The rapid polyphosphoinositide metabolism may be a triggering event for thrombin-mediated stimulation of human platelets

The metabolism of polyphosphoinositides was examined in human platelets activated by thrombin. The addition of thrombin to [3H]glycerol-labeled platelets induced an initial loss and a subsequent increase of the radioactivity in phosphatidylinositol-4,5-bisphosphate (TPI) without any significant change in phosphatidylinositol-4-phosphate (DPI). A marked enhancement of [32P]Pi incorporation into TPI occurred in parallel with an increase in this lipid content, which was accompanied with a conccurent decrease in phosphatidylinositol (PI). The rate of this subsequent increase in TPI was smaller than that observed in [3H]arachidonic acid-labeled platelets, suggesting that formed TPI in activated platelets may contain much greater amount of arachidonate than preexisting TPI in resting platelets. These data indicate that thrombin causes a rapid change in TPI metabolism (initial degradation of preexisting TPI and subsequent production of arachidonate-rich TPI), which might be a primary candidate to modulate thrombin-induced function in human platelets.     

Purification and properties of rat brain dipeptidyl aminopeptidase

Dipeptidyl aminopeptidase, which hydrolyzes the 7-(Gly-Pro)-4-methylcoumarinamide, has been purified from the brains of 3 week-old rats. It was purified about 2,600-fold by column chromatography on CM-cellulose, hydroxyapatite and Gly-Pro AH-Sepharose. This enzyme hydrolyzed Lys-Ala-beta-naphthylamide well with an optimum pH of 5.5. It was inhibited by diisopropyl fluorophosphate, phenyl-methanesulfonyl fluoride, some cations, and puromycin, but was not inhibited by p-chloromercuribenzoate, N-ethylmaleimide, dithiothreitol, EDTA, iodoacetic acid, and bacitracin, indicating that rat brain dipeptidyl aminopeptidase is a serine protease. This enzyme showed a molecular weight of 220,000 by gel filtration and of 51,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The properties of purified rat brain dipeptidyl aminopeptidase were similar to those of bovine pituitary dipeptidyl peptidase II, but the molecular weight and substrate specificity of these enzymes were different.     

Proton magnetic resonance studies of 7 Fe ferredoxins: Lower potential of the [4 Fe–4 S] redox center than the [3 Fe–3 S] cluster

Two types of iron-sulfur clusters, [3 Fe–3 S] and [4 Fe–4 S], were identified by ¹H-NMR in ferredoxins from Thermus thermophilus, Mycobacterium smegmatis and Pseudomonas ovalis. The [4 Fe–4 S] clusters always showed the redox couples which had potentials lower than that of the [3 Fe–3 S] clusters.     

Hydropiperoside,a novel coumaryl glycoside from the root of Polygonum hydropiper

From the methanol extract of the root of Polygonum hydropiper, a novel coumaryl glycoside hydropiperoside was isolated together with anthraquinone, ellagic acid 3,3′-di-O-methyl ether, gallic acid, two quercetin glycosides and an unidentified aromatic δ-lactone possessing antifertility activity. The structure of hydropiperoside was established as β-d-(1,3,6-tri-p-coumaryl)-fructofuranosyl-α-d-glucopyranoside by combination of extensive ¹H NMR and ¹³C NMR spectra, and the FD/MS spectrum.     

Evidence that cyclic AMP may regulate Ca2+-mobilization and phospholipases in thrombin-stimulated human platelets

The regulation of human platelet responses by cyclic AMP (cAMP) has been investigated by measuring thrombin-stimulated serotonin release, Ca²⁺ uptake and phospholipase activity. Thrombin-induced 1,2-diacylglycerol (DG) formation as a result of phospholipase C activation was inhibited by pretreatment with dibutyryl cAMP (dbcAMP) in a dose-dependent manner. Subsequent failure to produce phosphatidic acid (PA), which is converted from 1,2-DG by phosphorylation and would serve as intracellular Ca²⁺ ionophore, appeared to parallel the decrease in Ca²⁺ uptake activity. Phospholipase A₂ activity, monitored by the production of [³H]lysophosphatidylcholine and [³H]lysophosphatidylethanolamine, was also suppressed by dbcAMP. These data indicate that the intracellular cAMP level may be closely associated with Ca²⁺ uptake and phospholipases activation. In addition, it is suggested that alteration of intracellular cAMP regulates phospholipase activation and consequently platelet responses, perhaps by controlling available Ca²⁺ content.     

