[Preliminary assessment of fine needle aspiration biopsy in the diagnosis of parotid tumors].
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Journal Article↗
Biomedical subjects
F Li
Publications and source records attributed to F Li.
Consequences of cAMP and catalytic-subunit binding on the flexibility of the A-kinase regulatory subunit.
A combination of site-directed labeling and time-resolved fluorescence anisotropy was used to further elucidate the structure and underlying dynamic features of the type I regulatory (R(I)(alpha)) subunit of the cAMP-dependent protein kinase. Specifically, the consequences of cAMP and the catalytic (C)-subunit binding on the backbone flexibility around seven sites of cysteine substitution and fluorescein maleimide labeling (Thr(6)Cys, Leu(66)Cys, Ser(75)Cys, Ser(81)Cys, Ser(99)Cys, Ser(145)Cys, and Ser(373)Cys) in the R(I)(alpha) subunit were assessed. Focusing on the fast rotational correlation time, the results indicate that most of the interdomain segment connecting the dimerization/docking (D/D) and tandem cAMP-binding domains is probably weakly associated with the latter domain. Also, this segment becomes more tightly bound to the C subunit upon holoenzyme formation. The results also suggest that there is a short 'hinge' segment (around Leu(66)Cys) that could allow the structured interdomain/cAMP-binding and D/D domains to pivot about each other. Finally, cAMP binding dramatically reduces the backbone flexibility around only the two sites of cysteine substitution in the cAMP-binding domains, suggesting a selective structural stabilization caused by cAMP and a "tight" coupling of low-nanosecond fluctuations selectively within the tandem cAMP-binding domains.
Catalysis↗
Sulfolobus shibatae CCA-adding enzyme forms a tetramer upon binding two tRNA molecules: A scrunching-shuttling model of CCA specificity.
The tRNA CCA-adding enzyme adds CCA stepwise to immature transfer RNA molecules untemplated, but with high specificity. We examined the oligomerization state of the enzyme from Sulfolobus shibatae and its binding to transfer RNA molecules, using various biophysical and biochemical methods including size exclusion chromatography, multi-angle laser light scattering, small-angle X-ray scattering, and gel electrophoresis band mobility shift assay. The 48 kDa monomer forms a stable salt- resistant dimer in solution. Further dimerization of the dimeric enzyme to form a tetramer is induced by the binding of two tRNA molecules. The formation of a tetramer with only two bound tRNA molecules leads us to suggest that one pair of active sites may be specific for adding two C bases, which results in scrunching of the primer strand. An adjacent second pair of active sites may be specific for adding A after addition of two C bases which makes the 3' terminus long enough to reach the second pair of active sites.
Animals↗
Human glioma cell BT325 expresses a proteinase that converts human plasminogen to kringle 1-5-containing fragments.
Angiostatin, a specific angiogenesis inhibitor, is an internal fragment of plasminogen, and can be generated in many systems mediated by different enzymes in vitro. The mechanism of angiostatin generation in vivo has not been well defined. Here we demonstrated that human glioma cell line BT325 can express an enzyme that can convert purified plasminogen to angiostatin-like fragments with molecular masses of 65, 60, and 58 kDa, respectively. These fragments have an identical N-terminal as KVYLS, which starts from Lys(98) of the plasminogen precusor. According to their molecular mass, the three fragments should comprise kringle domain 1 to kringle domain 5 (kringle 1-5). The proteolytic fragments obtained as above can inhibit the growth of bovine aortic endothelial (BAE) cells specifically. The proteolysis process can be completely inhibited by serine proteinase inhibitors, and partially inhibited by EDTA. The molecular weight of the peptide, which contains an enzymatic activity responsible for the proteolysis, was 13 kD determined by gel filtration and SDS-PAGE. The present data suggest that glioma cell BT325 can produce a novel proteinase to generate kringle 1-5 of plasminogen as an angiogenesis inhibitor.
Amino Acid Sequence↗
Regulation of apoptosis at cell division by p34cdc2 phosphorylation of survivin.
