In vitro perfusion of whole bovine ovaries by freezing medium: effect of perfusion rate and elapsed time after extraction.

BACKGROUND: Cryopreservation and transplantation of the whole ovary with vascular pedicle would be helpful to prevent posttransplantation ischemia. In fact, perfusion of the intact mammalian ovary through arteries and veins is the most technically difficult part of the whole cryopreservation process because of its complexity. It is important to develop the technology of long-time perfusion of intact ovaries by cryoprotectants at low temperatures because it was established earlier that 24-hour cooling to 5 degrees C before cryopreservation is beneficial for the freezing of human ovarian tissue. The aim of this research was to study the effectiveness of perfusion of intact bovine ovaries with different rates of perfusion and elapsed time between extraction of these ovaries and beginning of perfusion. METHODS: Arteria ovarica was cannulated and ovaries were perfused with Leibovitz L-15 medium + 100 IU/mL heparin + 5% bovine calf serum + 6% dimethyl sulfoxide + 6% ethylene glycol + 0.15 M sucrose + Indian ink at room temperature (22 degrees C). In the first cycle of experiments, ovaries (n = 145) were perfused for 60 minutes during 1 to 1.5 hours after extraction of ovaries in the slaughter house at perfusion rates of 150 mL/hour (2.5 mL/minute), 100 mL/hour (1.67 mL/minute), 75 mL/hour (1.25 mL/minute), 50 mL/hour (0.83 mL/minute), 25 mL/hour (0.42 mL/minute), and 12.5 mL/hour (0.21 mL/minute) for groups 1, 2, 3, 4, 5, and 6, respectively. In the second cycle of experiments, ovaries (n = 29) were perfused with a rate of 25 mL/hour (0.42 mL/minute) for 60 minutes during the following time-periods elapsed after extraction of ovaries in the slaughter house: 3 hours (n = 18), 4 hours (n = 5), 5 hours (n = 3), and 6 hours (n = 3) for groups 1, 2, 3, and 4, respectively. Ovaries in luteal and follicular phase of development were distributed randomly into groups. Successful perfusion of blood vessels was detected visibly by a blue coloration of the vascular pedicle and ovarian tissues. The percentage of Indian ink-perfused tissues was detected. The intensity of the vascular leakage and tissue damage was scored microscopically and noted as follows: lack of disruption (-), weak disruption (+), moderate disruption (++), and strong disruption RESULTS: The first cycle of experiments shows that an optimal perfusion rate was established for groups 4 and 5 (50 and 25 mL/hour, respectively). In the second cycle of experiments, good perfusion of ovaries with the perfusion rate of 25 mL/hour was established only for ovaries of group 1 (3 hours after extraction). The effectiveness of perfusion in group 2 (4 hours after extraction) was sharply decreased. CONCLUSIONS: Effective perfusion of bovine intact ovaries with vascular pedicle with freezing medium (6% ethylene glycol + 6% dimethyl sulfoxide + 0.15 M sucrose) at room temperature includes a rate of perfusion 25 or 50 mL/ hour. Ovaries must be perfused no later than 3 hours after the death of animals.

Improved phylogenomic taxon sampling noticeably affects nonbilaterian relationships.

Despite expanding data sets and advances in phylogenomic methods, deep-level metazoan relationships remain highly controversial. Recent phylogenomic analyses depart from classical concepts in recovering ctenophores as the earliest branching metazoan taxon and propose a sister-group relationship between sponges and cnidarians (e.g., Dunn CW, Hejnol A, Matus DQ, et al. (18 co-authors). 2008. Broad phylogenomic sampling improves resolution of the animal tree of life. Nature 452:745-749). Here, we argue that these results are artifacts stemming from insufficient taxon sampling and long-branch attraction (LBA). By increasing taxon sampling from previously unsampled nonbilaterians and using an identical gene set to that reported by Dunn et al., we recover monophyletic Porifera as the sister group to all other Metazoa. This suggests that the basal position of the fast-evolving Ctenophora proposed by Dunn et al. was due to LBA and that broad taxon sampling is of fundamental importance to metazoan phylogenomic analyses. Additionally, saturation in the Dunn et al. character set is comparatively high, possibly contributing to the poor support for some nonbilaterian nodes.

