Cognition as a treatment target in depression.

Cognitive dysfunction in depression is associated with poorer clinical outcomes and impaired psychosocial functioning. However, most treatments for depression do not specifically target cognition. Neurocognitive deficits such as memory and concentration problems tend to persist after mood symptoms recover. Improving cognition in depression requires a better understanding of brain systems implicated in depression. A comprehensive approach is warranted for refined methods of assessing and treating cognitive dysfunction in depression.

Space-time adaptive numerical methods for geophysical applications.

In this paper we present high-order formulations of the finite volume and discontinuous Galerkin finite-element methods for wave propagation problems with a space-time adaptation technique using unstructured meshes in order to reduce computational cost without reducing accuracy. Both methods can be derived in a similar mathematical framework and are identical in their first-order version. In their extension to higher order accuracy in space and time, both methods use spatial polynomials of higher degree inside each element, a high-order solution of the generalized Riemann problem and a high-order time integration method based on the Taylor series expansion. The static adaptation strategy uses locally refined high-resolution meshes in areas with low wave speeds to improve the approximation quality. Furthermore, the time step length is chosen locally adaptive such that the solution is evolved explicitly in time by an optimal time step determined by a local stability criterion. After validating the numerical approach, both schemes are applied to geophysical wave propagation problems such as tsunami waves and seismic waves comparing the new approach with the classical global time-stepping technique. The problem of mesh partitioning for large-scale applications on multi-processor architectures is discussed and a new mesh partition approach is proposed and tested to further reduce computational cost.

Validation of the X-chromosomal STR DXS6809.

This paper presents sequence and population genetic data of the microsatellite marker DXS6809 (GDB 365492) obtained from a German population sample ( n=725 chromosomes). DXS6809 is a highly polymorphic X-linked tetranucleotide polymorphism presenting 12 alleles in our population. Sequencing of 77 PCR products covering 12 alleles (by length), characterised DXS6809 as a marker with a complex repeat sequence structure. A polymorphism information content (PIC) of 0.825 and a mean exclusion chance (MEC) of 0.815 were obtained. A deviation from the Hardy-Weinberg equilibrium (HWE) could not be detected and male and female samples exhibited a similar allele distribution. Kinship testing revealed a typical X-linked inheritance and 2 mutations were found in 394 meioses. DXS6809 is located 90.18 Mb, i.e. 102.3 cM, from the Xp-telomere (Xp-tel), corresponding to Xq21.33. The presented data qualify DXS6809 as a useful supplement to the known forensic ChrX marker panel.

Effects of natural intensities of visible and ultraviolet radiation on epidermal ultraviolet screening and photosynthesis in grape leaves.

Grape (Vitis vinifera cv Silvaner) vine plants were cultivated under shaded conditions in the absence of ultraviolet (UV) radiation in a greenhouse, and subsequently placed outdoors under three different light regimes for 7 d. Different light regimes were produced by filters transmitting natural radiation, or screening out the UV-B (280-315 nm), or screening out the UV-A (315-400 nm) and the UV-B spectral range. During exposure, synthesis of UV-screening phenolics in leaves was quantified using HPLC: All treatments increased concentrations of hydroxycinnamic acids but the rise was highest, reaching 230% of the initial value, when UV radiation was absent. In contrast, UV-B radiation specifically increased flavonoid concentrations resulting in more than a 10-fold increase. Transmittance in the UV of all extracted phenolics was lower than epidermal UV transmittance determined fluorimetrically, and the two parameters were curvilinearly related. It is suggested that curvilinearity results from different absorption properties of the homogeneously dissolved phenolics in extracts and of the non-homogeneous distribution of phenolics in the epidermis. UV-B-dependent inhibition of maximum photochemical yield of photosystem II (PSII), measured as variable fluorescence of dark-adapted leaves, recovered in parallel to the buildup of epidermal screening for UV-B radiation, suggesting that PSII is protected against UV-B damage by epidermal screening. However, UV-B inhibition of CO(2) assimilation rates was not diminished by efficient UV-B screening. We propose that protection of UV-B inactivation of PSII is observed because preceding damage is efficiently repaired while those factors determining UV-B inhibition of CO(2) assimilation recover more slowly.

AAA proteases of mitochondria: quality control of membrane proteins and regulatory functions during mitochondrial biogenesis.

An ubiquitous and conserved proteolytic system regulates the stability of mitochondrial inner membrane proteins. Two AAA proteases with catalytic sites at opposite membrane surfaces form a membrane-integrated quality control system and exert crucial functions during the biogenesis of mitochondria. Their activity is modulated by another membrane-protein complex that is composed of prohibitins. Peptides generated upon proteolysis in the matrix space are transported across the inner membrane by an ATP-binding cassette transporter. The function of these conserved components is discussed in the present review.

