Spatiotemporal regulation of cell-cycle genes by SHORTROOT links patterning and growth.

The development of multicellular organisms relies on the coordinated control of cell divisions leading to proper patterning and growth. The molecular mechanisms underlying pattern formation, particularly the regulation of formative cell divisions, remain poorly understood. In Arabidopsis, formative divisions generating the root ground tissue are controlled by SHORTROOT (SHR) and SCARECROW (SCR). Here we show, using cell-type-specific transcriptional effects of SHR and SCR combined with data from chromatin immunoprecipitation-based microarray experiments, that SHR regulates the spatiotemporal activation of specific genes involved in cell division. Coincident with the onset of a specific formative division, SHR and SCR directly activate a D-type cyclin; furthermore, altering the expression of this cyclin resulted in formative division defects. Our results indicate that proper pattern formation is achieved through transcriptional regulation of specific cell-cycle genes in a cell-type- and developmental-stage-specific context. Taken together, we provide evidence for a direct link between developmental regulators, specific components of the cell-cycle machinery and organ patterning.

Transcriptome-Wide Prediction and Measurement of Combined Effects Induced by Chemical Mixture Exposure in Zebrafish Embryos.

BACKGROUND: Humans and environmental organisms are constantly exposed to complex mixtures of chemicals. Extending our knowledge about the combined effects of chemicals is thus essential for assessing the potential consequences of these exposures. In this context, comprehensive molecular readouts as retrieved by omics techniques are advancing our understanding of the diversity of effects upon chemical exposure. This is especially true for effects induced by chemical concentrations that do not instantaneously lead to mortality, as is commonly the case for environmental exposures. However, omics profiles induced by chemical exposures have rarely been systematically considered in mixture contexts. OBJECTIVES: In this study, we aimed to investigate the predictability of chemical mixture effects on the whole-transcriptome scale. METHODS: We predicted and measured the toxicogenomic effects of a synthetic mixture on zebrafish embryos. The mixture contained the compounds diuron, diclofenac, and naproxen. To predict concentration- and time-resolved whole-transcriptome responses to the mixture exposure, we adopted the mixture concept of concentration addition. Predictions were based on the transcriptome profiles obtained for the individual mixture components in a previous study. Finally, concentration- and time-resolved mixture exposures and subsequent toxicogenomic measurements were performed and the results were compared with the predictions. RESULTS: This comparison of the predictions with the observations showed that the concept of concentration addition provided reasonable estimates for the effects induced by the mixture exposure on the whole transcriptome. Although nonadditive effects were observed only occasionally, combined, that is, multicomponent-driven, effects were found for mixture components with anticipated similar, as well as dissimilar, modes of action. DISCUSSION: Overall, this study demonstrates that using a concentration- and time-resolved approach, the occurrence and size of combined effects of chemicals may be predicted at the whole-transcriptome scale. This allows improving effect assessment of mixture exposures on the molecular scale that might not only be of relevance in terms of risk assessment but also for pharmacological applications. https://doi.org/10.1289/EHP7773.

Ca 2+ -specific removal of Z lines from rabbit skeletal muscle.

Removal of rabbit psoas strips immediately after death and incubation in a saline solution containing 1 mM Ca(2+) and 5 nM Mg(2+) for 9 hr at 37 degrees C and pH 7.1 causes complete Z-line removal but has no ultrastructurally detectable effect on other parts of the myofibril. Z lines remain ultrastructurally intact if 1 mM 1,2-bis-(2-dicarboxymethylaminoethoxy)-ethane (EGTA) is substituted for 1 mM Ca(2+) and the other conditions remain unchanged. Z lines are broadened and amorphous but are still present after incubation for 9 hr at 37 degrees C if 1 mM ethylenediaminetetraacetate (EDTA) is substituted for 1 mM Ca(2+) and 5 mM Mg(2+) in the saline solution. A protein fraction that causes Z-line removal from myofibrils in the presence of Ca(2+) at pH 7.0 can be isolated by extraction of ground muscle with 4 mM EDTA at pH 7.0-7.6 followed by isoelectric precipitation and fractionation between 0 and 40% ammonium sulfate saturation. Z-line removal by this protein fraction requires Ca(2+) levels higher than 0.1 mM, but Z lines are removed without causing any other ultrastructurally detectable degradation of the myofibril. This is the first report of a protein endogenous to muscle that is able to catalyze degradation of the myofibril. The very low level of unbound Ca(2+) in muscle cells in vivo may regulate activity of this protein fraction, or alternatively, this protein fraction may be localized in lysosomes.