The interface between apoptosis (programmed cell death) and the cell cycle is essential to preserve homeostasis and genomic integrity. Here, we show that survivin, an inhibitor of apoptosis over-expressed in cancer, physically associates with the cyclin-dependent kinase p34(cdc2) on the mitotic apparatus, and is phosphorylated on Thr(34) by p34(cdc2)-cyclin B1, in vitro and in vivo. Loss of phosphorylation on Thr(34) resulted in dissociation of a survivin-caspase-9 complex on the mitotic apparatus, and caspase-9-dependent apoptosis of cells traversing mitosis. These data identify survivin as a mitotic substrate of p34(cdc2)-cyclin B1 and suggest that survivin phosphorylation on Thr(34) may be required to preserve cell viability at cell division. Manipulation of this pathway may facilitate the elimination of cancer cells at mitosis.
Amino Acid Sequence↗
Broad-spectrum cation channel inhibition by LOE 908 MS reduces infarct volume in vivo and postmortem in focal cerebral ischemia in the rat.
Cation channels conduct calcium, sodium and potassium, cations that are likely deleterious in the evolution of focal ischemic injury. We studied the effects of a novel, broad-spectrum inhibitor of several cation channels, LOE 908 MS, on acute ischemic lesion development with diffusion-weighted magnetic resonance imaging (DWI) and on cerebral perfusion with perfusion imaging (PI) in vivo and on cerebral infarct size using 2,3,5-triphenyltetrazolium chloride (TTC) staining postmortem. A total of 18 male Sprague-Dawley rats underwent 90 min of middle cerebral artery occlusion (MCAO) and were randomly and blindly assigned to either LOE 908 MS or vehicle starting 30 min after inducing focal ischemia and continuing for 4 h. Whole-brain DWI and multislice PI were done before initiation of treatment and repeated frequently for the next 3.5 h. DWI-derived lesion volume at 4 h showed a significant difference in favor of the drug treated group (P=0.03), whereas PI-derived perfusion deficit volumes did not significantly differ between the groups. The postmortem infarct volume at 24 h was significantly attenuated in the treated group in comparison to controls (P=0.0001) and neurological score was significantly better in the treated group (P<0.02). Blocking several distinct cation channels with LOE 908 MS significantly reduced infarct size and improved neurological outcome without observable adverse effects in this focal ischemia model.
Acetamides↗
Radiation tolerance of complex oxides
The radiation performance of a variety of complex oxides is predicted on the basis of a material's propensity to accommodate lattice point defects. The calculations indicate that a particular class of oxides possessing the fluorite crystal structure should accept radiation-induced defects into their lattices far more readily than a structurally similar class of oxides based on the pyrochlore crystal structure. Preliminary radiation damage experiments substantiate the prediction that fluorites are inherently more radiation resistant than pyrochlores. These results may permit the chemical durability and radiation tolerance of potential hosts for actinides and radioactive wastes to be tailored.
Journal Article↗
Targeted inhibition of wound-induced PAI-1 expression alters migration and differentiation in human epidermal keratinocytes.
In the adult epidermis, keratinocytes do not normally express the type-1 inhibitor of plasminogen activator (PAI-1). Basal epithelial cell-specific PAI-1 synthesis, however, accompanies epidermal wound repair in vivo in which PAI-1 transcripts and immunoreactive protein are confined to epithelial cells in the migrating tongue and the hyperproliferative zone. A model system using human keratinocytes (HaCaT cells) was developed to assess functional relationships between epithelial growth state transitions and PAI-1 expression. PAI-1 synthesis was maximal in low population density, exponentially growing HaCaT cultures; relative PAI-1 mRNA and protein levels progressively declined as cells attained, and were maintained in, a postconfluent condition. While the fraction of PAI-1(+) keratinocytes remained stable (at approximately 85-90% of the population) throughout the culture period, both PAI-1 mRNA abundance and mean cell-associated PAI-1 protein declined by >90% during prolonged (i.e., 8-day) growth arrest. Similar to epidermal trauma in vivo, scrape wounding of HaCaT monolayers resulted in the rapid and location-specific induction of PAI-1 protein (an increase of 11- to 16-fold relative to unwounded cultures) in cells immediately bordering the injury site. PAI-1 expression was evident in keratinocytes that comprised the opposed migrating fronts and remained elevated until wound closure. Down-regulation of PAI-1 synthesis in HaCaT cells transfected with an inducible LacSwitch-based antisense vector system markedly impaired both the rate and the extent of wound closure. All injuries created in antisense PAI-1 monolayers remained unhealed at day 8 postinjury compared to the 3-day complete repair typical of control cultures. Vector-driven modulation of PAI-1 synthesis was also associated with an increase in the percentage of suprabasal-type keratinocytes within the wound field. PAI-1 expression by migrating HaCaT cells appears necessary to maintain the basal epidermal phenotype and/or appropriate cell-to-substrate adhesion during injury repair.
