Lymphocyte activation and serine-esterase induction following recombinant interleukin-2 infusion for lymphomas and acute leukaemias

Summary C57BL mice inoculated with radiation leukemia virus (RadLV) develop preleukemic cells long before the onset of leukemia. These cells are potentially immunogenic but fail to elicit an immune response in the host because of the appearance of virus-specific suppressor T cells. We have studied the effect of polysaccharide K (PSK) on the generation of RadLV-specific cell-mediated immune responses in vitro. Long-term exposure to PSK in culture potentiated the ability of immunized T cells to respond to a RadLV-induced lymphoma. It also abrogated the suppressive activity of suppressor T cells and simultaneously boosted the ability of reactive T cells to respond. The dual immunostimulating activity of PSK resulted in the generation of T cytotoxic lymphocytes that could lyse lymphoma cells in vitro. The results suggest that PSK could be used as a prophylactic immune response modifier in preleukemia.     

Differential and analytical subfractionation of rat liver components internalizing insulin and prolactin

Receptor-mediated endocytosis of 125I-insulin and 125I-prolactin into liver parenchymal cells has been studied by quantitative subcellular fractionation. Differential centrifugation yielded three particulate fractions, N (nuclear), ML (large granule), and P (microsomes), and a final supernatant (S). Quantitative differences in the extent and rates of accumulation of 125I-insulin and 125I-prolactin into the fractions were observed. The acidotropic agent chloroquine and the microtubule disrupting agent colchicine were administered separately to rats. The agents increased significantly the T 1/2 of hormone clearance from the liver and augmented the accumulation of both ligands in the low-speed ML fraction. However, differences in the rates of accumulation of insulin and prolactin into all cell fractions were still maintained. Analytical centrifugation of each of the particulate fractions was carried out in order to determine if different endocytic components were specific to insulin or prolactin internalization. This was not the case. An "early" endosomal component of density 1.11 was identified in microsomes. A "late" endosome of density 1.10 was identified in the large granule (ML) fraction. Both endosomal components appeared to accumulate insulin and prolactin but at different rates. Marker enzyme analysis identified the presumed plasma membrane component in microsomes (density approximately 1.155). This component showed a significant difference in the rate of loss of 125I-insulin (T 1/2 approximately 4.1 min) as compared to that of 125I-prolactin (T 1/2 approximately 12.7 min). A further difference in the handling of the ligands was observed in early endosomes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential kinetics and sensitivity to chloroquine of receptor-mediated insulin and prolactin endocytosis in liver parenchymal cells

Systemically injected [125I]prolactin or [125I]insulin was accumulated and cleared from rat liver at different rates. Quantitative subcellular fractionation indicated a predominant accumulation of [125I]insulin in liver microsomes while [125I]prolactin was found in both the light-mitochondrial and microsomal fractions. The acidotropic agent chloroquine diminished the rate and extent of loss of each ligand from liver homogenates. In chloroquine treated rats, radiolabeled insulin accumulated in both the light-mitochondrial and the microsomal fractions. Subfraction of microsomes on discontinuous sucrose gradients revealed "early' endosomes in which ligand uptake was maximal at 2-5 min. In contrast, comparable subfraction of the of light mitochondrial fraction revealed "late' endosomes in which ligand uptake was maximal at 10-20 min. Chloroquine-treated rats showed a more marked enhancement of insulin compared to prolactin uptake in the "early' endosomes. It is suggested that "early' endosomes found in the Golgi-intermediate and -heavy fractions floated from parent microsomes may selectively degrade insulin but not prolactin. This could account for the apparently different kinetics of insulin and prolactin uptake into liver parenchyma.     

A highly repeated DNA sequence in Arabidopsis thaliana

Summary Three members of a family of highly repeated DNA sequences from Arabidopsis thaliana have been cloned and characterized. The repeat unit has an average length of 180 bp and is tandemly repeated in arrays longer than 50 kb. This family represents more than one percent of the Arabidopsis genome. Sequence comparisons with tandemly repeated DNA sequences from other Cruciferae species show several regions of homology and a similar length of the repeat unit. Homologies are also found to highly repeated sequences from other plant species. When the sequence CCGG occurs in the repeated DNA, the inner cytosine is generally methylated.     

