Myocardial adaptations in aortic valve disease.

Injections of human chorionic gonadotropin (HCG) have been claimed to aid in weight reduction by reducing hunger, and affecting mood as well as aiding in localized (spot) reduction. We have tested these claims in a double-blind randomized trial using injections of HCG or placebo. Weight loss was identical between the two groups, and there was no evidence for differential effects on hunger, mood or localized body measurements. Placebo injections, therefore, appear to be as effective as HCG in the treatment of obesity.     

Radioimmunoassay for β-endorphin: Presence of immunoreactive “big-big” β-endorphin (“big” β-lipotropin) in human and rat pituitaries

A sensitive and specific radioimmunoassay for β-endorphin has been developed with an antiserum obtained in a rabbit immunized with β-endorphin contained in crude porcineACTH preparations. The minimal detectable quantity was 5 pg. The antiserum used reacted slightly with ovine β-lipotropin (5.5 %), but showed negligible cross-reactivity with other fragments of β-lipotropin, α-MSH and ACTH. Using this radioimmunoassay we have observed the presence of “big-big” β-endorphin (“big” β-lipotropin) with apparent molecular weights of 37,000 and 31,000 in human and rat pituitaries respectively, in addition to β-lipotropin and β-endorphin, by Sephadex gel-chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis.     

(+)-5,17-dehydromatrine N-oxide,a new alkaloid in Euchresta japonica

A new lupin alkaloid, (+)-5,17-dehydromatrine N-oxide, was isolated from the fresh aerial parts of Euchresta japonica. Its structure was confirmed by spectrometric data and by direct comparison with a synthetic sample, prepared from (+)-sophoranol ((+)-5-hydroxymatrine). It was also concluded that (+)-5,17-dehydromatrine N-oxide and (+)-matrine N-oxide possess the same configuration with respect to the asymmetric nitrogen by NMR spectra.     

Requirement of the core structure of a complex-type glycopeptide for the binding to immobilized lentil- and pea-lectins

Structural requirements for the binding of oligosaccharides and glycopeptides to immobilized lentil- and pea-lectins were investigated by use of radioactively-labeled glycopeptides and oligosaccharides. The results indicate that an intact 2- acetamido-2-deoxy-β-d-glucopyranosyl residue at the reducing end of a complex-type oligosaccharide is essential for high-affinity binding to lentil lectin-Sepharose but not to concanavalin A-Sepharose and that an asparagine residue is required for the binding of a complex-type glycopeptide to pea lectin-Sepharose. In addition, interaction of a complex-type oligosaccharide with lentil lectin-Sepharose was enhanced by exposure of nonreducing, terminal 2-acetamido-2-deoxy-β-d-glucopyranosyl groups, whereas interaction with pea lectin-Sepharose was enhanced only after exposure of nonreducing, terminal α-d-mannopyranosyl groups.     

Enhanced activity of deoxyuridine 5′-triphosphatase in regenerating rat liver

An enzyme, dUTPase, that catalyzes the conversion of dUTP to dUMP and PPi, was partially purified from regenerating rat livers. The molecular weight was estimated by gel filtration to be 60,000. The apparent Km for dUTP was 12 μM. No other deoxyribonucleoside triphosphates served as a substrate. This enzyme is active in the absence of added divalent cations or sulfhydryl reagents; the activity could be inhibited by EDTA and shows a broad pH optimum with no decrease in activity from pH 7 to 11. The specific activity of dUTPase in rat liver begins to rise 16 h after partial hepatectomy and reaches a maximum about 24 h after the operation, rising to at least 5 to 6 times the normal level.     

Temperature-sensitive mutants of adenovirus single-stranded DNA-binding protein. Inability to support DNA replication is associated with an altered DNA-binding activity of the protein.

The adenovirus single-stranded DNA-binding protein (DBP) is an essential factor in viral DNA replication. Three temperature-sensitive (ts) adenoviruses (Ad2+ND1ts23, Ad2ts111A, and Ad5ts125) are known to have single amino acid substitutions in their DBPs that result in defective DNA replication at the nonpermissive temperature. To elucidate the mechanism(s) involved in the ts phenotype, we purified the three mutant DBPs and studied their DNA-binding properties and their ability to support DNA replication in an in vitro system. The results confirm that the three ts DBPs were incapable of supporting DNA replication at the nonpermissive temperature (40 degrees C). The defect was found at both the initiation and elongation steps of DNA replication. The 2-fold stimulation of pTP.dCMP formation by the DBP was lost by prior heating of the ts DBPs. The pronounced effect of the DBP on the early elongation process was severely diminished, but not abolished, by prior heating to 40 degrees C. The functional change at 40 degrees C was irreversible, as the ts DBPs preincubated at 40 degrees C were no longer active when assayed at 30 degrees C. Upon heating to 40 degrees C, all three ts DBPs lost their ability to bind to oligonucleotides, although they still retained some binding activity for large single-stranded DNAs such as M13 DNA. Thus, the inability of these three ts DBPs to support DNA replication is attributable to their altered DNA-binding properties.     

