Arterial and portal venous liver perfusion using selective spin labelling MRI.

PURPOSE: To investigate the feasibility of selective arterial and portal venous liver perfusion imaging with spin labelling (SL) MRI, allowing separate labelling of each blood supply. METHODS: The portal venous perfusion was assessed with a pulsed EPISTAR technique and the arterial perfusion with a pseudo-continuous sequence. To explore precision and reproducibility, portal venous and arterial perfusion were separately quantified in 12 healthy volunteers pre- and postprandially (before and after meal intake). In a subgroup of 6 volunteers, the accuracy of the absolute portal perfusion and its relative postprandial change were compared with MRI flow measurements of the portal vein. RESULTS: The portal venous perfusion significantly increased from 63 ± 22 ml/100g/min preprandially to 132 ± 42 ml/100g/min postprandially. The arterial perfusion was lower with 35 ± 22 preprandially and 22 ± 30 ml/100g/min postprandially. The pre- and postprandial portal perfusion using SL correlated well with flow-based perfusion (r(2) = 0.71). Moreover, postprandial perfusion change correlated well between SL- and flow-based quantification (r(2) = 0.77). The SL results are in range with literature values. CONCLUSION: Selective spin labelling MRI of the portal venous and arterial blood supply successfully quantified liver perfusion. This non-invasive technique provides specific arterial and portal venous perfusion imaging and could benefit clinical settings where contrast agents are contraindicated. KEY POINTS: • Perfusion imaging of the liver by Spin Labelling MRI is feasible • Selective Spin Labelling MRI assessed portal venous and arterial liver perfusion separately • Spin Labelling based portal venous liver perfusion showed significant postprandial increase • Spin Labelling based portal perfusion correlated well with phase-contrast based portal perfusion • This non-invasive technique could benefit settings where contrast agents are contraindicated.

A new era of ventricular assist device surgery: less invasive procedures.

As the number of patients suffering of congestive heart failure is rising worldwide, the use of mechanical circulatory support to treat these patients has also grown enormously, surpassing the number of annual heart transplants. Moreover latest generation of left ventricular assist devices (LVADs) is characterized by improved technologies. Moreover the size of new LVAD systems is considerably reduced when compared to older generation devices. Therefore, less invasive surgery is now possible for the implantation, explantation, and exchange of LVADs. Although experience with these new techniques is still limited, minimally invasive procedures are thought to improve surgical outcomes by declining the rates of operative complications such as bleeding or wound infection. The miniaturization of LVADs will continue, so that minimally invasive techniques will be used for most LVAD-related procedures in the future. In this article, we summarize and describe minimally invasive surgical techniques, with a focus on the most common LVAD systems in adults.

Liver perfusion in dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI): comparison of enhancement in Gd-BT-DO3A and Gd-EOB-DTPA in normal liver parenchyma.

PURPOSE: Within-patient comparison of the enhancement patterns of normal liver parenchyma after gadobutrol and gadoxetate disodium, with emphasis on the start of hepatocytic uptake of gadoxetate disodium. MATERIALS AND METHODS: Twenty-one patients (12 female, 9 male) without chronic liver disease underwent 1.5-T contrast-enhanced MRI twice, once with an extracellular contrast agent (gadobutrol) and once with a hepatospecific agent (gadoxetate disodium), using a T1-weighted keyhole sequence. Fifteen whole-liver datasets were acquired up to 5 min for both contrast agents and two additional datasets, up to 20 min, for gadoxetate. Signal intensities (SI) of the parenchyma, aorta and portal vein were measured and analysed relative to pre-contrast parenchymal SI. RESULTS: After gadoxetate, in 29% of the patients the parenchymal SI decreased by ≥5% after the initial vascular-phase-induced peak, while in the other 71% the parenchymal SI remained stable or gradually increased until up to 20 min after the initial peak. The hepatocytic gadoxetate uptake started at a mean of 37.8 s (SD 14.7 s) and not later than 76 s after left ventricle enhancement. CONCLUSION: Parenchymal enhancement due to hepatocytic uptake of gadoxetate can start as early as in the late arterial phase. This may confound the assessment of lesion appearance as compared to extracellular contrast such as gadobutrol. KEY POINTS: Gadoxetate-enhanced liver MRI results in early enhancement of normal parenchyma in patients The start of the hepatobiliary phase coincides with the late arterial phase. This may confound the assessment of lesion appearance compared to extracellular contrast. Different parenchymal enhancement patterns after gadoxetate were found for normal parenchyma.