Structural requirements of Tom40 for assembly into preexisting TOM complexes of mitochondria.

Tom40 is the major subunit of the translocase of the outer mitochondrial membrane (the TOM complex). To study the assembly pathway of Tom40, we have followed the integration of the protein into the TOM complex in vitro and in vivo using wild-type and altered versions of the Neurospora crassa Tom40 protein. Upon import into isolated mitochondria, Tom40 precursor proteins lacking the first 20 or the first 40 amino acid residues were assembled as the wild-type protein. In contrast, a Tom40 precursor lacking residues 41 to 60, which contains a highly conserved region of the protein, was arrested at an intermediate stage of assembly. We constructed mutant versions of Tom40 affecting this region and transformed the genes into a sheltered heterokaryon containing a tom40 null nucleus. Homokaryotic strains expressing the mutant Tom40 proteins had growth rate defects and were deficient in their ability to form conidia. Analysis of the TOM complex in these strains by blue native gel electrophoresis revealed alterations in electrophoretic mobility and a tendency to lose Tom40 subunits from the complex. Thus, both in vitro and in vivo studies implicate residues 41 to 60 as containing a sequence required for proper assembly/stability of Tom40 into the TOM complex. Finally, we found that TOM complexes in the mitochondrial outer membrane were capable of exchanging subunits in vitro. A model is proposed for the integration of Tom40 subunits into the TOM complex.

A gene expression profile of embryonic stem cells and embryonic stem cell-derived neurons.

Embryonic stem (ES) cells have the ability to differentiate into a variety of cell lineages. We are examining ES cell differentiation in vitro by using cDNA microarrays to generate a molecular phenotype for each cell type. El4 ES cells induced by retinoic acid after forming embryoid bodies differentiate almost exclusively to neurons. We obtained expression patterns for about 8500 gene sequences by comparing mRNAs from undifferentiated ES cells and their differentiated derivatives in a competitive hybridization. Our results indicate that the genes expressed by ES cells change dramatically as they differentiate (58 gene sequences up-regulated, 34 down-regulated). Most notably, totipotent ES cells expressed high levels of a repressor of Hox expression (the polycomb homolog Mphl) and a co-repressor (CTBP2). Expression of these genes was undetectable in differentiated cells; the ES cell-derived neurons expressed a different set of transcriptional regulators, as weil as markers of neurogenesis. The gene expression profiles indicate that ES cells actively suppress differentiation by transcriptional repression; cell-cell contact in embryoid bodies and retinoic acid treatment may overcome this suppression, allowing expression of Hox genes and inducing a suite of neuronal genes. Gene expression profiles will be a useful outcome measure for comparing in vitro treatments of differentiating ES cells and other stem cells. Also, knowing the molecule phenotype of transplantable cells will allow correlation of phenotype with the success of the transplant.

Protein degradation in mitochondria.

The biogenesis of mitochondria and the maintenance of mitochondrial functions depends on an autonomous proteolytic system in the organelle which is highly conserved throughout evolution. Components of this system include processing peptidases and ATP-dependent proteases, as well as molecular chaperone proteins and protein complexes with apparently regulatory functions. While processing peptidases mediate maturation of nuclear-encoded mitochondrial preproteins, quality control within various subcompartments of mitochondria is ensured by ATP-dependent proteases which selectively remove non-assembled or misfolded polypeptides. Moreover; these proteases appear to control the activity- or steady-state levels of specific regulatory proteins and thereby ensure mitochondrial genome integrity, gene expression and protein assembly.

European field tests with HISTKOM telepathology equipment.

HISTKOM telemicroscopy equipment for telepathology is designed for the most challenging application in telepathology: intraoperational frozen section diagnosis. Adapted to this application, it is also excellently suited for all other telepathology modes requesting less sophisticated equipment. The technical concept and user interface are oriented to routine daily pathology. HISTKOM underwent heavy field-tests at several locations. The field-tests designs and the results of five of these are reported in this paper. Telepathology will exploit its advantages in networks hosting participants requesting and offering services. The solution of the interoperability problem caused by different equipment from different suppliers within such a network will be a major task, the solution for which is in progress. The new generation of HISTKOM equipment and software is designed in a modularized concept, allowing the integration of various hardware components from different manufacturers; thus special configurations can be realized easily. HISTKOM is offered as complete turnkey system, but can also be installed in yet existing configurations of the customer if they meet specifications.

