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Biomedical subjects

Y Fang

Publications and source records attributed to Y Fang.

At least 109 records · Page 6Linked to original sources
PubMed1999

Cationic silanes stabilize intermediates in DNA condensation.

In vitro condensation of DNA has been widely studied to gain insight into the mechanisms of DNA compaction in biological systems such as chromosomes and phage heads and has been used to produce nanostructured particles with novel material and functional properties. Here we report on the condensation of DNA in aqueous solutions by cationic silanes, which combine the condensing properties of polyamines with the cross-linking chemistry of silanes. DNA can be reversibly condensed into classical toroidal and rod-shaped structures with these agents. At low silane concentrations DNA forms a variety of looped structures with well-defined characteristics, including flower- and sausage-shaped forms. These structures suggest that at low silane concentrations a DNA-DNA contact in which the strands are at very large angles to each other is stabilized. Changes in these structures observed as a function of silane concentration suggest possible pathways for the formation of toroids and rods.

Cations↗
PubMed1999

Characterization of NFkappaB activation by detection of green fluorescent protein-tagged IkappaB degradation in living cells.

Activation of the transcription factor NFkappaB requires rapid degradation of its inhibitor, IkappaBalpha. To facilitate the study of IkappaBalpha degradation, we fused IkappaBalpha protein to enhanced green fluorescent protein to construct IkappaBalpha-enhanced green fluorescent protein (IG). We demonstrated by both flow cytometry and Western blot analysis that the half-life of IG in the presence of human tumor necrosis factor (TNF) alpha is approximately 5 min, which is similar to the half-life of native IkappaBalpha. The degradation coincided with NFkappaB translocation from the cytoplasm to the nucleus and NFkappaB-mediated induction of transcription. Phorbol 12-myristate 13-acetate (PMA), but not forskolin, also induces degradation of IG fusion protein. The half-life of IG in the presence of PMA is approximately 15 min, longer than when induced with TNFalpha. Co-treatment with TNFalpha and PMA did not result in a synergistic effect on IG degradation, although they stimulate different kinases in two different signaling pathways. Degradation of IG was inhibited by mutations at serine residues 32 and 36, which are the target sites of the phosphorylation modification that initiates degradation of IkappaBalpha. We also demonstrated that basal degradation of IG in the presence of cycloheximide is inhibited by such mutations, suggesting that basal degradation of IkappaBalpha also requires phosphorylation as the signal for degradation. Finally, we showed that the rate of TNFalpha-induced degradation of IG remains almost constant throughout the cell cycle, except at the mitotic phase, in which IG degrades more slowly.

DNA-Binding Proteins↗
PubMed1999

Controlled Site-Directed Assembly of Antibodies by Their Oligosaccharide Moieties onto APTES Derivatized Surfaces.

A convenient and efficient method for the site-directed incorporation of aldehydes generated on the oligosaccharide moieties at the C-terminal of immunoglobulin (IgG) using NaIO4 oxidation reaction is explored as a means of ensuring controlled assembly of IgG antibodies onto aminopropyltriethoxylsilane (APTES) derivatized silicon wafer surfaces. The orientation and antigen binding capacity (AgBC) of site-directly assembled IgG antibodies on derivatized surfaces were investigated using atomic force microscopy (AFM) and enzyme immunoassay (EIA), respectively. A major difference in preferential orientation is observed when the incubation of derivatized surfaces with oxidized IgG molecules is compared in two different kinds of buffer solutions. We obtained the stable and homogeneous IgG layer without loss of the AgBC on the APTES derivatized surface using the controlled incubation condition. Copyright 1999 Academic Press.

Journal Article↗
PubMed1999

Ethanol-induced structural transitions of DNA on mica.