Cytological evidence for holocentric chromosomes of the silkworms,Bombyx mori and B. mandarina, (Bombycidae,Lepidoptera)

The nature of the centromere and the orientation in meiosis of silkworm chromosomes were investigated using the trivalent of the F₁ hybrid between the wild and domestic silkworm and X-ray-induced aberrant chromosomes as well as normal silkworm chromosomes. The results of the experiments were as follows: (1) Pro-metaphase chromosomes showed no distinct primary constriction even after treatment with hypotonic solution, (2) sister chromatids separated in parallel along the entire length of the chromosome at mitotic anaphase, (3) chiasmata underwent complete terminalization during diakinesis and thus chromosome dyads were always connected end-to-end by a terminal chiasma at metaphase I, (4) radiation-induced aberrant chromosomes were stably transmitted throughout a number of cell generations, and (5) although the homomorphic bivalents generally orientated axially at metaphase I and equatorially at metaphase II, this normal sequence tended to be inverted or modified in the X-ray-induced aberrant chromosomes and in the trivalent of the F₁ hybrid silkworms. These observations may be best interpreted by assuming the holocentric nature of silkworm chromosomes.     

Scanning and transmission electron microscopy of human ejaculate spermatozoa with special reference to their abnormal forms

Summary Spermatozoa from fertile and infertile human ejaculates were observed under the scanning electron microscope. A parallel study of sections was performed by transmission electron microscope.The normal head shows under the scanning electron microscope vesicular elevations in the region of the acrosome and a smooth and rigid appearance corresponding to the postnuclear cap whose occurrence is confirmed under the transmission electron microscope. Immediately anterior to this cap a shallow furrow transverses the head. Duplicated, unusually large or small and deformed heads are found under the scanning electron microscope. Most of these abnormal heads show no surface structure suggesting an acrosome.The neck and middle piece are occasionally, though frequently in abnormal spermatozoa, covered by a cytoplasmic droplet. Otherwise, the mitochondrial sheath is recognized under the scanning electron microscope as a beaded thickening in the middle piece. The lack of mitochondria is manifested by a smooth middle piece thinner than the principal portion. Transmission electron microscopy of sections reveals various types of anomalies in the number of cores, core filaments and mitochondria embedded in the cytoplasmic droplets.Abnormalities in the principal portion of the tail such as duplication, unusual thickness and length are shown under the scanning electron microscope.The investigation indicates that scanning electron microscopy is suited for the clinical as well as cytological examination of human ejaculate spermatozoa.     

Electron microscopic observations of lung alveolar epithelial cells of normal young mice,with special reference to formation and secretion of osmiophilic lamellar bodies

Summary Using the electron microscopy and electron microscopic histochemistry the authors studied the lung alveolar epithelial cell of normal young mice.Type II cell of the alveolar epithelium has characteristically numerous osmiophilic lamellar bodies. The lamellar boies are formed in the cytoplasmic vesicle, and never originate from the mitochondrion. These bodies have abundant acid phosphatase activity in their limiting membrane therefore it is considered to be lysosomal origin, but the mitochondria have no such enzyme activity.The body which is newly formed in the cytoplasmic vesicle grows up to the large lamellar body as a result of an accumulation of the fibrous dense substance, migrates to the free margin of the type II cell of alveolar epithelium, and then is discharged into the alveolar lumen as a merocrine type secretion.Acknowledgement is given to Professor Dr. Y. Sano and Professor Dr. H. Fujita, Department of Anatomy, and Assistant Professor Dr. S. Fujita, Department of Pathology, for their kind advice and criticism.     

Nuclear transplantation of male primordial germ cells in the mouse

We examined the developmental ability of enucleated eggs receiving embryonic nuclei and male primordial germ cells (PGCs) in the mouse. Reconstituted eggs developed into the blastocyst stage only when an earlier 2-cell nucleus was transplanted (36%) but very rarely if the donor nucleus was derived from a later 2-cell, 8-cell, or inner cell mass of a blastocyst (0-3%). 54-100%, 11-67%, 6-43% and 6-20% of enucleated eggs receiving male PGCs developed to 2-cell, 4-cell, 8-cell and blastocyst stage, respectively, in culture. The overall success rate when taking into account the total number of attempts at introducing germ cells was actually 0-6%. Live fetuses were not obtained after transfer of reconstituted eggs to recipients, although implantation sites were observed. The developmental ability of reconstituted eggs in relation to embryonic genome activation and genomic imprinting is discussed.