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At least 1,081 records · Page 60Linked to original sources
PubMed1995

Didemnin B induces cell death by apoptosis: the fastest induction of apoptosis ever described.

Didemnin B, a cyclic N-methylated peptolide induces apoptosis in human HL-60 cells. When incubated with 1 microM didemnin B, unsynchronized HL-60 cultures undergo apoptosis to 100% within 140 minutes. Apoptosis has been assessed by the typical apoptotic morphology, the presence of double-stranded DNA fragments within the cytosol and the generation of DNA ladders. None of these characteristics of apoptosis are seen when HL-60 cells are pretreated with 1mM Zn2+ immediately before treatment with didemnin B.

Antineoplastic Agents↗
PubMed1996

Evidence for a TSH-controlled ectophosphotyrosine phosphatase in pig thyroid cultured cells.

In porcine thyroid cells in primary culture, thyrotropin (TSH) provokes important morphological changes associated with specific thyroidal function recovery. TSH influences cell-cell interactions and follicular morphogenesis. In the present report, we identify an ectotyrosine phosphatase activity controlled by chronic treatment with TSH. Specific substrates and various inhibitors allowed us to characterize this activity. This tyrosine phosphatase is located at the outer surface of thyroid cells as demonstrated by phosphotyrosylhistone hydrolysis. Control of cell viability and trypsin treatment confirm this extracellular localization. These data show that TSH activates ectotyrosine phosphatase activity, suggesting a role for surface protein phosphorylation in regulating specific functions of porcine thyroid cells in suspension.

Animals↗
PubMed1997

Characteristics of glutathione-capped ZnS nanocrystallites.

The titration of increasing equivalents of inorganic sulfide into preformed Zn-glutathione led to the appearance of UV/VIS spectral features attributable to ZnS nanocrystallites. Glutathione-ZnS complexes upon irradiation caused reduction of methylviologen confirming their semiconductor properties. Size-fractionation of glutathione-ZnS samples on a gel filtration column showed the formation of a range of complexes whose spectral properties were correlated with the sulfide content. The stoichiometry of Zn:glutathione increased from 1:2 to a maximum of about 7:1 as the sulfide/Zn ratios increase from 0 to approximately 1.0 in Zn-glutathione complex indicating up to 14-fold increase in the Zn-binding capacity of glutathione upon sulfide incorporation. Spectral characteristics of GSH-capped ZnS nanocrystallites were significantly influenced by pH and by the stoichiometry of Zn, sulfide and glutathione in the complex. Samples containing least glutathione and highest sulfide showed maximal luminescence at pH 6, whereas those with higher glutathione and lower sulfide content showed maximal luminescence at pH 11.

Chromatography, Gel↗
PubMed1999

A novel endopeptidase with a strict specificity for threonine residues at the P1' position.

An endopeptidase was purified from Archachatina ventricosa by chromatography on columns of gel filtration, DEAE-Sepharose and phenyl-Sepharose. The preparation was shown to be homogeneous by polyacrylamide gel electrophoresis and capillary electrophoresis. The purified enzyme displayed two protein bands on SDS-polyacrylamide gel electrophoresis with estimated molecular weights of 90,000 and 121,000. The protease exhibited maximum proteolytic activity at 55 degrees C and at pH 8.0, but it retained more than 85% of its activity in the pH range 7.5 to 8.5. It was completely inactivated by the chelating agents EDTA and 1,10-phenanthroline which are metalloprotease inhibitors. Studies on substrate specificity showed that only the amide bonds of peptide substrates having a threonine residue at the P1' position were hydrolyzed by the purified protease. This endopeptidase constitutes a novel tool for the study of proteins in view of its narrow and unique substrate specificity.

Animals↗
PubMed2000

High levels of zinc ions induce loss of mitochondrial potential and degradation of antiapoptotic Bcl-2 protein in in vitro cultivated human prostate epithelial cells.

