Molecular cloning and characterization of RGC-32, a novel gene induced by complement activation in oligodendrocytes

Sublytic complement activation on oligodendrocytes (OLG) down-regulates expression of myelin genes and induces cell cycle in culture. Differential display (DD) was used to search for new genes whose expression is altered in response to complement and that may be involved in cell cycle activation. DD bands showing either increased or decreased mRNA expression in response to complement were identified and designated Response Genes to Complement (RGC) 1-32. RGC-1 is identical with heat shock protein 105, RGC-2 with poly(ADP-ribose) polymerase, and RGC-10 with IP-10. A new gene, RGC-32, that encodes a protein of 137 amino acids was cloned. RGC-32 has no homology with other known proteins, and contains no motif that would indicate its function. In OLG, the mRNA expression was increased by complement activation and by terminal complement complex assembly. RGC-32 protein was localized in the cytoplasm and co-immunoprecipitated with cdc2 kinase. Overexpression of RGC-32 increased DNA synthesis in OLGxC6 glioma cell hybrids. These results suggest that RGC-32 may play a role in cell cycle activation.     

The role of spatial working memory in inhibition of return: evidence from divided attention tasks

Inhibition of return (IOR) refers to a bias against returning attention to a location that has been recently attended. In the present experiments, we examined the role of working memory in IOR by introducing secondary tasks (in the temporal interval between the cue and the target) that involved a working memory component. When the secondary task was nonspatial in nature (monitoring odd digits or adding digits), IOR was present, although overall reaction times were greater in the presence of the secondary task. When the task involved a spatial working memory load (remembering the directionality of arrows or the orientation of objects), IOR was eliminated. However, when the participants had incentive to process the directionality of an arrow but did not have to use any memory system, IOR persisted at peripheral locations. Overall, the results suggest that IOR is partially mediated by a spatial working memory system.     

Chemical synthesis of the precursor molecule of the Aequorea green fluorescent protein, subsequent folding, and development of fluorescence

The present paper describes the total chemical synthesis of the precursor molecule of the Aequorea green fluorescent protein (GFP). The molecule is made up of 238 amino acid residues in a single polypeptide chain and is nonfluorescent. To carry out the synthesis, a procedure, first described in 1981 for the synthesis of complex peptides, was used. The procedure is based on performing segment condensation reactions in solution while providing maximum protection to the segment. The effectiveness of the procedure has been demonstrated by the synthesis of various biologically active peptides and small proteins, such as human angiogenin, a 123-residue protein analogue of ribonuclease A, human midkine, a 121-residue protein, and pleiotrophin, a 136-residue protein analogue of midkine. The GFP precursor molecule was synthesized from 26 fully protected segments in solution, and the final 238-residue peptide was treated with anhydrous hydrogen fluoride to obtain the precursor molecule of GFP containing two Cys(acetamidomethyl) residues. After removal of the acetamidomethyl groups, the product was dissolved in 0.1 M Tris. HCl buffer (pH 8.0) in the presence of DTT. After several hours at room temperature, the solution began to emit a green fluorescence (lambdamax = 509 nm) under near-UV light. Both fluorescence excitation and fluorescence emission spectra were measured and were found to have the same shape and maxima as those reported for native GFP. The present results demonstrate the utility of the segment condensation procedure in synthesizing large protein molecules such as GFP. The result also provides evidence that the formation of the chromophore in GFP is not dependent on any external cofactor.     