The effect of ethanol on the structure of DNA confined to mica in the presence of Mg2+was examined by varying the ethanol concentration and imaging the DNA by atomic force microscopy. Contour length measurements of the DNA show a transition from all-B-form at 0% ethanol to all-A-form at >25% ethanol. At intermediate ethanol concentrations, contour lengths suggest that individual molecules of air-dried DNA are trapped with mixed compositions of A-form and B-form. The relative composition depends on the ethanol concentration. Fitting the length distributions at intermediate ethanol concentrations to a simple binomial model results in an upper bound estimate for the A-form and B-form domains of approximately 54 bp in the individual molecules. In addition to length changes, the apparent persistence length of DNA decreases with increasing ethanol concentration. At high concentrations of ethanol (>20%), DNA formed several higher order structures, including flower shaped condensates and toroids.

Aluminum Silicates↗
PubMed1999

Heptameric structures of two alpha-hemolysin mutants imaged with in situ atomic force microscopy.

Atomic force microscopy has been used to study self-assembled structures of two alpha-hemolysin mutants. For a mutant (alphaHL-H5) that was locked into the prepore state on fluid phase egg-PC membranes, we visualized, for the first time, heptameric prepores and showed that the 7-fold axis in the prepore lies perpendicular to the membrane surface. For another mutant (TCM) with the transmembrane domain, the self-assembled oligomer that assumes the conformation of the fully assembled pore is also a heptamer. These results show that heptamers are the preferred oligomerization state of alpha-hemolysin.

Bacterial Toxins↗
PubMed1999

The FKBP12-rapamycin-binding domain is required for FKBP12-rapamycin-associated protein kinase activity and G1 progression.

The immunosuppressant rapamycin, in complex with its cellular receptor FKBP12, targets the cellular protein FKBP12-rapamycin-associated protein/mammalian target of rapamycin/rapamycin and FKBP12 target 1 (FRAP/mTOR/RAFT1) and inhibits/delays G1 cell cycle progression in mammalian cells. As a member of the novel phosphatidylinositol kinase-related kinase family, FRAP's kinase activity is essential for its signaling function. The FKBP12-rapamycin binding (FRB) domain in FRAP is also speculated to play an important role in FRAP function and signaling. However, the biochemical and physiological functions of FRB, as well as the mechanism for rapamycin inhibition, have been unclear. The present study focuses on investigation of FRB's role and the functional relationship between FRB domain and kinase domain in FRAP. Microinjection of purified FRB protein into human osteosarcoma MG63 cells results in a drastic blockage of the G1 to S cell cycle progression; such a dominant negative effect is reversed by a point mutation (Trp2027 --> Phe). The same mutation also abolishes kinase activity of FRAP without affecting ATP binding, and truncation studies suggest that upstream sequences including FRB are required for kinase activity in vitro. Given these data, we propose a model for FRAP function, in which the FRB domain is required for activation of the kinase domain, possibly through the interaction with an upstream activator. In addition, our observations provide direct evidence linking FRAP function to G1 cell cycle progression.

Amino Acid Sequence↗
PubMed1999

Synthesis of novel chelating agents and their effect on cadmium decorporation.

A series of novel dithiocarbamates, disodium salts of N-glucamyl-N-dithiocarboxyl-amino acids, were synthesized, and their usefullness as an antagonist of cadmium intoxication was investigated. These chelating agents were found to be effective in both acute and repeated exposure cadmium poisoning. The results showed that the cadmium mobilizing properties of disodium N-(2,3,4,5, 6-pentahydroxylhexyl)-N-dithiocarbamate-L-threoninate and disodium N-(2,3,4,5,6-pentahydroxylhexyl)-N-dithiocarbamate-L-cysteinate are clearly superior to those of sodium N-(4-methoxybenzyl)-D-glucamine-N-carbodithioate (MeOBGDTC) revealed in the experiments described here. The toxicity of these novel compounds is modest, and their effect on the concentrations of essential metal ions in the renal cortex is quite small in comparison with that of a group treated with cadmium only. The new dithiocarbomates were identified by MS, rather than by elemental analysis, as they were extremely hygroscopic.

Animals↗
PubMed1999

Hierarchy in the expression of the locus of enterocyte effacement genes of enteropathogenic Escherichia coli.