An ecotoxicological view on neurotoxicity assessment.

The numbers of potential neurotoxicants in the environment are raising and pose a great risk for humans and the environment. Currently neurotoxicity assessment is mostly performed to predict and prevent harm to human populations. Despite all the efforts invested in the last years in developing novel in vitro or in silico test systems, in vivo tests with rodents are still the only accepted test for neurotoxicity risk assessment in Europe. Despite an increasing number of reports of species showing altered behaviour, neurotoxicity assessment for species in the environment is not required and therefore mostly not performed. Considering the increasing numbers of environmental contaminants with potential neurotoxic potential, eco-neurotoxicity should be also considered in risk assessment. In order to do so novel test systems are needed that can cope with species differences within ecosystems. In the field, online-biomonitoring systems using behavioural information could be used to detect neurotoxic effects and effect-directed analyses could be applied to identify the neurotoxicants causing the effect. Additionally, toxic pressure calculations in combination with mixture modelling could use environmental chemical monitoring data to predict adverse effects and prioritize pollutants for laboratory testing. Cheminformatics based on computational toxicological data from in vitro and in vivo studies could help to identify potential neurotoxicants. An array of in vitro assays covering different modes of action could be applied to screen compounds for neurotoxicity. The selection of in vitro assays could be guided by AOPs relevant for eco-neurotoxicity. In order to be able to perform risk assessment for eco-neurotoxicity, methods need to focus on the most sensitive species in an ecosystem. A test battery using species from different trophic levels might be the best approach. To implement eco-neurotoxicity assessment into European risk assessment, cheminformatics and in vitro screening tests could be used as first approach to identify eco-neurotoxic pollutants. In a second step, a small species test battery could be applied to assess the risks of ecosystems.

Tetrameric structure of the nonactivated glucocorticoid receptor in cell extracts and intact cells.

Mouse lymphoma cells contain a nonactivated glucocorticoid receptor of Mr approximately 330,000 which is heteromeric in nature and is unable to bind to DNA. Following affinity labeling of the steroid-binding subunit and subsequent cross-linking with dimethyl suberimidate at various times either in cell extracts or in intact cells, a series of labeled bands was detected in SDS gels. From the molecular masses of completely and partially cross-linked complexes we conclude that the large nonactivated receptor is a tetramer composed of two 90 kDa subunits, one 50 kDa polypeptide and one steroid-binding subunit.

Subunit dissociation and activation of wild-type and mutant glucocorticoid receptors.

Apparent molecular weights of wild-type and nti ('increased nuclear transfer') mutant glucocorticoid receptors were obtained from Stokes radii and sedimentation coefficients. At low salt concentrations molecular forms of Mr 328,000 and 298,000 of the wild-type and mutant, respectively, were predominant. Increasing ionic strength resulted in receptor dissociation. Dissociated forms of Mr 130,000 and 63,000 of the wild-type and mutant, respectively, were obtained at 300 mM KCl and above. Some metal oxi-anions prevented dissociation. Receptor activation to allow DNA binding produced the dissociated forms which could be separated from non-activated receptors by filtration through DNA-cellulose or by DEAE-cellulose chromatography. Non-activated wild-type and nti receptors eluted from DEAE-cellulose under identical conditions while activated wild-type and nti receptors eluted differently. Partially proteolyzed wild-type receptors behaved identically to nti receptors. We conclude that the large forms of wild-type and nti receptors are heteromeric and contain only one hormone-building polypeptide per complex.

