Effect of angiotensin coverting enzyme inhibitor on regression in cardiac hypertrophy

Cardiac hypertrophy in rats was produced by aortic banding for 6 weeks and regression of hypertrophy in these experimental animals was induced by administration of angiotensin converting enzyme inhibitor, enalapril (10 mg/kg/ day) for 6 weeks. The left ventricular muscle mass and systolic pressure were decrease upon treating the hypertrophied rats with enalapril. This drug also decreased the number of 1-adrenoceptors in hypertrophyied myocardium without any changes in -adrenoceptors. The regression of cardiac hypertrophy in spontaneously hypertensive rats by enalapril for 10 weeks was not associated with any alterations in 1-adrenoceptors in hypertrophied myocardium, but was decreased in -adrenoceptors. Effects of enalapril on extracellular matrix in the myocardium was also observed in regression of hypertrophy in which the type III collagen mRNA expression and collagen contents were reduced in comparison with those of hypertrophied myocardium. These results indicate that regression of cardiac hypertrophy is not alway associated with a decrease in the number of 1-adrenergic receptors and that the beneficial effects of enalapril in the hypertrophied heart in aortic banding animals may be of some specific nature.     

Rapid Changes in Polyphosphoinositide Metabolism in Pea in Response to Fungal Signals

Effects of the elicitor and/or suppressor from Mycosphaerellapinodes on polyphosphoinositide metabolism (PI metabolism) inpea were examined both in vivo and in vitro. The elicitor induceda rapid and biphasic increase in levels of phosphatidylinositol-4,5-bisphosphate(PtdInsP₂) and inositol 1,4,5-trisphosphate (IP₃) in epicotyltissues that was apparent within 15 min. A transient increasein levels of PtdInsP₂ and IP₃ was detected immediately in elicitor-treatedplasma membranes. However, the concomitant presence of suppressorwith elicitor resulted in inhibition of these increases bothin vivo and in vitro. These findings suggest that the elicitorrapidly activates phosphatidylinositol kinase, phosphatidylinositol-4-monophosphatekinase and phospholipase C, which are involved in PI metabolism,whereas the suppressor markedly inhibits these enzymes. Neomycin,a known inhibitor of phospholipase C, blocked the elicitor-inducedaccumulation both of IP₃ and pisatin and it also induced localsusceptibility in pea tissues that resembled that of the fungalsuppressor. From these results, it appears that rapid changesin PI metabolism are indispensable in the signal transductionrelated to defense responses of pea plants. (Received January 18, 1993; Accepted May 13, 1993)     

Characteristics of cross-flow filtration of pullulan broth

Cross-flow filtration of culture broth from Aureobasidium pullulans, which elaborates pullulan, was done with a thin channel-type module and microfiltration membranes made of different materials and with different pore sizes. Various factors affecting the results of the filtration were studied. The specific resistance of the microbial cake was found to be higher than that of bakers yeast, the cells of which are about the same size as an A. pullulans cell, and resistance increased with cultivation time. The flux and transmission of pullulan through the membrane decreased with cultivation time as the specific resistance increased. The flux and transmission ] of pullulan depended on the structure and pore size of the membrane and also on the pH of the broth. With a polysulphone membrane with a nominal pore size of 2.0 m, transmission was nearly 100% with negligible leakage of cells and the flux was high when the pH of the broth was adjusted to 2.0.On leave from Hayashibara Co., Ltd., Amase-minamimachi, Okayama 700 Japan Correspondence to: K. Nakanishi     

Improvement of performance for cross-flow membrane filtration of pullulan broth

To improve the performance of cross-flow membrane filtration of pullulan broth from Aureobasidium pullulans, the effect of the cultivation conditions was examined. In particular, the sucrose concentration in the medium was changed over a wide range. By decreasing the sucrose concentration the distribution of morphology of the microbial cells in the broth changed; the yeast-like form became predominant and, as a result, the specific resistance of the microbial cake was lowered. When the broth was fermented with a sucrose concentration of 2.5% or lower, the filtration characteristics were greatly improved by periodic closure of permeation during cross-flow filtration.On leave from Hayashibara Co., Ltd., Amase-minamimachi, Okayama 700, Japan Correspondence to: K. Nakanishi     

Isolation and characterization of mouse CD7 cDNA

The human CD7 antigen is a glycoprotein, M _r 40 000, expressed on the surface of peripheral blood T-lymphocytes and thymocytes, and is the earliest surface antigen to appear on T-cell lineage cells. In this study, putative mouse CD7 cDNA was identified based on its similarities with human CD7. Five independent clones originating from the same mRNA species were isolated (designated as mCD7) by screening a mouse thymocyte cDNA library with human CD7 cDNA, J61, under moderate stringency. The longest insert of a 995 base pair had an open reading frame of 210 amino acids. Northern blot analysis using the mouse CD7 cDNA probe demonstrated a single 1.2 kilobase mRNA ni the thymus, spleen, bone marrow, and small intestine. The protein deduced from mCD7 cDNA consisted of the leader, extracellular, transmembrane, and cytoplasmic domains of 24, 126, 21, and 39 amino acids, respectively, based on the hydrophobicity plot and the structure of human CD7. The extracellualr domain contained three potential N-glycosilation sites, while the cytoplasmic domain contained one potential protein kinase C phosphorylation site. The amino acid sequence had 45.5% similarity with human CD7, while the similarities for the individual domains ranged from 49.2% to 63.2%. The six highly conserved regions, which may possibly be involved with still unknown CD7-mediated functions, were located in the extracellular and cytoplasmic domains.The nucleotide sequence data reported in this paper have seen submitted to the GenBank, DDBJ, and EMBL nucleotide sequence database and have been assigned the accession number D10329.     