Contrast agents and applications to assess tumor angiogenesis in vivo by magnetic resonance imaging.

Angiogenesis plays a key role in the development of cancer and is precondition for tumor growth, invasion and spread. Therefore, numerous angiogenesis inhibitors have been developed, of which some show potential to defeat cancer in preclinical and clinical trials. However, response to antiangiogenic treatments is often delayed and marked by high interindividual variability making a closely mashed and efficient observation of the patient necessary. Therefore, surrogate markers which specifically catch early response to tumor therapy are highly desirable. Functional parameters like relative blood volume, perfusion and vessel permeability can be assessed using T(1) and T(2)*-weighted dynamic contrast-enhanced (DCE) MRI. Various reports are available on this topic but results are controversial. During antiangiogenic therapies some authors describe pronounced changes in blood volume: others find effects only on vessel permeability or perfusion. These conflictive observations can be attributed to the different tumor models, therapies, measurement techniques and contrast agents (CA). Particularly the choice of the optimal CA is considered to be essential for a successful characterization of tumor angiogenesis. Often therapy effects on vessel permeability only become apparent, when blood pool CA are used. This article reviews the current state of DCE and molecular MRI of angiogenesis. Besides a general introduction of the different measurement and postprocessing methods and its previous applications, design, structure and use of different types of CA are the main focus of this article.

TICO: a tool for postprocessing the predictions of prokaryotic translation initiation sites.

Exact localization of the translation initiation sites (TIS) in prokaryotic genomes is difficult to achieve using conventional gene finders. We recently introduced the program TICO for postprocessing TIS predictions based on a completely unsupervised learning algorithm. The program can be utilized through our web interface at http://tico.gobics.de/ and it is also freely available as a commandline version for Linux and Windows. The latest version of our program provides a tool for visualization of the resulting TIS model. Although the underlying method is not based on any specific assumptions about characteristic sequence features of prokaryotic TIS the prediction rates of our tool are competitive on experimentally verified test data.

Stability and inactivation of SARS coronavirus.

The SARS-coronavirus (SARS-CoV) is a newly emerged, highly pathogenic agent that caused over 8,000 human infections with nearly 800 deaths between November 2002 and September 2003. While direct person-to-person transmission via respiratory droplets accounted for most cases, other modes have not been ruled out. Faecal shedding is common and prolonged and has caused an outbreak in Hong Kong. We studied the stability of SARS-CoV under different conditions, both in suspension and dried on surfaces, in comparison with other human-pathogenic viruses, including human coronavirus HCoV-229E. In suspension, HCoV-229E gradually lost its infectivity completely while SARS-CoV retained its infectivity for up to 9 days; in the dried state, survival times were 24 h versus 6 days. Thermal inactivation at 56 degrees C was highly effective in the absence of protein, reducing the virus titre to below detectability; however, the addition of 20% protein exerted a protective effect resulting in residual infectivity. If protein-containing solutions are to be inactivated, heat treatment at 60 degrees C for at least 30 min must be used. Different fixation procedures, e.g. for the preparation of immunofluorescence slides, as well as chemical means of virus inactivation commonly used in hospital and laboratory settings were generally found to be effective. Our investigations confirm that it is possible to care for SARS patients and to conduct laboratory scientific studies on SARS-CoV safely. Nevertheless, the agents tenacity is considerably higher than that of HCoV-229E, and should SARS re-emerge, increased efforts need to be devoted to questions of environmental hygiene.

[Analysis of Foetal Heart Rate Data using Complex Software: Comparison of Recurrence Plot of Foetal Heart Rate with the Course of Pregnancy -]. / Weitergehende Analyse von Kardiotokogramm-Daten mittels komplexer Software: Vergleich von "Recurrence Plot", fetaler Herzfrequenz und Schwangerschaftsverlauf - Eine Pilotstudie -

