Germline ATM variants predispose to melanoma: a joint analysis across the GenoMEL and MelaNostrum consortia.

PURPOSE: Ataxia-Telangiectasia Mutated (ATM) has been implicated in the risk of several cancers, but establishing a causal relationship is often challenging. Although ATM single-nucleotide polymorphisms have been linked to melanoma, few functional alleles have been identified. Therefore, ATM impact on melanoma predisposition is unclear. METHODS: From 22 American, Australian, and European sites, we collected 2,104 familial, multiple primary (MPM), and sporadic melanoma cases who underwent ATM genotyping via panel, exome, or genome sequencing, and compared the allele frequency (AF) of selected ATM variants classified as loss-of-function (LOF) and variants of uncertain significance (VUS) between this cohort and the gnomAD non-Finnish European (NFE) data set. RESULTS: LOF variants were more represented in our study cohort than in gnomAD NFE, both in all (AF = 0.005 and 0.002, OR = 2.6, 95% CI = 1.56-4.11, p < 0.01), and familial + MPM cases (AF = 0.0054 and 0.002, OR = 2.97, p < 0.01). Similarly, VUS were enriched in all (AF = 0.046 and 0.033, OR = 1.41, 95% CI = 1.6-5.09, p < 0.01) and familial + MPM cases (AF = 0.053 and 0.033, OR = 1.63, p < 0.01). In a case-control comparison of two centers that provided 1,446 controls, LOF and VUS were enriched in familial + MPM cases (p = 0.027, p = 0.018). CONCLUSION: This study, describing the largest multicenter melanoma cohort investigated for ATM germline variants, supports the role of ATM as a melanoma predisposition gene, with LOF variants suggesting a moderate-risk.

Low frequency of VHL germline mutations in Norwegian patients presenting with isolated central nervous system hemangioblastomas--a population-based study.

OBJECTIVES: Explore the genetic and clinical incidence of von Hippel-Lindau disease in patients presenting with isolated central nervous system hemangioblastomas. RESULTS: We report a 3.2% (1/31) and 25% (8/32) incidence of genetic and clinical VHL, respectively. One patient tested positive for a VHL mutation that has not previously been reported. This genotype phenotypically predicts VHL type 2B. We had seven patients with renal cysts. In a total follow-up of 33 person years, none of these cysts progressed to renal cell carcinoma. CONCLUSION: von Hippel-Lindau disease anchored in germline mutations of the VHL gene is rare in the Norwegian population as opposed to clinical VHL disease, which appears to be relatively common in patients with apparently sporadic hemangioblastomas. There exists insufficient data regarding the natural history of patients with renal cysts, which makes it difficult to include or disregard these lesions as an entity of VHL disease.

Can morphology predict 1p/19q loss in oligodendroglial tumours?

AIMS: To investigate the relationship between phenotype and genotype in oligodendroglial tumours and evaluate whether 1p/19q status can be reliably predicted from histological findings. METHODS AND RESULTS: Three neuropathologists reviewed the association between 10 histological variables, location and genetic losses at 1p, 19q and 17p13 in 63 oligodendroglial tumours (cohort 1). Based on these findings, a multiple logistic regression model for prediction of 1p/19q status was constructed. The ability of this model to predict 1p/19q status was tested on cohort 2, comprising 20 oligodendroglial tumours. Loss of heterozygosity at 1p, 19q and 17p13 was analysed using polymerase chain reaction. Combined 1p/19q loss and losses at 17p13 were mutually exclusive (P < 0.001). The variable H1a (more or <50% of cells with round, uniform nuclei and perinuclear halos) demonstrated the strongest association with 1p/19q status (odds ratio 11.9, 95% confidence interval 3.6, 39.6, P < 0.001). Calcifications, absence of gemistocytic cells and a non-temporal/non-insular location were also associated. The correct 1p/19q status was predicted in 80% of cases in cohort 2. CONCLUSIONS: There is a strong association between phenotype and genotype in oligodendroglial tumours. However, even when all significant variables are accounted for, perfect prediction (100%) of 1p/19q status cannot be obtained.