Prostate epithelial cells contain the highest levels of zinc among all organs and tissues in the human body. Zinc is accumulated primarily in the mitochondria, where it is responsible for inhibition of mitochondrial aconitase activity, thereby increasing citrate production. The present study was designed to clarify the role of zinc for human prostate epithelial cell growth and apoptosis. Apoptosis of in vitro cultivated human prostate epithelial cells exposed to ZnCl(2) was analyzed by determination of phospholipid membrane asymmetry, nuclear fragmentation, DNA strand breaks, changes of mitochondrial potential and cellular pro/antiapoptotic proteins. Zinc induced apoptosis without involvement of p53 by decreasing mitochondrial transmembrane potential (DeltaPsi(m)) and Bcl-2 protein levels in proliferating epithelial cells. Thus, the high local concentrations of zinc ions in the prostatic lumen seem to be necessary to regulate proliferative activities and to enforce epithelial differentiation processes.

Actins↗
PubMed2001

Spatial organization of Ca(2+) entry and exocytosis in mouse pancreatic beta-cells.

Secretion from single pancreatic beta-cells was imaged using a novel technique in which Zn(2+), costored in secretory granules with insulin, was detected by confocal fluorescence microscopy as it was released from the cells. Using this technique, it was observed that secretion from beta-cells was limited to an active region that comprised approximately 50% of the cell perimeter. Using ratiometric imaging with indo-1, localized increases in intracellular Ca(2+) concentration ([Ca(2+)](i)) evoked by membrane depolarization were also observed. Using sequential measurements of secretion and [Ca(2+)](i) at single cells, colocalization of exocytotic release sites and Ca(2+) entry was observed when cells were stimulated by glucose or K(+). Treatment of cells with the Ca(2+) ionophore 4-Br-A23187 induced large Ca(2+) influx around the entire cell circumference. Despite the nonlocalized increase in [Ca(2+)](i), secretion evoked by 4-Br-A23187 was still localized to the same region as that evoked by secretagogues such as glucose. It is concluded that Ca(2+) channels activated by depolarization are localized to specific membrane domains where exocytotic release also occurs; however, localized secretion is not exclusively regulated by localized increases in [Ca(2+)](i), but instead involves spatial localization of other components of the exocytotic machinery.

Animals↗
PubMed1998

Oncoprotein v-Myb and retinoic acid receptor alpha are mutual antagonists.

The process of hematopoiesis is critically dependent on correct interactions of multiple regulatory molecules and transcription factors. We have studied the interactions of the v-Myb and retinoic acid receptor proteins which have opposing effects on hematopoiesis. While v-myb acts as a transforming oncogene preventing differentiation of monoblasts to macrophages, RAR alpha functions as an anti-oncogene arresting the growth of v-myb-transformed cells and allowing their final myeloid differentiation steps to occur. We found that the retinoic acid receptor alpha inhibits v-Myb transformation by suppressing the expression of v-Myb target genes typified by the mim-1 gene. Conversely, v-Myb protein interferes with RAR alpha transactivation function as well as with retinoic acid-induced apoptosis of HL-60 cells. These results demonstrate that retinoic acid receptor and v-Myb proteins act in antagonistic ways and reciprocally modify each other's functions.

Acetyltransferases↗
PubMed1997

Differential role of calcium in tumour necrosis factor-mediated apoptosis and secretion of granulocyte-macrophage colony-stimulating factor in a T cell hybridoma.

The authors investigated the dependence on extracellular and intracellular free Ca2+ in the induction of apoptosis and secretion of granulocyte-macrophage colony-stimulating factor (GM-CSF) by tumour necrosis factor (TNF) in a rat/mouse T cell hybridoma PC60 R55/R75, using the Ca2+ chelators EGTA and BAPTA/AM, respectively. TNF-induced apoptosis still occurred in the absence of free Ca2+, while GM-CSF production required the continuous presence of Ca2+. The latter was also true for GM-CSF production driven by interleukin 1 (IL-1). The dependence on Ca2+ in the induction of GM-CSF, but not of apoptosis, was further confirmed by the inhibition of TNF- or IL-1-induced cytokine production by cyclosporin A or FK506, drugs that block the Ca2+/calmodulin-dependent protein Ser/Thr phosphatase calcineurin. This differential requirement for Ca2+ illustrates the partial functional redundancy between TNF and IL-1, showing the activation of cytokine gene expression through a Ca(2+)-dependent activation of calcineurin, and a Ca(2+)-independent activation of apoptosis, exerted solely by TNF.

