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1.
Reassessment of fluid-phase endocytosis and diacytosis in monolayer cultures of human fibroblasts 总被引:3,自引:0,他引:3
We have investigated the kinetics of fluid-phase endocytosis and diacytosis in confluent monolayers of human fibroblasts by comparing the behavior of three markers that have been previously used to study this process: [14C]sucrose, 125I-labeled polyvinylpyrrolidone ([125I]PVP), and Lucifer Yellow. Three distinct kinetic compartments were observed with all markers. The first was relatively large (10-60 fl/cell), reached steady state within 15 min at 37 degrees C, and was rapidly lost from monolayers after removing the markers at 37 degrees C but not at 0 degree C. These properties indicate that this compartment is the same as that previously proposed to be the major intracellular compartment involved in diacytosis. However, this compartment is probably extracellular fluid trapped between cells since it is rapidly lost into the medium when the cells are either scraped or enzymatically removed from the culture dishes at 0 degree C. In addition, it very slowly undergoes both filling and emptying at 0 degree C. However, we did observe a second, much smaller, kinetic compartment (approximately 2 fl/cell) undergoing rapid diacytosis that does seem to be intracellular. A third compartment that we observed accumulates markers at a linear rate (10-20 fl cell-1 hr-1) and is not lost from cells even after incubation periods greater than 6 hr. The markers [14C]sucrose and [125I]PVP displayed very similar behavior with respect to all three compartments and yielded nearly linear long-term uptake rates, thus indicating that there is little if any absorbed component in their uptake. However, Lucifer Yellow displayed significantly higher incorporation rates and its uptake rate was strongly nonlinear, indicating its uptake in fibroblasts is predominantly adsorptive. Our observations indicate that the rate of fluid-phase endocytosis in fibroblasts is significantly less than previously reported and that any compartment involved in diacytosis is very small and turns over very rapidly. Significantly, we estimate that the constitutive internalization of clathrin-coated pits is sufficient to account for the majority of fluid-phase endocytosis and thus represents a major mechanism of membrane retrieval in these cells. 相似文献
2.
Physical and Chemical Correlates of Microbial Activity and Biomass in Composting Municipal Sewage Sludge 总被引:8,自引:2,他引:6
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Various physical and chemical parameters were monitored to evaluate their influence on the microbial communities present in composting municipal sewage sludge. Temperature, moisture content, depth, pH, protein content, total nitrogen, total carbon, lipid phosphate biomass, and the rates of microbial incorporation of substrates into lipids were measured at several times throughout the 17- to 19-day composting runs. Temperature was found to have the most consistent and dramatic effect on microbial activity and biomass. When temperatures exceeded 55 to 60°C, microbial activity fell dramatically, usually by more than 1 order of magnitude. Microbial activity was generally greatest in samples taken from the 35 to 50°C areas of the composting piles. Changes in the composition of the compost over time included increased pH, increased protein content, and decreased total organic content. The changes in these parameters appeared to reflect the microbial activity and biomass present. The results of this study indicate that the rate of composting may best be optimized by controlling the composting temperatures, provided that the other parameters fall within reasonable limits in the starting material. 相似文献
3.
E Tarjan D A Denton M J McKinley J F Nelson R S Weisinger 《Journal de physiologie》1984,79(6):466-470
Wild rabbits Oryctolagus cuniculus (L) introduced to Australia over a century ago successfully colonized diverse environments in a large part of the continent varying from arid desert, alps, to lush grasslands and coastline where water and salt may be either abundant or very scarce. Wild rabbits caught in Northern Victoria were studied under laboratory conditions, where they adapted to dry pelleted food and drank regularly water and a cafeteria of electrolyte solutions offered. Intracerebroventricular (IVT) infusion of angiotensin II (AII) in doses 10, 50 and 500 ng/h did not increase their water drinking, but increased salt appetite, although it was delayed one or more days after the beginning of AII infusion. IVT infusion of AII 500 ng/h for one day caused a halving in water intake and a tenfold increase in sodium excretion. These were followed by compensatory changes in water and 0.5 M NaCl intake on the consecutive days. IVT infusion of AII 50 ng/h for one day induced an increased urinary sodium excretion, a negative sodium balance which was not followed by an increased salt appetite. IVT infusion of AII 10 ng/h for five days caused a progressive increase in sodium excretion and salt appetite which were significant on the fourth day of infusion and both remained eight-ten times greater than control levels for three days after the cessation of infusion. Water intake was unchanged. IVT infusion of 0.3 M Na-CSF for two days reduced water and food intake, and caused a negative sodium balance on the second day of infusion which was not followed by increase in salt appetite.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
4.
