Free and polymerized tubulin in cultured bone cells and Chinese hamster ovary cells: the influence of cold and hormones

A low pH method of liposome-membrane fusion (Schneider et al., 1980, Proc. Natl. Acad. Sci. U. S. A. 77:442) was used to enrich the mitochondrial inner membrane lipid bilayer 30-700% with exogenous phospholipid and cholesterol. By varying the phospholipid-to- cholesterol ratio of the liposomes it was possible to incorporate specific amounts of cholesterol (up to 44 mol %) into the inner membrane bilayer in a controlled fashion. The membrane surface area increased proportionally to the increase in total membrane bilayer lipid. Inner membrane enriched with phospholipid only, or with phospholipid plus cholesterol up to 20 mol %, showed randomly distributed intramembrane particles (integral proteins) in the membrane plane, and the average distance between intramembrane particles increased proportionally to the amount of newly incorporated lipid. Membranes containing between 20 and 27 mol % cholesterol exhibited small clusters of intramembrane particles while cholesterol contents above 27 mol % resulted in larger aggregations of intramembrane particles. In phospholipid-enriched membranes with randomly dispersed intramembrane particles, electron transfer activities from NADH- and succinate-dehydrogenase to cytochrome c decreased proportionally to the increase in distance between the particles. In contrast, these electron- transfer activities increased with decreasing distances between intramembrane particles brought about by cholesterol incorporation. These results indicate that (a) catalytically interacting redox components in the mitochondrial inner membrane such as the dehydrogenase complexes, ubiquinone, and heme proteins are independent, laterally diffusible components; (b) the average distance between these redox components is effected by the available surface area of the membrane lipid bilayer; and (c) the distance over which redox components diffuse before collision and electron transfer mediates the rate of such transfer.     

The fate of spargana inoculated into the cat brain and sequential changes of anti-sparganum IgG antibody levels in the cerebrospinal fluid

To establish an animal model of intracranial sparganosis, the fate and behavior of the experimentally inoculated spargana were observed. A total of 102 scolices of spargana were injected into 22 cat brains, and the cats were sacrificed at 2 weeks, 1 month, 3 months and 6 months after the inoculation. Neurosparganosis was established in 77% of the cats. Of 43 recovered worms, 19 (44%) were located in the subdural or subarachnoid space, 16 (37%) in the brain parenchyme, and 2 (5%) in the lateral ventricle. One was detected at the diploic space of the skull and 5 were outside the cranial cavity. All but one were alive, and had grown tails. They were distributed in the brain parenchyme randomly. There was no place which they could not invade. No adult was found in the intestine. Cerebrospinal fluid (CSF) was collected before inoculation, 1 week, 2 weeks, 1 month, 3 months and 6 months after inoculation. The level of anti-sparganum IgG antibody in CSF measured by ELISA began to increase above the criteria of positivity 1 month after inoculation. Three months after inoculation, the values markedly increased. The present findings reveal that intracranial inoculation of spargana into the brains of cats would be a good animal model of experimental neurosparganosis.     

The changes of histopathology and serum anti-sparganum IgG in experimental sparganosis of mice

The present study is intended to observe the chronologic changes of experimental sparganosis by histopathological observation and detection of circulating anti-sparganum IgG antibody using ELISA. Each of 25 mice was infected with five spargana, and they were examined after 1, 2, 4, 10 weeks or 6 months from infection. The followings are summarized results. 1. The plerocercoids were detected in the subcutaneous tissue of the trunk, neck or axilla, but a few often extended into the skeletal muscle. The recovery rates were 72% at the first week, 80% at the second week, 95% at the fourth week, 92% at the tenth week and 100% at the sixth month. The larvae grew slowly in both length and weight until 6 months. 2. Histopathologically, most of the larvae were observed alive in the soft tissue or skeletal muscle. Numerous eosinophils, neutrophils, lymphocytes and plasma cells were infiltrated focally around the worms by the second week, but they surrounded the worms to form a layer of inflammatory reaction after 4 weeks of infection. Also histiocytes and fibroblasts began to appear around the inflammatory cells at 4 weeks. After 10 weeks, the worms encircled by a thin fibrous layer were found. After 6 months, the worms were surrounded by either fibrous tissue or active inflammatory cells. The inflammation looked more severe in the tracks left by the worms, rather than around the worms. 3. The level of anti-sparganum IgG antibody in the serum showed an increase by the fourth week, and a rapid and continuous increase was observed thereafter by the tenth week after infection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ginkgolide B production in cultured cells derived from Ginkgo biloba L. leaves

Callus cultures and cell suspension cultures derived from Ginkgo biloba L. leaves produced ginkgolidc B. In cell suspension cultures, the production reached a maximum by the 13th day of subculture and followed by a sharp decrease. The medium of Murashige and Skoog induced the highest ginkgolide B content in cultures while the medium of Schenk and Hildebrandt promoted cell growth. For the maximal production of ginkgolide B, cells were cultured in Murashige and Skoog medium modified to contain 1.0 mg/l of -naphthaleneacetic acid, 0.1 mg/1 of kinetin, 30 g/1 sucrose and 1.25 mM potassium phosphate with a molar ratio of ammonium to nitrate ions of 1 3.Abbreviations B5 Gamborg et al (1968) medium - GKB Ginkgolide B - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic aicd - SH Schenk and Hildebrandt (1972) medium     

A metal ion-binding site in the kringle region of bovine prothrombin fragment 1.

