Determination of the rate of microthrombosis in experimentally induced disseminated intravascular coagulation (DIC)]

A procedure is described for the determination of the rate of microthrombosis in rats. It is based on the quantitative estimation of I125-fibrin deposits by measurement of the accumulated radioactivity in the organs after substitution of blood.     

Radioimmunoassays for free and conjugated trienbolone and for trienbolone acetate in bovine tissue and plasma samples.

A specific, sensitive, precise and accurate radioimmunoassay has been developed for the quantitation of the synthetic anabolic steroid trienbolone acetate (TBA) and its major metabolites, free and conjugated trienbolone (TBOH) in bovine tissues and plasma. With the extraction procedure described unspecific interference with the antigen-antibody reaction could be ruled out. The assay can significantly detect amounts of more than 40 pg TBOH and 70 pg TBA. 0.1 - 2.0 g tissue and 0.1 - 1 ml plasma are sufficient for 1 determination. Residues (range 0.1 -2.0 ng/g) were still present in calves 69 days after implantation of "Revalor" (20 mg estradiol-17beta and 140 mg TBA) with the highest concentrations found in liver and TBA could only be quantitated in fat.     

Measurements of fast light-induced light-scattering and -absorption changes in outer segments of vertebrate light sensitive rod cells

Flash-induced changes of light-absorption and of light-scattering of vertebrate rod outer segments (ROS) from frog and cattle in suspension were measured at 380 and 800 nm. The photometer used allows the observation of light intensity changes under well defined angles. We studied the successive decrease of the signal amplitude in series of flashes. One flash bleaches about 1% rhodopsin. The following results are discussed:

1. The signal at 380 nm is a superposition of the absorption change caused by formation of metarhodopsin II and of a biphasic additional signal. The latter exists only for the initial range of bleaching (15 to 25% rhodopsin).
2. At 800 nm three scattering signals are observed which are characterized by their successive amplitude decrease and time course:

N: A small signal with time course and successive amplitude decrease comparable to the metarhodopsin II absorption change, probably arising from a structural change within the disc membrane. Ni: A slow signal, disappearing with the first flash, which may be understood as an outer membrane effect. P: A biphasic signal with a successive decrease rate, by a factor of 10 to 20 higher than that of the metarhodopsin II signal. The two kinetically different components are separated by variation of the observation angle. Two regions of different extension appear to change structurally with different time course. “P” may reflect an influence of the light-induced transmitter release on disc shape and/or mass.     

Distribution of alpha-1-antitrypsin phenotypes in two Dutch population groups

Summary The ₁-antitrypsin phenotypes of two Dutch population groups (consisting of 672 and 802 individuals) were determined by the isoelectric focusing technique, which due to its recent development, has been used for the first time in large-scale phenotyping.As in other population studies on the ₁-antitrypsin phenotype distribution, Pi M is the most frequently occurring allele. The two investigated groups exhibit remarkable differences, both to other studied groups as well as to each other. The most interesting results are probably the high frequencies of the alleles Pi- and of the recently discovered Pi M_N. Comparison with phenotype studies carried out in other populations is also presented.     

Macrophage requirements of CR- and CR+ B lymphocytes for antibody production in vitro.

A Sephadex G-10 column coated with antigen-antibody complexes and complement retains complement receptor-bearing (CR+) mouse spleen cells. The effluent is rich in thymus-derived cells (T cells), and contains bone marrow-derived cells (B cells) which carry surface immunoglobulin (Ig), Ir-associated antigen (Ia), and Fc receptors, but no complement receptors (CR-). Although both unfractionated and CR- B cell populations are capable of producing antibody to red cell antigens, they differ in their requirements for the initiation of the response. Unfractionated B cells cooperate with primed as well as unprimed helper T cells; macrophages are required for this cooperation but can be replaced by 2-mercaptoethanol. CR- B cells cooperate with primed but not with unprimed T cells provided macrophages are added to cultures. After addition of culture supernatant from BCG-activated macrophages CR- B cells cooperate with both unprimed and primed T helper cells.     

Regulatory gene INO4 of yeast phospholipid biosynthesis is positively autoregulated and functions as a transactivator of fatty acid synthase genes FAS1 and FAS2 from Saccharomyces cerevisiae.

