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Background & Aims: Bacterial infections, in particular a spontaneous bacterial peritonitis (SBP), are a major threat in patients with liver cirrhosis. Recently, it has been shown that the impact on mortality might be underestimated by established risk-scores. Onset of infection was suggested to define a distinct stage of cirrhosis. However, it remains unclear whether all stages of decompensated cirrhosis are equally affected. Moreover, if there is such a distinct stage, it must be determined whether it is reversible after the infection has resolved. In this study we aimed to further analyze the impact of a current as well as a resolved SBP in different stages of decompensated liver cirrhosis., Methods: A number of 579 patients with liver cirrhosis and ascites were included. MELD-score was used to determine the stage of liver disease. Low (<15), intermediate (15-25) and high (>25) MELD-groups were compared. Patients were followed up for 90 days. Primary endpoint was overall mortality. Statistical analyses were performed using the log-rank test, Cox regression and competing risk analysis., Results: Mortality was significantly higher in patients with nosocomial-acquired SBP (nSBP) compared to patients without SBP (p<0.001;HR = 2.05). However, the most prominent difference in mortality was documented in the intermediate MELD-group (nSBP: p = 0.02;HR = 2.10). Importantly, mortality in nSBP patients remained increased even after the initial nSBP episode had resolved (p<0.01;HR = 1.90). Again, this was only significant in those with intermediate MELD-scores (p = 0.02;HR = 2.28). While a current as well as a resolved nSBP were significantly linked to a higher mortality, neither of them did increase the likelihood for liver transplantation., Conclusions: Development of nSBP is independently associated with increased mortality supporting the concept of a distinct status of cirrhosis. Importantly, the prognosis remains unfavorable even after resolution of nSBP. This could be particularly relevant for patients with intermediate MELD-scores, who have limited chances for a donor liver., Competing Interests: The authors have declared that no competing interests exist.

The Logical Observation Identifiers, Names and Codes (LOINC) is a common terminology used for standardizing laboratory terms. Within the HiGHmed consortium, LOINC is used as a central terminology for health data sharing across all university hospital sites. Therefore, linking the LOINC codes to the site-specific tests and measures is one crucial step to reach this goal. In this work we report our ongoing work in implementing LOINC to the laboratory information system, our challenges and lessons learned.

Introduction: Type 2 diabetes mellitus (DM) is a frequent comorbidity among patients with liver cirrhosis. However, data regarding the impact of DM on spontaneous bacterial peritonitis (SBP) are quite limited. Our aim was to analyze the impact of DM and HbA1c values on the incidence of SBP and mortality in patients with liver cirrhosis and ascites., Methods: A number of 475 consecutive patients with liver cirrhosis and ascites were analyzed. Presence of DM as well as HbA1c was assessed at the time of the first paracentesis. Patients were followed up for a mean of 266 days. Primary endpoints were SBP development and mortality., Results: Overall, 118 (25%) patients were diagnosed with DM. DM patients had an increased risk for developing a SBP during follow-up (HR: 1.51; p = 0.03). SBP incidence was particularly high in DM patients with HbA1c values ≥6.4%, significantly higher than in DM patients with HbA1c values <6.4% (HR: 4.21; p = 0.0002). Of note, DM patients with HbA1c <6.4% at baseline had a similar risk for SBP as those without DM (HR: 0.93; p = 0.78, respectively). After excluding all patients who were eligible for secondary antibiotic prophylaxis, HbA1c ≥6.4% but neither bilirubin nor ascites protein level were associated with primary SBP development in the multivariate analysis (p = 0.003)., Conclusions: Individuals with liver cirrhosis and concomitant DM have a higher risk for developing a SBP. HbA1c values may be useful to further stratify the risk for SBP among DM patients, which may help to identify those who benefit from antibiotic prophylaxis.