BACKGROUND: Is it useful to further analyse foetal heart rate to improve the prediction of pregnancy complications? The analysis of the foetal heart rate is usually based on the variability of the heart rate, i. e. the more variable the heart rate presents - except a decrease - the better the condition of the foetus is. The same concept is applied in our own analysis which differs only in the presentation of the data. MATERIAL AND METHODS: We analysed 25 non-stress-tests from unselected third trimester pregnancies using sophisticated software. The recurrence plot (RP) is able to rearrange data from foetal heart rate monitoring in order to make the heart rate variability visible. We developed criteria for a normal and an abnormal test result describing the structure of the diagram to predict an uneventful and a high-risk pregnancy, respectively. RESULTS: 11 out of 11 patients with uneventful course and outcome of pregnancy showed a coarse and blurred RP pattern. 12 out of 14 (86 %) patients developing either intrauterine growth retardation or preeclampsia and requiring caesarean section because of foetal heart rate abnormalities showed a fine and clear RP pattern. CONCLUSIONS: Our preliminary results show that it makes sense to further evaluate foetal heart rate variability in order to predict pregnancy complications. Computer programs including the algorithms needed (calculation of the recurrence plot) are not expensive and easy to handle. A widespread use of these programs represents the basis requirement for large controlled clinical trials.

[Primary diagnosis of hormone-secreting pituitary adenoma during pregnancy and after birth -- a rare occurrence]. / Die Erstdiagnose von hormonproduzierenden Hypophysenadenomen in Schwangerschaft und Wochenbett - eine Rarität.

About seven to ten percent of all brain tumours are neoplasias of the pituitary gland. Pituitary gland tumours can cause different clinical symptoms often making it difficult to come to the correct diagnosis. They can lead to severe complications such as hypopituitarism with secondary hypogonadism, hypothyroidism, and adrenocortical insufficiency, compression of the optic tract or obstructive hydrocephalus. We report on two patients with hormone-secreting pituitary tumours that were unknown prior to pregnancy. The first woman suffered from a growth hormone-producing pituitary adenoma, causing persistent headaches after birth. The second woman showed a significant loss of vision and visual field defects in the 32nd week of gestation, caused by a prolactin-producing pituitary tumour.

Infection of cultured intestinal epithelial cells with severe acute respiratory syndrome coronavirus.

To identify a model for the study of intestinal pathogenesis of severe acute respiratory syndrome (SARS) we tested the sensitivity of six human intestinal epithelial cell lines to infection with SARS coronavirus (SARS-CoV). In permissive cell lines, effects of SARS-CoV on cellular gene expression were analysed using high-density oligonucleotide arrays. Caco-2 and CL-14 cell lines were found to be highly permissive to SARS-CoV, due to the presence of angiotensin-converting enzyme 2 as a functional receptor. In both cell lines, SARS-CoV infection deregulated expression of cellular genes which may be important for the intestinal pathogenesis of SARS.

Wilms' tumor and neuroblastoma.

Significant differences exist between the European and North American treatment protocols for Wilms' tumor and neuroblastoma. There are variations in biopsy technique, timing and extent of initial surgery, chemotherapy protocols and dosage routines, as well as the type of salvage therapy. With the consolidation of the two major North American study groups into a single entity (Children's Oncology Group), the European and North American study groups represent the only remaining large-scale venues for treatment comparison. It is important to study and understand the variation in treatment protocols in order to maintain an open forum of scientific investigation that will lead to improving the care and outcome of children with cancer. It is anticipated that the unification of the North American groups will lead to greater interest and scientific cooperation with the European study group. This paper will serve as a forum for such a discussion at a local level.

Treatment of SARS with human interferons.

Effective antiviral agents are needed to treat severe acute respiratory syndrome-associated coronavirus (SARS-CoV) infection. We assessed the antiviral potential of recombinant interferons against two clinical isolates of SARS-CoV--FFM-1, from Frankfurt patients, and Hong Kong--replicated in Vero and Caco2 cells. Interferon beta was five to ten times more effective in Caco2 cells. Interferon alpha effectively inhibited SARS-CoV replication, but with a selectivity index 50-90 times lower than that for interferon beta. Interferon gamma was slightly better than interferon alpha in Vero cell cultures, but was completely ineffective in Caco2 cell cultures. Interferon beta could be useful alone or in combination with other antiviral drugs for the treatment of SARS.

Glycyrrhizin, an active component of liquorice roots, and replication of SARS-associated coronavirus.