The sequence motif 5' TYTTCACATGY 3' functions as an upstream activation site common to both yeast fatty acid synthase genes, FAS1 and FAS2. In addition, this UASFAS element is shared by all so far characterized genes of yeast phospholipid biosynthesis. We have investigated the influence of a functional INO4 gene previously described as a regulator of inositol biosynthesis on the expression of FAS1 and FAS2. In a delta ino4 null allele strain, both genes are expressed at only 50% of wild type level. Using individual UASFAS sequence motifs inserted into a heterologous test system, a drastic decrease of reporter gene expression to 2-10% of the wild type reference was observed in the delta ino4 mutant. In gel retardation assays, the protein-DNA complex involving the previously described FAS binding factor 1, Fbf1, was absent when using a protein extract from the delta ino4 mutant. On the other hand, this signal was enhanced with an extract from cells grown under conditions of inositol/choline derepression. Subsequent experiments demonstrated that INO4 expression is itself affected by phospholipid precursors, mediated by an UASFAS element in the INO4 upstream region. Thus, in addition of being an activator of phospholipid biosynthetic genes, INO4 is also subject to a positive autoregulatory loop in its own biosynthesis.     

DNA methylation affecting the expression of murine leukemia proviruses.

The endogenous, vertically transmitted proviral DNAs of the ecotropic murine leukemia virus in AKR embryo fibroblasts were found to be hypermethylated relative to exogenous AKR murine leukemia virus proviral DNAs acquired by infection of the same cells. The hypermethylated state of the endogenous AKR murine leukemia virus proviruses in these cells correlated with the failure to express AKR murine leukemia virus and the lack of infectivity of cellular DNA. Induction of the endogenous AKR murine leukemia virus proviruses with the methylation antagonist 5-azacytidine suggested a causal connection between DNA methylation and provirus expression. Also found to be relatively hypermethylated and noninfectious were three of six Moloney murine leukemia virus proviral DNAs in an unusual clone of infected rat cells. Recombinant DNA clones which derived from a methylated, noninfectious Moloney provirus of this cell line were found to be highly active upon transfection, suggesting that a potentially active proviral genome can be rendered inactive by cellular DNA methylation. In contrast, in vitro methylation with the bacterial methylases MHpaII and MHhaI only slightly reduced the infectivity of the biologically active cloned proviral DNA. Recombinant DNA clones which derived from a second Moloney provirus of this cell line were noninfectious. An in vitro recombination method was utilized in mapping studies to show that this lack of infectivity was governed by mechanisms other than methylation.     

Helper cell factors restore antibody responses of allogeneic bone marrow chimeras: evidence for ineffective cellular interactions

We investigated the nature of deficient antibody responses to SRBC in stable, fully allogeneic bone marrow chimeras. No evidence for a suppressor cell-mediated mechanism was found. Chimera spleens possessed adequate numbers of antigen-reactive B cells to produce a normal antibody response. Using separated chimera cell populations and soluble helper factors, we assessed the functional capabilities of chimera B cells, T cells, and macrophages. Our data suggest that the failure of allogeneic chimeras to produce antibody is not the result of impaired B cell, T cell, or macrophage function, but rather that it is due in ineffective cellular interactions that normally result in the generation of helper factors. In vitro stimulation of chimera macrophages with LPS, and of chimera spleen cells with Con A, resulted in the release of soluble helper factors that were capable of fully restoring chimera B cell responses.     

Anther culture as a breeding tool in rape

Summary Progeny analysis of androgenetic plants from inbred rape-seed (Brassica napus) shows that selective growth of microspores can occur in cultured anthers. The property of privileged growth in culture seems to be linked to such characters as flowering time and seed glucosinolate content which can be analyzed in regenerated plants. This type of selection and the fact that more variability is visible in regenerants from different microspores than in the progeny of the highly inbred anther donor line, demonstrates the higher degree of homozygosity in the doubled amphihaploids of B. napus. Furthermore, it is shown that haploid genomes of rape may be mutable. Thus it is possible to obtain several different homozygous lines from a single microspore. A system of haploid embryoids arising from single cells of the primary microspore regenerant has also been used to produce experimentally induced mutants. It is demonstrated that recessive mutations can be obtained in a homozygous state in doubled haploid regenerants from mutagenized haploid single cells.Part of the material analyzed in this work was produced under the guidance of Emrys Thomas when he was one of the project leaders of the Projektgruppen Haploide in der Pflanzenzüchtung at the Max-Planck-Institut für Pflanzengenetik in Ladenburg. His untimely death at the age of 36 (May 23, 1981), which overtook him at his new place of work at Harpenden, prevented him seeing the completion of this paper. It is dedicated to the memory of this distinguished and restless researcher and highly-valued colleague     

Isolation and identification of three ecdysteroid conjugates with a C-20 hydroxy group in eggs of locusta migratoria

Three ecdysteroids conjugates with a hydroxy group at C-20 were isolated from developing eggs of Locusta migratoria and identified as 22-phosphate conjugates of 2-deoxy-20-hydroxyecdysone, 20-hydroxyecdysone and 20-hydroxyecdysone acetate.