Topological-chiral magnetic interactions driven by emergent orbital magnetism.

Two hundred years ago, Ampère discovered that electric loops in which currents of electrons are generated by a penetrating magnetic field can mutually interact. Here we show that Ampère's observation can be transferred to the quantum realm of interactions between triangular plaquettes of spins on a lattice, where the electrical currents at the atomic scale are associated with the orbital motion of electrons in response to the non-coplanarity of neighbouring spins playing the role of a magnetic field. The resulting topological orbital moment underlies the relation of the orbital dynamics with the topology of the spin structure. We demonstrate that the interactions of the topological orbital moments with each other and with the spins form a new class of magnetic interactions [Formula: see text] topological-chiral interactions [Formula: see text] which can dominate over the Dzyaloshinskii-Moriya interaction, thus opening a path for realizing new classes of chiral magnetic materials with three-dimensional magnetization textures such as hopfions.

Gene targeting reveals a critical role for neurturin in the development and maintenance of enteric, sensory, and parasympathetic neurons.

Neurturin (NTN) is a neuronal survival factor that activates the Ret tyrosine kinase in the presence of a GPI-linked coreceptor (either GFR alpha1 or GFR alpha2). Neurturin-deficient (NTN-/-) mice generated by homologous recombination are viable and fertile but have defects in the enteric nervous system, including reduced myenteric plexus innervation density and reduced gastrointestinal motility. Parasympathetic innervation of the lacrimal and submandibular salivary gland is dramatically reduced in NTN-/- mice, indicating that Neurturin is a neurotrophic factor for parasympathetic neurons. GFR alpha2-expressing cells in the trigeminal and dorsal root ganglia are also depleted in NTN-/- mice. The loss of GFR alpha2-expressing neurons, in conjunction with earlier studies, provides strong support for GFR alpha2/Ret receptor complexes as the critical mediators of NTN function in vivo.

Differential effect of drought regimes on the seedling performance of six floodplain grassland species.

The performance of seedlings is crucial for the survival and persistence of plant populations. Although drought frequently occurs in floodplains and can cause seedling mortality, studies on the effects of drought on seedlings of floodplain grasslands are scarce. We tested the hypotheses that drought reduces aboveground biomass, total biomass, plant height, number of leaves, leaf area and specific leaf area (SLA), and increases root biomass and root-mass fraction (RMF) and that seedlings from species of wet floodplain grasslands are more affected by drought than species of dry grasslands. In a greenhouse study, we exposed seedlings of three confamilial pairs of species (Pimpinella saxifraga, Selinum carvifolia, Veronica teucrium, Veronica maritima, Sanguisorba minor, Sanguisorba officinalis) to increasing drought treatments. Within each plant family, one species is characteristic of wet and one of dry floodplain grasslands, confamilial in order to avoid phylogenetic bias of the results. In accordance with our hypotheses, drought conditions reduced aboveground biomass, total biomass, plant height, number of leaves and leaf area. Contrary to our hypotheses, drought conditions increased SLA and decreased root biomass and RMF of seedlings. Beyond the effects of the families, the results were species-specific (V. maritima being the most sensitive species) and habitat-specific. Species indicative of wet floodplain grasslands appear to be more sensitive to drought than species indicative of dry grasslands. Because of species- and habitat-specific responses to reduced water availability, future drought periods due to climate change may severely affect some species from dry and wet habitats, while others may be unaffected.

Molecular cloning, expression, and chromosome 19 localization of a human Ro/SS-A autoantigen.