Risk for trisomy 21 in offspring of individuals who have relatives with trisomy 21.

This study was performed to determine if sibs and other relatives of individuals with trisomy 21 are themselves at increased risk for having offspring with trisomy 21. The results suggest that the reproductive risk to these relatives is not increased beyond the risk to the general population.

Subunit structure of the glucocorticoid receptor and activation to the DNA-binding state.

Glucocorticoid receptors of S49.1 mouse lymphoma cells were analyzed under a variety of conditions. The complexes with an agonist or a steroidal antagonist can be formed in cytosolic extracts, they are of high molecular weight, Mr approximately 330,000 and have a Stokes radius of 82 A. Cross-linking by several agents stabilized this structure against subunit dissociation which produces the activated receptor form of 60 A and DNA-binding ability. Careful analysis of intermediate cross-linked forms lead to the conclusion that the large receptor structure is a hetero-tetramer consisting of one hormone-bearing polypeptide of Mr approximately 94,000, two 90 kDa subunits and a protein component of Mr approximately 50,000. The 90 kDa subunits are the heat shock protein hsp90. The high molecular weight receptor form also exists in intact cells as revealed again by cross-linking. The cytosolic complex with the antagonist can become activated to the DNA-binding form upon warming but simultaneously looses the ligand. Ligand rebinding does not occur subsequent to receptor dissociation. Upon incubation of intact cells at 37 degrees C with agonist or antagonist the respective receptor-ligand complexes are formed. The agonist complex is immediately activated, however, the antagonist complex remains stable in the undissociated state. This explains the biological effect of the antagonist.

Ciprofloxacin in the treatment of chronic bacterial prostatitis: a prospective, non-comparative multicentre clinical trial with long-term follow-up. The German Prostatitis Study Group.

The efficacy and safety of ciprofloxacin 500 mg bd for 28 days were assessed in 65 adult males with symptomatic bacterial prostatitis, from eight centres in Germany. Urine and prostatic secretions were obtained for culture. Clinical signs and symptoms were evaluated at 12-18 days during treatment, end of treatment (4-9 days post-treatment), and 1, 3, 6 and 9 months post-treatment. Safety was monitored during and at the end of treatment. E. coli was the most frequent pathogen causing infection (35/62) and Enterococcus faecalis the second most frequent (13/62). The combined bacteriological response by patient determined at return visits 4-9 days post and 1 month post-treatment, respectively, was eradication 48/54 (88.9%), persistence 3/54 (5.6%), eradication with recurrence 1/54 (1.9%) and eradication with reinfection 2/54 (3.7%). The clinical response at 1 month post-treatment was resolution 53/54 (98. 1%) and 1/54 (1.9%) failure. The rates for continued eradication in the extended follow-up were 32/39 (82.1%) after 3 months, 26/34 (76. 4%) after 6 months and 13/22 (59.1%) after 9 months. Nineteen patients experienced at least one adverse event. In two patients the trial was prematurely discontinued due to adverse events.

The importance of complementary cytology and colposcopy in a detection program.

Since 1956, we have used cytology and colposcopy routinely as complementary methods and the percentage of positive results has brought us much closer to the ideal 100 per cent. In order to achieve this, it is essential to put to good use the experience acquired by cytologists and colposcopists and to constantly update and improve the technique employed. The population must be made aware of the benefits of the simultaneous use of these detection methods through health education. The possibilities of diagnostic prediction of cytology are already well known. By means of colposcopy, we may classify the findings as follows: 1) normal, 2) with benign pathology, 3) suspicious, requiring additional frequent control, 4) highly suspicious of neoplasm, 5) malignant. Using the combined methodology, we were able to detect the flaws in use of both methods separately. Considering the carcinoma in situ and microcarcinoma, the results amounted to 16.6 per cent of cytologic false negatives and 12.9 per cent of colposcopic false negatives, whereas applying both methods simultaneously, we obtained only 2.7 per cent of false negatives. As we mentioned previously, this percentage of error may be attributable to technical faults, associated pathology and/or anatomic-histologic localization. We emphasize our reference to the simplicity of our method, making it possible to dispense with exaggerated human and material requirements and, at the same time, obtain highly successful results.