Tyrosinaemia type I--de novo mutation in liver tissue suppressing an inborn splicing defect.

Many patients with tyrosinaemia type 1 have a mosaic pattern of fumarylacetoacetase (FAH) immunopositive or immunonegative nodules in liver tissue. This phenomenon has been explained by a spontaneous reversion of the mutation in one allele to a normal genotype, but only a few nodules have been examined. We now report on a Norwegian patient, compound heterozygous for the mutations IVS12g(+5)-->a and G(1009-->)A, with liver mosaicism, but with an immunopositive nodule in which both primary mutations were intact. In the immunopositive hepatocytes of this nodule, genetic analyses showed a new mutation, C(1061-->)A, 6 bp upstream of the primary mutation IVS12g(+5)-->a in the FAH gene. The splicing defect caused by the primary mutation is most likely suppressed by the new mutation due to improvement of the splicing site. In the same liver we demonstrate another nodule of regenerating immunopositive tissue due to reversion of one of the primary mutations to a normal genotype. Together with the original cells this makes a triple mosaicism of hepatocytes with one, two or three point mutations in the FAH gene.

Modulation of COX-2 expression in peripheral blood cells by increased intake of fruit and vegetables?

BACKGROUND: Enhanced cyclooxygenase-2 (COX-2) expression is associated with carcinogenesis, ischemia, angiogenesis, inflammation, and neurodegeneration. The preventing effect of aspirin and nonsteroidal anti-inflammatory drugs is partly due to inhibition of the COX-2 enzyme. Fruit and vegetables (FVs) contain numerous compounds that may decrease disease risk by several different mechanisms, for example through the inhibition of COX-2 activity. OBJECTIVE: We tested the hypothesis that an increased intake of FVs would modulate the COX-2 expression in peripheral blood cells. DESIGN: A strictly controlled dietary crossover study (n = 39). After 1 week run-in period with no FVs in the diet, one group was given two portions of FVs (2 FV), while another group was given five portions (5 FV) daily for 14 days. Following a 2 weeks washout period and 1 week run-in, the regimens were switched between the groups. Gene expression analysis of COX-2 mRNA in blood samples was performed by quantitative real-time-PCR. RESULTS: No significant treatment effect of diet intervention was found in the crossover analyses (P = 0.74). However, the individual variation in response may seem large. CONCLUSIONS: These data does not contradict the recommendations for an increased intake of FVs. Further studies on expression directly and indirectly, through analysis of factors regulating and being regulated by COX-2, should be carried out. A first step would be to evaluate the correspondence between COX-2 mRNA expression and products of the COX pathway, like prostaglandins. Naturally occurring polymorphisms of COX-2 promoters and coding regions might contribute to functional variations and response to different diets. SPONSORSHIP: Norwegian Research Council, National Nutrition Council, Throne Holst Foundation for Nutrition Research, Freia Chokoladefabriks Medisinske Fond and the Norwegian Cancer Society.

Quantitative effects of medium hardness and nutrient availability on the swarming motility of Serratia liquefaciens.

We report the first controlled measurements of expansion rates for swarming colonies of Serratia liquefaciens under different growth conditions, combined with qualitative observations of the organization of the colony into regions of differentiated cell types. Significantly, the results reveal that swarming colonies of S. liquefaciens can have an increasing expansion rate with time. We compare and contrast the expansion rate results with predictions from a recent mathematical model which coupled key hydrodynamical and biological mechanisms. Furthermore, we investigate whether the swarming colonies grow according to a power law or exponentially (for large times), as suggested by recent theoretical results.

Non-rigid registration by geometry-constrained diffusion.

Assume that only partial knowledge about a non-rigid registration is given: certain points, curves or surfaces in one 3D image are known to map to certain points, curves or surfaces in another 3D image. In trying to identify the non-rigid displacement field, we face a generalized aperture problem since along the curves and surfaces, point correspondences are not given. We will advocate the viewpoint that the aperture and the 3D interpolation problem may be solved simultaneously by finding the simplest displacement field. This is obtained by a geometry-constrained diffusion, which in a precise sense yields the simplest displacement field. The point registration obtained may be used for segmentation, growth modeling, shape analysis or kinematic interpolation. The algorithm applies to geometrical objects of any dimensionality. We may thus keep any number of fiducial points, curves and/or surfaces fixed while finding the simplest registration. Examples of inferred point correspondences in a synthetic example and a longitudinal growth study of the human mandible are given.