Effects of cationic liposome-DNA complexes on pulmonary surfactant function in vitro and in vivo.

Cationic liposome-DNA complexes are being evaluated as potential gene therapy agents for the lung. Cations have strong effects on the biophysical functions of lung surfactant. Therefore, we assessed whether cationic liposomes [composed of N-(1-(2,3-dioleyloxy) propyl)-N,N,N-trimethyl-ammonium chloride and dioleylphosphatidylethanolamine] with or without DNA affect behavior of four types of surfactant in vitro. Experiments were carried out using a modified Wilhelmy surface balance. The ability of surfactants that contain protein and anionic lipids to lower surface tension was inhibited in the presence of cationic liposomes. Inactivation was less when DNA was preincubated with cationic liposomes. Surfactant that contained neither protein nor anionic lipids was not inactivated. Mechanical properties of the lung were studied to assess in vivo surfactant function after intratracheal instillation of a cationic liposome-DNA complex into adult rats. Pressure-volume deflation curves were shifted by 18% compared with those from normal (untreated) animals, but this effect was transient and not different from that observed in animals who received a similar volume of saline. These findings indicate that cationic liposomes alone may have deleterious effects on behavior of some surfactants possibly by disrupting charge interactions between negatively charged phospholipids and surfactant proteins. When DNA is added to liposomes before exposure to surfactants, the adverse charge interactions may be obviated by charge neutralization of liposomes by DNA.

Inhibition of bacterial growth by synthetic SP-B1-78 peptides.

Residues 12-34 of mature human pulmonary surfactant protein B (SP-B1-78) are 68% homologous to residues 48-72 of the frog peptide antibiotic dermaseptin b I. We examined the effects of SP-B1-78 on the growth of Escherichia coli in order to find whether full length SP-B1-78 might act as a peptide antibiotic. We found that SP-B1-78 peptide inhibited growth of E. coli (MIC = 210 micrograms.ml-1), but that the SP-B variant [R/K-->S]SP-B1-78 was less potent (MIC = 500 micrograms.ml-1).

Comparison between epidermal growth factor, transforming growth factor-alpha and EGF receptor levels in regions of adult rat brain.

Examination of adult rat brain regions by specific radioimmunoassays revealed a widespread distribution of transforming growth factor-alpha (TGF-alpha), but not epidermal growth factor (EGF), the peptide that had previously been reported to be present in rodent brain. Polyadenylated RNA samples from the different regions of rat brain were analyzed by Northern blot to identify mRNA species encoding precursor proteins for EGF (preproEGF), TGF-alpha (preproTGF-alpha), and the EGF/TGF-alpha receptor. The results indicate that TGF-alpha is the most abundant ligand for the EGF/TGF-alpha receptor in most parts of the brain analyzed. Message for preproEGF was only detectable after prolonged autoradiographic exposure; levels of preproEGF mRNA were between two and three orders of magnitude lower in brain than those expressed in control tissue (kidney), and one to two orders of magnitude lower than preproTGF-alpha mRNA levels in all brain regions. These results were confirmed by analysis of mRNA by RT/PCR, and support the hypothesis that expression of preproEGF mRNA in the brain is limited to smaller discrete areas, whereas preproTGF-alpha gene expression is almost ubiquitous.

Ontogeny of epidermal growth factor, transforming growth factor-alpha, epidermal growth factor receptor, and thyroid hormone receptor RNA levels in rat kidney and changes in those levels induced by early thyroxine treatment.

Ontogenic changes in the mRNA levels of epidermal growth factor (EGF), transforming growth factor-alpha, EGF receptor, and thyroid hormone receptor (r-erbA) were examined in developing rat kidneys. The mRNA levels of both EGF and thyroid hormone receptor rose dramatically during the postnatal period with the rise in thyroid hormone receptor message preceding the rise in EGF message. In addition, we examined renal mRNA levels in 1-wk-old rats treated with thyroxine (T4) from birth through d 6. Neonatal T4 treatment augmented the renal mRNA levels of EGF but decreased the levels of EGF-receptor and transforming growth factor-alpha. T4 treatment did not significantly affect the levels of renal mRNA for thyroid hormone receptor. Although the EGF and transforming growth factor-alpha peptides are similar and interact with the same receptor, our findings indicate that these homologous growth factors are regulated differently during development. In addition, hormones that influence growth and development, such as T4, may function both as positive and negative regulators of growth factor expression.

Expression of exogenous c-myc oncogene does not initiate DNA synthesis in primary rat hepatocyte cultures.