Moxifloxacin in acute exacerbations of chronic bronchitis: clinical evaluation and assessment by patients.

The clinical success of a 5-day course of oral moxifloxacin (administered once daily at a dose of 400 mg) was evaluated in 328 patients with acute exacerbations of chronic bronchitis (Anthonisen type 1) in a non-comparative study conducted by chest physicians in private practice. Results were assessed on the basis of clinical parameters and, for the first time in a trial involving oral moxifloxacin, by the surrogate marker of patient satisfaction. Improvement in (and severity of) cough, dyspnoea, chest pain and sputum were scored daily by patients. Cough, chest pain and purulent sputum production improved rapidly within the first 5 days of treatment. At least 90% of patients were satisfied with the antibiotic. The clinical success rate (cure and improvement) for all patients involved (intent-to-treat analysis) was 90.5%. The most commonly experienced adverse events were gastrointestinal related, with diarrhoea the most frequent of these (2.7% of all patients).

[The microflora of the peritoneal exudate in patients with disseminated peritonitis]. / Izuchenie mikroflory peritoneal'nogo ékssudata u bol'nykh s rasprostranennym peritonitom.

In 11 patients with diffuse peritonitis and 39 patients with general peritonitis the microflora of peritoneal exudate and the sensitivity of isolated microorganisms to antimicrobial preparations were studied. The study revealed that in the total number of microorganisms isolated from peritoneal exudate enterobacteria prevailed in patients with diffuse peritonitis and bacteroids, in patients with general peritonitis. Enterobacteria were most sensitive to ciprofloxacin and claforan, while bacteroids, to ciprofloxacin; at the same time 15.4% of the bacteroid strains under study were resistant to ciprofloxacin.

[The effect of total gnotobiological isolation and antimicrobial preparations on the survival of mice with acute radiation sickness]. / Izuchenie vliianiia obshchei gnotobiologicheskoi izoliatsii i antimikrobnykh preparatov na vyzhivaemost' myshei pri ostroi luchevoi bolezni.

In experiments on 406 CBA mice the effect of antimicrobial preparations, used in combination with total gnotobiotic isolation, on the survival rate of totally irradiated animals was studied. The use of antimicrobial preparations was shown to considerably enhance the survival rate of these animals. Cyprofloxacin, a new preparation of the kinolone row, exhibited a good effect ensuring selective decontamination of the intestine. The use of cyprofloxacin prior to irradiation with the subsequent isolation of the animals in gnotobiotic chambers increased the survival rate of these animals by 2.7 times. But the maximum survival rate of irradiated mice was observed in those cases when antimicrobial preparations were used both before and after irradiation in combination with the total gnotobiotic isolation of the animals.

[The effect of different antimicrobial preparations on the normalization of the intestinal microflora in patients with disseminated peritonitis]. / Vliianie razlichnykh antimikrobnykh preparatov na normalizatsiiu mikroflory kishechnika u bol'nykh s rasprostranennym peritonitom.

These investigations revealed that the treatment of patients with generalized peritonitis with ampiox, gentamicin and metronidazole, as well as with cephalosporins and metronidazole, led to the aggravation of dysbiotic disturbances in normal intestinal microflora, observed in these patients before treatment. Ciprofloxacin had a selective decontaminating effect on the intestine, which facilitated the decrease of suppuration in postoperative wounds.

[The efficacy of different antimicrobial preparations in treating patients with disseminated peritonitis]. / Izuchenie éffektivnosti razlichnykh antimikrobnykh preparatov pri lechenii bol'nykh s rasprostranennym peritonitom.