Enteropathogenic Escherichia coli (EPEC) elicit changes in host cell morphology and cause actin rearrangement, a phenotype that has commonly been referred to as attaching/effacing (AE) lesions. The ability of EPEC to induce AE lesions is dependent upon a type III protein secretion/translocation system that is encoded by genes clustered in a 35.6 kb DNA segment, named the locus of enterocyte effacement (LEE). We used transcriptional fusions between the green fluorescent protein (gfp) reporter gene and LEE genes rorf2, orf3, orf5, escJ, escV and eae, together with immunoblot analysis with antibodies against Tir, intimin, EspB and EspF, to analyse the genetic regulation of the LEE. The expression of all these LEE genes was strictly dependent upon the presence of a functional integration host factor (IHF). IHF binds specifically upstream from the ler (orf1) promoter and appears to activate expression of ler, orf3, orf5 and rorf2 directly. The ler-encoded Ler protein was involved in activating the expression of escJ, escV, tir, eae, espB and espF. Expression of both IHF and Ler was needed to elicit actin rearrangement associated with AE lesions. In conclusion, IHF directly activates the expression of the ler and rorf2 transcriptional units, and Ler in turn mediates the expression of the other LEE genes.

Actins↗
PubMed1999

A glucocorticoid-inducible transcription system causes severe growth defects in Arabidopsis and induces defense-related genes.

A glucocorticoid-inducible transcription system was employed to control the expression of AtEBP, an Arabidopsis transcription factor. A number of the transgenic AtEBP lines had developmental and growth defects when grown on dexamethasone (DEX), a strong synthetic glucocorticoid. However, these growth defects were not confined to the AtEBP lines but were observed with other transgenic lines that were generated using the same system, including empty vector lines. In about 25% of the AtEBP or empty vector transgenic lines, these growth defects were severe and in some cases led to death. As AtEBP has been linked to the plant defense response, the expression of specific defense-related genes, including a number of pathogenesis-related (PR) genes was also examined. PDF1.2, a plant defensin gene, was strongly induced in all transgenic lines examined following treatment with DEX, including empty vector lines that did not show any observable DEX-induced growth defect. PR-5 was induced to a lesser extent in all the lines, while the expression of PR-1, PR-2 and phenylalanine ammonia-lyase 3 (PAL3) did not change significantly. While the induction of the AtEBP transgene and PDF1.2 had similar DEX concentration requirements, the kinetics of induction differed significantly, with the AtEBP transgene being induced within 1 h and PDF1.2 only being induced between 24 and 48 h. Although the molecular mechanisms underlying the growth defects and changes in gene expression remain to be determined, these changes appear to result from the glucocorticoid-inducible system itself, and may therefore limit the usefulness of this system for controlling gene expression in Arabidopsis.

Arabidopsis↗
PubMed1999

Secretion of phospholipid transfer protein by human hepatoma cell line, Hep G2, is enhanced by sodium butyrate.

Hep G2 cells were used to study the synthesis and secretion of phospholipid transfer protein (PLTP). Upon incubation of the cells at confluence with serum-free Dulbecco's modified Eagle's medium (DMEM), phosphatidylcholine (PC) transfer activity was found to accumulate in the culture media. The PC transfer activity in the media was effectively inhibited by rabbit anti-human PLTP immunoglobulin (Ig)G, thus indicating that the PC transfer activity was due to secreted PLTP. The molecular weight of Hep G2 PLTP was approximately 78 kDa by Western blot analysis, in agreement with the molecular weight obtained for purified human plasma PLTP. The PLTP secreted by Hep G2 also possessed an HDL conversion activity similar to that of human plasma PLTP. The addition of butyrate to the cell culture media resulted in a marked increase in the secretion of PLTP. After 24 h incubation with 4 mmol/L sodium butyrate, a more than twofold increase (P < 0.01) of PC transfer activity in the cell-conditioned media was obtained. The dose-dependent increase in the PC transfer activity in the media upon butyrate treatment was well correlated (r = 0.80, P < 0.01) with that of PLTP mass as determined by immuno-slot blot analysis of cell-conditioned media. The increased secretion of PLTP by Hep G2 treated with sodium butyrate was accompanied by a greater increase in the level of PLTP mRNA in the cells as determined by ribonuclease protection assay. In the presence of 4 mmol/L sodium butyrate, a fourfold increase (P < 0. 01) in mRNA level was obtained at 24 h. No stabilizing effect of butyrate on PLTP mRNA was apparent upon treatment of the cultured cells with the RNA synthesis inhibitor, actinomycin D. Thus, the up-regulatory effect of butyrate on PLTP gene expression seemed to have occurred at the transcriptional level.

