Comparative morphology of the hemolymph vascular system in Uropygi and Amblypygi (Arachnida): Complex correspondences support Arachnopulmonata.

Although the circulatory system of arthropods has long been considered as rather simple, recent studies have demonstrated that in certain arthropod taxa, such as Malacostraca, some Chilopoda and also many Chelicerata, the vascular systems in particular are rather complex. Furthermore, a recent study has revealed that the prosomal ganglion of scorpions and spiders is supplied by an intricate network of arteries, the complexity of which bears a close resemblance to that of vertebrate capillary systems. In this study, we analyzed the hemolymph vascular systems of various species of Pedipalpi (i.e., Amblypygi and Uropygi). By combining modern techniques, such as MicroCT and cLSM, with computer-based 3D-reconstruction, we were able to produce comprehensive visualizations and descriptions of the vascular systems. Despite the lack of well-corroborated phylogenetic hypotheses on arachnid relationships and the controversial assertion of relationships between the pulmonate arachnids, we aim to elucidate the evolution of complex vascular systems in Arachnida. By comparing these highly complex vascular systems not only with each other, but also with other pulmonate arachnids, we found numerous detailed correspondences in the general branching pattern as well as in the supply patterns of the prosomal ganglion. We argue that these numerous and detailed correspondences by their absence in other arachnids i.e. aplumonates, support Arachnopulmonata. J. Morphol. 277:1084-1103, 2016. © 2016 Wiley Periodicals, Inc.

Characterization and purification of an outer membrane metalloproteinase from Pseudomonas aeruginosa with fibrinogenolytic activity.

A membrane proteinase from Pseudomonas aeruginosa, called insulin-cleaving membrane proteinase (ICMP), was located in the outer membrane leaflet of the cell envelope. The enzyme is expressed early in the logarithmic phase parallel to the bacterial growth during growth on peptide rich media. It is located with its active center facing to the outermost side of the cell, because its whole activity could be measured in intact cells. The very labile membrane proteinase was solubilized by non-ionic detergents (Nonidet P-40, Triton X-100) and purified in its amphiphilic form to apparent homogeneity in SDS-PAGE by copper chelate chromatography and two subsequent chromatographic steps on Red-Sepharose CL-4B (yield 58.3%, purification factor 776.3). It consisted of a single polypeptide chain with a molecular mass of 44.6 kDa, determined by mass spectrometry. ICMP was characterized to be a metalloprotease with pH-optimum in the neutral range. The ICMP readily hydrolyzed Glu(13)-Ala(14) and Tyr(16)-Leu(17) bonds in the insulin B-chain. Phe(25)-Tyr(26) and His(10)-Leu(11) were secondary cleavage sites suggesting a primary specificity of the enzyme for hydrophobic or aromatic residues at P'(1)-position. The ICMP differed from elastase, alkaline protease and LasA in its cleavage specificity, inhibition behavior and was immunologically diverse from elastase. The amino acid sequence of internal peptides showed no homologies with the known proteinases. This outer membrane proteinase was capable of specific cleavage of alpha and beta fibrinogen chains. Among the p-nitroanilide substrates tested, substrates of plasminogen activator, complement convertase and kallikrein with arginine residues in the P(1)-subsite were the substrates best accepted, but they were only cleaved at a very low rate.

The cell envelope-bound metalloprotease (camelysin) from Bacillus cereus is a possible pathogenic factor.