Animals↗
PubMed1997

Neuropsychiatric syndromes and occupational exposure to zinc phosphide in Egypt.

Eighty-six workers exposed to zinc phosphide (Zn3P2) pesticide were studied for evidence of neuropsychiatric manifestations. They were evaluated clinically, by electroencephalography (EEG), and, in some cases, by electromyography (EMG). All were males (mean age, 35.8 years; mean duration of exposure to zinc phosphide, 11.3 years). Most presented with one (or more) neuropsychiatric symptom(s), including fear of poisoning, anxiety, impotence, and easy fatigue. About half showed evidence of neuropsychiatric signs, including hyperreflexia, polyneuropathy, lumber radiculopathy, and cervical myelopathy, as well as anxious mood, impaired attention, and psychomotor stimulation. EEG recordings showed abnormal findings in 17.4% of the subjects. The mean age in that group was 39.1 years; mean duration of exposure to Zn3P2 was 15.1 years. EMG studies showed evidence of partial denervation of the anterior tibial group of muscles and flexor digiti minimi in 2 of the 30 workers (6.7%) who underwent EMG examination. Serum levels of zinc (Zn) and cadmium (Ca) were significantly higher in exposed workers than in controls (P < 0.005). Serum copper (Cu), iron (Fe), phosphorus (P), and magnesium (Mg) were significantly lower in exposed workers than in controls. Electrophoretic pattern of globulin showed that gammaglobulin fraction was significantly increased (P < 0.005); alpha2 and beta-globulin were decreased (P < 0.005) in exposed workers. Lipoprotein pattern showed that the total lipids, B-lipoprotein, and B/alpha ratio were significantly increased (P < 0.005) in exposed workers; the alpha1 lipoprotein was decreased. Triglycerides and cholesterol were significantly increased (P < 0.001), and phospholipids and phospholipid/cholesterol ratio were significantly decreased (P < 0.005) in exposed workers compared to controls. The study findings indicated that exposure to Zn3P2 not only caused mild acute and subacute liver cell damage, but also affected renal function and perhaps B-cells of the pancreas. A total of 68.6% of the exposed workers had chest symptoms; only 24.4% presented with chest or cardiac signs. Ventilatory functions were abnormal in 70% of the exposed workers; abnormal ECG findings were present in 12.8%.

Adult↗
PubMed2000

Detection of apoptosis induced by topoisomerase inhibitors and serum deprivation in syrian hamster embryo cells.

The sensitivity of normal diploid Syrian hamster embryo (SHE) cells to apoptosis was tested after treatment with the topoisomerase inhibitors camptothecin and etoposide and after serum withdrawal. Programmed cell death (PCD) was identified through morphological, biochemical, and molecular changes and compared with that of HL60 cell line. The results showed that topoisomerase inhibitors, which were shown to be potent PCD inducers in the HL60 cell line, induced a weaker apoptotic response in SHE cells than after growth factor deprivation. In addition, serum-free medium, which rapidly induced apoptosis in SHE cells, did not affect the HL60 cell line. In both cell types, PCD was expressed by condensed chromatin, fragmented nuclei, and DNA laddering on electrophoretic gels, an indisputable sign of apoptosis. In apoptotic HL60 cells, the cleavage of 113-kDa poly(ADP-ribose)polymerase (PARP) resulted in the so-called apoptotic 89-kDa fragment and was associated with increased caspase-3 activity. In apoptotic SHE cells, PARP degraded early but the degradation profile was not characterized by the appearance of an 89-kDa fragment. Moreover, no activation of caspase-3 was noted. ZnCl(2), which is known to prevent protease activity responsible for apoptosis features, inhibited PARP cleavage and nuclear modifications induced by apoptotic stimuli in both cell types, but with a higher sensitivity in SHE cells. Apoptosis induced by serum deprivation was linked with c-myc negative regulation in SHE cells, but not with p53 protein accumulation, while topoisomerase inhibitors led to p53 stabilization without any change in c-myc expression. Serum-free medium and topoisomerase inhibitors did not modify c-myc expression in the HL60 cell line. The overall results demonstrated that apoptosis, which is a carefully regulated process of cell death, may proceed through mechanisms varying according to cell type or apoptosis inducer. In addition, markers which are generally considered hallmarks of apoptosis may fail to appear in some cell types.