Background & Aims: Spontaneous bacterial peritonitis is a severe complication in patients with cirrhosis leading to acute kidney injury, hepatic encephalopathy and a high mortality. In this study, we aimed to investigate the impact of proton pump inhibitors and the potential relevance of the taken dosage on the incidence and clinical course of spontaneous bacterial peritonitis., Methods: Overall, 613 consecutive patients with decompensated cirrhosis were included. All patients were carefully evaluated for proton pump inhibitors intake including the applied dosage and were further followed up for spontaneous bacterial peritonitis development as well as for the incidence of clinical complications like hepatic encephalopathy, acute kidney injury and mortality., Results: Cumulative spontaneous bacterial peritonitis incidence did neither differ between the proton pump inhibitors and the no-proton pump inhibitors group nor between those taking the high (>40 mg/d) and the low (10-40 mg/d) proton pump inhibitors' dose. However, proton pump inhibitors' intake was associated with an impaired clinical course of spontaneous bacterial peritonitis reflected by a higher likelihood for acute kidney injury (71% vs 43%; P = .002), severe hepatic encephalopathy (15% vs 0%; P = .04) and an increased mortality (24% vs 0%; P = .008) within 28 days after spontaneous bacterial peritonitis diagnosis. In particular, patients with proton pump inhibitors dosages >40 mg/d had an increased short-term risk for acute kidney injury (adjusted hazard ratio: 1.86; P = .009) and mortality (adjusted hazard ratio: 2.05; P = .02). In contrast, there was no effect of proton pump inhibitors on acute kidney injury, hepatic encephalopathy and mortality in patients without spontaneous bacterial peritonitis irrespective of the applied proton pump inhibitors dosage., Conclusions: High dosages of proton pump inhibitors are associated with an adverse outcome in patients with spontaneous bacterial peritonitis. Thus, indication for high-dosage proton pump inhibitors therapy should be evaluated carefully in these patients., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Scheimpflug photography and densitometric image analysis are very precise techniques for light scattering measurement and biometry in the anterior segment of the eye. They provide reproducible data on the characteristics of the anterior eye segment in clinical and experimental studies and the set of data obtained allows discrimination of light scattering changes because of ageing, disease or toxic effects. The techniques can also be used to determine no-effect levels or maximally tolerable dosages of physical and chemical noxious factors. Several Scheimpflug cameras have been marketed, but the only cameras commercially available today are the Nidek EAS 1000 and the Oculus Pentacam. This review outlines the development of the technique and its introduction into ophthalmology. Furthermore, the application of the technique in clinical and experimental ophthalmology as well as in ocular toxicology are presented and discussed.

Protein-protein interactions are critical to most biological processes, and locating protein-protein interfaces on protein structures is an important task in molecular biology. We developed a new experimental strategy called the 'absence of interference' approach to determine surface residues involved in protein-protein interaction of established yeast two-hybrid pairs of interacting proteins. One of the proteins is subjected to high-level randomization by error-prone PCR. The resulting library is selected by yeast two-hybrid system for interacting clones that are isolated and sequenced. The interaction region can be identified by an absence or depletion of mutations. For data analysis and presentation, we developed a Web interface that analyzes the mutational spectrum and displays the mutational frequency on the surface of the structure (or a structural model) of the randomized protein. Additionally, this interface might be of use for the display of mutational distributions determined by other types of random mutagenesis experiments. We applied the approach to map the interface of the catalytic domain of the DNA methyltransferase Dnmt3a with its regulatory factor Dnmt3L. Dnmt3a was randomized with high mutational load. A total of 76 interacting clones were isolated and sequenced, and 648 mutations were identified. The mutational pattern allowed to identify a unique interaction region on the surface of Dnmt3a, which comprises about 500-600 A(2). The results were confirmed by site-directed mutagenesis and structural analysis. The absence-of-interference approach will allow high-throughput mapping of protein interaction sites suitable for functional studies and protein docking.

DNA methyltransferase 1 methylates hemi-methylated CG sites generated during DNA replication. Serine 515 of this enzyme has been shown to be phosphorylated. To explore the importance of S515 phosphorylation, we generated mutants of Dnmt1 which removed the phosphorylation potential (S515A) or mimic phosphoserine (S515E), purified the proteins from insect cells and analyzed their DNA methylation activity in vitro. The S515E mutant was found to be active, while S515A mutant had severe loss in activity when compared to the wild type protein. The loss of activity of the S515A variant was not due to loss of DNA binding capacity. Furthermore, we show that a phosphorylated peptide whose sequence mimics the surrounding of Ser515 (EKIYIS(P)KIVVE) inhibited the activity of wild type Dnmt1 ten-fold more than the non-phosphorylated peptide. The inhibition was specific for Dnmt1 and for the particular peptide sequence. Our data suggest that phosphorylation of Ser515 is important for an interaction between the N-terminal domain of Dnmt1 and its catalytic domain that is necessary for activity and that this interaction is specifically disrupted by the phosphorylated peptide. We conclude that phosphorylation of Dnmt1 at Ser515 could be an important regulator of Dnmt1 activity during cell cycle and after proliferative stimuli.

