plants and play pivotal roles in controlling nitrogen assimilatory metabolism. This study reports on biochemical properties of the P-II-homologue GlnK (originally termed NrgB) in Bacillus subtilis (BsGlnK). Like other P-II proteins, the native BsGlnK protein has a trimeric structure and readily binds ATP in the absence of divalent cations, whereas 2-oxoglutarate is only weakly bound. In contrast to other P-II-like proteins, Mg2+ severely affects its ATP-binding properties. BsGlnK forms a tight complex with the membrane-bound ammonium transporter AmtB (NrgA), from which it can be relieved by millimolar concentrations of ATP. Immunoprecipitation and co-localization experiments identified a novel interaction between the BsGlnK-AmtB complex and the major transcription factor of nitrogen metabolism, TnrA. In vitro in the absence of ATP, TnrA is completely tethered to membrane (AmtB)-bound GlnK, whereas in extracts from BsGlnK- or AmtB-deficient cells, TnrA is entirely soluble. The presence of 4 mM ATP leads to concomitant solubilization of BsGlnK and TnrA. This ATP-dependent membrane re-localization of TnrA by BsGlnK/AmtB may present a novel mechanism to control the global nitrogen-responsive transcription regulator TnrA in B. subtilis under certain physiological conditions
NEW GRADING DEVICE WITH NOVEL STRUCTURES INCREASES SELFGRADING SUCCESS OF WHITELEG SHRIMPS L. VANNAMEI IN AQUACULTURE CULTIVATION SYSTEMS
No full textUneven growth of aquacultured shrimp leads to an increase in cannibalism, untargeted feeding and uneven product size. Size grading can prevent these effects. However, current sorting methods are highly invasive and cause
high stress levels and flight responses. The animals can injure themselves and may even jump out of the tank.
In this study, a size grading device was developed that allows the animals to sort themselves voluntarily according to size in the cultivation tank. For this purpose, innovative structures were developed that are specifically adapted
to the body shape and behaviour of the animals. These structures are conceptualised to either encourage or discourage passing of the grading device. Results demonstrate strong grading effects (75 %) on the
distribution of shrimps between different tank compartments and the ability to separate shrimps of different size voluntarily (up to 85%). Small animals
can be separated from the cohort without stress or any personnel effort and uniform size distribution of the shrimps can be guaranteed. This voluntary grading system is expected to increase animal welfare and growth performance in shrimp farms while it reduces labour
3D Printing of Piezoelectric Barium Titanate-Hydroxyapatite Scaffolds with Interconnected Porosity for Bone Tissue Engineering
No full textThe prevalence of large bone defects is still a major problem in surgical clinics. It is, thus, not a surprise that bone-related research, especially in the field of bone tissue engineering, is a major issue in medical research. Researchers worldwide are searching for the missing link in engineering bone graft materials that mimic bones, and foster osteogenesis and bone remodeling. One approach is the combination of additive manufacturing technology with smart and additionally electrically active biomaterials. In this study, we performed a three-dimensional (3D) printing process to fabricate piezoelectric, porous barium titanate (BaTiO3) and hydroxyapatite (HA) composite scaffolds. The printed scaffolds indicate good cytocompatibility and cell attachment as well as bone mimicking piezoelectric properties with a piezoelectric constant of 3 pC/N. This work represents a promising first approach to creating an implant material with improved bone regenerating potential, in combination with an interconnected porous network and a microporosity, known to enhance bone growth and vascularization
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3D Printing of Piezoelectric Barium Titanate-Hydroxyapatite Scaffolds with Interconnected Porosity for Bone Tissue Engineering
No full textThe prevalence of large bone defects is still a major problem in surgical clinics. It is, thus, not a surprise that bone-related research, especially in the field of bone tissue engineering, is a major issue in medical research. Researchers worldwide are searching for the missing link in engineering bone graft materials that mimic bones, and foster osteogenesis and bone remodeling. One approach is the combination of additive manufacturing technology with smart and additionally electrically active biomaterials. In this study, we performed a three-dimensional (3D) printing process to fabricate piezoelectric, porous barium titanate (BaTiO3) and hydroxyapatite (HA) composite scaffolds. The printed scaffolds indicate good cytocompatibility and cell attachment as well as bone mimicking piezoelectric properties with a piezoelectric constant of 3 pC/N. This work represents a promising first approach to creating an implant material with improved bone regenerating potential, in combination with an interconnected porous network and a microporosity, known to enhance bone growth and vascularization
Bone Morphogenetic Protein-7 Enhances Degradation of Osteoinductive Bioceramic Implants in an Ectopic Model
No full textBackground:
The aim of the present study was to evaluate the degradation pattern of highly porous bioceramics as well as the bone formation in presence of bone morphogenetic protein 7 (BMP-7) in an ectopic site.
