Pre- and post-implantation losses of embryos in aging female IVCS mice

Mated female mice of the IVCS strain, aged 90 (control group), 180, 210, 240, 270, 300, 330, 360 and 420 days old, were studied for pre- and post-implantation loss of embryos. The percentage of pre-implantation loss in mice aged 90 to 210 days was 1.7/11.8 (14.4%) to 2.7/11.7 (23.1%). In mice aged 240 to 300 days it increased significantly as compared to the controls (46.5-90.2% versus 14.4%). It reached 100% in 300-days-old mice. The post-implantation loss of embryos and/or fetuses in mice aged 90 to 240 days was 1.0/10.2 (9.8%) to 2.5/9.0 (27.8%). In mice aged 270 to 300 days it increased significantly compared to the controls (100% versus 9.8%). The decrease in reproductive activity appeared first in a decrease in litter size, followed by a decrease in the number of blastocyst and implantation sites, and finally by anovulation during the process of aging in IVCS mice.     

Immunological detection of propolypeptide of von Willebrand factor on platelet surface

Recently we have found that propolypeptide of von Willebrand factor (pp-vWF) obtained from platelets binds to type I collagen. It is known that pp-vWF is present in platelet alpha-granules and is secreted upon activation. In this paper, we demonstrate the two following evidences to show that it is also present on the surface of resting platelets. [1] The antibody against pp-vWF bound to the surface of platelets. [2] The antibody induced aggregation of platelets. The binding of the antibody and the antibody-induced aggregation of platelets were inhibited in a dose-dependent manner by Fab fragment of the antibody. Platelets from von Willebrand disease patients bound less of the antibody and responded weakly to the antibody.     

Protection against acute paraquat toxicity by ambroxol

An expression vector for G-CSF, pASLB3-3, was constructed and introduced into Namalwa KJM-1 cells (Hosoi et al., 1988), and cells resistant to 100 nM of methotrexate (MTX) were obtained. Among them, the highest producer, clone SC57, was selected and the productivity of this clone was further characterized. The maximal production of G-CSF was at the most 1.8 g/ml/day using a 25 cm² tissue culture flask, even though the cell number was above 7×10⁵ cells/ml. The limiting factors at high density were analyzed as the deficiency of nutrients, such as glucose, cysteine and serine, and pH control. The depression of specific G-CSF productivity per cell under the batch culture conditions was overcome by using a perfusion culture system, Biofermenter^TM (Sato, 1983) with modifications of nutrients supplementation by a dialysis membrane and/or dissolved oxygen (DO) supplementation by microsilicone fibers. ITPSGF medium was modified to elevate concentrations of amino acids and glucose by 2.0- and 2.5-times, respectively. Under the control of pH at 7.4 and DO at 3 ppm, the specific G-CSF productivity was not depressed even at high cell density (above 1×10⁷ cells/ml), and the amount of G-CSF reached 41 g/ml. These results indicated the possibility of finding the optimum culture conditions for the production of recombinant proteins by Namalwa KJM-1 cells.Abbreviations ABTS 2,2-Azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid - BSA Bovine Serum Albumin - BSA-PBS Phosphate-buffered Saline without Ca²⁺ and Mg²⁺ containing Bovine Serum Albumin - dhfr Dihydrofolate Reductase - DO Dissolved Oxygen - G-CSF Granulocyte Colony-stimulating Factor - HEPES 4-(2-Hydroxyethyl)-1-piperazineethansulfonic Acid - IFN Interferon - MTX Methotrexate - PBS(-) Phosphate-buffered saline without Ca²⁺ and Mg²⁺ - Tween-PBS Phosphate-buffered saline without Ca²⁺ and Mg²⁺ containing 0.05% of Tween 20     

Synthetic sialyl glycolipids (sialo-cholesterol and sialo-diglyceride) induce granulocytic differentiation of human myelogenous leukemia cell line HL-60

When HL-60 cells were cultivated with synthetic sialyl glycolipids, sialo-cholesterol and sialo-diglyceride, the cells were found to be differentiated into mature granulocytes on morphological and functional criteria. The differentiation of cells was accompanied by inhibition of cell proliferation. The differentiation-inducing activity of sialo-cholesterol was greater than that of sialo-diglyceride on a molar basis, and the alpha-anomer of each compound was more potent than the beta-anomer, suggesting that the stereospecific structure of the compounds is important for the differentiation-inducing activity.     

Ca2(+)-independent secretory mechanism of thyrotropin-releasing hormone (TRH) involves protein kinase C in rat pituitary cells

Thyrotropin-releasing hormone (TRH) stimulates biphasic prolactin (PRL) secretion from rat pituitary GH3 cells. The pretreatment of cells with EGTA (100 microM) plus arachidonic acid (15 microM), a condition which decreased TRH-responsive intracellular Ca2+ pools, eliminated the activity of TRH on burst PRL secretion (2 min) but did not alter that on sustained PRL secretion (30 min). However, the treatment of cells with EGTA, arachidonic acid and H-7 (300 microM), a potent inhibitor of protein kinase C (PKC), almost completely suppressed the activity of TRH for sustained PRL secretion. In cells down-modulated for PKC, TRH abolished this Ca2(+)-independent sustained PRL secretion. These results suggest that TRH acts through a separate, Ca2(+)-independent secretory mechanism, besides by modulating the Ca2(+)-dependent mechanism and that PKC is involved in this Ca2(+)-independent secretory pathway.     