Slow Wallerian degeneration (Wld(S)) mutant mice express a chimeric nuclear protein that protects sick or injured axons from degeneration. The C-terminal region, derived from NAD(+) synthesizing enzyme Nmnat1, is reported to confer neuroprotection in vitro. However, an additional role for the N-terminal 70 amino acids (N70), derived from multiubiquitination factor Ube4b, has not been excluded. In wild-type Ube4b, N70 is part of a sequence essential for ubiquitination activity but its role is not understood. We report direct binding of N70 to valosin-containing protein (VCP; p97/Cdc48), a protein with diverse cellular roles including a pivotal role in the ubiquitin proteasome system. Interaction with Wld(S) targets VCP to discrete intranuclear foci where ubiquitin epitopes can also accumulate. Wld(S) lacking its N-terminal 16 amino acids (N16) neither binds nor redistributes VCP, but continues to accumulate in intranuclear foci, targeting its intrinsic NAD(+) synthesis activity to these same foci. Wild-type Ube4b also requires N16 to bind VCP, despite a more C-terminal binding site in invertebrate orthologues. We conclude that N-terminal sequences of Wld(S) protein influence the intranuclear location of both ubiquitin proteasome and NAD(+) synthesis machinery and that an evolutionary recent sequence mediates binding of mammalian Ube4b to VCP.

Proteasome inhibitors such as lactacystin were first isolated when assaying their ability to stimulate neurite outgrowth in neuronal-like cell lines; however, their effect on neurites in primary culture has been largely neglected. We report here that lactacystin causes immediate arrest of nerve growth factor (NGF)-stimulated neurite outgrowth in sympathetic and sensory explant cultures. This is followed by neurite degeneration that in sympathetic cultures has a distinctive "dying-back" morphology. Remarkably, this occurs even at concentrations below that required to induce neurite outgrowth in PC12 cells. Thus, lactacystin opposes rather than potentiates the effect of NGF on sympathetic neurite outgrowth and the role of the ubiquitin proteasome pathway in growth and long-term maintenance of axons and dendrites differs from that in neuritogenesis in neuronal-like cell lines. Retrograde degeneration caused by blocking of the ubiquitin proteasome pathway may mimic some aspects of gracile axonal dystrophy, a dying-back axonopathy in mice caused by ubiquitin hydrolase (Uch-l1) deficiency, and may be relevant to human neurodegenerative diseases involving ubiquitination or proteasome abnormalities., (Copyright 2003 Wiley-Liss, Inc.)

The paper focuses on possible effects of the botanical composition and weather conditions on the extend of zearalenone and ochratoxin A concentrations in various grass swards in winter pasture systems. Zearalenone is only detectable in pure stands ofLolium perenne or inLolio-Cynosuretum plant communities, respectively. The occurrence of ochratoxin A is more frequent and less specific concerning the botanical composition. This mycotoxin was found in both,Lolium perenne andFestuca arundinacea in varying years. The incidence of ochratoxin A depends on year and is apparently related to the weather conditions in autumn and winter. There was no evidence that particular locatio8/22/2008 12:57PMns have a higher or a lower risk for high ochratoxin A or zearalenone concentrations than others. Peak values in individual swards are not intermittent over the years.

The split-ubiquitin assay detects protein interactions in vivo. To identify proteins interacting with Gal4p and Tup1p, two transcriptional regulators, we converted the split-ubiquitin assay into a generally applicable screen for binding partners of specific proteins in vivo. A library of genomic Saccharomyces cerevisiae DNA fragments fused to the N-terminal half of ubiquitin was constructed and transformed into yeast strains carrying either Gal4p or Tup1p as a bait. Both proteins were C-terminally extended by the C-terminal half of ubiquitin followed by a modified Ura3p with an arginine in position 1, a destabilizing residue in the N-end rule pathway. The bait fusion protein alone is stable and enzymatically active. However, upon interaction with its prey, a native-like ubiquitin is reconstituted. RUra3p is then cleaved off by the ubiquitin-specific proteases and rapidly degraded by the N-end rule pathway. In both screens, Nhp6B was identified as a protein in close proximity to Gal4p as well as to Tup1p. Direct interaction between either protein and Nhp6B was confirmed by coprecipitation assays. Genetic analysis revealed that Nhp6B, a member of the HMG1 family of DNA-binding proteins, can influence transcriptional activation as well as repression at a specific locus in the chromosome of the yeast S. cerevisiae.