Animals↗
PubMed2002

Surface chemical studies on sphalerite and galena using extracellular polysaccharides isolated from Bacillus polymyxa.

Adsorption, electrokinetic, microflotation, and flocculation studies have been carried out on sphalerite and galena minerals using extracellular polysaccharides (ECP) isolated from Bacillus polymyxa. The adsorption density of ECP onto galena is found to be higher than that onto sphalerite. The adsorption of ECP onto sphalerite is found to increase from pH 3 to about pH 7, where a maximum is attained, and thereafter continuously decreases. With respect to galena, the adsorption density of ECP steadily increases with increased pH. The addition of ECP correspondingly reduces the negative electrophoretic mobilities of sphalerite and galena in absolute magnitude without shifting their isoelectric points. However, the magnitude of the reduction in the electrophoretic mobility values is found to be greater for galena compared to that for sphalerite. Microflotation tests show that galena is depressed while sphalerite is floated using ECP in the entire pH range investigated. Selective flotation tests on a synthetic mixture of galena and sphalerite corroborate that sphalerite could be floated from galena at pH 9-9.5 using ECP as a depressant for galena. Flocculation tests reveal that in the pH range 9-11, sphalerite is dispersed and galena is flocculated in the presence of ECP. Dissolution tests indicate release of the lattice metal ions from galena and sphalerite, while co-precipitation tests confirm chemical interaction between lead or zinc ions and ECP. Fourier transform infrared spectroscopic studies provide evidence in support of hydrogen bonding and chemical interaction for the adsorption of ECP onto galena/sphalerite surfaces.

Bacillus↗
PubMed1997

Conformational flexibility and polymerization of vesicular stomatitis virus matrix protein.

The matrix protein of vesicular stomatitis virus (VSV) plays a pivotal role in viral assembly. We previously demonstrated the ability of M protein to self-associate at low salt concentrations. Now, we show the ability of M protein to polymerize in the presence of ZnCl2 in a nucleation-dependent manner. Analysis of kinetics revealed that the nuclei are probably made of three or four molecules of M. These results are consistent with the idea that in vitro self association of M protein is not due to amorphous aggregation but rather reflects an intrinsic ability of M to polymerize. Using attenuated total reflectance Fourier transform infrared spectroscopy, we showed that M polymerization is associated with an increase in the beta-sheet content of the protein. We propose a model explaining both the apparent M protein solubility in infected cells and how M polymerization could promote viral assembly. Data available for other negative strand viruses suggest that M polymerization may be the general basis of viral assembly.

Chlorides↗
PubMed1993

Characterization of non-opioid [3H]dynorphin A-(1-13) binding sites in the rat heart.

