Biochemistry and functional aspects of human glandular kallikreins

Human urinary kallikrein was purified by gel filtration on Sephacryl S-200 and affinity chromatography on aprotinin-Sepharose, followed by ion exchange chromatography on DEAE-Sepharose. In dodecylsulfate gel electrophoresis two protein bands with molecular weights of 41,000 and 34,000 were separated. The amino acid composition and the carbohydrate content of the kallikrein preparation were determined; isoleucine was identified as the only aminoterminal amino acid. The bimolecular velocity constant for the inhibition by diisopropyl fluorophosphate was determined as 9±2 l mol–1 min–1. The hydrolysis of a number of substrates was investigated and AcPheArgOEt was found to be the most sensitive substrate for human urinary kallikrein. Using this substrate an assay method for kallikrein in human urine was developed. It was shown by radioimmunoassay that pig pancreatic kallikrein can be absorbed in the rat intestinal tract. Furthermore, in dogs the renal excretion of glandular kallikrein from blood was demonstrated by radioimmunological methods

Introduction of a guideline for measurements of greenhouse gas fluxes from soils using non-steady-state chambers

Method Soils represent a major global source and sink of greenhouse gases (GHGs). Many studies of GHG fluxes between soil, plant and atmosphere rely on chamber measurements. Different chamber techniques have been developed over the last decades, each characterised by different requirements and limitations. In this manuscript, we focus on the non-steady-state technique which is widely used for manual measurements but also in automatic systems. Although the measurement method appears very simple, experience gained over the years shows that there are many details which have to be taken into account to obtain reliable measurement results. Aim This manuscript aims to share lessons learnt and pass on experiences in order to assist the reader with possible questions or unexpected challenges, ranging from the planning of the design of studies and chambers to the practical handling of the chambers and the quality assurance of the gas and data analysis. This concise introduction refers to a more extensive Best Practice Guideline initiated by the Working Group Soil Gases (AG Bodengase) of the German Soil Science Society (Deutsche Bodenkundliche Gesellschaft). The intention was to collect and aggregate the expertise of different working groups in the research field. As a compendium, this Best Practice Guideline is intended to help both beginners and experts to meet the practical and theoretical challenges of measuring soil gas fluxes with non-steady-state chamber systems and to improve the quality of the individual flux measurements and thus entire GHG studies by reducing sources of uncertainty and error

In-vitro hemolysis and its financial impact using different blood collection systems

Background: Hemolytic specimens are among the most challenging preanalytical issues in laboratory diagnostics. The type of blood collection tube in use is claimed to influence in vitro hemolysis. We aimed to examine this hypothesis and estimate the respective financial impact, evaluating routine blood samples from the past 4 years. Methods: A total of 47,820 hemolysis index (HI) values from five different time intervals (IV1-IV5) were compared against each other, representing the following tubes: IV1-Sarstedt Monovette; IV2-8 mL/16×100 mm Greiner BioOne (GBO) Vacuette; IV3/IV4-5 mL/16×100 mm GBO Vacuette; IV5-4.5 mL/13×75 mm GBO Vacuette. For estimation of the economic impact, material, personnel and analytical costs were calculated. Results: HI mean values in time interval IV2 were significantly higher than in all other intervals, while mean values amongst all other intervals were comparable. The number of moderately and severely hemolyzed samples increased with incrementing vacuum. Overall comparable costs between intervals IV1 and IV5 were €11,370, €14,045, €12,710, €11,213 and €8138 per 10,000 samples, respectively. Conclusions: Aspiration tubes and low vacuum tubes show comparable hemolysis rates. Increasing vacuum levels are associated with higher hemolysis rates. By decreasing in vitro hemolysis, financial savings up to €5907 per 10,000 samples could be gained

First-in-Human Phase I Study of MP0250, a First-in-Class DARPin Drug Candidate Targeting VEGF and HGF, in Patients With Advanced Solid Tumors.

PURPOSE: A first-in-human study was performed with MP0250, a DARPin drug candidate. MP0250 specifically inhibits both vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) with the aim of disrupting the tumor microenvironment. PATIENTS AND METHODS: A multicenter, open-label, repeated-dose, phase I study was conducted to assess the safety, tolerability, and pharmacokinetics of MP0250 in 45 patients with advanced solid tumors. In the dose-escalation part, 24 patients received MP0250 as a 3-hour infusion once every 2 weeks at five different dose levels (0.5-12 mg/kg). Once the maximum tolerated dose (MTD) was established, 21 patients were treated with a 1-hour infusion (n = 13, 8 mg/kg, once every 2 weeks and n = 8, 12 mg/kg, once every 3 weeks) of MP0250 in the dose confirmation cohorts. RESULTS: In the dose-escalation cohort, patients treated with 12 mg/kg MP0250 once every 2 weeks experienced dose-limiting toxicities. Therefore, MTD was 8 mg/kg once every 2 weeks or 12 mg/kg once every 3 weeks. The most common adverse events (AEs) were hypertension (69%), proteinuria (51%), and diarrhea and nausea (both 36%); hypoalbuminemia was reported in 24% of patients. Most AEs were consistent with inhibition of the VEGF and HGF pathways. Exposure was dose-proportional and sustained throughout the dosing period for all patients (up to 15 months). The half-life was about 2 weeks. Signs of single-agent antitumor activity were observed: 1 unconfirmed partial response with a time to progression of 23 weeks and 24 patients with stable disease, with the longest duration of 72 weeks and a median duration of 18 weeks. CONCLUSION: MP0250 is a first-in-class DARPin drug candidate with suitable tolerability and appropriate pharmacokinetic properties for further development in combination with other anticancer therapies