The outbreak of SARS warrants the search for antiviral compounds to treat the disease. At present, no specific treatment has been identified for SARS-associated coronavirus infection. We assessed the antiviral potential of ribavirin, 6-azauridine, pyrazofurin, mycophenolic acid, and glycyrrhizin against two clinical isolates of coronavirus (FFM-1 and FFM-2) from patients with SARS admitted to the clinical centre of Frankfurt University, Germany. Of all the compounds, glycyrrhizin was the most active in inhibiting replication of the SARS-associated virus. Our findings suggest that glycyrrhizin should be assessed for treatment of SARS.

MIPS: a database for genomes and protein sequences.

The Munich Information Center for Protein Sequences (MIPS-GSF, Neuherberg, Germany) continues to provide genome-related information in a systematic way. MIPS supports both national and European sequencing and functional analysis projects, develops and maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences, and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the databases for the comprehensive set of genomes (PEDANT genomes), the database of annotated human EST clusters (HIB), the database of complete cDNAs from the DHGP (German Human Genome Project), as well as the project specific databases for the GABI (Genome Analysis in Plants) and HNB (Helmholtz-Netzwerk Bioinformatik) networks. The Arabidospsis thaliana database (MATDB), the database of mitochondrial proteins (MITOP) and our contribution to the PIR International Protein Sequence Database have been described elsewhere [Schoof et al. (2002) Nucleic Acids Res., 30, 91-93; Scharfe et al. (2000) Nucleic Acids Res., 28, 155-158; Barker et al. (2001) Nucleic Acids Res., 29, 29-32]. All databases described, the protein analysis tools provided and the detailed descriptions of our projects can be accessed through the MIPS World Wide Web server (http://mips.gsf.de).

Impact of total body water errors on Kt/V estimates in children on peritoneal dialysis.

Determining Kt/V in peritoneal dialysis (PD) requires estimation of total body water (TBW). The Dialysis Outcomes Quality Initiative (DOQI) guidelines recommend use of the Mellits and Cheek (MC) formulas for the estimation of TBW in children. However, the MC formulas were developed from healthy children and may not apply to children on PD. Re-evaluating the MC data with additional, recent data from healthy infants has led to the development of new formulas. In addition, and as part of a prospective study of children initiating PD, the Pediatric Peritoneal Dialysis Study Consortium (PPDSC) has directly measured TBW using H2[18O]. To assess the impact of various TBW estimates, KPDt/V values prospectively collected in 24 children were calculated using H2[18O]-measured TBW (O18), MC-derived TBW (MCD), and new-formula TBW (NEW). The mean weekly KPDt/V by O18 was 2.2; by MCD, it was 2.0; and by NEW, it was 2.0. The results derived using the O18 method varied from both the MCD and the NEW results (p < 0.001). The mean deviation from the measured KPDt/V using O18 was 9.5% (maximum: 16%) using the MCD estimate and 7.8% (maximum: 18%) using the NEW formulas. Determinations of KPDt/V are significantly affected by the method of estimating TBW. The PPDSC formulas for children on PD based on the use of H2[18O] offer the most accurate means of calculating TBW and should replace formulas derived from healthy children. The use of Kt/V itself as a marker of adequacy in children will be validated only in prospective studies.

The HIB database of annotated UniGene clusters.

SUMMARY: The HumanInfoBase (HIB) is a database of putative human gene transcripts. UniGene clusters are assembled, and the resulting consensus sequences are submitted to the PEDANT software system (Frishman,D., Albermann,K., Hani,J., Heumann,K., Metanomski,A., Zollner,A. and Mewes,H.-W., 2001, Bioinformatics, 17, 44--57) for fully automatic sequence analysis and annotation. Predicted transcripts are classified using a variety of functional and structural categories, and hyperlinks to various databases are provided for additional information. A WWW-based graphical user interface represents the assembly process as well as functionally important sites in the putative transcripts.

A space-efficient algorithm for aligning large genomic sequences.

SUMMARY: In the segment-by-segment approach to sequence alignment, pairwise and multiple alignments are generated by comparing gap-free segments of the sequences under study. This method is particularly efficient in detecting local homologies, and it has been used to identify functional regions in large genomic sequences. Herein, an algorithm is outlined that calculates optimal pairwise segment-by-segment alignments in essentially linear space. AVAILABILTIY: The program is available at the Bielefeld Bioinformatics Server (BiBiServ) at http://bibiserv.techfak. uni-bielefeld.de/dialign/

Palliative treatment of recurrent obstructing gastric cancer. A case report of successful treatment with three self-expanding metallic stents.