45Ca(II) binding studies (equilibrium dialysis) on the kringle domain of bovine prothrombin fragment 1 were conducted using a mixture of peptides (residues 43-156 and 46-156) resulting from limited alpha-chymotryptic hydrolysis of fragment 1. Analysis of the Scatchard plot of these data indicates a single, low affinity Ca(II)-binding site to be present. Similar results were obtained from studies on the decarboxylated fragment 1 derivative, 10-gamma-MGlu-fragment 1. Acetylation of bovine fragment 1 in the absence of Ca(II) or Mg(II) ions results in the loss of the metal ion-promoted quenching of the intrinsic Trp fluorescence of the protein and the Ca(II)-mediated binding to phosphatidylserine/phosphatidylcholine (PS/PC) vesicles. The acetylation of the NH2 alpha-group of Ala-1 has been shown (Welsch, D. J., and Nelsestuen, G. L. (1988) Biochemistry 27, 4946-4952) to abolish the PS/PC binding property of fragment 1. The present study demonstrates that acetylation of a second site possibly Ser-79 or Thr-81 using the conditions described in the preceding paper results in loss of both the fluorescence transition and the Ca(II)-mediated PS/PC binding of the resulting protein derivative. Removal of the O-acetyl group at the Ser-79/Thr-81 site is accomplished by aminolysis with 0.2 M hydroxylamine, pH 10, 50 degrees C; the fluorescence transition is partially restored. PS/PC binding is partially restored if the NH2 alpha-group of Ala-1 is trinitrophenylated but is not restored if the NH2 alpha-group of Ala-1 is acetylated. We conclude that the Ser-79/Thr-81 site may represent a portion of the metal ion-binding site within the kringle domain of fragment 1. Occupancy of this site by a Ca(II) ion appears to be important in the binding of the protein to PS/PC vesicles.     

Thermal history of reef-associated environments during a record cold-air outbreak event

Summary Several polar continental air masses intruding into the south Florida/northern Bahama Bank region during January 1981 caused record low air temperatures and rapid chilling of extensive shallow-water carbonate systems. Numerous coral kills along the Florida reef tract and massive fish mortalities in Florida Bay were attributable to unusually cold waters generated at this time. Thermal evolution of Florida Bay/Florida reef tract and northern Bahama Bank waters from 8 to 21 January was assessed from thermal infrared data acquired by the NOAA-6 environmental satellite, in situ water temperatures, local meteorological data, and a computerized heat flux model. Field observations and laboratory experiments identify 16°C as a thermal stress threshold for most reef corals (Mayor 1915; Davis 1981). Temperaturecorrected digital satellite data indicated that water temperatures below 16°C were generated in Florida Bay and on Little and Great Bahama Banks during a 10-day period in January. Lowest temperatures on the Florida reef tract resulted from offshelf transport of Florida Bay water through major tidal channels. Offshelf movement of bay water is driven primarily by strong northerly winds, density gradients, and tidal pumping. Absence of reef development opposite major tidal passes along the Florida reef tract (Ginsburg and Shinn 1964) and aperiodic coral kills along bank margins can be attributed to this process, which has probably had a limiting influence on Holocene reef development in these areas.     

The Drosophila caspase Ice is important for many apoptotic cell deaths and for spermatid individualization, a nonapoptotic process

Caspase family proteases play important roles in the regulation of apoptotic cell death. Initiator caspases are activated in response to death stimuli, and they transduce and amplify these signals by cleaving and thereby activating effector caspases. In Drosophila, the initiator caspase Nc (previously Dronc) cleaves and activates two short-prodomain caspases, Dcp-1 and Ice (previously Drice), suggesting these as candidate effectors of Nc killing activity. dcp-1-null mutants are healthy and possess few defects in normally occurring cell death. To explore roles for Ice in cell death, we generated and characterized an Ice null mutant. Animals lacking Ice show a number of defects in cell death, including those that occur during embryonic development, as well as during formation of adult eyes, arista and wings. Ice mutants exhibit subtle defects in the destruction of larval tissues, and do not prevent destruction of salivary glands during metamorphosis. Cells from Ice animals are also markedly resistant to several stresses, including X-irradiation and inhibition of protein synthesis. Mutations in Ice also suppress cell death that is induced by expression of Rpr, Wrinkled (previously Hid) and Grim. These observations demonstrate that Ice plays an important non-redundant role as a cell death effector. Finally, we demonstrate that Ice participates in, but is not absolutely required for, the non-apoptotic process of spermatid differentiation.     

Morphological Changes in the Midgut of Aedes aegypti L. (Diptera: Culicidae) Larvae Following Exposure to an Annona coriacea (Magnoliales: Annonaceae) Extract

Bioinsecticides are important in the control of disease vectors, but data regarding their physiological effects on target insects are incomplete. This study describes morphological changes that occur in the midgut of third instar Aedes aegypti L. (Diptera: Culicidae) following treatment with a methanolic extract of Annona coriacea (Magnoliales: Annonaceae). Dissected midguts were subdivided into anterior and posterior regions and analyzed by light and scanning electron microscopy. Insects exposed to the extract displayed intense, destructive cytoplasmic vacuolization in columnar and regenerative midgut cells. The apical surfaces of columnar cells exhibited cytoplasmic protrusions oriented toward the lumen, suggesting that these cells could be involved in apocrine secretory processes and/or apoptosis. We report that A. coriacea extracts induced morphological alterations in the midgut of A. aegypti midgut larvae, supporting the use of plant extracts for control of the dengue vector.