Pituitary Adenylate Cyclase Activating Polypeptide (PACAP)-like immunoreactivity in human hypothalamus: co-localization with arginine vasopressin

Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel hypothalamic peptide consisting of 38 amino acids (PACAP1–38) with a potent stimulatory action on adenylate-cyclase in rat pituitary. The presence of PACAP-like immunoreactivity in human brain was studied by radioimmunoassay. Co-localization of PACAP with arginine vasopressin and oxytocin was investigated by immunocytochemistry in the human hypothalamus. Immunoreactive PACAP was detected in all regions of human brain (cortex, thalamus, hypothalamus, pons and hemisphere of cerebellum) with the highest levels found in the hypothalamus (8.5±1.9 pmol/g wet weight, n=w, mean±S.E.M.). High performance liquid chromatography of the human hypothalamic ex approximately 50% of the immunoreactive PACAP was eluted in the position of PACAP1–38. Immunocytochemical studies showed the presence of PACAP immunoreactive neurons in the paraventricular and supraoptic nuclei of human hypothalamus. PACAP co-localized with arginine vasopressin in magnocellular cells of these nuclei. These findings suggest that PACAP1-38 plays important physiological roles in the human hypothalamus.     

Ras oncogene enhances the production of a recombinant protein regulated by the cytomegalovirus promoter in BHK-21 cells

In order to enhance recombinant protein productivity in animal cells, we developed the oncogene activated production (OAP) system. The OAP system is based on the premise that oncogenes are able to enhance promoter activity. To this end, we constructed reported plasmids by fusing various promoters to the human interleukin-6 (hIL-6) cDNA, and the effector plasmids by inserting individual oncogenes, for example c-myc, c-fos, v-jun, v-myb and c-Ha-ras, downstream from the human cytomegalovirus immediate early (CMV) promoter. Results of transient expression experiments with BHK-21 cells suggest that the CMV promoter is the most potent promoter examined and that theras product is able to transactivate the -actin, CMV and SR promoters. Recombinant BHK-21 cells producing hIL-6 under the control of the CMV promoter were contransfected with theras oncogene and dihydrofolate reductase gene, then selected with 50 nM methotrexate to coamplify theras oncogene. We were able to rapidly establish a stable and highly productive clone which exhibited a 35-times higher production rate as compared to the control value.     

Identification of DNA topoisomerases involved in immediate and transient DNA relaxation induced by heat shock inEscherichia coli

The linking number of plasmid DNA in exponentially growingEscherichia coli increases immediately and transiently after heat shock. The purpose of this study was to search for DNA topoisomerases that catalyze this relaxation of DNA. Neither introduction of atopA deletion mutation nor treatment of cells with DNA gyrase inhibitors affected the DNA relaxation induced by heat shock. Thus, DNA topoisomerase I and DNA gyrase are apparently not involved in the process. However, the reaction was inhibited by nalidixic acid or by oxolinic acid in thetopA mutant and the reaction was resistant to nalidixic acid in atopA mutant carrying, in addition, thenalA26 mutation. These results are interpreted as indicating that both DNA topoisomerase I and DNA gyrase are involved in the DNA relaxation induced by heat shock.     

A sensitive determination of α-keto acids by gas-liquid chromatography and its application to the assay of l-glutamate dehydrogenase and aminotransferases

A sensitive and selective determination of α-keto acids was established by the use of a gas chromatograph equipped with an electron capture detector. α-Keto acids (pyruvic, oxaloacetic, α-ketobutyric, and α-ketoglutaric acids) were reacted with pentafluorophenylhydrazine, and the derivatives were extracted with ethyl ether, reacted with diazomethane, and were subjected to gas-liquid chromatography with an electron capture detector. In the course of the reaction, oxaloacetic acid was decarboxylated, and yielded pyruvic acid. In the case of pyruvic (oxaloacetic) and α-ketobutyric acids two peaks corresponding to the syn and anti forms of the hydrazone appeared, and in the case of α-ketoglutaric acid, two peaks corresponding to the hydrazone and the cyclization compound produced from the hydrazone. The sum of the two peaks was taken for the determination. The present method was applicable to the assay of l-glutamate dehydrogenase, aspartate: 2-oxoglutarate, and l-alanine: 2-oxoglutarate aminotransferases.     

Dysmyelination of Shiverer Mutant Mice In Vivo and In Vitro

Demyelination in the CNS of shiverer mutant mice was studied in vivo and in vitro. By immunohistochemical reaction with glial fibrillary acidic protein antibody, hypertrophy of the fibrous astrocytes was observed in the white matter of shiverer cerebella. The cerebella of shiverer mice in primary culture from the day of birth showed very poor myelination under optical microscopy. Axons of Purkinje cells are thought to be the main myelinated axons in the primary culture of the cerebellum. Purkinje cells from shiverer appeared normal with regard to Bodian silver impregnation, hematoxylin and eosin staining, and P400 protein characterization of Purkinje cells. Addition of the conditioned culture medium of shiverer to the control culture did not interfere with myelination. We concluded that the demyelination in the CNS of shiverer could be caused by an intrinsic defect of the oligodendrocyte rather than by hypertrophy of the astrocytes or by diffusible factors.