Tissue and subcellular distributions of an inhibitory GDP/GTP exchange protein (GDI) for smg p25A by use of its antibody

We have recently purified from bovine brain cytosol to near homogeneity a GDP/GTP exchange protein for smg p25A, named smg p25A GDI, that inhibits the dissociation of GDP from and the subsequent binding of GTP to smg p25A. In the present study, we made an antiserum against smg p25A GDI and studied its tissue distribution in rat and its subcellular distribution in rat cerebrum by use of this antiserum. smg p25A GDI was found in secretory cells with both regulated and constitutive secretion types. Since smg p25A was previously found in only secretory cells with a regulated secretion type, this result suggests that small GTP-binding proteins different from smg p25A but recognized by smg p25A GDI are present in secretory cells with a constitutive secretion type, and that smg p25A GDI is involved in both regulated and constitutive secretory processes. In subcellular fractionation analysis of rat cerebrum, smg p25A GDI was mostly found in the cytosol fraction of neuron body and synaptosome. In synaptosome, it was mainly found in the synaptic cytosol.     

Microbial sensor system for nondestructive evaluation of fish meat quality

A microbial sensor system consisting of the bacterium (Alteromonas putrefaciens) immobilized within membranes, a flow cell, an oxygen electrode, peristaltic pumps, a buffer tank, a thermostatically controlled bath and a recorder, was constructed for the nondestructive quality evaluation of bluefin tuna. The chemical compounds on fish meat surfaces which are the indicators of fish meat quality were rapidly determined by using the proposed sensor system. Fish meat quality was determined from the rate of current decrease of the sensor. Good correlations were obtained between fish meat quality and sensor response. One assay could be completed within one minute.     

Light and electron microscopical combined staining by immunohistochemical and enzyme histochemical methods using rat prolactin-secreting pituitary tumours

Summary Combined immunohistochemical staining (IHCS) and enzyme histochemical staining (EHCS) methods for light microscopy (LM) and electron microscopy (EM) are reported, using oestrogeninduced rat pituitary tumours. For LM, combined staining for alkaline phosphatase and acid phosphatase by EHCS, using the azo dye method, and for prolactin and ACTH by IHCS, using the enzyme-labelled antibody method, gave the best results on 1 m glycol methacrylate sections. For EM, combined staining by EHCS on 30 m tissue sections followed by IHCS for prolactin on ultrathin Epon sections (enzyme-labelled antibody method) provided acceptable results. By these combined staining methods, the neoplastic prolactin cells were shown to have close affinity to rich alkaline phosphatase-positive capillaries and to possess an alkaline phosphatase-positive cell membrane. Furthermore, they revealed acid phosphatase-positive lysosomal and secretory granules. These combined staining methods may be valuable in studies on the actual functional status of cells.     

An improved method based on the Porter-Silber reaction for determining 17-hydroxycorticosteroids in urine

This paper describes a highly specific method for determining urinary 17-hydroxycorticosteroids, which has been developed by (i) changing the composition of the Porter-Silber reagent and (ii) removing contaminants interfering with the color reaction by addition of sodium bisulfite to β-glucuronidase-hydrolyzed urine before extraction with solvent. For a reference method the Norymberski-Riondel (J. K. Norymberski and A. Riondel 1970, Biochem. J. 120, 493–498) gas chromatography (glc) was used: Correlation coefficient between the present method and GLC = 0.988, deviation from the theoretical regression LINE = 6.8%, and coefficient of SIMILARITY = 0.56. These results are much better than those obtained by I. Ernest, B. Håkansson, J. Lehmann, and B. Sjögren (1964, Acta Endocrinol. 46, 552–562) for the original Porter-Silber method in comparison with the chromatographic measurement of grouped and individual steroids.     

Genetic variations in humans associated with differences in the course of hepatitis C

The outcome of hepatitis C virus (HCV) infection varies among individuals, but the genetic factors involved remain unknown. We conducted a population-based association study in which 238 Japanese individuals positive for anti-HCV antibody were genotyped for 269 single nucleotide polymorphisms (SNPs) in 103 candidate genes that might influence the course of infection. Altogether, 50 SNPs in 32 genes were listed. Genetic polymorphisms in IL4, IL8RB, IL10RA, PRL, ADA, NFKB1, GRAP2, CABIN1, IFNAR2, IFI27, IFI41, TNFRSF1A, ALDOB, AP1B1, SULT2B1, EGF, EGFR, TGFB1, LTBP2, and CD4 were associated with persistent viremia (P < 0.05), whereas those in IL1B, IL1RL1, IL2RB, IL12RB1, IL18R1, STAT5A, GRAP2, CABIN1, IFNAR1, Mx1, BMP8, FGL1, LTBP2, CD34, and CD80 were associated with different serum alanine aminotransferase levels in HCV carriers (P < 0.05). The sorted genes allow us to draw novel hypotheses for future studies of HCV infection to ultimately identify bona fide genes and their variations.     

A novel gene family defined by human dihydropyrimidinase and three related proteins with differential tissue distribution

We have isolated cDNA clones encoding dihydropyrimidinase (DHPase) from human liver and its three homologues from human fetal brain. The deduced amino acid (aa) sequence of human DHPase showed 90% identity with that of rat DHPase, and the three homologues showed 57–59% aa identity with human DHPase, and 74–77% aa identity with each other. We tentatively termed these homologues human DHPase related protein (DRP)-1, DRP-2 and DRP-3. Human DRP-2 showed 98% aa identity with chicken CRMP-62 (collapsin response mediator protein of relative molecular mass of 62 kDa) which is involved in neuronal growth cone collapse. Human DRP-3 showed 94–100% aa identity with two partial peptide sequences of rat TOAD-64 (turned on after division, 64 kDa) which is specifically expressed in postmitotic neurons. Human DHPase and DRPs showed a lower degree of aa sequence identity with Bacillus stearothermophilus hydantoinase (39–42%) and Caenorhabditis elegans unc-33 (32–34%). Thus we describe a novel gene family which displays differential tissue distribution: i.e., human DHPase, in liver and kidney; human DRP-1, in brain; human DRP-2, ubiquitously expressed except for liver; human DRP-3, mainly in heart and skeletal muscle.     

Direct comparison of the pharmacodynamics of four antifungal drugs in a mouse model of disseminated candidiasis using microbiological assays of serum drug concentrations

The aim of this study was to compare the pharmacodynamics of the azole antifungal drugs fluconazole, itraconazole and ketoconazole, and the polyene antifungal amphotericin B, in a mouse model of disseminated Candida albicans infection. In order to directly compare effective serum concentrations of these antifungals, drug concentrations were assayed microbiologically by measuring inhibition of C. albicans mycelial growth (mMIC) in a mouse serum-based assay (serum antifungal titer). Efficacy in the mouse infection model was determined using an organ-based (kidney burden) endpoint. For all four drugs, the serum antifungal titers, 8 hr after administration of single doses of drugs at a range of drug concentrations, correlated closely with C. albicans kidney fungal burden in the mouse model. The results showed that determining serum antifungal titer may be used to accurately represent kidney fungal burden in a mouse model of disseminated candidiasis and allowed direct comparison of the pharmacodynamics of differing classes of antifungal drugs.