Improvements in and Environmental Applications of Double-Vial Radiorespirometry for the Study of Microbial Mineralization 总被引:4,自引:3,他引:1
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Several variations in the scintillation mixture and the filter paper arrangements for double-vial radiorespirometry were compared. Improved efficiencies (44%) and shorter response times were found by adding wetting agents and methanolic NaOH to the scintillation mixture in the filter paper. The scintillation chemicals used did not contain dioxane and were found to be nontoxic to the test microbiota in this system. Covering the inner reaction vial with aluminum foil minimized the reduction in counting efficiency when testing colored or dense environmental samples. Mineralization rates were determined with 14C-labeled glucose, acetate, and glutamate and [14C]cellulose- and [14C]lignin-labeled lignocellulose for composting cow manure, forest soil, and arctic lake sediment microbiota. This improved method can be used in a variety of procedures involving the measurement of microbial mineralizations of organic compounds. Since no liquid scintillation cocktail is used for counting, the radioactive wastes are aqueous or can be incinerated, making disposal easy. 相似文献
5.
D Sarantakis W A McKinley N H Grant 《Biochemical and biophysical research communications》1973,55(2):538-542
The tetradecapeptide H-Ala-Gly-Ala-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-Ala-OH (Ala3, 14-somatostatin) an analog of the somatotropin release inhibiting factor (somatostatin SRIF) was synthesized by solid phase peptide methods. It shows somatotropin release inhibiting activity at 5 μg/ml concentration. 相似文献
6.
Conservation of an intact human immunodeficiency virus type 1 vif gene in vitro and in vivo. 总被引:5,自引:5,他引:0
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P Sova M van Ranst P Gupta R Balachandran W Chao S Itescu G McKinley D J Volsky 《Journal of virology》1995,69(4):2557-2564
Replication of vif-negative human immunodeficiency virus type 1 (HIV-1) is attenuated in certain cell lines and highly impaired in peripheral blood lymphocytes in vitro. To determine whether intact vif is positively selected during natural HIV-1 infection and to determine vif sequence variability, we employed PCR amplification, cloning, and sequencing to investigate the vif region of replicating virus in short-term-passage HIV-1 primary isolates from five asymptomatic individuals and from five persons with AIDS. A total of 46 vif clones were obtained and analyzed. Recombinant proviruses were constructed from selected vif clones from one patient and found to be fully infectious. We found that 38 of the 46 clones sequenced carried open vif reading frames and that there was a low degree of heterogeneity of vif genes within isolates from the same individual and among isolates from different donors. The cysteines previously found to be essential for vif protein function were conserved in all clones. A phylogenetic tree constructed from all available vif nucleotide sequences resulted in a virus grouping similar to those of gag and env. Direct sequencing of vif amplified by PCR from uncultured lymphocytes of 15 individuals at various stages of progression toward AIDS demonstrated vif open reading frames in 13 of 15 samples tested. There was no obvious correlation between disease status and the presence of an intact vif within this sample group at the time of sample procurement. The conservation of the vif open reading frame in vitro and in vivo and its limited variability following virus transmission in vitro are consistent with a role for vif in natural HIV-1 infection. 相似文献
7.