The results of our investigations revealed that in 68% of cases the treatment of patients with diffuse peritonitis with ampiox, gentamicin and metronidazole led to arresting the infectious process and to a considerable decrease in microbial contamination of the abdominal cavity. The use of cephalosporins and metronidazole proved to be effective in 85% of cases. In this treatment a considerable decrease in the amount of aerobic and anaerobic bacteria in peritoneal exudate was registered. In the process of treatment with cyprophloxacin positive clinical and microbiological dynamics were observed in all patients.

[The experimental and clinical effect of ciprofloxacin on the microflora of the gastrointestinal tract]. / Vliianie tsiprofloksatsina na mikrofloru zheludochno-kishechnogo trakta v éksperimente i klinike.

The study of the influence of cyprofloxacin on the microflora of the gastrointestinal tract has been made under experimental and clinical conditions. As revealed in this study, cyprofloxacin produces a corrective effect on the intestinal microflora; the action of this preparation, in contrast to that of other antimicrobial preparations, is retained for a long time. In patients having duodenal ulcer with bacteriosis caused by Campylobacter pylori and with intestinal dysbacteriosis the combination of cyprofloxacin and cimetidine yields a higher therapeutic effect than the use of cimetidine alone.

[Behavioral response of cows after intrauterine application of disinfectants for treatment of endometritis]. / Das Verhalten von Kühen nach intrauteriner Applikation von Desinfizienzien zur Behandlung der Endometritis.

Seventy-five animals were investigated in an attempt to study the chronological process of response to vaginal irritation by application of disinfectants for endometritis therapy. Aqueous peracetic acid solutions and Lugol solution were used. Cows reacted to intrauterine application of disinfectant drugs with tail lifting, pressing and sometimes groaning as expression of pain. We tried to quantify this response by registration of these different signs longer than two hours after treatment. The responses are caused by reflux of solution from the uterus into the vagina. They were different and depending on reflux volume, used drugs and their concentration. In comparison to peracetic acid solutions, the application of Lugol with 0.5% iodine seemed to induce a more intensive response to application.

[A phagocytosis test for the evaluation of the local immunity status during endometritis therapy with special regard to the use of uterofertil in cattle]. / Ein Phagozytosetest zur Einschätzung der lokalen Abwehrsituation bei Endometritistherapie unter besonderer Berücksichtigung des Uterofertileinsatzes beim Rind.

After the influence of organic acids or peroxide acids with low concentration upon mucous membranes results a higher activity of the topical defence-situation. The application of a 0.2 per cent solution of peroxyethane acid (drug Uterofertil) into the uterus for endometritis-therapy directs an increase of phagocytosis-activity not only topical but also in the peripheric circulation of blood. The phagocytosis-activity was measured with a test by granulocytes. Other drugs were also tested, which will needed by endometritis-treatment of cattle.

Germline Mutations in RASA1 Are Not Found in Patients with Klippel-Trenaunay Syndrome or Capillary Malformation with Limb Overgrowth.

The RASA1 gene encodes p120RASGAP, a multidomain cytoplasmic protein that acts as a negative regulator of the RAS signalling pathway. Heterozygous loss-of-function RASA1 mutations were identified in patients with Parkes Weber syndrome and multifocal capillary malformations. This syndrome is characterised by a capillary blush on an extremity, arteriovenous microfistulas, and bony and soft tissue hypertrophy. The aim of this study was to test RASA1 in 2 disorders characterised by asymmetric limb enlargement and vascular malformations, namely Klippel-Trenaunay syndrome and regional capillary malformation with overgrowth. We did not identify any clear pathogenic change in these patients. Thus, besides clinical and radiological criteria, RASA1 testing constitutes an additional tool to differentiate Parkes Weber syndrome of capillary malformation-arteriovenous malformation (CM-AVM) from overlapping disorders.