Simultaneous quantification of etoposide and etoposide phosphate in human plasma by capillary electrophoresis using laser-induced native fluorescence detection.

A method using capillary electrophoresis with UV laser-induced native fluorescence detection was developed as a sensitive and selective assay for the simultaneous determination of etoposide and etoposide phosphate in human plasma. Laser-induced native fluorescence detection with a frequency-doubled argon ion laser at an excitation wavelength of 257 nm was used for the simultaneous assay of etoposide and etoposide phosphate in plasma to improve the sensitivity compared to that obtained with UV absorption. The detection system consists of an imaging spectrograph and an intensified CCD camera which views an illuminated 1.5-mm section of the capillary. This setup is able to record the whole emission spectra of the analytes to achieve additional wavelength-resolved electropherograms. In the concentration range of 200 microg/L-50 mg/L in plasma for etoposide and 100 microg/L-20 mg/L for etoposide phosphate, coefficients of correlation were better than 0.998. Within-day variation determined with three different concentrations showed accuracies ranging from 91.0 to 109.3% for etoposide and from 91.2 to 109.9% for etoposide phosphate (n = 6) with a precision of about 8%. Day-to-day variation presented accuracies ranging from 91.8 to 107.9% for etoposide and from 94.4 to 109.3% for etoposide phosphate with a relative standard deviation less than 6% (n = 5). To our knowledge, this is the first method for the simultaneous quantification of etoposide and etoposide phosphate in plasma samples.

High efficiency beam splitter for multifocal multiphoton microscopy.

In this article we present the development of a multibeam two-photon laser scanning microscope. A new type of beam splitter to create the multitude of laser beams is described. This type of beam splitter has higher transmission and generates more uniform beams than can be achieved with the microlens approach used by other groups. No crosstalk exists between the different foci due to small temporal delays between the individual beams. The importance of dispersion compensation to obtain maximum efficiency of the microscope is discussed. With optimum compensation the fluorescence signal was raised by a factor of 14. Different modes of detecting the fluorescence signals and their effect on imaging speed and resolution are discussed.

[Genetic analysis in familial adenomatous polyposis]. / Genetiske analyser ved familiaer adenomatøs polypose.

BACKGROUND: Familial adenomatous polyposis (FAP) is an autosomal dominantly inherited disorder caused by germline mutations in the APC gene. FAP is characterised by a variable, but normally large number of colorectal adenomas and variations in extracolonic manifestations. These variations are associated with specific mutations of the APC gene. MATERIAL AND METHODS: Representatives from 70 Norwegian families are under molecular investigation. Analyses have so far been concentrated on the part of the APC gene associated with classic FAP. RESULTS: Germline mutations causing FAP have been identified in 36 of the 70 families examined. All mutations identified are confined to the first half of the gene and correlate to classic FAP. INTERPRETATION: Because of the mutation heterogeneity in FAP, the size of the APC gene and variations in phenotype, it is a laborious task to identify the causative mutations. Better approaches to the analysis of the whole APC have now been established and will result in a higher degree of mutation detection independent of phenotype. Family history and phenotype-genotype correlations are still important guidelines for efficient molecular genetic analysis of the APC gene. Genetic surveillance, personal and socio-economic benefits from presymptomatic and predictive testing of members of FAP families are discussed.

Direct chiral assay of tramadol and detection of the phase II metabolite O-demethyl tramadol glucuronide in human urine using capillary electrophoresis with laser-induced native fluorescence detection.