A novel membrane proteinase of the nosocomial important bacteria species Bacillus cereus (synonyms: camelysin, CCMP) was purified up to homogeneity as was shown by mass spectrometry in its amphiphilic form. Camelysin is a neutral metalloprotease with a molecular mass of 19 kDa. Its unique N-terminus Phe-Phe-Ser-Asp-Lys-Glu-Val-Ser-Asn-Asn-Thr-Phe-Ala-Ala-Gly-Thr-Leu-Asp-Leu-Thr-Leu-Asn-Pro-Lys-Thr-Leu-Val-Asp-(Ile-Lys-Asp)- was not detected in the protein data bases during BLAST searches, but in the partially sequenced genome of Bacillus anthracis, coding for an unknown protein. Cleavage sites of the membrane proteinase for the insulin A- and B-chains were determined by mass spectrometry and N-terminal sequencing. Camelysin prefers cleavage sites in front of aliphatic and hydrophilic amino acid residues (-OH, -SO3H, amido group), avoiding bulky aromatic residues. The internally quenched fluorogenic substrates of the matrix metalloproteases 2 and 7 were cleaved with the highest efficiency at the Leu-decrease-Gly or Leu-decrease-Ala bond with the smaller residue in the P1' position. The protein specificity is broad--all various kinds of casein were cleaved as well as acid-soluble collagen, globin and ovalbumin; intact insulin was destroyed only to a low extent. Actin, collagen type I, fibrinogen, fibrin, alpha2-antiplasmin and alpha1-antitrypsin were cleaved. The protease formed SDS-stable complexes with Glu-plasminogen and antithrombin III, visible after SDS electrophoresis by gold staining and Western blot. The CCMP-plasminogen complex caused a partial activation of plasminogen to plasmin. Camelysin interacts with proteins of the blood coagulation cascade and could facilitate the penetration of fibrin clots and of the extracellular matrix during bacterial invasion.

Nerve fibers innervating the cranial and spinal meninges: morphology of nerve fiber terminals and their structural integration.

Pachymeninx and leptomeninx of cranial cavity and spine are considerably different in their collagenous fiber texture, cellular composition, vascularization, and innervation. The majority of meningeal nerve fibers terminate as free nerve endings whereas encapsulated and lamellated nerve terminals additionally occur in higher vertebrates including man. With respect to nerve fiber classification, arborization pattern, topography, and organization of the microenvironment at the termination site afferent and efferent nerve terminals are differentiated. Only the dura mater and the pial subcompartment of the leptomeninx possess the morphological prerequisites for neurogenic inflammation. In the current review, the results of morphological studies regarding the meningeal innervation including the sites of CSF (cerebrospinal fluid) production and absorption are discussed with emphasis on their structure-function relationships.

Virtual image navigation: a new method to control intraoperative bleeding in neuroendoscopic surgery. Technical note.

In this neuroendoscopic study the authors tested the newly developed "red-out module" of their visual navigation system that enables the neurosurgeon to achieve hemostasis if total visualization is lost due to hemorrhage ("red out") within the visual field. An optical position measurement system connected to the endoscope guarantees that digitized endoscopic images are coupled with the accurate endoscopic position. Computerized images are simultaneously stored with their respective position data, and this creates a virtual anatomical landscape. The system was tested in in vivo bleeding conditions in a rat model. Artificial endoscopic cavities were created in the inguinal, pelvic, and jugular regions in rats to imitate the conditions of the human ventricular system. Two experimental settings were tested: Technique I, in which a computer landmark has been previously determined at the point where the vessel will be lesioned; and Technique II, in which a landmark has been previously set in the surrounding area of the vessel. Immediately after hemorrhage obscures the visual field (red out), the computer automatically displays the virtual images on a separate monitor. The previously set landmarks and the graphic overlay of the coagulation fiber enable the surgeon to navigate within the operative field based on the virtual images and to perform coagulation at the site of the lesion. A total of 175 vessels were coagulated: 43 arteries and 132 veins. In using Technique I, 130 (90.9%) of 143 vessels and in using Technique II, 26 (81.2%) of 32 arteries were successfully coagulated. The authors' data revealed that virtual image guidance has the potential to be a helpful tool in neuroendoscopy.

Unusual chromatographic behaviour and one-step purification of a novel membrane proteinase from Bacillus cereus.