IL-10 is an important regulatory cytokine. The recent characterization of the 5'-flanking region of IL-10 led to the identification of the promoter region. The infection of B cells with EBV induces IL-10 production which may contribute to EBV-induced transformation. In the present report, IL-10 promoter elements involved in the constitutive expression of IL-10 in EBV-positive lymphoma cells are described. The AT-rich region between -54/-66 from the transcriptional start site was found to be important for IL-10-promoter activity in BL36. A point mutation at position -60 (T/A) was associated with over 90% reduction of luciferase activity in the cell lines BL36 and BL74. The conversion of A/T at position -57 led to enhanced promoter activity. In addition the AT-rich region could serve as an enhancer for the beta-globin basic promoter. In BJAB cells (EBV-negative), sequences between -205/-139 rather than the AT-rich region were involved in IL-10 promoter regulation. This underlines the importance of the AT-rich region for EBV-associated IL-10 promoter regulation. Our results further the understanding of how the IL-10 gene could be regulated in B cell lymphomas.

Some years ago, Nidek developed a new imaging system for the anterior eye segment, which offers the possibility of recording Scheimpflug and retroillumination images. The system consists of 2 different technical units, a camera unit for image recording and a computer unit for storage, system operation and image analysis. To evaluate the clinical use of the system and its reproducibility, a study with 31 volunteers/patients was performed that were photographed with the Nidek EAS 1000 at the meridians 0 degree, 45 degrees and 180 degrees. Two examinations with a 1-week interval were performed. In addition, a direct comparison of the EAS 1000 with the Topcon SL-45 was carried out with 32 volunteers that were photographed at 0 degree and 45 degrees; a retroillumination photo was additionally recorded. The video images were evaluated with the computer software, the SL-45 negatives were standardly measured with a Joyce-Loebl densitometer. The handling of the EAS 1000 in clinical use was found to be easy and comfortable for the patient, as the alignment is done with infrared light. Difficulties occur only with the retroillumination photography in that the infrared light for imaging may outshine the red fixation light during exposure. However, shadowing problems due to the eyelashes occur in oblique slit positions, especially in those patients with a pronounced front head. In studying the reproducibility, the mean values of the individual coefficients of variation for light scattering (density) were in a range between 3.6 and 5.06%, but more than 35% of the single values show a variation coefficient above 5% (maximum 20.3%).(ABSTRACT TRUNCATED AT 250 WORDS)

A double-masked study was performed to investigate a possible side effect of the antiallergic substance Picumast dihydrochloride on lens transparency. 372 patients with Picumast dihydrochloride- or Ketotifen-treatment regularly underwent ophthalmologic examinations and Scheimpflug photography of the anterior eye segment. Results of slitlamp examinations as well as microdensitometric analyzes of the film negatives after one year treatment did not reveal any evidence that Picumast dihydrochloride compromised lens transparency in humans.

Topcon Tokyo manufactured the first series of SL-45 in 1980 and a second in 1982/83, which were completed in Bonn with all technical improvements developed so far. In 1987, the SL-45B was manufactured which included all previously added improvements. To compare both camera types, we carried out a test with 10 volunteers and the following technical variables for the SL-45: (1) miosis; (2) mydriasis; (3) flashbulb 1; (4) flashbulb 2; (5) film batch 1; (6) film batch 2, and (7) different types of film. Afterwards, 5 of these volunteers were photographed with the SL-45B. Photographer and reader were identical in all cases. Based on microdensitometry it could be demonstrated that, with the exception of a different film type, all variables can be compensated for by using the corneal peak as an external standard.

Experimental cataract development can be objectively monitored in rats by Scheimpflug slit-image photography and microdensitometric image analysis. Zeiss (Oberkochen, Federal Republic of Germany) has developed a new computerized slit-lamp measuring system that works according to the Scheimpflug principle; we have very successfully applied it to rats with streptozotocin-induced cataracta diabetica vera and with naphthalene-induced opacities. With these cataract models we tested the gyrase inhibitor compound, Ciprofloxacin (Bayer 09867), after 6 weeks' daily peroral application (20 mg/kg body weight) for differences in cataract progression indicating a possible cocataractogenic effect of the compound. When we compared the two different Scheimpflug methods, we found that the Zeiss-Scheimpflug measuring system SLC has some handling advantages with respect to animal cataract studies. If the equipment is to be fully used, complements to the program of the integrated computer part are necessary. The image-analytical results from the animals treated with Ciprofloxacin did not show any indication of cocataractogenic potential with respect to cataracta diabetica vera or naphthalene cataract.