Methods:
Novel calcium phosphate ceramic cylinders sintered at 1,300ºC with a total porosity of 92–94 vol%, 45 pores per inch, and sized 15 mm (Ø) × 5 mm were grafted on the musculus latissimus dorsi bilaterally in 10 Göttingen minipigs: group I (n = 5):
hydroxyapatite (HA) versus biphasic calcium phosphate (BCP), a mixture of HA and tricalcium phosphate (TCP) in a ratio of 60/40 wt%; group II (n = 5): TCP versus
BCP. A test side was supplied in situ with 250 μg BMP-7. Fluorochrome bone labeling
and computed tomography were performed in vivo. Specimens were evaluated 14
weeks after surgery by environmental scanning electron microscopy, fluorescence
microscopy, tartrate-resistant acid phosphatase, and pentachrome staining.
Results:
Bone formation was enhanced in the presence of BMP-7 in all ceramics (P = 0.001).
Small spots of newly formed bone were observed in all implants in the absence of BMP-7.
Degradation of HA and BCP was enhanced in the presence of BMP-7 (P = 0.001). In those
ceramics, osteoclasts were observed. TCP ceramics were almost completely degraded independently
of the effect of BMP-7 after 14 weeks (P = 0.76), osteoclasts were not observed
BMP-7 Preserves Surface Integrity of Degradable-ceramic Cranioplasty in a Göttingen Minipig Model
No full textBackground:
The aim of the study was to evaluate the integrity of a craniotomy grafted site in a minipig model using different highly porous calcium phosphate ceramic scaffolds either loaded or nonloaded with bone morphogenetic protein-7 (BMP-7).
Methods:
Four craniotomies with a diameter of 15 mm (critical-size defect) were grafted with different highly porous (92–94 vol%) calcium phosphate ceramics [hydroxyapatite (HA), tricalcium phosphate (TCP), and biphasic calcium phosphate (BCP; a mixture of HA and TCP)] in 10 Göttingen minipigs: (a) group I (n = 5): HA versus BCP; (b) group II (n = 5): TCP versus BCP. One scaffold of each composition was supplied with 250 μg of BMP-7. In vivo computed tomography scan and fluorochrome bone labeling were performed. Specimens were evaluated 14 weeks after surgery by environmental scanning electron microscopy, fluorescence microscopy, and Giemsa staining histology.
Results:
BMP-7 significantly enhanced bone formation in TCP (P = 0.047). Slightly enhanced bone formation was observed in BCP (P = 0.059) but not in HA implants. BMP-7 enhanced ceramic degradation in TCP (P = 0.05) and BCP (P = 0.05) implants but not in HA implants. Surface integrity of grafted site was observed in all BMP-7-loaded implants after successful creeping substitution by the newly formed bone. In 9 of 10 HA implants without BMP-7, partial collapse of the implant site was observed. All TCP implants without BMP-7 collapsed. Fluorescent labeling showed bone formation at week 1 in BMP-7-stimulated implants.
