International registry on the use of the CytoSorb® adsorber in ICU patients : Study protocol and preliminary results.

INTRODUCTION: The aim of this clinical registry is to record the use of CytoSorb® adsorber device in critically ill patients under real-life conditions. METHODS: The registry records all relevant information in the course of product use, e. g., diagnosis, comorbidities, course of the condition, treatment, concomitant medication, clinical laboratory parameters, and outcome (ClinicalTrials.gov Identifier: NCT02312024). Primary endpoint is in-hospital mortality as compared to the mortality predicted by the APACHE II and SAPS II score, respectively. RESULTS: As of January 30, 2017, 130 centers from 22 countries were participating. Data available from the start of the registry on May 18, 2015 to November 24, 2016 (122 centers; 22 countries) were analyzed, of whom 20 centers from four countries provided data for a total of 198 patients (mean age 60.3 ± 15.1 years, 135 men [68.2%]). In all, 192 (97.0%) had 1 to 5 Cytosorb® adsorber applications. Sepsis was the most common indication for CytoSorb® treatment (135 patients). Mean APACHE II score in this group was 33.1 ± 8.4 [range 15-52] with a predicted risk of death of 78%, whereas the observed mortality was 65%. There were no significant decreases in the SOFA scores after treatment (17.2 ± 4.8 [3-24]). However interleukin-6 levels were markedly reduced after treatment (median 5000 pg/ml before and 289 pg/ml after treatment, respectively). CONCLUSIONS: This third interim report demonstrates the feasibility of the registry with excellent data quality and completeness from 20 study centers. The results must be interpreted with caution, since the numbers are still small; however the disease severity is remarkably high and suggests that adsorber treatment might be used as an ultimate treatment in life-threatening situations. There were no device-associated side effects.

Hypercalciuria in osteogenesis imperfecta type I.

BACKGROUND: In Osteogenesis Imperfecta severity of disease and reduced physical activity have been considered the main factors contributing to hypercalciuria; however, its pathogenesis in Osteogenesis Imperfecta Type I, in which mobility is normal, is still unclear. PATIENT, METHODS AND RESULTS: We describe a patient with Osteogenesis Imperfecta Type I and hypercalciuria, in whom measurement of calcium intake, plasma 1 - 25(OH) (2) Vitamin D, fasting calciuria and tubular proteinuria led us to exclude an absorptive or renal component in the pathogenesis of hypercalciuria. CONCLUSIONS: We believe that hypercalciuria is determined by bone disease in Osteogenesis Imperfecta Type I. This condition should be added to the causes of normocalcemic hypercalciuria in children and the mildest forms should be differentiated from Idiopathic Hypercalciuria.

The thermosome: alternating alpha and beta-subunits within the chaperonin of the archaeon Thermoplasma acidophilum.

The thermosome of the archaeon Thermoplasma acidophilum is composed of two subunits, alpha and beta, which are arranged in two stacked, eight-membered rings. Electron cryo-microscopy in conjunction with image analysis revealed a 4-fold symmetry in the heterooligomeric alpha + beta thermosome isolated from Thermoplasma, but not in the homooligomeric alpha-only thermosome expressed in Escherichia coli. This indicates that alpha and beta-subunits alternate within the rings of the Thermoplasma thermosome rather than forming two different homooligomeric rings. In addition, a small subpopulation of 9-fold symmetric complexes was found among the recombinant alpha-only thermosomes, and a central mass most likely representing bound substrate molecules was observed in about half of the native and recombinant thermosome particles.

Electron Tomography of Single Ice-Embedded Macromolecules: Three-Dimensional Alignment and Classification

From 3-D reconstructions of automatically recorded tilt series of ice-embedded macromolecules, several hundred 3-D images of single particles can be extracted. Here we describe correlation-based techniques to align the particles with respect to translation and orientation in 3-D and the calculation of an averaged reconstruction after application of the correct weighting function to the particle projections. Multivariate statistical analysis and classification are applied to the set of three-dimensionally reconstructed particles to investigate interimage variations on the 3-D level. Copyright 1997 Academic Press. Copyright 1997 Academic Press

Purification and structural characterization of the thermosome from the hyperthermophilic archaeum Methanopyrus kandleri.

From Methanopyrus kandleri, the most thermophilic methanogen known so far, we have purified to homogeneity a protein complex of high molecular mass. Image analysis of transmission electron micrographs revealed a barrel-shaped particle composed of two rings with 8-fold symmetry. Only one type of subunit could be detected. The corresponding gene has been cloned and sequenced. The deduced amino acid sequence shows high homology with the members of group II chaperonins. The structure of the projection and the sequence homology suggest that this particle is the first thermosome isolated from a methanogen.

Expression of an archaeal chaperonin in E. coli: formation of homo- (alpha, beta) and hetero-oligomeric (alpha+beta) thermosome complexes.