Use of stomatal conductance and pre-dawn water potential to classify terroir for the grape variety Kékfrankos

Background and Aims: A 3‐year study was carried out in order to evaluate the ecophysiology, yield and quality characteristics of Vitis vinifera L. cv. Kékfrankos (syn. Limberger) at Eger‐Nagyeged hill (steep slope) and at Eger‐K?lyuktet? (flat) vineyard sites located in the Eger wine region, Hungary. The aim of this paper was to analyse the effect of ‘vintage’ and ‘terroir’ on the seasonal changes of Kékfrankos ecophysiology and its possible relationship with yield and wine composition. Methods and Results: Grapevine physiological responses (midday‐ and pre‐dawn water potential, pressure–volume analysis and gas‐exchange), growing stages, yield and wine composition of each vineyard were studied. Lower grapevine water supply was detected at Eger‐Nagyeged hill in each season due to its steep slope and soil characteristics. Pressure‐volume curves indicated that there was no osmotic adjustment in the leaves of this variety. Higher osmotic concentration was measured at turgor loss and full turgor in the leaves of the unstressed vineyard (Eger‐K?lyuktet?) presumably due to higher photosynthetic activity. Differences in soil water content of the vineyards resulted in a slightly altered cell wall elasticity. Stomatal conductance, transpiration rate and photosynthetic production per unit leaf area were affected by water availability. Lower yield in Eger‐Nagyeged hill was partly associated with decreased photosynthetic production of the canopy. Improved wine quality of Eger‐Nagyeged hill was due to moderate water stress which induced higher concentration of anthocyanins and phenolics in the berries. The duration of the phenological stages was dependent on vintage temperature characteristics rather than on vineyard site. Conclusion: There was a close relationship between environmental conditions, Kékfrankos gas‐exchange, water relations, yield and wine composition. Water deficit plays an important role in creating a terroir effect, resulting in decreased yield, better sun exposure of leaves and clusters and thus higher concentration of phenolics and anthocyanins. Although quality is mainly influenced by vintage differences, vineyard characteristics are able to buffer unfavourable vintage effects even within a small wine region. Significance of the Study: Stomatal conductance, pre‐dawn water potential and climatic data may be reliable parameters for terroir classification, although variety–terroir interactions must always be considered.     

Intrinsic human immunodeficiency virus type 1 resistance of hematopoietic stem cells despite coreceptor expression

Interactions of human immunodeficiency virus type 1 (HIV-1) with hematopoietic stem cells may define restrictions on immune reconstitution following effective antiretroviral therapy and affect stem cell gene therapy strategies for AIDS. In the present study, we demonstrated mRNA and cell surface expression of HIV-1 receptors CD4 and the chemokine receptors CCR-5 and CXCR-4 in fractionated cells representing multiple stages of hematopoietic development. Chemokine receptor function was documented in subsets of cells by calcium flux in response to a cognate ligand. Productive infection by HIV-1 via these receptors was observed with the notable exception of stem cells, in which case the presence of CD4, CXCR-4, and CCR-5, as documented by single-cell analysis for expression and function, was insufficient for infection. Neither productive infection, transgene expression, nor virus entry was detectable following exposure of stem cells to either wild-type HIV-1 or lentivirus constructs pseudotyped in HIV-1 envelopes of macrophage-tropic, T-cell-tropic, or dualtropic specificity. Successful entry into stem cells of a vesicular stomatitis virus G protein-pseudotyped HIV-1 construct demonstrated that the resistance to HIV-1 was mediated at the level of virus-cell membrane fusion and entry. These data define the hematopoietic stem cell as a sanctuary cell which is resistant to HIV-1 infection by a mechanism independent of receptor and coreceptor expression that suggests a novel means of cellular protection from HIV-1.     

Arterial disease in lupus and secondary antiphospholipid syndrome: association with anti-beta2-glycoprotein I antibodies but not with antibodies against oxidized low-density lipoprotein