Cell envelopes of Bacillus cereus contain a casein-cleaving membrane proteinase (CCMP) and an insulin-cleaving membrane proteinase (ICMP), which differ in their substrate and inhibitor specificity from all Bacillus proteinases described previously. They remained localized in the cytoplasmic membrane after treatment with lysozyme and mutanolysin and they are strongly attached to the membrane compared with other known membrane proteinases. Only high a concentration of the Zwitterionic detergent sulfobetain SB-12 enabled an effective solubilization of both membrane proteinases. The usual conventional purification methods, such as chromatofocusing, ion-exchange chromatography and hydrophobic interaction chromatography in the presence of detergent concentrations beyond their critical micelle concentration, could not be applied to the purification, because the solubilized membrane proteinases bound strongly and irreversibly to the chromatographic matrix. In the search for other purification methods, we used a tentacle ion-exchanger (EMD trimethylaminoethyl-Fractogel) to reduce the hydrophobic interactions between the proteinases and the matrix. All contaminating proteins could be removed by a first gradient of sodium chloride without elution of CCMP; a second gradient with isopropanol and a decreasing salt concentration resulted in an efficiently purified CCMP. The ICMP was irreversibly denaturated. Purified CCMP is a member of the metalloproteinase family with a pH optimum in the neutral range and a temperature optimum of 40 degrees C, whose properties differ from the serine-type membrane proteinase of Bacillus subtilis described by Shimizu et al. [Agric. Biol. Chem., 47 (1983) 1775]. It consists of two subunits in sodiumdodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions (Mr 53,000 and 65,000); however, the molecular mass of the purified enzyme could not be determined by size exclusion or SDS-PAGE, because the purified enzyme aggregated at the top of the gel matrix. CCMP solubilized before the purification process, could be eluted in the presence of 0.1% octylphenol-poly(ethyleneglycol ether)9-10 (Triton X-100) in two peaks of Mr 56,000 and 128,000, respectively. We discuss this special chromatographic behaviour of the CCMP from Bacillus cereus, with regard to the strong hydrophobic interactions of the enzyme with the chromatographic matrix and additional self-aggregation, which could only be dissolved by solvents such as isopropanol.

The curious genomic path from leaky red cell to nephrotic kidney.

The human red cell has proved to be an invaluable model cell for the study of many aspects of membrane structure and function. It has a series of transport pathways which mediate the movements of the univalent cations Na and K, which are either identical or similar to systems in other human tissues, including the human kidney. The balance between the energy-consuming NaK pump and a 'passive leak' component maintains a net deficit of cations within the cell, which defends the cell volume against osmotic swelling. There exist a series of dominantly inherited human red cell conditions, gathered under the generic title 'hereditary stomatocytoses', in which the so-called 'passive leak' to Na and K is pathologically increased. In the more severe variants this compromises the integrity of the cell and the patients suffer haemolytic anaemia. Some less severe variants present with pseudohyperkalaemia caused by loss of K from red cells on storage of blood at room temperature. The most severe variants show a deficiency in a widely distributed 'raft' protein known as stomatin. The stomatin protein is homologous to the 'podocin' protein, the gene for which is mutated in a recessively inherited form of nephrotic syndrome. Among other possible functions, both proteins could be involved in the trafficking of membrane proteins to and from the plasma membrane.

[Cyclosporin A in therapy of chronic uveitis]. / Cyclosporin A in der Therapie der chronischen Uveitis.