Co-expression of the two genes encoding the alpha- and beta-subunit of the Thermoplasma acidophilum thermosome in Escherichia coli yielded fully assembled hetero-oligomeric complexes (alpha+beta). Surprisingly, also separate expression of both genes resulted in formation of hexadecameric complexes (alpha, beta) in the bacterial cytoplasm. On electron micrographs these complexes were indistinguishable from each other and from the native thermosome. The recombinant alpha-complex as well as the native thermosome could be reconstituted in vitro from their dissociated subunits in the presence of Mg-ATP.

The use of children's drawings in the evaluation and treatment of child sexual, emotional, and physical abuse.

Primary care physicians can be instrumental in the initial identification of potential sexual, emotional, and physical abuse of children. We reviewed the use of children's artwork as a method of communicating individual and family functioning. A quantitative method of analyzing children's artwork provides more reliability and validity than some methods used previously. A new scoring system was developed that uses individual human figure drawings and kinetic family drawings. This scoring system was based on research with 842 children (341 positively identified as sexually molested, 252 positively not sexually molested but having emotional or behavioral problems, and 249 "normal" public school children). This system is more comprehensive than previous systems of assessment of potential abuse.

The thermosome of Thermoplasma acidophilum and its relationship to the eukaryotic chaperonin TRiC.

A high molecular-mass protein complex from the archaebacterium Thermoplasma acidophilum, referred to here as the 'thermosome', is built from two subunits (M(r) 58 and 60). The thermosome has been purified to homogeneity. The molecular mass of the native complex was determined to be 1061 +/- 30 Da by scanning transmission electron microscopy. It shows a weak ATPase activity and is able to bind denatured polypeptides. Averages obtained from electron micrographs of negatively stained molecules in the end-on and side-on orientations, respectively, were compared with those of the t-complex polypeptide 1 ring complex (TRiC), isolated from bovine testes. Both molecules consist of two stacked pseudo eightfold symmetric rings which build up a cylindrical particle with a large cavity in the center. Sequence alignments of peptides generated from both subunits of the thermosome and different subunits of TRiC reveal a high partial similarity to each other and to the archaebacterial chaperonin thermophilic factor 55 from Sulfolobus shibatae as well as to eukaryotic TCP1 proteins. These striking structural similarities confirm the proposition that all these molecules belong to a single protein family which is structurally and functionally related to the GroEL class of molecular chaperones.

Tetrabrachion: a filamentous archaebacterial surface protein assembly of unusual structure and extreme stability.

The surface (S-) layer of the hyperthermophilic archaebacterium Staphylothermus marinus was isolated, dissected into separate domains by chemical and proteolytic methods, and analyzed by spectroscopic, electron microscopic and biochemical techniques. The S-layer is formed by a poorly ordered meshwork of branched, filiform morphological subunits resembling dandelion seed-heads. A morphological subunit (christened by us tetrabrachion) consists of a 70 nm long, almost perfectly straight stalk ending in four straight arms of 24 nm length that provide lateral connectivity by end-to-end contacts. At 32 nm from the branching point, tetrabrachion carries two globular particles of 10 nm diameter that have both tryptic and chymotryptic protease activity. Tetrabrachion is built by a tetramer of M(r) 92,000 polypeptides that form a parallel, four-stranded alpha-helical rod and separate at one end into four strands. These strands interact in a 1:1 stoichiometry with polypeptides of M(r) 85,000 to form the arms. The arms are composed entirely of beta-sheets. All S-layer components contain bound carbohydrates (glucose, mannose, and glucosamine) at a ratio of 38 g/100 g protein for the complete tetrabrachion-protease complex. The unique structure of tetrabrachion is reflected in an extreme thermal stability in the presence of strong denaturants (1% (w/v) SDS of 6M guanidine): the arms, which are stabilized by intramolecular disulphide bridges, melt around 115 degrees C under non-reducing conditions, whereas the stalk sustains heating up to about 130 degrees C. Complete denaturation of the stalk domain requires treatment with 70% (v/v) sulfuric acid or with fuming trifluoromethanesulfonic acid. The globular protease can be heated to 90 degrees C in 6M guanidine and to 120 degrees C in 1% SDS and represents one of the most stable proteases characterized to date.

Separation of granule subpopulations in human polymorphonuclear leukocytes.

Human polymorphonuclear leukocytes were isolated, disrupted by sonification and the nuclei and unbroken cells removed by centrifugation. The supernatant was applied on top of an optimised discontinuous Percoll gradient. After centrifugation we found nine gradient bands of distinct density. Both the nine bands and the whole fractionated gradient material were assayed for granule marker enzymes. Granule fractions of distinct density, enclosing different enzyme concentrations demonstrated the existence of granule subpopulations. There were three subpopulations of azurophil granules, about four subpopulations of specific granules, one granule fraction perhaps representing the C-particles, and a fraction of plasma membrane vesicles.