Ro/SS-A antibodies are found in a number of human autoimmune disorders including Sjogren's syndrome and several systemic lupus erythematosus-related disorders. These heterogeneous autoantibodies are known to recognize several distinct cellular antigens. With synthetic oligonucleotides corresponding to amino acid sequence information we have isolated a full-length cDNA clone which encodes a human Ro ribonucleoprotein autoantigen. The 1,890-base pair clone contains an open reading frame that encodes a 417-amino acid, 48-kD polypeptide that migrates aberrantly at 60 kD by SDS-PAGE. Rabbit antibodies raised against this protein's recently described amino-terminal epitope react with a previously identified 52-kD human Ro protein and immunoprecipitate the human cytoplasmic RNAs. Ultraviolet light cross-linking studies suggest that this Ro protein binds each of the four major human cytoplasmic RNAs. The deduced amino acid sequence is 63% homologous to an Onchocerca volvulus antigen. Southern filter hybridization analysis indicates that this gene is not highly polymorphic and exists as a single copy in the human genome. Chromosomal localization studies place this gene on the short arm of chromosome 19 near the gene encoding the low density lipoprotein receptor.

Identification and characterization of an Alu-containing, T-cell-specific enhancer located in the last intron of the human CD8 alpha gene.

Expression of the human CD8 alpha gene is restricted to cells of the lymphoid lineage and developmentally regulated during thymopoiesis. As an initial step towards understanding the molecular basis for tissue-specific expression of this gene, we surveyed the surrounding chromatin structure for potential cis-acting regulatory regions by DNase I hypersensitivity mapping and found four hypersensitive sites, three of which were T cell restricted. By using a reporter-based expression approach, a T-cell-specific enhancer was identified by its close association with a prominent T-cell-restricted hypersensitive sites in the last intron of the CD8 alpha gene. Deletion studies demonstrated that the minimal enhancer is adjacent to a negative regulatory element. DNA sequence analysis of the minimal enhancer revealed a striking cluster of consensus binding sites for Ets-1, TCF-1, CRE, GATA-3, LyF-1, and bHLH proteins which were verified by electrophoretic mobility shift assays. In addition, the 5' end of the enhancer was composed of an Alu repeat which contained the GATA-3, bHLH, and LyF-1 binding sites. Site-directed mutation of the Ets-1 and GATA-3 sites dramatically reduced enhancer activity. The functional importance of the other binding sites only became apparent when combinations of mutations were analyzed. Taken together, these results suggest that the human CD8 alpha gene is regulated by the interaction of multiple T-cell nuclear proteins with a transcriptional enhancer located in the last intron of the gene. Comparison of the CD8 alpha enhancer with other recently identified T-cell-specific regulatory elements suggests that a common set of transcription factors regulates several T-cell genes.

The ubiquitous octamer-binding protein(s) is sufficient for transcription of immunoglobulin genes.

All immunoglobulin genes contain a conserved octanucleotide promoter element, ATGCAAAT, which has been shown to be required for their normal B-cell-specific transcription. Proteins that bind this octamer have been purified, and cDNAs encoding octamer-binding proteins have been cloned. Some of these proteins (referred to as OTF-2) are lymphoid specific, whereas at least one other, and possibly more (referred to as OTF-1), is found ubiquitously in all cell types. The exact role of these different proteins in directing the tissue-specific expression of immunoglobulin genes is unclear. We have identified two human pre-B-cell lines that contain extremely low levels of OTF-2 yet still express high levels of steady-state immunoglobulin heavy-chain mRNA in vivo and efficiently transcribe an immunoglobulin gene in vitro. Addition of a highly enriched preparation of OTF-1 made from one of these pre-B cells or from HeLa cells specifically stimulated in vitro transcription of an immunoglobulin gene. Furthermore, OFT-1 appeared to have approximately the same transactivation ability as OTF-2 when normalized for binding activity. These results suggest that OTF-1, without OTF-2, is sufficient for transcription of immunoglobulin genes and that OTF-2 alone is not responsible for the B-cell-specific regulation of immunoglobulin gene expression.

NonO, a non-POU-domain-containing, octamer-binding protein, is the mammalian homolog of Drosophila nonAdiss.

We have cloned the ubiquitous form of an octamer-binding, 60-kDa protein (NonO) that appears to be the mammalian equivalent of the Drosophila visual and courtship song behavior protein, no-on-transient A/dissonance (nonAdiss). A region unprecedently rich in aromatic amino acids containing two ribonuclear protein binding motifs is highly conserved between the two proteins. A ubiquitous form of NonO is present in all adult tissues, whereas lymphocytes and retina express unique forms of NonO mRNA. The ubiquitous form contains a potential helix-turn-helix motif followed by a highly charged region but differs from prototypic octamer-binding factors by lacking the POU DNA-binding domain. In addition to its conventional octamer duplex-binding, NonO binds single-stranded DNA and RNA at a site independent of the duplex site.