48 patients with chronic severe uveitis of pressumed noninfectious origin resistent to corticosteroids have been treated with Cyclosporin A. All patients had an initial loading dose of 5 mg/kg/day followed by a dose reduction according to ocular inflammatory activity and tolerability and according the guidelines by BenEzra, Nussenblatt and Timonen. Most of the patients received additional corticosteroids in a low dose. 35 out of these 48 patients (13 suffering from intermediate uveitis, 11 from retinal vasculitis, 5 panuveitis, 4 "pressumed histoplasmosis" and 2 sympathetic ophthalmia) were treated for 1 year and observed after withdrawing of Sandimmun for at least 6 months. The majority of these patients have manifested a positive therapeutic response to Cyclosporine, in particular patients suffering from vasculitis, panuveitis and sympathetic ophthalmia. All 35 patients were treated before for a long time with steroids without success, 19 out of these 35 in addition with cytotoxic agents. The immunosuppressive effect of Cyclosporine was not permanent, frequently the inflammation relapsed on reduction of dosage or withdrawing of the drug. Guidelines for combined regimen (Cyclosporine and corticosteroids and vitrectomy) were given. Although a large variety of side effects were reported the compliance was good.

Development of an endoscopic navigation system based on digital image processing.

We developed a new system to couple the endoscope to an optical position measurement system (OPMS) so that the image frames from the endoscope camera can be labeled with the accurate endoscopic position. This OPMS is part of the EasyGuide Neuro navigation system, which is used for microsurgery and neuroendoscopy. Using standard camera calibration techniques and a newly developed system calibration, any 3-dimensional (3-D) world point can be mapped onto the view from the endoscope. In particular, we can display the coordinates of any anatomical landmark of the patient as it is viewed from the current position of the camera. This and other image-processing techniques are applied to the labeled frame sequence in order to offer the neurosurgeon a variety of control modules that increase the safety and flexibility of neuroendoscopic operations. Several modules, including a new motion alarm system and the "tracking" and "virtual map" modules, were tested in a human cadaveric model using the frontal and occipital approaches. A failure rate of 8.6% was experienced during testing of the first version of the software, but the second version was 100% successful. Thus, an endoscopic navigation system based on digital image processing has been developed that could be a revolutionary advance in image-guided surgery.

Evolutionary morphology of the hemolymph vascular system in scorpions: a character analysis.

Phylogenetically informative characters from the internal anatomy of scorpions were first reported more than 150 years ago, but the subject received little attention after the mid-1920s. Recent investigations, using traditional dissection, illustration and histological sectioning, microscopy, and innovative new methods for investigating complex soft tissue anatomy identified a new wealth of variation. Additionally, these investigations confirmed the phylogenetic significance of previously identified structures. Building on earlier work we present a more detailed description of the hemolymph vascular system (HVS) in scorpions, based on comparison of the vascular structures of the heart and the branching pattern of the prosomal arteries among 45 exemplar species representing most of the major scorpion lineages. Using corrosion-casting, MicroCT in combination with computer-aided 3D-reconstruction, and scanning electron microscopy, we conceptualize a series of phylogenetically informative characters for the anterior aorta system and characters of the heart and associated structures (e.g. arrangement of the ostia) of scorpions. Furthermore, we optimize the possible evolution of these new characters on a previous hypothesis of scorpion phylogeny, and discuss alternative character state transformations, their evolutionary consequences, and possible underlying evolutionary mechanisms acting on the HVS.

The excitation energies, ionization potentials, and oscillator strengths of neutral and ionized species of Uuq (Z=114) and the homolog elements Ge, Sn, and Pb.

Multiconfiguration Dirac-Fock method is employed to calculate the excitation energies, ionization potentials, oscillator strengths, and radii for all neutral and up to four times ionized species of element Uuq, as well as the homolog elements Ge, Sn, and Pb. Using an extrapolative scheme, improved ionization potentials of Uuq were obtained with an uncertainty of less than 2000 cm(-1). Two relatively stronger resonance transitions are predicted for the element Uuq. In particular, the strongest line in Uuq, corresponding to the [6d(10)7s(2)7p(3/2)8s(1/2)](1)-->[6d(10)7s(2)7p(3/2)(2)](2) transition at 22 343 cm(-1), just lies in the prime energy region of experimental measurement.

Intermetallic compounds of the heaviest elements and their homologs: the electronic structure and bonding of MM', where M=Ge, Sn, Pb, and element 114, and M'=Ni, Pd, Pt, Cu, Ag, Au, Sn, Pb, and element 114.

