Repeatability of Standardized and Normalized Relative CBV in Patients with Newly Diagnosed Glioblastoma.

BACKGROUND AND PURPOSE: For more widespread clinical use advanced imaging methods such as relative cerebral blood volume must be both accurate and repeatable. The aim of this study was to determine the repeatability of relative CBV measurements in newly diagnosed glioblastoma multiforme by using several of the most commonly published estimation techniques. MATERIALS AND METHODS: The relative CBV estimates were calculated from dynamic susceptibility contrast MR imaging in double-baseline examinations for 33 patients with treatment-naïve and pathologically proved glioblastoma multiforme (men = 20; mean age = 55 years). Normalized and standardized relative CBV were calculated by using 6 common postprocessing methods. The repeatability of both normalized and standardized relative CBV, in both tumor and contralateral brain, was examined for each method with metrics of repeatability, including the repeatability coefficient and within-subject coefficient of variation. The minimum sample size required to detect a parameter change of 10% or 20% was also determined for both normalized relative CBV and standardized relative CBV for each estimation method. RESULTS: When ordered by the repeatability coefficient, methods using postprocessing leakage correction and ΔR2*(t) techniques offered superior repeatability. Across processing techniques, the standardized relative CBV repeatability in normal-appearing brain was comparable with that in tumor (P = .31), yet inferior in tumor for normalized relative CBV (P = .03). On the basis of the within-subject coefficient of variation, tumor standardized relative CBV estimates were less variable (13%-20%) than normalized relative CBV estimates (24%-67%). The minimum number of participants needed to detect a change of 10% or 20% is 118-643 or 30-161 for normalized relative CBV and 109-215 or 28-54 for standardized relative CBV. CONCLUSIONS: The ΔR2* estimation methods that incorporate leakage correction offer the best repeatability for relative CBV, with standardized relative CBV being less variable and requiring fewer participants to detect a change compared with normalized relative CBV.

Transcriptome analysis of ozone-responsive genes in leaves of European beech (Fagus sylvatica L.).

Suppression subtractive hybridization (SSH) was performed to isolate cDNAs representing genes that are differentially expressed in leaves of Fagus sylvatica upon ozone exposure. 1248 expressed sequence tags (ESTs) were obtained from 2 subtractive libraries containing early and late ozone-responsive genes. Sequences of 1139 clones (91 %) matched the EBI/NCBI database entries. For 578 clones, no putative function could be assigned. Most abundant transcripts were O-methyltransferases, representing 7 % of all sequenced clones. ESTs were organized into 12 functional categories according to the MIPS database. Among them, 12 % (early)/15 % (late) were associated with disease and defence, 19/11 % with cell structure, 4/10 % with signal transduction, and 9/6 % with transcription. The expression pattern of selected ESTs (ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit [rbcS], WRKY-type transcription factor, ultraviolet-B-repressible protein, aquaporine, glutathione S-transferase, catalase, caffeic acid O-methyltransferase, and pathogenesis-related protein 1 [PR1]) was analysed by quantitative real-time RT-PCR (qRT-PCR) which confirmed changed transcript levels upon ozone treatment of European beech saplings. The ESTs characterized will contribute to a better understanding of forest tree genomics and also to a comparison of ozone-responsive genes in woody and herbaceous plants.

Adenosine triphosphate-evoked currents in cultured dorsal root ganglion neurons obtained from rat embryos: desensitization kinetics and modulation of glutamate release.

Sensory neurons express purinergic P2X receptors on their central and peripheral terminals as well as their cell bodies. ATP activation of these receptors drives action potential firing and glutamate release with potentially important consequences for sensory function. Here we show ATP-gated currents activated in cultured embryonic dorsal root ganglion neurons have heterogeneity of time-courses comparable to those observed in different subpopulations of acutely dissociated adult dorsal root ganglion neurons. The distribution of time-courses across the population of cultured neurons is strongly influenced by culture conditions. Heterogeneity in ATP current kinetics occurs even though immunocytochemical staining reveals a relatively homogeneous and widespread expression of the P2X2 and P2X3 subunits. We show that the time-courses of ATP-gated currents recorded at the cell bodies are mirrored by the time-courses of transmitter release from the dorsal root ganglion nerve terminals, indicating similar P2X receptor properties on the soma and their associated terminals. Our results illustrate a functional heterogeneity of P2X receptor-mediated currents that is strongly influenced by external factors. This heterogeneity in current kinetics may have implications for neuronal function as it constrains the time-course of ATP-mediated modulation of neurotransmitter release at sensory nerve terminals.