Cultured hepatocytes from adult rats stimulated with combinations of growth factors enter into S phase but do not undergo multiple rounds of DNA synthesis nor mitosis. We have examined the potential of an introduced oncogene to induce alterations in the DNA synthetic activity of the cultured hepatocytes in response to epidermal growth factor (EGF). Overexpression of c-myc did not initiate significant DNA synthesis in rat hepatocyte cultures alone, although it cooperated with added EGF to super-induce thymidine incorporation into DNA. From our results, it is suggested that EGF is also necessary to initiate hepatocyte DNA synthesis probably by inducing a battery of cell cycle-related genes if incubated with c-myc transfected cultures for only 5 hours. Hepatocyte polypeptides reacting with anti-MYC antisera were found to migrate between 55-67 KDa in SDS-PAGE; only the 64-67 KDa species were found to be phosphorylated, and the observed size heterogeneity may be due to proteolytic degradation or may reflect presently unknown posttranslational modifications.

Hepatocyte conditioned medium modulates the response of primary rat hepatocyte cultures to epidermal growth factor.

Primary hepatocytes may produce autocrine growth trigger(s) with or without a mitogenic stimulus. We explored the potential of hepatocyte conditioned medium--from untreated quiescent cultures--to modulate the DNA synthetic responses induced by EGF. The EGF-induced responses were similar when EGF was continuously or transiently (3 h) present. Conditioned medium (CM) from 48 h hepatocyte culture was the most effective in eliciting thymidine incorporation into hepatocyte DNA. At the same time the conditioned medium from hepatocyte cultures stimulated lymphocyte DNA synthesis at levels much lower than those observed using PHA, a specific lymphocyte inducer. The maximal EGF-binding by intact hepatocytes was also significantly increased in the presence of conditioned medium (48 h). We therefore suggest that hepatocytes produce autocrine growth trigger(s) which might be in part responsible for the regulation of the in vitro and/or in vivo hepatocyte proliferation.

Prostaglandins E2 and F2a mediate the increase in c-myc expression induced by EGF in primary rat hepatocyte cultures.

Epidermal Growth Factor (EGF) and prostaglandins (PGs) E2 and F2a, have been shown to stimulate primary hepatocyte proliferation. Verapamil (5-20 microM), a calcium channel inhibitor, inhibited hepatocyte DNA synthesis and c-myc expression, induced by EGF (50 ng/dish) and prostaglandins (1-12 micrograms/dish). Indomethacin (20-100 microM) decreased significantly the EGF-induced hepatocyte DNA synthesis and c-myc expression. Addition of PGs (1-9 micrograms) in hepatocyte cultures treated with EGF+indomethacin (100 microM) restored the capacity of EGF to increase c-myc expression and DNA synthesis. We propose that arachidonic acid derivatives and calcium channel blockers modulate c-myc expression in primary hepatocytes.

Prothrombin mRNA is expressed by cells of the nervous system.

Thrombin, a serine protease of the blood coagulation system, has additional effects on cells in vitro. It is mitogenic for fibroblasts and astrocytes and contributes to the regulation of neurite outgrowth and astrocyte stellation. Until now the expression of thrombin or its precursor prothrombin in tissues other than liver has not been demonstrated conclusively because of difficulty in avoiding serum contamination. Using sensitive mRNA detection methods, we show here that prothrombin is expressed not only in the liver, but also in the brain throughout development. Polymerase chain reaction, Northern, and in situ hybridization studies demonstrate the presence of prothrombin transcripts in the olfactory bulb, the cortex, the cerebellum, and other regions of the rat and human nervous system, as well as in neural cell lines. These results support an involvement of (pro)thrombin in the regulation of cellular events in the nervous system.

Beryllium toxicity. The selective inhibition of casein kinase 1.

1. Cyclic AMP-independent casein kinase 1 in liver cytoplasm and nuclei was inhibited by Be2+ in vitro (Ki 2.5 microM and 29 microM respectively). Casein kinase 2 (phosvitin kinase) and cyclic AMP-dependent protein kinase were unaffected. 2. The inhibition of casein kinase 1 by Be2+ was competitive with respect to the protein substrate; at non-saturating concentrations of casein, inhibition was non-competitive with respect to ATP. 3. In rats given LD50 doses of Be2+ 24 h before death, the activities of cytoplasmic and nuclear casein kinase 1 in livers from partially hepatectomized animals were diminished approx. 50%; with intact rats, nuclear casein kinase 1 was inhibited at concentrations of casein less than the Km.