Fully relativistic (four-component) density-functional theory calculations were performed for intermetallic dimers MM', where M=Ge, Sn, Pb, and element 114, and MM'=group 10 elements (Ni, Pd, and Pt) and group 11 elements (Cu, Ag, and Au). PbM and 114M, where M are group 14 elements, were also considered. The results have shown that trends in spectroscopic properties-atomization energies D(e), vibrational frequencies omega(e), and bond lengths R(e), as a function of MM', are similar for compounds of Ge, Sn, Pb, and element 114, except for D(e) of PbNi and 114Ni. They were shown to be determined by trends in the energies and space distribution of the valence ns(MM')atomic orbitals (AOs). According to the results, element 114 should form the weakest bonding with Ni and Ag, while the strongest with Pt due to the largest involvement of the 5d(Pt) AOs. In turn, trends in the spectroscopic properties of MM' as a function of M were shown to be determined by the behavior of the np(1/2)(M) AOs. Overall, D(e) of the element 114 dimers are about 1 eV smaller and R(e) are about 0.2 a.u. larger than those of the corresponding Pb compounds. Such a decrease in bonding of the element 114 dimers is caused by the large SO splitting of the 7p orbitals and a decreasing contribution of the relativistically stabilized 7p(1/2)(114) AO. On the basis of the calculated D(e) for the dimers, adsorption enthalpies of element 114 on the corresponding metal surfaces were estimated: They were shown to be about 100-150 kJ/mol smaller than those of Pb.

Adsorption of superheavy elements on metal surfaces.

Fully relativistic four-component density functional theory with the general gradient approximation calculations have been performed to determine the adsorption energy and position of the superheavy element 112 on a Au surface. Extended cluster as well as embedded cluster calculations were used to simulate the surface which allow for the top, bridge, and hollow adsorption sites without losing the advantage of symmetry considerations. Comparison with analogous calculations of the adsorption of the homologue element Hg allows to predict the adsorption of element 112 at a bridge site with a binding energy of 0.67 eV.

[The cortisol awakening response in patients with acute and chronic low back pain. Relations with psychological risk factors of pain chronicity]. / Die Cortisol-Aufwachreaktion bei Patienten mit akuten und chronischen Rückenschmerzen. Zusammenhänge mit psychologischen Risikofaktoren der Schmerzchronifizierung.

BACKGROUND: Peculiarities of the hypothalamic-pituitary-adrenal axis activity in stress-related pain-disorders and potential relations with psychological risk factors of pain chronicity have been discussed controversially. MATERIAL AND METHODS: The cortisol awakening responses of 31 low back pain patients (14 acute, 17 chronic) and 14 healthy controls were compared. In addition the interrelations between awakening response and chronic stress as well as depressive mood and - for the first time - maladaptive painprocessing and -copingstrategies were investigated. RESULTS: The groups did not differ in their cortisol awakening responses. Chronic stress, depressive mood and maladaptive cognitive painprocessing did not correlate with the awakening response. There were, however, significant interrelations between awakening responses and the behavioral paincoping-strategies. CONCLUSIONS: Behavioral paincoping-strategies should be considered as a potentially important contributing psychological factor in the relation between the activity of the hypothalamic-pituitary-adrenal axis and stress-related pain disorders.

Relativistic effects on the electronic structure and volatility of group-8 tetroxides MO4, where M=Ru, Os, and element 108, Hs.

The influence of relativistic effects on properties and volatility of the group-8 tetroxides MO4, where M=Ru, Os, and element 108, Hs, was studied on the basis of results of the fully relativistic (four component) and nonrelativistic density functional theory calculations. Relativistic effects were shown to increase bond strengths and decrease bond lengths in these molecules. They are responsible for a decrease in molecular polarizabilities and an increase in ionization potentials. The effects are much stronger in HsO4 than in the lighter congeners. Relativistic effects were also shown to slightly decrease dispersion interaction energies of RuO4, OsO4, and HsO4 with an inert (quartz or silicon nitride) surface, i.e., they increase volatility of these compounds as studied in the "one-atom-at-a-time" gas-phase chromatography experiments. They do, however, not influence the trend in group 8: both relativistically and nonrelativistically, volatility should change as RuO4

Application of a newly developed visual navigation system in humans. First results.