Antineuronal antibodies in patients with neuroblastoma and paraneoplastic opsoclonus-myoclonus.

PURPOSE: To identify serologic markers in children with paraneoplastic opsoclonus-myoclonus (POM). MATERIALS AND METHODS: We examined the sera of 64 children with neuroblastoma (16 with POM and 48 age-matched and stage-matched controls) by immunohistochemistry of rat brain and human cerebellum, and by Western blot analysis of protein extracts from human Purkinje cells, cortical neurons, neuroblastoma cell lines, and HuD. RESULTS: Using immunohistochemistry, IgG reactivity against neurons was identified in 13 of 16 POM sera (81%), and 12 of 48 non-POM sera (25%; P<0.001). IgM antineural antibodies were present in 3 of 16 POM sera (19%) and 11 of 48 (23%) non-POM sera. Except for anti-Hu antibodies detected in 10 sera (4 with POM), no other specific reactivities were identified by Western blot analysis of neuronal or of neuroblastoma protein extracts. CONCLUSIONS: We conclude that: 1) patients with neuroblastoma and POM are more likely to harbor antineuronal antibodies than patients without POM; 2) no specific serologic marker of POM was identified, but the frequent presence of antineuronal antibodies suggests that POM is immune-mediated; and 3) anti-Hu antibodies are present in some sera from patients with neuroblastoma, irrespective of the presence of POM.

A serologic marker of paraneoplastic limbic and brain-stem encephalitis in patients with testicular cancer.

BACKGROUND: In patients with cancer, symptoms of limbic and brain-stem dysfunction may result from a paraneoplastic disorder. Paraneoplastic limbic or brain-stem encephalitis occurs more frequently with testicular cancer than with most other cancers. We sought antineuronal antibodies that might be used in a diagnostic test for this syndrome. METHODS: Immunohistochemical and immunoblotting techniques were used to detect serum and cerebrospinal fluid antibodies. Serologic screening of a complementary DNA library and Northern blotting were used to clone the target antigen and determine which tissues expressed it. RESULTS: Of 13 patients with testicular cancer and paraneoplastic limbic or brain-stem encephalitis (or both), 10 had antibodies in serum and cerebrospinal fluid against a 40-kd neuronal protein. These antibodies were used to clone a gene that we call Ma2, which codes for a protein (Ma2) that was recognized by serum from the 10 patients, but not by serum from 344 control subjects. Ma2 was selectively expressed by normal brain tissue and by the testicular tumors of the patients. Ma2 shares homology with Ma1, a "brain-testis-cancer" gene related to other paraneoplastic syndromes and tumors. CONCLUSIONS: The serum of patients with subacute limbic and brain-stem dysfunction and testicular cancer contains antibodies against a protein found in normal brain and in testicular tumors. Detection of these antibodies supports the paraneoplastic origin of the neurologic disorder and could be of diagnostic importance.

Ma1, a novel neuron- and testis-specific protein, is recognized by the serum of patients with paraneoplastic neurological disorders.

The identification of antineuronal antibodies has facilitated the diagnosis of paraneoplastic neurological disorders and the early detection of the associated tumours. It has also led to the cloning of possibly important neuron-specific proteins. In this study we wanted to identify novel antineuronal antibodies in the sera of patients with paraneoplastic neurological disorders and to clone the corresponding antigens. Serological studies of 1705 sera from patients with suspected paraneoplastic neurological disorders resulted in the identification of four patients with antibodies that reacted with 37 and 40 kDa neuronal proteins (anti-Ma antibodies). Three patients had brainstem and cerebellar dysfunction, and one had dysphagia and motor weakness. Autopsy of two patients showed loss of Purkinje cells, Bergmann gliosis and deep cerebellar white matter inflammatory infiltrates. Extensive neuronal degeneration, gliosis and infiltrates mainly composed of CD8+ T cells were also found in the brainstem of one patient. In normal human and rat tissues, the anti-Ma antibodies reacted exclusively with neurons and with testicular germ cells; the reaction was mainly with subnuclear elements (including the nucleoli) and to a lesser degree the cytoplasm. Anti-Ma antibodies also reacted with the cancers (breast, colon and parotid) available from three anti-Ma patients, but not with 66 other tumours of varying histological types. Preincubation of tissues with any of the anti-Ma sera abrogated the reactivity of the other anti-Ma immunoglobulins. Probing of a human complementary DNA library with anti-Ma serum resulted in the cloning of a gene that encodes a novel 37 kDa protein (Mal). Recombinant Mal was specifically recognized by the four anti-Ma sera but not by 337 control sera, including those from 52 normal individuals, 179 cancer patients without paraneoplastic neurological symptoms, 96 patients with paraneoplastic syndromes and 10 patients with non-cancer-related neurological disorders. The expression of Mal mRNA is highly restricted to the brain and testis. Subsequent analysis suggested that Mal is likely to be a phosphoprotein. Our study demonstrates that some patients with paraneoplastic neurological disorders develop antibodies against Mal, a new member of an expanding family of 'brain/testis' proteins.