Cell Differentiation↗
Interaction of albumin mRNA with proteins from rat liver with CCl(4)-induced injury.
Acute phase responses to intragastric administration of a single dose of CCl(4) were examined with albumin mRNA turnover as a marker. Based on the combination of the changes in stability of albumin mRNA and activity of transcription of its gene, the entire course of liver injury was classified into three stages, the first stage for aggravation of injury until 9 h, the second from 9 to 24 h, and the third for repair of injury or regeneration of liver after 48 h. Liver S100 fractions from normal and CCl(4)-treated rats contained, in total, 11 polypeptides cross-linked with part of albumin mRNA, although they did not appear to be specific to this mRNA. Their profiles were altered together with the changes in stability of albumin mRNA in different stages. These findings suggest that the polypeptides with distinct properties play roles in physiologically significant processes involved in utilization and turnover of albumin mRNA, apparently altering its stability.
Acute-Phase Reaction↗
A survey of acid catalysts for use in two-step, one-flask syntheses of meso-substituted porphyrinic macrocycles.
[reaction: see text] Diverse Lewis acids and Bronsted acids were examined in the two-step, one-flask synthesis of meso-tetraphenylporphyrin, N-confused tetraphenylporphyrin, and tetraphenylsapphyrin. The scope of acid catalysis was found to be very broad, with 35 of 45 acids providing TPP in yields ranging from 5% to 58%. NC-TPP was also widely observed in yields of 1-40%, and TPS was infrequently observed in yields of <1%. Additionally, conditions were found for direct preparation of magnesium TPP and copper TPP.
Acids, Noncarboxylic↗
Adsorption of Gemini and Conventional Cationic Surfactants onto Montmorillonite and the Removal of Some Pollutants by the Clay.
The adsorption of a series of gemini surfactants, [C(n)H(2n+1)N(+) (CH(3))(2)-CH(2)CH(2)](2).2Br(-), where n = 10, 12, 14, and 16, on clay (Na-montmorillonite) from their aqueous solution in 0.01 M KBr and the effect of this adsorption on the removal of 2-naphthol and 4-chlorophenol have been studied. Compared to those of conventional cationic surfactants with similar single hydrophilic and hydrophobic groups (C(n)H(2n+1)N(+)(CH(3))(3).Br(-), where n = 10, 12, 14, and 16), the molar adsorptions of the gemini and conventional surfactants are almost identical. This indicates that only one of the hydrophilic groups in the gemini molecule is adsorbed onto the clay and that the second hydrophilic is presumably oriented toward the aqueous phase, in contrast to the adsorption of the conventional surfactants, where the hydrophobic group is oriented toward the aqueous phase. Stability studies on dispersions of clay treated with the two types of surfactants confirm this. The slight increase in the moles of surfactant to values above the CEC of the clay with an increase in the carbon number of the hydrophobic chain indicates that adsorption through hydrophobic group interaction occurs in addition to the major ion exchange. Adsorption studies of the pollutants onto the clay treated by either the gemini or the conventional surfactants show that the former are both more efficient and more effective at removing the pollutants from the aqueous phase. Copyright 2000 Academic Press.
Journal Article↗
The relative abilities of TCDD and its congeners to induce oxidative stress in the hepatic and brain tissues of rats after subchronic exposure.