High mobility group box 1 (HMGB1) acts as an “alarmin” to promote acute myeloid leukaemia progression

High mobility group box 1 (HMGB1) is a non-histone protein localised in the cell nucleus, where it interacts with DNA and promotes nuclear transcription events. HMGB1 levels are elevated during acute myeloid leukaemia (AML) progression followed by participation of this protein in triggering signalling events in target cells as a pro-inflammatory stimulus. This mechanism was hypothesised to be employed as a survival pathway by malignant blood cells and our aims were therefore to test this hypothesis experimentally. Here we report that HMGB1 triggers the release of tumour necrosis factor alpha (TNF-?) by primary human AML cells. TNF-? induces interleukin 1 beta (IL-1?) production by healthy leukocytes, leading to IL-1?-induced secretion of stem cell factor (SCF) by competent cells (for example endothelial cells). These results were verified in mouse bone marrow and primary human AML blood plasma samples. In addition, HMGB1 was found to induce secretion of angiogenic vascular endothelial growth factor (VEGF) and this process was dependent on the immune receptor Tim-3. We therefore conclude that HMGB1 is critical for AML progression as a ligand of Tim-3 and other immune receptors thus supporting survival/proliferation of AML cells and possibly the process of angiogenesis

Highly specific targeting of human acute myeloid leukaemia cells using pharmacologically active nanoconjugates

In this study we used 5 nm gold nanoparticles as delivery platforms to target cancer cells expressing the immune receptor Tim-3 using single chain antibodies. Gold surfaces were also covered with the cytotoxic drug rapamycin which was immobilised using a glutathione linker. These nanoconjugates allowed highly specific and efficient delivery of cytotoxic rapamycin into human malignant blood cells

The use of the Airtraq® optical laryngoscope for routine tracheal intubation in high-risk cardio-surgical patients

<p>Abstract</p> <p>Background</p> <p>The Airtraq^®optical laryngoscope (Prodol Ltd., Vizcaya, Spain) is a novel disposable device facilitating tracheal intubation in routine and difficult airway patients. No data investigating routine tracheal intubation using the Airtaq^®in patients at a high cardiac risk are available at present. Purpose of this study was to investigate the feasibility and hemodynamic implications of tracheal intubation with the Aitraq^®optical laryngoscope, in high-risk cardio-surgical patients.</p> <p>Methods</p> <p>123 consecutive ASA III patients undergoing elective coronary artery bypass grafting were routinely intubated with the Airtraq^®laryngoscope. Induction of anesthesia was standardized according to our institutional protocol. All tracheal intubations were performed by six anesthetists trained in the use of the Airtraq^®prior.</p> <p>Results</p> <p>Overall success rate was 100% (n = 123). All but five patients trachea could be intubated in the first attempt (95,9%). 5 patients were intubated in a 2nd (n = 4) or 3rd (n = 1) attempt. Mean intubation time was 24.3 s (range 16-128 s). Heart rate, arterial blood pressure and SpO₂were not significantly altered. Minor complications were observed in 6 patients (4,8%), i.e. two lesions of the lips and four minor superficial mucosal bleedings. Intubation duration (p = 0.62) and number of attempts (p = 0.26) were independent from BMI and Mallampati score.</p> <p>Conclusion</p> <p>Tracheal intubation with the Airtraq^®optical laryngoscope was feasible, save and easy to perform in high-risk patients undergoing cardiac surgery. In all patients, a sufficient view on the vocal cords could be obtained, independent from BMI and preoperative Mallampati score.</p> <p>Trial Registration</p> <p>DRKS 00003230</p

Methane-carbon flow into the benthic food web at cold seeps – a case study from the Costa Rica subduction zone

Cold seep ecosystems can support enormous biomasses of free-living and symbiotic chemoautotrophic organisms that get their energy from the oxidation of methane or sulfide. Most of this biomass derives from animals that are associated with bacterial symbionts, which are able to metabolize the chemical resources provided by the seeping fluids. Often these systems also harbor dense accumulations of non-symbiotic megafauna, which can be relevant in exporting chemosynthetically fixed carbon from seeps to the surrounding deep sea. Here we investigated the carbon sources of lithodid crabs (Paralomis sp.) feeding on thiotrophic bacterial mats at an active mud volcano at the Costa Rica subduction zone. To evaluate the dietary carbon source of the crabs, we compared the microbial community in stomach contents with surface sediments covered by microbial mats. The stomach content analyses revealed a dominance of epsilonproteobacterial 16S rRNA gene sequences related to the free-living and epibiotic sulfur oxidiser Sulfurovum sp. We also found Sulfurovum sp. as well as members of the genera Arcobacter and Sulfurimonas in mat-covered surface sediments where Epsilonproteobacteria were highly abundant constituting 10% of total cells. Furthermore, we detected substantial amounts of bacterial fatty acids such as i-C15:0 and C17:1ω6c with stable carbon isotope compositions as low as −53‰ in the stomach and muscle tissue. These results indicate that the white microbial mats at Mound 12 are comprised of Epsilonproteobacteria and that microbial mat-derived carbon provides an important contribution to the crab's nutrition. In addition, our lipid analyses also suggest that the crabs feed on other 13C-depleted organic matter sources, possibly symbiotic megafauna as well as on photosynthetic carbon sources such as sedimentary detritus