Significantly enhanced lung metastasis and reduced organ NK cell functions in diet-induced obese rats.

BACKGROUND: Obesity was identified as a major risk factor for malignant diseases, but underlying mechanisms remain unclear. Natural killer (NK) cells, a pivotal aspect of innate immunity, are capable of identifying and killing virally infected and tumor cells. Previous studies have shown altered NK cell functions in obesity, and the current study aimed to investigate the relationship between altered NK cell functions and increased cancer risk in obesity. METHODS: To induce obesity male F344-rats received a high-fat diet (34% fat) or a control diet (4% fat). Thereafter, syngeneic mammary adenocarcinoma cells (MADB106) or a vehicle were intravenously (i.v.) injected. 15 min after injection, half of each group of rats were killed, lungs removed and immunohistochemically stained. Numbers of NK cells, MADB106 cells and NK cell-tumor cell interactions were quantified. Twenty-one days after tumor-cell injection the other half group of rats was killed and lung metastases were counted and relative mRNA concentrations of different NK cell receptors were determined. RESULTS: After short-term MADB106-challenge, DIO fed animals showed significantly decreased NK cell numbers in the blood and NK cell-tumor cell interactions in the lung as compared to their control littermates. Twenty-one days after MADB106 injection, the lungs of the DIO fed rats showed significantly more lung metastases compared to control animals, accompanied by reduced relative mRNA concentrations of the activating NK cell receptor NKG2D. CONCLUSIONS: We conclude that induction of obesity in F344-rats leads to reduced lung NK cell function against tumor cells and results in significantly enhanced lung metastasis as compared to lean animals. It can be hypothesized that obesity-induced altered NK cell functions play an important role in cancer growth and metastasis.

Expression of activated CDC42 induces T cell apoptosis in thymus and peripheral lymph organs via different pathways.

CDC42, a Ras-related small GTP binding protein, is involved in diverse cellular functions in lymphocytes. We generated transgenic mice expressing constitutively active murine CDC42 (Q61L) under the control of the human CD2 promoter. Transgenic mice showed smaller thymi with a dramatic reduction of CD4+CD8+, CD4+ and CD8+ thymocytes and with increase of CD4-CD8- thymocytes at CD25-CD44+ and CD25+ stage. A high percentage of the transgenic thymocytes were apoptotic, explaining the reduction of cellularity and size of the thymus. Mature T cells (TCR alphabeta+) in peripheral lymph organs, spleen and lymph node, were also dramatically reduced, and exhibited massive apoptosis. Expression of Fas and Fas ligand on both thymocytes and peripheral T cells was upregulated in transgenic mice, but the increased apoptosis in the thymus was independent of Fas (CD95), whereas peripheral spleen and lymph node T cell apoptosis was Fas dependent. Thus, activated CDC42 triggers distinct apoptotic pathways in thymocytes and peripheral T cells.

Repetitive Alu elements form a cruciform structure that regulates the function of the human CD8 alpha T cell-specific enhancer.

We previously identified a T cell-specific enhancer in the last intron of the human CD8 alpha gene that is adjacent to a sequence element that significantly represses enhancer function. This negative regulatory region consists of a half-Alu sequence that has potential to base-pair with a downstream Alu element, which is part of the fully active enhancer, to form a cruciform structure. The activity of this half-Alu silencer sequence is position and orientation-dependent, suggesting that DNA structure plays an important role in its function. Using site-directed mutational analysis and P1 nuclease mapping, we directly demonstrate that formation of a cruciform structure is required for repression of enhancer function in transient transfection assays. Finally, a P1 nuclease-sensitive site is present in the endogenous CD8 alpha gene in T cell lines providing indirect evidence that the stem-loop may form in vivo. Taken together, these results suggest that Alu elements may contribute to the regulation of the CD8 alpha gene enhancer through the formation of secondary structure that disrupts enhancer function.

Merging extracellular domains: fold prediction for laminin G-like and amino-terminal thrombospondin-like modules based on homology to pentraxins.