The binding characteristics of [3H]Dynorphin A-(1-13) ([3H]Dyn A-(1-13) were examined in membrane preparations of rat heart. Saturation binding studies with increasing concentrations between 2.5 and 500 nM indicated that [3H]Dyn A-(1-13) binds to a single population of sites with a Kd of 285 nM and a Bmax of 215 pmol/mg protein. [3H]Dyn A-(1-13) binding is sensitive to trypsin treatment and it is inhibited by Zn2+ and Mg2+ with IC50 values of 159 and 310 microM, respectively. Dyn A and related peptides competes with the binding of [3H]Dyn A-(1-13) with the following order of potency: Dyn A-(1-13) > Dyn A > Dyn B > alpha-neo-endorphin > Dyn A-(1-8). The non-opioid peptides Dyn A-(2-13), Dyn A-(3-13) and Dyn A-(5-13) are as potent (Ki of 0.35, 0.44 and 0.59 microM, respectively) as Dyn A-(1-13) (Ki of 0.36 microM) in inhibiting [3H]Dyn A-(1-13) binding while Leu-enkephalin (Leu-Enk) exhibits no inhibitory effect at 100 microM. Selective ligands for kappa (kappa: U-50,488H, U-69,593), mu (mu: [D-Ala2, MePhe4, Glyol5]Enk) and delta (delta: [D-Ser2, Thr6]Leu-Enk) opioid receptors as well as for phencyclidine (PCP: MK-801, TCP) and sigma (sigma: (+)-SKF-10047, DTG, 3(+)-PPP) receptors show little or no inhibition of [3H]Dyn A-(1-13) binding at 100 microM. These results indicate that the heart contains a low affinity high capacity binding site for Dyn A and related peptides, distinct from opioid, PCP and sigma receptors.

Animals↗
PubMed1977

Thiolation and disulphide cross-linking of insulin to form macromolecules of potential therapeutic value.

Macromolecules have been prepared containing native insulin carried by a modified insulin skeleton made by partially thiolating the insulin hexamer and forming intermolecular cross-links through disulphide bridges. Oxidation of partially thiolated insulin (0.5-0.7 SH group/mole), formed by reacting insulin with AHTL, with, (a) potassium ferricyanide, (b) Cu++-oxygen gave water soluble macromolecules containing 20-26 and 410-708 monomer units respectively which had rod-random coil shape (light scattering). The larger molecules formed by (b) contained 8g-atom CU++/hexamer unit and insulin. The insulin was firmly bound within the marcomolecules and was probably bound within an insulin-modified insulin hexamer through coordination to copper.

Chemical Phenomena↗
PubMed1987

Luminescence emission from the Cu(I)-thiolate complex in metallothioneins.

The luminescence emission properties of Cu-metallothioneins (from Neurospora crassa, Agaricus bisporus and livers of Bedlington terriers affected by copper toxicosis) as well as of (Cu,Zn)-metallothionein from bovine fetal liver are reported. Upon excitation in the U.V., these proteins emit a largely red-shifted luminescence with a maximum at 565 nm attributable to the Cu(I)-thiolate chromophores of the proteins. Differences in the shapes of the spectra and the emission intensity are observed with (Cu,Zn)-metallothionein probably due to the influence of Zn ions or to a different coordination geometry of the Cu ions. The emissive properties of the Cu(I)-thiolate chromophore are compared with those of metallo-organic Cu(I)-mercaptide complexes.

Agaricus↗
PubMed1991

Optimization of growth conditions for the production of proteolytically-sensitive proteins in the periplasmic space of Escherichia coli.

The expression of many secreted recombinant proteins in Gram-negative bacteria is limited by degradation in the periplasmic space. We have previously shown that the production of protein A-beta-lactamase, a secreted fusion protein highly sensitive to proteolysis in Escherichia coli, can be increased in mutant strains deficient in up to three cell-envelope-associated proteolytic activities. In this work we investigated the effect of fermentation conditions on suppressing any residual proteolytic activity in various protease-deficient strains. Optimal production of the fusion protein was observed in cells grown under mildly acidic conditions (5.5 less than or equal to pH less than or equal to 6.0) and a low temperatures. These conditions were shown to specifically decrease the rate of proteolysis. In addition, a further increase in production was observed in cultures supplemented with 0.5 to 0.75 mM zinc chloride. This may relate to the inhibition of a cell envelope protease by Zn2+ ions.

Bacterial Proteins↗