The effects of acid on the microbial decomposition of the dominant aquatic macrophyte (Carex sp.) in Toolik Lake, Alaska were studied in microcosms during the ice-free season of 1980. Toolik Lake is slightly buffered, deep, and very oligotrophic. Microbial activities, as determined by 14C-acetate incorporation into extractable lipids, associated with Carex litter were significantly (P < 0.01) reduced within 2 days at pHs of 3.0 and 4.0, but not 5.0, 5.5, or 6.0, as compared with ambient controls (pH 7.4). ATP levels were significantly reduced at pH 3.0, but not at the other pHs tested. After 18 days, microbial activity significantly correlated with weight loss (P < 0.05), nitrogen content (P < 0.01), and C/N ratios (P < 0.01) of the litter, but did not correlate with ATP levels. Scanning electron microscopy of the litter surface revealed that the fungi present at ambient pH did not become dominant at pHs below 5.5, diatoms were absent below pH 4.0, and bacterial numbers and extracellular slime were greatly reduced at pH 4.0 and below. Mineralization of Carex14C-lignin-labeled or 14C-cellulose-labeled lignocellulose was reduced at pH 2.0, but not at pH 4.0, 5.0, or 6.0, compared with controls (pH 7). We concluded that if the pH of the water from this slightly buffered lake was sufficiently reduced, rates of litter decomposition would be significantly reduced. 相似文献
8.
Summary Permeability properties and the effects of a changed membrane potential on Ca2+ release of sarcoplasmic reticulum vesicles of rabbit skeletal muscle were investigated by Millipore filtration. The relative permeability of sarcoplasmic reticulum to solutes determined under conditions of isotope exchange at equilibrium and/or under conditions of net flow of solute and water into the vesicles was as follows: sucrose, Ca2+, Mn2+–, choline+, Tris+–+, Na+, Li+, Cl–. Transient membrane potentials were induced by rapidly changing the ionic environment of the vesicles. Knowledge of the relative permeation rates of the above ions allowed prediction of the direction and extent of membrane polarization. Osmotic effects in the polarization measurements due to the rapid influx of solute and water into the vesicles were minimized by using media containing a fast (K+ or Cl–) and a relatively slow (gluconate– or choline+) penetrating ion.45Ca2+ efflux from vesicles derived from different parts of the sarcoplasmic reticulum structure was not appreciably changed when vesicles were made more positive inside (choline chloride potassium gluconate) or more negative inside (potassium gluconate choline chloride). These studies suggest that part or all of the ion-induced changes in sarcoplasmic reticulum membrane permeability, previously interpreted to indicate depolarization-induced Ca2+ release, may be due to osmotic effects. 相似文献
9.
Between 1971 and 1975, Fascioloides magna was found in 46 of 67 (69%) feral swine (Sus scrofa) in southern Texas. Flukes were recovered from swine in areas where F. magna commonly has been recovered from white-tailed deer and cattle. One to 12 flukes were recovered from each infected animal. Their presence was indicated by black hematin pigment on the liver and various other internal organs. Eggs were not detected in the gallbladder or feces of infected animals although mature flukes and eggs were recovered in the livers suggesting that, like cattle, feral swine can be infected but are aberrant hosts for the parasite and do not disseminate eggs. 相似文献
10.
Developmental expression of prion protein gene in brain 总被引:8,自引:0,他引:8
M P McKinley B Hay V R Lingappa I Lieberburg S B Prusiner 《Developmental biology》1987,121(1):105-110
Synthesis of the cellular isoform of the prion protein (PrPC) was found to be regulated during development of the hamster brain. PrP poly A(+) RNA was readily detectable 10 days postpartum; after 20 days of age, no change in its level could be detected through 13 months of age. Low levels of PrP poly A(+) RNA were detectable 1 day after birth. By contrast, myelin basic protein poly A(+) RNA was found at high levels in brain at 30 days of age and thereafter declined steadily. Using monospecific PrP antisera, immunoprecipitable cell-free translation products were detected at low levels 2 days after birth and increased progressively through 10 days of age. How the levels of PrP mRNA participate in brain development and function remains to be established. 相似文献