Patients with advanced or recurrent gastric cancer affecting the upper and lower gastrointestinal tract usually experience obstructive symptoms, causing a severe compromise in their quality of life. Surgery may not be feasible because of the patient's precarious medical condition and multilevel tumor infiltration. When faced with these circumstances, surgeons have few options. Parenteral nutrition and comfort measures are utilized when surgical bypass is not a tenable option. We herein describe a unique case of multilevel upper and lower gastrointestinal obstruction secondary to recurrent gastric cancer. The patient was treated palliatively through a combined surgical, radiological, and endoscopic approach by implanting a series of self-expanding metallic stents. To our knowledge, there are no previous reports of successful management of simultaneous strictures of the upper and lower gastrointestinal tract using this technique.

Evolution of bHLH transcription factors: modular evolution by domain shuffling?

Multidomain proteins usually contain several conserved and apparently independently evolved domains. As a result, classifications based on only a single small domain may obscure the true evolutionary relationships of the proteins. The current classification of basic helix-loop-helix (bHLH) domain-containing proteins is based on the conserved bHLH domain alone. Herein, we explore whether sequence homology and, therefore, evolutionary relationships can be detected among the flanking or non-bHLH components of the amino acid sequences of 122 bHLH proteins. These 122 proteins were the same proteins previously used to construct the existing classification of the bHLH-domain-containing proteins. Several possible scenarios are examined in order to explain the observed patterns of sequence divergence, including (1) monophyly, (2) convergent evolution, (3) addition of functional components to the bHLH domain, and (4) modular evolution with domain shuffling. Drawing on several lines of evidence, we suggest that modular evolution by domain shuffling may have played an important role in the evolution of this large group of transcriptional regulators.

Disseminated varicella infection in pediatric renal transplant recipients treated with mycophenolate mofetil.

BACKGROUND: Mycophenolate mofetil (MMF) is a new immune suppressive agent, effective in the prevention of acute rejection after renal transplantation. METHODS: The study was a retrospective review of records of pediatric renal transplant recipients from 1985 to the present. RESULTS: Since October 1995, the immune suppression protocol for pediatric renal transplant recipients at Mayo Eugenio Litta Children's Hospital has included MMF, prednisone, and cyclosporine A. During that time, 19 children and adolescents have received renal allografts, 17 of whom were seropositive for varicella antibody before transplantation, while 2 were seronegative. Varicella infection occurred in 3 of 19 patients (15.8%), all 3 of whom had serologically documented immunity to varicella virus before transplantation. All episodes occurred within 12 months of transplantation. All had generalized vesicular lesions without dermatomal distribution. None of the patients developed fever, respiratory, mucocutaneous, or central nervous system manifestations. All were managed with oral acyclovir, and had an uncomplicated recovery without neuralgia. By contrast, of 74 consecutive patients transplanted before use of MMF, only 1 patient (1.4%) had varicella infection after transplantation (P=0.026). CONCLUSION: The enhanced immunosuppression achieved with MMF appears to be associated with increased susceptibility to varicella infection.

An exact solution for the segment-to-segment multiple sequence alignment problem.

MOTIVATION: In molecular biology, sequence alignment is a crucial tool in studying the structure and function of molecules, as well as the evolution of species. In the segment-to-segment variation of the multiple alignment problem, the input can be seen as a set of non-gapped segment pairs (diagonals). Given a weight function that assigns a weight score to every possible diagonal, the goal is to choose a consistent set of diagonals of maximum weight. We show that the segment-to-segment multiple alignment problem is equivalent to a novel formulation of the Maximum Trace problem: the Generalized Maximum Trace (GMT) problem. Solving this problem to optimality, therefore, may improve upon the previous greedy strategies that are used for solving the segment-to-segment multiple sequence alignment problem. We show that the GMT can be stated in terms of an integer linear program and then solve the integer linear program using methods from polyhedral combinatorics. This leads to a branch-and-cut algorithm for segment-to-segment multiple sequence alignment. RESULTS: We report on our first computational experiences with this novel method and show that the program is able to find optimal solutions for real-world test examples.