Using a new method for construction and database searches of sequence consensus strings, we have identified a new superfamily of protein modules comprising laminin G, thrombospondin N and the pentraxin families. The conserved patterns correspond mainly to hydrophobic core residues located in central beta strands of the known three-dimensional structures of two pentraxins, the human C-reactive protein and the serum amyloid P-component. Thus, we predict a similar jellyroll fold for all members of this superfamily. In addition, the conservation of two exposed aspartate residues in the majority of superfamily members suggests hitherto unrecognised functional sites.

Prediction of nonsynonymous single nucleotide polymorphisms in human disease-associated genes.

Analysis of human genetic variation can shed light on the problem of the genetic basis of complex disorders. Nonsynonymous single nucleotide polymorphisms (SNPs), which affect the amino acid sequence of proteins, are believed to be the most frequent type of variation associated with the respective disease phenotype. Complete enumeration of nonsynonymous SNPs in the candidate genes will enable further association studies on panels of affected and unaffected individuals. Experimental detection of SNPs requires implementation of expensive technologies and is still far from being routine. Alternatively, SNPs can be identified by computational analysis of a publicly available expressed sequence tag (EST) database following experimental verification. We performed in silico analysis of amino acid variation for 471 of proteins with a documented history of experimental variation studies and with confirmed association with human diseases. This allowed us to evaluate the level of completeness of the current knowledge of nonsynonymous SNPs in well studied, medically relevant genes and to estimate the proportion of new variants which can be added with the help of computer-aided mining in EST databases. Our results suggest that approx. 50% of frequent nonsynonymous variants are already stored in public databases. Computational methods based on the scan of an EST database can add significantly to the current knowledge, but they are greatly limited by the size of EST databases and the nonuniform coverage of genes by ESTs. Nevertheless, a considerable number of new candidate nonsynonymous SNPs in genes of medical interest were found by EST screening procedure.

Self-organizing hierarchic networks for pattern recognition in protein sequence.

We present a method based on hierarchical self-organizing maps (SOMs) for recognizing patterns in protein sequences. The method is fully automatic, does not require prealigned sequences, is insensitive to redundancy in the training set, and works surprisingly well even with small learning sets. Because it uses unsupervised neural networks, it is able to extract patterns that are not present in all of the unaligned sequences of the learning set. The identification of these patterns in sequence databases is sensitive and efficient. The procedure comprises three main training stages. In the first stage, one SOM is trained to extract common features from the set of unaligned learning sequences. A feature is a number of ungapped sequence segments (usually 4-16 residues long) that are similar to segments in most of the sequences of the learning set according to an initial similarity matrix. In the second training stage, the recognition of each individual feature is refined by selecting an optimal weighting matrix out of a variety of existing amino acid similarity matrices. In a third stage of the SOM procedure, the position of the features in the individual sequences is learned. This allows for variants with feature repeats and feature shuffling. The procedure has been successfully applied to a number of notoriously difficult cases with distinct recognition problems: helix-turn-helix motifs in DNA-binding proteins, the CUB domain of developmentally regulated proteins, and the superfamily of ribokinases. A comparison with the established database search procedure PROFILE (and with several others) led to the conclusion that the new automatic method performs satisfactorily.

EST comparison indicates 38% of human mRNAs contain possible alternative splice forms.

Expressed sequence tag (EST) databases represent a large volume of information on expressed genes including tissue type, expression profile and exon structure. In this study we create an extensive data set of human alternative splicing. We report the analysis of 7867 non-redundant mRNAs, 3011 of which contained alternative splice forms (38% of all mRNAs analysed). From a total of 12572 ESTs 4560 different possible alternative splice forms were detected. Interestingly, 70% of the alternative splice forms correspond to exon deletion events with only 30% exonic insertions. We experimentally verified 19 different splice forms from 16 genes in a total subset of 20 studied; all of the respective genes are of medical relevance.

Neonatal thyroxine treatment: changes in the number of corticotropin-releasing-factor (CRF) and neuropeptide Y (NPY) containing neurons and density of tyrosine hydroxylase positive fibers (TH) in the amygdala correlate with anxiety-related behavior of wistar rats.