Isolation and Characterization of Pyrophosphate:D-Fructose-6-phosphate 1-Phosphotransferase from Cucumber Seeds

Pyrophosphate:D-fructose-6-phosphate 1-phosphotransferase waspurified over 700-fold from germinating cucumber (Cucumis sativuscv. Fletcher) seeds. The purified enzyme has a specific activityof 5.2 µmol.min^–1.mg protein^–1 in the presenceof 1 µM fru-2,6-P₂. The pH optima is similar for boththe forward and reverse reactions (pH 7.5–7.8). Magnesium,manganese and cobalt activate the enzyme, with the highest affinitybeing for magnesium. The enzyme exhibits normal Michaelis-Mentenkinetics in both the presence and absence of fru-2,6-P₂. Half-maximumactivation of the enzyme was obtained with 35 nM fru-2,6-P2.Fru-2,6-P₂ stimulates activity by increasing V_max and increasingthe affinity for fru-6-P, fru-1,6-P₂ and PPi. Phosphate causesnoncompetitive inhibition with respect to both fru-6-P and PPi.On the basis of the steadystate substrate interaction and Piinhibition data a sequential ternary complex mechanism is proposed. (Received April 28, 1986; Accepted July 9, 1986)     

Method for improving the detection of viruses in fecal samples.

A method is described that improved the detection of viruses in fecal samples by electron microscopy. The virus particles were concentrated, and much of the background debris was removed by adsorption of viruses on meat protein added to the fecal sample at a low pH and a low salt concentration. Viruses were eluted by raising the pH and the salt concentration. Further concentration was achieved by acid precipitation and vacuum dialysis.     

Sex ratio and intrasexual kin competition in mammals

Summary Asymmetries in both intrasexual competition and generation overlap occur in Antechinus (Dasyuridae; Marsupialia). We show that the range of interpopulation variation in the sex ratio of pouch young spans and exceeds the range of sex ratios at birth hitherto recorded from eutherians (Clutton-Brock and Albon 1983). Although postweaning dispersal and male mortality are similar among all Antechinus populations, interpopulation variance in female longevity leads to variable duration of mother/daughter interaction. As this duration increases, parental investment is increasingly biased toward males, supporting the view that local competition among female kin for resources may influence mammalian sex ratios.     

Restriction fragment polymorphisms as probes for plant diversity and their development as tools for applied plant breeding

Summary Maize and tomato cDNA clones have been hybridized in Southern blotting experiments to plant genomic DNA prepared from different lines to detect restriction fragment polymorphisms (RFPs). In maize we have found that a high degree of genetic variability is present, even among domestic inbred lines. Most randomly chosen maize cDNA clones can be used to detect elements of this variability. Similar levels of polymorphism are observed when genomic DNA is digested with any of a number of different restriction enzymes and probed with individual clones. When a clone is hybridized to genomic DNAs prepared from several different maize lines, a number of different alleles are often detected at a single locus. At the same time one clone can often detect more than one independently segregating locus by cross hybridization to related sequences at other loci. As expected these markers are inherited as simple codominant Mendelian alleles from one generation to the next and colinkage of these markers can be demonstrated in the progeny from a heterozygous parent. In similar studies with tomato, remarkably different results were found. Few RFPs were demonstrable among domestic Lycopersicon esculentum lines although a higher level of variability could be detected when comparing esculentum with its wild Lycopersicon relatives. These results are discussed in relation to the applied uses of RFPs in plant breeding as well as the inherent variability of different plant genomes.This work was supported in part by funds from Sandoz Ltd. (Basel, Switzerland) and its subsidiary company, Northrup King Co. (Minneapolis, Minn., U.S.A.) as well as by NSF SBIR grant #BSR-8360870.     

General method for the derivation and numerical solution of epithelial transport models

Summary A general method is presented for the formulation and numerical evaluation of mathematical models describing epithelial transport. The method is based on the principles of conservation of mass, and maintenance of electroneutrality within the cells and bathing solutions. It is therefore independent of the specific membrane transport mechanisms, and can be used to evaluate different models describing arbitrary transport processes (including passive, active and cotransport processes). Detailed numerical methods are presented that allow computation of steady-state and transient responses under open-circuit, current-clamp and voltage-clamp conditions, using a general-purpose laboratory minicomputer. To evaluate the utility of this approach, a specific model is presented that is consistent with the Koefoed-Johnson and Ussing hypothesis of sodium transport in tight epithelia (Acta Physiol. Scand. 42:298–308, 1958). This model considers passive transport of an arbitrary number of permeant solutes, active transport of sodium and potassium, and osmotically induced water transport across the apical and basolateral membranes. Results of the model are compared to published experimental measurements in rabbit urinary bladder epithelium.