The prevalence and clinical significance of antibodies against beta2-glycoprotein I (anti-beta2GPI) and antibodies against oxidized low-density lipoprotein (anti-ox-LDL) were evaluated as potential indicators of arterial disease in patients with systemic lupus erythematosus (SLE) and SLE with secondary antiphospholipid syndrome (APS). IgG anti-beta2GPI and IgG anti-ox-LDL were measured by enzyme-linked immunosorbent assay (ELISA) in serum samples from 118 patients with SLE, including 40 with secondary APS. IgG anti-beta2GPI were positive in 17% (20/118) of SLE patients. The presence and titres of IgG anti-beta2GPI were strongly associated with a history of arterial thrombosis. Haemolytic anaemia was also significantly associated with the presence of IgG anti-beta2GPI. The prevalence of IgG anti-ox-LDL was 53% (63/118), but there was no association with arterial thrombosis. No correlation between the values of anti-ox-LDL and those of anti-beta2GPI was found. These results suggest that IgG anti-beta2GPI could be a marker for arterial thrombosis in SLE patients, while IgG anti-ox-LDL were not associated with arterial disease in this group of lupus patients.     

The 75 g oral glucose tolerance in pregnancy

The first case of the human immunodeficiency virus (HIV) infection was detected in Singapore in 1985 and the first case of the acquired immunodeficiency syndrome (AIDS) in 1986. Since then, the number of infections had increased. By the end of 1993, there were 222 residents with HIV infection, including 75 cases of AIDS. In view of the rapidly increasing magnitude of HIV infection, a microcomputer-based surveillance system was designed and developed in 1992 to better monitor epidemiological trends of HIV infection in Singapore. OBJECTIVE--The objective was to define a composite model of a successful HIV and AIDS registry that included: (a) patient data forms, (b) patient's contact data forms, (c) data analysis, and (d) report generation. METHODOLOGY--An IBM-compatible desk-top microcomputer was used for the project. The main software used for computer programming and data analysis were DBase IV (Version 1.5) and Epi Info (Version 5.0), respectively. Security features were incorporated into the programme to ensure confidentiality of information and that only authorized personnel could gain access to the programme. MAIN FINDINGS--The system functioned as the National HIV Notification Registry and was able to track notifications, analyse data and enabled prompt dissemination of information. The system was also linked to another database system for tuberculosis to enhance surveillance of both HIV infection and tuberculosis. CONCLUSION--The authors believe that this system would enhance surveillance and provide timely information for national AIDS control programmes. However, the effectiveness of this computer-based surveillance system is dependent on an established notification structure with notifications of sufficient completeness for both HIV infection and AIDS.     

Localization of dopamine receptor subtypes in corpus striatum and nucleus accumbens septi of rat brain: comparison of D1-, D2-, and D4-like receptors

The purpose of this study was to determine whether bradykinin mediates ovalbumin-induced increase in macromolecular efflux from the nasal mucosa of ovalbumin-sensitized hamsters in vivo and, if so, whether the L-arginine/nitric oxide biosynthetic pathway transduces, in part, this response. We found that suffusion of ovalbumin onto the in situ nasal mucosa of ovalbumin-sensitized hamsters, but not of controls, elicited a significant time- and concentration-dependent increase in clearance of fluorescein isothiocyanate-labeled dextran (mol mass, 70 kDa; P < 0.05). HOE-140, but not des-Arg9,[Leu8]-bradykinin, and NG-L-arginine methyl ester (L-NAME), but not NG-D-arginine methyl ester, significantly attenuated ovalbumin-induced responses. L-Arginine, but not D-arginine, abolished the effects of L-NAME. L-NAME also significantly attenuated bradykinin-, but not adenosine-induced increase in macromolecular efflux from the in situ nasal mucosa. Overall, these data suggest that ovalbumin increases macromolecular efflux from the in situ nasal mucosa of ovalbumin-sensitized hamsters, in part, by producing bradykinin with subsequent activation of the L-arginine/ nitric oxide biosynthetic pathway.     

Interferon-inducing and antiviral activity of levamisole

It was shown that levamisol administered orally to mice induced production of interferon with its maximum level in 4-6 hours and prolonged subsequent circulation in the host (the observation period of 5 days). Antiviral activity of levamisol in experimental forest-spring encephalitis was shown (protection of 35-40 per cent). When levamisol were used in combination with polyguacyl, an additive effect was recorded.