Phospholipid monolayers at the triolein-saline interface: production of microemulsion particles and conversion of monolayers to bilayers

Interfacial tensions of phospholipid monolayer at the triolein (TO)-saline interface were measured. The adsorption isotherms and the interfacial pressure-molecular area curves were evaluated on the basis of the measurements. Phosphatidylcholine (PC) forms a highly condensed monolayer, with a large lateral attractive interaction; phosphatidylethanolamine (PE) and phosphatidylserine (PS) form expanded monolayers with smaller lateral interaction energies. At the lowest interfacial tension (the highest interfacial pressure), the mole fractions of PC, PE, and PS in the monolayers are estimated as 0.95, 0.73, and 0.88, respectively. Therefore, PC forms the most stable monolayer at the interface. These results are consistent with the finding that the stable TO particles in aqueous solution were produced by using PC as an emulsifier, and PE and PS did not stabilize the particles. The phase diagram of TO and PC mixtures in saline obtained from theoretical considerations predicts the equilibrium conversion of the monolayers on TO particles to bilayers. This process may be closely related to the transformations of very low density lipoproteins and chylomicrons to high-density lipoproteins in plasma. The particle sizes of the emulsion are calculated theoretically as a function of PC mole fraction in the TO-PC mixture and compared with the experimental values obtained from quasi-elastic light scattering (QLS) measurements.     

Extraction method for preparing pyridylamino sugar derivatives and application to porcine gastric mucus glycoprotein analysis

For the sensitive detection of free sugars and oligosaccharides, the use of their pyridylamino derivatives has now found general acceptance. To remove excess 2-aminopyridine from this derivative in a reaction mixture, gel filtration and ion-exchange chromatography were conducted. It was found in the present study that contaminated 2-aminopyridine could be selectively removed from the reaction mixture by adjusting the pH with saturated sodium bicarbonate at above 8.5 followed by extraction with benzene. By using this method, fewer purification steps and less time are required, with minimum loss of pyridylamino sugar derivatives.     

Genetic studies on site-specific endodeoxyribonucleases in Bacillus subtilis: multiple modification and restriction systems in transformants of Bacillus subtilis 168

Summary We transformed B. subtilis 168 with DNA from B. subtilis IAM1231, IAM1192 and ATCC6633. When we examined the restriction activities of the transformants in vivo and in vitro using phage 105C we found the following: (1) Cells of either IAM1231 or IAM1192 have two modification and restriction systems (Bsu1231(1)-system and Bsu1231(II)-system in IAM1231, and Bsu1192(I)-system and Bsu1192(II)-systems in IAM1192), and cells of ATCC6633 have only one system (Bsu6633-system). (2) The restriction enzymes of all of these five systems are site-specific endonucleases. (3) The nucleotide sequence specificities of the enzymes involved in Bsu1231(I)-system, Bsu1192(I)-system and Bsu6633-system are the same; and those of Bsu1231(II)-system and Bsu1192(II)-system are the same. The sequence specificities of these two groups are different from each other and also different from those of the Bsu168-system of B. subtilis 168, the BsuR-system of B. subtilis R and the Bsu1247(I)-and Bsu1247(II)-systems which are systems of B. subtilis IAM1247. (4) Transformants possessing four different modification and restriction systems (Bsu1231(I)-, Bsu1247(I)-, BsuR- and Bsu168-systems) were constructed. (5) Transformation of two derivatives of 168 that were m _R ⁺ r _R ⁺ by DNA from IAM1231 produced 16 transformants that had the Bsu1231(II) restriction system, but had lost the BsuR system. Transformation of a derivative of 168 that was m _1247(II) ⁺ r _1247(II) ⁺ by DNA from m _1231(II) ⁺ r _1231(II) ⁺ -or m _R ⁺ r _R ⁺ -derivative of 168 produced about 100 each of transformants that had the Bsu1231(II)-restriction system or the BsuR-restriction system. But all these transformants lost the Bsu1247(II)-system.     

A new site-specific endonuclease StuI from Streptomyces tubercidicus

A new sequence-specific endonuclease, StuI, produced by Streptomyces tubercidicus KCC S-0054, was identified and partially purified. StuI recognizes the hexanucleotide "palindromic" sequence (Formula: see text), and cleaves it at the middle, producing blunt ends.     

Relationship of ploidy and chromatin condensation in the rat liver,moreover a comparison of the nuclear texture in sections and touch preparations

Summary In touch preparations and tissue sections from normal rat hepatocytes and -amanitine- and actinomycind-poisoned liver cells the extent of chromatin condensation relatively to the degree of ploidy and the chromatin distribution were studied by means of computer aided cytophotometry. It could be found, that the relationship of the condensed and decondensed chromatin is independent of the degree of ploidy and first of all dependent upon the intoxicant. Therefore, the extent of chromatin condensation can be utilized also in sections as additional parameter for automated cell screening.Supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 105