The abilities of single doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to induce oxidative stress in hepatic and some extra-hepatic tissues of animals, are well documented. In this study we have investigated the induction of oxidative stress in hepatic and brain tissues of rats after subchronic (13 weeks) exposure to TCDD and two of its congeners, namely 2,3,4,7,8-pentachlorodibenzofuran (PeCDF) and 3,3',4,4',5-pentachlorobiphenyl (PCB126). TCDD, PeCDF and PCB126 were administered daily to groups of rats at various doses, for 13 weeks, and biomarkers of oxidative stress, including the production of superoxide anion, lipid peroxidation and DNA-single strand breaks (SSBs), were determined in the hepatic and brain tissues at the end of the exposure period. The three congeners caused dose-dependent increases in the production of superoxide anion, lipid proxidation and DNA-SSBs, with maximal effects achieved at doses ranging between 10-100, 20-92, and 300-550 ng/kg per day for TCDD, PeCDF and PCB126, respectively. The doses that produce 50% of maximal responses by each of the xenobiotics in the hepatic and brain tissues were found to be within the ranges of 7-34, 13-32, and 137-400 ng/kg per day for TCDD, PeCDF and PCB126, respectively. The results of the study suggest that subchronic exposures to TCDD, PeCDF and PCB126 induce significant oxidative damage in the hepatic and brain tissues of rats, with more damage observed in the brain as compared to the hepatic tissues. Also, as inducers of oxidative stress in the hepatic and brain tissues, TCDD is the most potent among the three congeners and PCB126 being the least potent.
Animals↗
Angiopoietin-1 inhibits endothelial cell apoptosis via the Akt/survivin pathway.
A productive angiogenic response must couple to the survival machinery of endothelial cells to preserve the integrity of newly formed vessels. Angiopoietin-1 (Ang-1) is an endothelium-specific ligand essential for embryonic vascular stabilization, branching morphogenesis, and post-natal angiogenesis, but its contribution to endothelial cell survival has not been completely elucidated. Here we show that Ang-1 acting via the Tie 2 receptor induces phosphorylation of the survival serine-threonine kinase, Akt (or protein kinase B). This is associated with up-regulation of the apoptosis inhibitor, survivin, in endothelial cells and protection of endothelium from death-inducing stimuli. Moreover, dominant negative survivin negates the ability of Ang-1 to protect cells from undergoing apoptosis. The activation of anti-apoptotic pathways mediated by Akt and survivin in endothelial cells may contribute to Ang-1 stabilization of vascular structures during angiogenesis, in vivo.
Angiopoietin-1↗
Participation in the cooperative family registry for breast cancer studies: issues of informed consent.
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Australia↗
Inhibition of chronic rejection of aortic allografts by dietary glycine.
BACKGROUND: Chronic rejection is influenced by a variety of risk factors, including histoincompatibility and ischemia. Glycine, a cytoprotective agent, has been shown to protect against ischemia-reperfusion injury in the liver, inactivate hepatic resident macrophages, minimize cyclosporin A-induced nephrotoxicity, and exhibit immunosuppressive properties in vitro. The aim of this study was to investigate whether dietary glycine could reduce development of chronic rejection. METHODS: Lewis recipients of Fisher-344 abdominal aortic allografts received diets that contained either 5% glycine plus 15% casein or 20% casein as control for 10 weeks. Vascular lesions of aortic isografts and allografts were evaluated quantitatively with image analysis and cell counting. RESULTS: No significant vascular changes were observed in isografts (mean medial areas of 3.3 +/- 0.3x0(5) microm2). However, dramatic intimal thickening (neointimal area 2.1+/-0.3) and medial thinning (1.5+/-0.3) were observed in allografts from rats fed control diet. In contrast, glycine significantly reduced the neointima by 45% (1.2+/-0.3) and protected the media (3.5+/-0.2). This led to intima to media area ratios almost twice as large in the control group as in glycine-fed rats (2.2+/-0.4 vs. 1.1+/-0.3, P<0.05). Moreover, infiltrating leukocytes, especially macrophages, were reduced significantly in the adventitia by glycine. In addition, glycine inhibited proliferation and migration of rat aortic smooth muscle cells in culture by 45 and 60%, respectively. CONCLUSION: These results indicate that dietary glycine minimizes histopathological changes of chronic rejection by reducing the immune response and, in part, by minimizing proliferation and migration of smooth muscle cells.
Animals↗
Cell division regulation by BIR1, a member of the inhibitor of apoptosis family in yeast.