A chiral separation using carboxymethyl-beta-cyclodextrin and methyl-beta-cyclodextrin for the direct assay of tramadol in human urine by capillary electrophoresis (CE) with laser-induced native fluorescence detection was developed. Furthermore, the phase II metabolite O-demethyl tramadol glucuronide was determined from the urine samples and the ratio of the diasteromers was determined. The chiral method was validated. Correlation coefficients were higher than 0.999. Within day variation showed accuracy in the range 96.1-105.8% with a RSD less than 6.00%. Day to day variation present an accuracy ranging from 100.2 to 103.5% with a RSD less than 5.4%. After oral administration of 150 mg tramadol hydrochloride to a healthy volunteer, the urinary excretion was monitored during 24 h. About 11.4% of the dose was excreted as 1S,2S-tramadol, 16.4% as 1R,2R-tramadol and 23.7% as O-demethyl tramadol glucuronide. The amount of 1S,2S O-demethyl tramadol glucuronide was more than three fold higher as IR,2R-O-demethyl tramadol glucuronide. The enantiomeric ratio of tramadol and the diastereomeric ratio of O-demethyl tramadol glucuronide was deviated from 1.0 showing that a stereoselective metabolism of tramadol occurs.

Capillary electrophoresis with laser-induced fluorescence in clinical drug development routine application and future aspects.

The clinical bioanalytical setting is characterized by sample volumes of < 1 ml biological fluid (e.g. plasma, urine), a range of 3-4 decades of concentrations to be quantified and a limit of quantitation in the microg/l-ng/l range for sets of 100-5000 individual samples. Setup of capillary electrophoresis (CE) for routine application in this analytical field was successful for analytes accessible to fluorescence detection by using laser-induced fluorescence (LIF) detection. Empowerment of CE-LIF for routine serial analysis of thousands of samples includes improvement in autosampler techniques, thorough procedures for capillary treatment and particularly more advanced detection technology. Introduction of multi-capillary systems with charge-coupled device cameras and frequency doubled Ar-ion laser (lambda = 257 nm) offers this technique the chance of superiority over classical analytical assays - especially in the field of (new) low volume samples e.g. capillary blood or microdialysate encouraging clinicians to search for meaningful non-invasive samples.

Comment on "Flow-distributed oscillations: stationary chemical waves in a reacting flow"

In a recent paper by Kaern and Menzinger [Phys. Rev. E 60, R3471 (1999)] a successful verification of the stationary space-periodic structures predicted by Andresen et al. [Phys. Rev. E 60, 297 (1999)] was reported. Kaern and Menzinger suggest a mechanism for the formation of such structures that yields a linear relationship between the selected wavelength and the flow rate. We find this mechanism too simple and produce numerical simulations that support the original interpretation of these structures.

Assay of tramadol in urine by capillary electrophoresis using laser-induced native fluorescence detection.

Capillary electrophoresis (CE) with UV laser-induced native fluorescence detection was developed as a sensitive and selective assay for the direct determination of tramadol in human urine without extraction or preconcentration. The main problem in CE is the small inner diameter of the capillary which causes a low sensitivity with instruments equipped with a UV detector. Laser-induced native fluorescence with a frequency doubled argon ion laser at an excitation wavelength of 257 nm was used for the direct assay of tramadol in urine to enhance the limit of detection about 1,000-fold compared to UV absorption detection. The detection system consists of an imaging spectrograph and an intensified CCD camera, which views an illuminated 1.5 mm section of the capillary. This set-up is able to record the whole emission spectra of the analytes to achieve additionally wavelength-resolved electropherograms. In the concentration range of 20 ng/ml-5 microg/ml in human urine coefficients of correlation were better than 0.998. Within-day variation determined on four different concentrations showed accuracies ranging from 90.2 to 108.4%. The relative standard deviation (RSD) was determined to be less than 10%. Day-to-day variation presented accuracies ranging from 90.9 to 103.1% with an RSD less than 8%.

[Leflunomide for active rheumatoid arthritis]. / Leflunomid til aktiv reumatoid arthritis.