A visual navigation system (VN) was developed which uses intraoperatively stored endoscopic images together with their specific 3D-address. A special calibration enables one to recalculate the distortion of the endoscopic images. Several modules (e. g., landmark tracking, virtual back-movement) are offered to the neurosurgeon. The system was tested in 12 human subjects during neuroendoscopic interventions and worked without problems in nearly all cases. The possibilities of digital image navigation can be used especially for control of instrument movement in case of red-out situations or blurred vision. Many further developments of the VN system are possible in order to increase the safety of neuroendoscopic interventions.

Consideration of ergonomic aspects in the development of a new endoscopic navigation system.

During the development of new navigation systems, the ergonomic aspects of the accompanying software are seldom taken into account. The VN system is a navigation system working with real and previously stored neuroendoscopic images. The latter enable the module to execute virtual back movement, e.g. in case of bleeding. Several other modules are offered. The aim of the study was the ergonomic analysis of different modules and their learning curves in cadaveric heads. The endoscope was navigated by the neurosurgeon while another person operated the computer. A total of 128 experiments were performed with two software versions. When the landmark tracking module was used, a real learning curve could be observed. By contrast, testing the measurement module did not produce a learning curve. A significant reduction of the time required by the three modules investigated could be observed with software version 2. The module for virtual back movement works best with a minimum time of 20 s for image storage. During machine-human interactions ergonomic software use is important, especially if operative procedures are performed.

Phase separation of nonionic detergents by salt addition and its application to membrane proteins.

By adding salts (sodium chloride, ammonium sulfate), it is possible to induce phase separation in membrane-protein solubilisates containing Triton X-100 or Nonidet P-40 at temperatures between 0 and 20 degrees C. Other nonionic detergents of the Brij, Lubrol, and Tween series can also be used for this procedure. The salt concentration required for induction of phase separation is dependent on the hydrophobicity of the detergent used. For detergents of the Triton series it seems that detergents with lower hydrophilic lipophilic balance numbers need lower salt concentrations to separate the phases than those with larger hydrophilic parts. Ammonium sulfate precipitation as an initial purification step for membrane proteins should be avoided in the presence of the nonionic detergents tested. Instead of this procedure, phase separation induced by sodium chloride or ammonium sulfate can be recommended, as was proved for membrane-bound proteases of Pseudomonas aeruginosa and Bacillus cereus and for bacteriorhodopsin.

Quantitative determination of zwitterionic detergents using salt-induced phase separation of Triton X-100.

Zwitterionic detergents interfere with the salt-induced phase separation for nonionic detergents in a concentration-dependent manner by shifting the normal cloud point of nonionic detergents to a higher ionic strength at room temperature. This phenomenon was used to determine the concentration of the zwitterionic detergents CHAPS, CHAPSO, and sulfobetaine SB-12 in solution by titration with ammonium sulfate in the presence of Triton X-100. Among the ionic detergents tested, the method was only applicable to sodium cholate. The assay can be used to control the removal of zwitterionic detergents during the reconstitution of membrane proteins in liposomes. However, it cannot be used to determine the specific binding of zwitterionic detergents to highly diluted, pure membrane proteins because of the limited sensitivity. Neither proteins nor phospholipids interfered with this method at concentrations up to 20 mg/ml of test solution (human serum albumin) or 10 mg/ml (phospholipids), respectively. Since the assay is based on the competition between salts and nonionic detergents for water molecules, it is important to equalize the ionic strength of samples and calibration standards.