Scheimpflug photography of the anterior eye segment with densitometric image analysis to measure lens transparency has proven its value in long-term follow-up studies of lens changes. The necessary technical conditions of the SL 45 Topcon camera guarantee high reproducibility. The technical control with respect to quality in case of repeated photography is ensured by an internal standard, the camera being equipped with a five-step scale of known density values. External standardization is done by taking the densitometer height of the cornea as standard. The coefficients of variation of internal and external standards were nearly equal (approximately 5%) over the 9-month period of observation of 100 patients (3,200 photos). The coefficients of variation correspond to the standard error found with repeated lens photos of volunteers' eyes evaluated by analysis of variance of the respective data sets. The standard error for distance measurements on the x-axis (cornea thickness, depth of anterior chamber, lens thickness, etc.) is less than 3%. Evaluation of follow-up examinations may most successfully be done by densitometric planimetry, which, in case of follow-up studies on opacified lenses, is superior to the usual measurements of densitometer height. The light scatter registered by Scheimpflug photography is highly individual. This is demonstrated by densitograms of normal lenses of persons of different age groups. Besides linear densitometry, various other procedures of densitometric planimetry are possible. The best results so far were obtained by multilinear densitometry, but the enormous amount of data obtained in this way renders routine application rather difficult.

The recovery of clarity in frozen, lyophilized human tissue lenticules used for epikeratophakia has been a matter of controversy. According to some authors, several months are necessary for the lenticules to resume normal transparency; others report clinical experience of much shorter times. Up to now, objective documentation of such findings has not been substantiated. We used a Topcon LS-45 camera to photograph four eyes that underwent epikeratophakia, preoperatively and at 2, 4, 6, and 8 weeks after surgery, and analyzed the film negatives by linear microdensitometry. The degree of film blackening, that is the density of the tissue, was expressed graphically by peaks of various height measured in millimeters. Best spectacle corrected Snellen visual acuity was also recorded. Similar measurements of corneal transparency were performed in 20 healthy adult volunteers whose best corrected spectacle visual acuity was 20/20. In the four operated eyes, (two keratoconus, one aphakia, and one myopia), the light scattering was increased in both the host cornea and the donor lenticule. The lenticule had a considerably increased light scattering in all patients at both 2 and 4 weeks after surgery, but was comparable to that of unoperated corneas in three of four patients at 6 weeks.

Development of experimental cataract can be objectively monitored in rats by application of Scheimpflug photography with the SL 45 Topcon camera and subsequent densitometric image analysis. The method has been used to study naphthalene cataract in Brown-Norway rats (BN-CPB), as well as diabetic cataract induced by streptozotocin injection in Sprague-Dawley rats. The values obtained by linear microdensitometric image analysis allowed precise characterization of the opacification with respect to size, topography, and time progress so that statistical evaluation of the efficacy of certain drugs in prevention or delay of experimental cataracts was possible.

An hypothesis is presented to explain the red cell lysis which accompanies an acute malarial infection, as well as the mode of action of certain schizonticidal drugs in the quinoline and acridine series. Quinine and a number of other antimalarial drugs have been found to counteract the inhibition by protein of fatty acid-induced lysis, when tested in an in vitro system. It is suggested that these schizonticides exert their chemotherapeutic effect by inducing the premature lysis of the parasitized red cell, as a result of relieving the inhibition by protein of haemolysis.

Quinine activates the hydrolysis of phosphatidyl choline suspensions by phospholipase C (E.C. 3.1.4.3) obtained from Clostridium welchii. Low levels of calcium are an absolute requirement for this activation: Mg2+, Ba2+, Sr2+, and Zn2+ are ineffective. The induction period, or lag phase for this enzyme is dependent upon both calcium concentration and substrate interfacial surface area. At low concentrations (less then 50 muM) calcium ions affect the induction period but not the maximal rate of hydrolysis, whereas guinine predominantly affects the rate of hydrolysis by alterations in the surface charge carried by the substrate.