Testing a new Monte Carlo algorithm for protein folding.

We demonstrate that the recently proposed pruned-enriched Rosenbluth method (PERM) (Grassberger, Phys. Rev. E 56:3682, 1997) leads to extremely efficient algorithms for the folding of simple model proteins. We test it on several models for lattice heteropolymers, and compare it to published Monte Carlo studies of the properties of particular sequences. In all cases our method is faster than the previous ones, and in several cases we find new minimal energy states. In addition to producing more reliable candidates for ground states, our method gives detailed information about the thermal spectrum and thus allows one to analyze thermodynamic aspects of the folding behavior of arbitrary sequences.

Differences between induction effects of 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene and phenobarbitone.

The inductive effects of phenobarbitone (PB) and 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) were compared in C57BL/6J mice. Induction parameters included six substrates: ethylmorphine (EM), benzphetamine (Bph), biphenyl, ethoxycoumarin (EtoC), pentoxyresorufin and dichloro-p-nitroanisole (DPNA). In order to validate this descriptive approach the comparison was extended to diazepam, rifampicin, warfarin, and pregnenolone-16 alpha-carbonitrile (PCN). All inducers were clearly distinguishable from each other. Warfarin was similar to PB, rifampicin was similar to PCN. TCPOBOP differed significantly from PB in relative liver weight, cytochrome P-450 content of liver microsomes, EM-, Bph- and DPNA-demethylations, biphenyl-hydroxylations, EtoC de-ethylation and absorption maximum of reduced CO-cytochrome P-450. TCPOBOP, as an inducer, was less "specific" than PB: total metabolic rates were excessively increased due to microsomal protein (1.5 times) and cytochrome P-450 (4 times) augmentation, whereas cytochrome P-450-related metabolic rates were less increased than those after PB. Thus TCPOBOP does not seem to be as similar to PB as was suggested in the first description of its inducing potency.

Preparation of 11 alpha-hemisuccinyl-progesterone for immunological purposes and structural determination of the hapten.

Progress in reproductive endocrinology to some extent is due to the application of immunogens produced from synthetic haptens. In this preliminary study the synthesis of the hapten 11 alpha-hemisuccinyl-progesterone (11 alpha-hemisuccinyl-pregn-4-ene-3,20-dione) from 11 alpha-hydroxy-progesterone and its complete structure elucidation by u.v./VIS, i.r., [1H]NMR-spectroscopy and E.I.-mass-spectrometry is described. Estimates of sample purity are given.

[A new mandelonitrile lyase from the cherrylaurel (Prunus laurocerasus) (author's transl)]. / Eine neue Mandelsäurenitril-Lyase (D-Oxynitrilase) aus Prunus laurocerasus (Kirschlorbeer)

Mandelonitrile lyase has been isolated from the seeds of Prunus laurocerasus and characterized. The enzyme is a glycoprotein and contains FAD as prosthetic group. It has an absorption spectrum of the hydrophobic type. The molecular weight is 60000. The new mandelonitrile lyase catalyses both formation and cleavage of D-(+)-benzaldehyde cyanohydrin. Despite the existence of marked morphologic and biochemical differences between P. laurocerasus and P. amygdalus (var. sativa) (sweet almond) the enzymes isolated from the seeds of the two Prunoideae species are closely related, as judged from their immunological properties. However they exhibit specific differences in the isoelectric points and quantitative distribution of the three isoenzymes.