Conclusions:
BMP-7 supports bone formation, ceramic degradation, implant integration, and surface integrity of the grafted site
Nitrogen regulation of protein–protein interactions and transcript levels of GlnK PII regulator and AmtB ammonium transporter homologs in Archaea
No full textGene homologs of GlnK PII regulators and AmtB-type ammonium transporters are often paired on prokaryotic genomes, suggesting these proteins share an ancient functional relationship. Here, we demonstrate for the first time in Archaea that GlnK associates with AmtB in membrane fractions after ammonium shock, thus, providing a further insight into GlnK-AmtB as an ancient nitrogen sensor pair. For this work, Haloferax mediterranei was advanced for study through the generation of a pyrE2-based counterselection system that was used for targeted gene deletion and expression of Flag-tagged proteins from their native promoters. AmtB1-Flag was detected in membrane fractions of cells grown on nitrate and was found to coimmunoprecipitate with GlnK after ammonium shock. Thus, in analogy to bacteria, the archaeal GlnK PII may block the AmtB1 ammonium transporter under nitrogen-rich conditions. In addition to this regulated protein–protein interaction, the archaeal amtB-glnK gene pairs were found to be highly regulated by nitrogen availability with transcript levels high under conditions of nitrogen limitation and low during nitrogen excess. While transcript levels of glnK-amtB are similarly regulated by nitrogen availability in bacteria, transcriptional regulators of the bacterial glnK promoter including activation by the two-component signal transduction proteins NtrC (GlnG, NRI) and NtrB (GlnL, NRII) and sigma factor σN (σ54) are not conserved in archaea suggesting a novel mechanism of transcriptional control
Electrically Conductive and 3D-Printable Oxidized Alginate-Gelatin Polypyrrole : PSS Hydrogels for Tissue Engineering
No full textElectroactive hydrogels can be used to influence cell response and maturation by electrical stimulation. However, hydrogel formulations which are 3D printable, electroactive, cytocompatible, and allow cell adhesion, remain a challenge in the design of such stimuli-responsive biomaterials for tissue engineering. Here, a combination of pyrrole with a high gelatin-content oxidized alginate-gelatin (ADA-GEL) hydrogel is reported, offering 3D-printability of hydrogel precursors to prepare cytocompatible and electrically conductive hydrogel scaffolds. By oxidation of pyrrole, electroactive polypyrrole:polystyrenesulfonate (PPy:PSS) is synthesized inside the ADA-GEL matrix. The hydrogels are assessed regarding their electrical/mechanical properties, 3D-printability, and cytocompatibility. It is possible to prepare open-porous scaffolds via bioplotting which are electrically conductive and have a higher cell seeding efficiency in scaffold depth in comparison to flat 2D hydrogels, which is confirmed via multiphoton fluorescence microscopy. The formation of an interpenetrating polypyrrole matrix in the hydrogel matrix increases the conductivity and stiffness of the hydrogels, maintaining the capacity of the gels to promote cell adhesion and proliferation. The results demonstrate that a 3D-printable ADA-GEL can be rendered conductive (ADA-GEL-PPy:PSS), and that such hydrogel formulations have promise for cell therapies, in vitro cell culture, and electrical-stimulation assisted tissue engineering
Encapsulation of Mesenchymal Stem Cells Improves Vascularization of Alginate-Based Scaffolds
No full textNew Insights in the Occurrence of Venous Thromboembolism in Critically Ill Patients with COVID-19—A Large Postmortem and Clinical Analysis
No full textCritically ill COVID-19 patients are at high risk for venous thromboembolism (VTE), namely deep vein thrombosis (DVT) and/or pulmonary embolism (PE), and death. The optimal anticoagulation strategy in critically ill patients with COVID-19 remains unknown. This study investigated the ante mortem incidence as well as postmortem prevalence of VTE, the factors predictive of VTE, and the impact of changed anticoagulation practice on patient survival. We conducted a consecutive retrospective analysis of postmortem COVID-19 (n = 64) and non-COVID-19 (n = 67) patients, as well as ante mortem COVID-19 (n = 170) patients admitted to the University Medical Center Hamburg-Eppendorf (Hamburg, Germany). Baseline patient characteristics, parameters related to the intensive care unit (ICU) stay, and the clinical and autoptic presence of VTE were evaluated and statistically compared between groups. The occurrence of VTE in critically ill COVID-19 patients is confirmed in both ante mortem (17%) and postmortem (38%) cohorts. Accordingly, comparing the postmortem prevalence of VTE between age- and sex-matched COVID-19 (43%) and non-COVID-19 (0%) cohorts, we found the statistically significant increased prevalence of VTE in critically ill COVID-19 cohorts (p = 0.001). A change in anticoagulation practice was associated with the statistically significant prolongation of survival time (HR: 2.55, [95% CI 1.41–4.61], p = 0.01) and a reduction in VTE occurrence (54% vs. 25%; p = 0.02). In summary, in the autopsy as well as clinical cohort of critically ill patients with COVID-19, we found that VTE was a frequent finding. A change in anticoagulation practice was associated with a statistically significantly prolonged survival time