The inhibitor of apoptosis (IAP) gene family comprises molecules that block the activity of pro-apoptotic caspase proteases. Paradoxically, yeasts contain IAP proteins but no caspases and no apoptotic program. To determine the function of these proteins in vivo, we disrupted the BIR1 gene, encoding the only known IAP in yeast Saccharomyces cerevisiae. Sporulation of heterozygous diploids yielded no viable mutant haploids, indicating that BIR1 is an essential gene. By flow cytometry, some heterozygous mutants were polyploid accumulating >4 N DNA content. These cells exhibited a 20-40% reduction in growth rate, which was rescued by plasmid-borne over-expression of BIR1 but not by its human counterpart, survivin. Deletion analysis revealed that the N-terminal domain of Bir1, containing the conserved baculovirus IAP repeat, was able to partially complement the cell growth defect caused by BIR1 deletion. Moreover, the full-length and truncated forms of Bir1 accelerated cell division in wild-type cells. Finally, BIR1 heterozygous mutants exhibited grossly altered cell morphology with misshapen or abnormally long buds connected to an unusually large mother cell. These findings identify a novel function of IAP proteins in the pleiotropic control of cell division, in addition to their role in the suppression of apoptosis.
Apoptosis↗
Temporal evolution of ischemic injury evaluated with diffusion-, perfusion-, and T2-weighted MRI.
OBJECTIVE: Ischemic lesions seen on diffusion-weighted imaging (DWI) are reversible if reperfusion is performed within minutes after the onset of ischemia. This study was designed to determine whether acute reversibility of DWI abnormalities is transient following brief temporary focal brain ischemia and to characterize the temporal evolution of in vivo ischemic lesions. METHODS: Eight rats were subjected to 30 minutes of temporary middle cerebral artery occlusion and underwent diffusion-, perfusion-, and T2-weighted MRI during occlusion; immediately after reperfusion; 30, 60, and 90 minutes after reperfusion; and 12, 24, 48, and 72 hours after reperfusion. Average apparent diffusion coefficient (ADCav) values and the cerebral blood flow index (CBFi) ratio were calculated in both the lateral caudoputamen and overlying cortex at each time point. The size of the in vivo ischemic abnormalities was calculated from the ADCav and the T2 maps. Postmortem triphenyltetrazolium chloride (TTC) staining was used to verify ischemic injury. RESULTS: Both the CBFi ratio and ADCav values declined significantly in the two regions during occlusion. The CBFi ratio recovered immediately after reperfusion and remained unchanged over 72 hours. However, ADCav values returned to normal at 60 to 90 minutes and secondarily decreased at 12 hours after reperfusion as compared with those in the contralateral hemisphere. The extent of the in vivo ischemic lesions maximized at 48 hours and was highly correlated with TTC-derived lesion size. CONCLUSIONS: Acute recovery of initial ADCav-defined lesions after reperfusion is transient, and secondary ADCav-defined lesions develop in a slow and delayed fashion.
Animals↗
Recombinant subunit ORF2.1 antigen and induction of antibody against immunodominant epitopes in the hepatitis E virus capsid protein.
A recombinant subunit antigen (ORF2.1), representing the carboxy-terminal 267 amino acids of the 660-amino-acid hepatitis E virus (HEV) capsid protein, was expressed in Escherichia coli and used for the immunisation of rats. Purified antigen formulated with either Aluminium Hydroxide Gel Adjuvant (Alum) or Titermax gave high and equivalent levels of antibody after three doses. Responses to two doses of 15, 75, or 150 microg antigen, formulated with Alum and given at 0 and 4 weeks, were also equivalent by 17 weeks after immunisation. Rats initially developed antibody to a wide range of linear epitopes in the ORF2.1 region, but by 27 weeks the predominant response detected by Western immunoblotting was restricted to the conformational epitope unique to ORF2.1 [Li et al. (1997) Journal of Medical Virology 52:289-300], a pattern that was also observed when comparing acute-phase patient serum samples with serum samples from convalescing patients. Antibody from immunised rats blocked the majority of patients' serum reactivity in enzyme-linked immunosorbent assay against both ORF2.1 (57-92% inhibition) and virus-like particles of HEV produced using the baculovirus system (74-97% inhibition). Together, these results suggest that the ORF2.1 subunit vaccine induces an antibody response against immunodominant, conformational epitopes in the viral capsid, which largely mimics that seen in convalescent patients, who are presumed to be immune to HEV infection.
Adjuvants, Immunologic↗