Neonatal hyperthyroidism induces persisting alterations in the adult brain, e.g. in spatial learning and hippocampal morphology. In the present study, the relationship between anxiety-related behavior and amygdala morphology was investigated in the adult rat after transient neonatal hyperthyroidism (daily s.c. injections of 7.5 microg L-thyroxine in 0.5 ml 0.9% NaCl solution from postnatal day p1 to p12). The behavioral tests used to study anxiety-related behavior were the motility test, elevated plus-maze and fear-sensitized acoustic startle response. In the amygdala, the number of neurons containing the anxiogenic peptide corticotropin releasing factor (CRF-ir and CRF mRNA) and anxiolytic neuropeptide Y (NPY-ir), the total number of neurons and the density of tyrosine hydroxylase immunoreactive (TH-ir) fibers were quantified. Thyroxine-treated pups presented an accelerated development including opening of eyes and snout elongation as typical signs of hyperthyroidism. Thyroxine-treated adult animals displayed a reduced anxiety in the motility box and elevated plus maze, a reduction in the number of CRF-ir neurons in the central nucleus of the amygdala, as well as an increase in the number of NPY-ir neurons and density of TH-ir fibers in nuclei of the basolateral complex of the amygdala. Moreover, there was a reduction in the total number of neurons in all nuclei of the basolateral complex (despite the higher number of NPY-ir neurons), but not central nucleus of the amygdala. The number of CRF-ir neurons in the central nucleus correlated positively with anxiety-related behavior, and the number of NPY-ir neurons and the density of TH-ir fibers in the basolateral complex correlated inversely with anxiety-related behavior. The findings suggested a shift toward an anxiolytic rather than anxiogenic distribution of peptidergic neurons and fibers in the amygdala at adult age following transient neonatal hyperthyroidism.

Mapping the Trypanosoma cruzi genome: analyses of representative cosmid libraries.

In order to generate contiguous cosmid coverage of the genome of the protozoan parasite Trypanosoma cruzi for large-scale sequence analysis, a cosmid library of 36864 individual, primary clones was generated. Total genomic DNA of the reference strain CL Brener was fragmented both by partial digestion with MboI and by physical shearing. For cloning, a modified cosmid vector was used that simplifies analyses such as restriction mapping. The library's representation is about 25 genome equivalents, assuming a size of 55 Mb per haploid genome. No chimerism of inserts in the clones could be detected. The colinearity between cosmid inserts and genomic DNA was verified. Also, hybridizations to the gel-separated karyotype of the organism were carried out as a quality check. Gridded onto two nylon filters, the library was analyzed with a variety of probes. Apart from being used for combined physical and transcriptional mapping of the genome, library filters and clones are also available to interested parties.

Fluorine-18-FDG and iodine-131-iodide uptake in thyroid cancer.

UNLABELLED: We conducted a prospective study to define the sensitivity of 131I scintigraphy and 18FDG PET whole-body scanning in the detection of thyroid cancer and metastases. METHODS: Forty-one patients with differentiated thyroid carcinoma who underwent thyroidectomy and 131I elimination of the remaining thyroid were studied by 18FDG whole-body PET in 52 examinations and by 131I whole-body scanning. RESULTS: Combined 18FDG and 131I imaging resulted in a sensitivity of about 95%, with alternating uptake of 131I and 18FDG in the metastases: 131I trapping metastases with no 18FDG uptake and 18FDG trapping metastases with no 131I uptake. Five uptake types were differentiated. Alternating uptake was found in about 90% of the patients, which was nearly identical to the sensitivity of the combined 131I/18FDG investigation. In six patients with increasing human thyroglobulin levels, we found that 18FDG whole-body PET localized positive neck metastases of papillary thyroid carcinomas that were histologically confirmed after extirpation. CONCLUSION: Combination 18FDG and 131I whole-body imaging protocol enables detection of local recurrence or metastases on whole-body scans that are often not shown by other imaging methods. Biochemical grading of thyroid cancer may also be possible with this method: Tumors with remaining functional differentiation for hormone synthesis and iodine uptake have low glucose metabolism in more than 95%; tumors without this functional differentiation of 131I uptake show high, glucose metabolism. Fluorine-18-FDG uptake seems to be an indicator of poor functional differentiation, and possibly higher malignancy, in thyroid cancer.