An improved approach for transformation of plant cells by microinjection: molecular and genetic analysis

A new culture method for the injection of tobacco mesophyll protoplasts has been established. The protoplasts are embedded in a thin layer of alginate and are nourished from the medium in the underlying basislayer. In the alginate layer the protoplasts regenerate to calli at a frequency of up to 80%. Embedded protoplasts can be selected either with 50 mg l⁻¹ kanamycin or 5 mg l⁻¹ paromomycin. Single resistant cells can be recovered from about 10 000 sensitive cells in one alginate layer. Injection of theneo gene (coding for neomycin phosphotransferase II) into protoplast derived single cells in the alginate layer results in kanamycin resistant colonies that can be regenerated to mature plants. These plants express the neomycin phosphotransferase as shown by enzyme activity assay. The integration of the transgene into the plant genome could be proved by Southern hybridization to high molecular weight DNA. With this culture method 100 cells can be injected per hour. Transformation frequencies range from 2 to 20%. In crossing experiments, it was shown that the foreign gene is transmitted to the next generation in a Mendelian fashion.     

Immunogenic properties of mannose-containing ceramide disaccharide and immunochemical detection of its hapten in the two kinds of crustacean, Euphausia superba and Macrobrachium nipponense.

Antiserum against Man beta 1-4Glc beta 1-1Ceramide (MIOse2Cer), a mannolipid isolated from spermatozoa of the fresh-water bivalve, Hyriopsis schlegellii, has been elicited in rabbits by repeated injection of a mixture of hapten-bovine serum albumin (1:1, mg/ml) with Freund's adjuvant. The specificity of the affinity-purified antibody (immunoglobulin G type) obtained from the serum was examined, using other glycosphingolipids and glyco-proteins structurally related to MIOse2Cer, by means of ELISA and TLC-immunostaining. The purified antibody was highly specific to MIOse2Cer and lacked reactivity with other glycolipids and glycoproteins including glucosylceramide, lactosylceramide, dimannosylglucosylceramide (MIOse3Cer), glucosaminylmannosylglucosylceramide (ArOse3Cer), thyroglobulin and alpha 1-acid glycoprotein. The antibody was found to bind, although less efficiently, to certain other compounds containing the group Man beta 1-4Glc and/or Man beta 1-4GlcNAc at their termini, such as MIOse2-sphingosine and Man beta 1-4GlcNAc beta 1-p-aminobenzoic acid ethylester derivatives. The present antibody was applied to the detection of the natural hapten in crustacean glycolipids. The purified antibody reacted with a neutral glycosphingolipid present in the two kinds of crustacean, Euphausia superba (antarctic krill) and Macrobrachium nipponense (fresh-water shrimp) as shown by TLC-immunostaining. The crustacean glycolipid antigen was isolated and characterized to be the Man beta 1-4Glc-Cer. This is the first report on the presence of a mannose-containing glycosphingolipid in the crustacean.     

Development of the Formula-Detoxification System during Adaptation to Air of Submerged Rice Seedlings

Changes in activities of enzymes and levels of antioxidant substratesinvolved in the -detoxification system in seedlings of rice (Oryza sativa L. cv. Yamabiko) inresponse to variations in the oxygen environment were studied.Activities of superoxide dismutase, ascorbate peroxidase, monodehydroascorbatereductase, dehydroascorbate reductase, glutathione reductaseand catalase, expressed either on the basis of fresh weightof shoots or relative to levels of soluble protein were muchlower in seedlings germinated under water for 6 days than inthose germinated in air for the same period of time. When submergedseedlings were exposed to air, the activities of these six enzymesincreased to or exceeded the levels in aerobically grown controlsduring 24 h of adaptation to air. Ascorbate and glutathione,which act as antioxidant substrates in the -detoxification system, were present in submergedseedlings at nearly the same levels as those found in aerobicallygrown controls. On exposure of submerged seedlings to air, thelevel of ascorbate in creased slightly, but the level of glutathioneshowed a rapid increase, reaching 7 times that in aerobicallygrown controls within 12 h of adaptation to air. Levels of allsix antioxidative enzymes and of two substrates involved inthe detoxification of the superoxide radical increased withincreases in oxygen tension in the environment. Moreover, thedevelopment of this system consisted of two steps, namely, arapid increase in the level of glutathione and a subsequentslow increase in the activities of antioxidative enzymes. ¹ Present address: Research Institute for Bioresources, OkayamaUniversity, Kurashiki, 710 Japan.     

Search for MHC-associated genes in human: five new genes centromeric to HLA-DP with yet unknown functions

Taking advantage of five mouse genomic or cDNA probes [KE5(probe 14), KE4 (probe11), KE3 (probe7), KE2 (probe5), and SET] mapped on the H-2K region in mouse, we have identified and localized homologues of these five genes in the human major histocompatibility complex region (HKE5, HKE4, HKE3, HKE2, and HSET, respectively). Cosmid cloning and pulsed field gel electrophoresis analyses indicated that a human homologous gene, HKE5, is located 10 kilobases (kb) centromeric of the 2(XI) collagen (COL11A2) gene followed by HKE4. HKE3, closely linked to HKE2, is located 170 kb centromeric of HKE4. Furthermore, HSET is located 50 kb centromeric of HKE2. This gene organization outside the DP subregion is completely identical to that of the mouse H-2K region centromeric of I-Pb 3, a mouse homologue of the DPB gene, except the lack of genes corresponding to the H-2K and -K2 genes in human.