In 1999, leflunomide was introduced for the treatment of active rheumatoid arthritis. Leflunomide is a reversible inhibitor of "de novo" synthesis of pyrimidine, resulting in a restriction of lymphocyte proliferation. The pharmacodynamics are characterized by slow elimination. Leflunomide has shown an efficacy and a pattern of mild and serious side effects similar to methotrexate and sulphasalazine. Treatment is started with a loading dose of 100 mg orally for three days followed by 20 (10) mg daily. Regular haematological measurements, determination of serum chemistry and blood pressure should be performed every second week the first six months, subsequently every sixth week. If the treatment is to be terminated abruptly, a drug elimination procedure must be performed. Leflunomide will find its place in the treatment of rheumatoid arthritis. Leflunomide should follow an unsuccessful treatment attempt with methotrexate or sulphasalazine.

The interaction of thin-film flow, bacterial swarming and cell differentiation in colonies of Serratia liquefaciens.

The rate of expansion of bacterial colonies of S. liquefaciens is investigated in terms of a mathematical model that combines biological as well as hydrodynamic processes. The relative importance of cell differentiation and production of an extracellular wetting agent to bacterial swarming is explored using a continuum representation. The model incorporates aspects of thin film flow with variable suspension viscosity, wetting, and cell differentiation. Experimental evidence suggests that the bacterial colony is highly sensitive to its environment and that a variety of mechanisms are exploited in order to proliferate on a variety of surfaces. It is found that a combination of effects are required to reproduce the variation of bacterial colony motility over a large range of nutrient availability and medium hardness.

Surface-bounded growth modeling applied to human mandibles.

From a set of longitudinal three-dimensional scans of the same anatomical structure, we have accurately modeled the temporal shape and size changes using a linear shape model. On a total of 31 computed tomography scans of the mandible from six patients, 14,851 semilandmarks are found automatically using shape features and a new algorithm called geometry-constrained diffusion. The semilandmarks are mapped into Procrustes space. Principal component analysis extracts a one-dimensional subspace, which is used to construct a linear growth model. The worst case mean modeling error in a cross validation study is 3.7 mm.

[Efficacy and tolerance of a calcium-magnesium-aspartate solution in the treatment of hypocalcemic parturient paresis in cows]. / Zur Wirksamkeit und Verträglichkeit einer Kalzium-Magnesium-Aspartat-Lösung bei der Behandlung der hypokalzämischen Gebärparese des Rindes.

The systemic tolerance of a solution of calcium aspartate and magnesium aspartate was studied in 7 cows. Intravenously administered dosages of 500 ml per cow were well tolerated. A twofold increase of the serum calcium concentration was measured. In 2 cows which were treated with 1000 ml of the solution a threefold increased calcium concentration and heart arrhythmia were found. The clinical efficacy of the solution was demonstrated in a study with 44 hypocalcemic cows. A long lasting increase of the serum calcium as well as an enhanced phosphorus concentration were measurable. In conclusion, the calcium-magnesium-aspartate solution seems to be an efficacious and well tolerated alternative for the treatment of hypocalcemia in cows.

A possible contributory role of BK virus infection in neuroblastoma development.

The tumor suppressor protein p53 is aberrantly localized to the cytoplasm of neuroblastoma cells, compromising the suppressor function of this protein. Such tumors are experimentally induced in transgenic mice expressing the large tumor (T) antigen of polyomaviruses. The oncogenic mechanisms of T antigen include complex formation with, and inactivation of, the tumor suppressor protein p53. Samples from 18 human neuroblastomas and five normal human adrenal glands were examined. BK virus DNA was detected in all neuroblastomas and none of five normal adrenal glands by PCR. Using DNA in situ hybridization, polyomaviral DNA was found in the tumor cells of 17 of 18 neuroblastomas, but in none of five adrenal medullas. Expression of the large T antigen was detected in the tumor cells of 16 of 18 neuroblastomas, but in none of the five adrenal medullas. By double immunostaining BK virus T antigen and p53 was colocalized to the cytoplasm of the tumor cells. Immunoprecipitation revealed binding between the two proteins. The presence and expression of BK virus in neuroblastomas, but not in normal adrenal medulla, and colocalization and binding to p53, suggest that this virus may play a contributory role in the development of this neoplasm.