Animals↗
PubMed1999

Signaling between the placenta and the uterus involving the mitogen-regulated protein/proliferins.

The aim of this investigation was to examine signaling between the placenta and uterus during pregnancy. To do this, we determined the tissue messenger RNA and protein levels of members of a glycopeptide hormone family known to stimulate the proliferation of uterine cells and related these levels to the growth of the uterus during pregnancy in the mouse. This hormone family is known as mitogen-regulated protein (MRP); alternatively proliferin (PLF). Three mrp/plf genes, plf1, mrp3 and mrp4, are expressed by the placenta with different developmental profiles. The major increase of about 4-fold in DNA content of the uterus occurs between days 9 and 14 when MRP/PLFs are present in the placenta. By contrast, the gestational changes in estradiol-17beta levels in placental and uterine tissues and in circulation do not correlate with the period of uterine growth. The previously reported mitogenic activity of the MRP/PLFs and their gestational profiles suggest that one or more of these proteins stimulates uterine proliferation during gestation. Evidence is also presented that expression of MRP3 and/or PLF1, but not MRP4, is negatively regulated by feedback from the uterus. Our results are consistent with the hypothesis that MRP/PLFs stimulate uterine proliferation in vivo and that a uterine factor shuts off PLF1 and/or MRP3 synthesis in the latter half of gestation.

Amniotic Fluid↗
PubMed1999

[Cloning the 5 ' end fragment of ST13 cDNA by nested PCR].

OBJECTIVE: To clone and sequence the 5' -end fragment of ST13 cDNA. METHODS: The 5' -end fragment of the ST13cDNA was amplified directly from cDNA library by nested PCR method, and cloned into pGEM-T. easy vector, then the sequencing of the inserted PCR product was performed. RESULTS: After primary and secondary PCR, two PCR products obtained with the size of about 550bp and 480bp, respectively, were both proved to be the 5' -end of ST13 cDNA by sequencing. CONCLUSION: the method presented is very simple and effective for cloning and sequencing the 5'-end of a target gene.

Cloning, Molecular↗
PubMed1999

[Clinical analysis of percutaneous balloon mitral valvotomy in 1063 patients].

This article reports the clinical study of 1063 patients who underwent percutaneous balloon mitral valvotomy (PBMV) and 1043 patients among them who did successfully (98.2%). After the procedure, left atrial mena pressure fell from 3.19 +/- 1.06 kPa to 2.00 +/- 0.93 kPa (P < 0.001), mitral pressure gradient decreased from 2.79 +/- 1.20 kPa to 1.33 +/- 1.06 kPa (P < 0.001). Disappearance of diastolic murmur and significant improvement of cardiac function were achieved. Among the 20 failed cases, there were cardiac perforation and/or pericardial tamponade (8 cases), cerebral embolism (3 cases), severe mitral regurgitation (3 cases), atrial septal defect (2 cases) and discontinuance of the procedure due to various causes. The short-term outcome of PBMV is satisfying, so it can replace the surgical closed valvotomy as an effective treatment for selected patients with mitral stenosis.

Adolescent↗
PubMed1999

[Changes and clinical significance of serum thyroxine level in patients with congestive heart failure].

In order to explore the relationship between congestive heart failure (CHF) and serum thyroxine, we assayed serum thyroxine level in 52 patients with CHF and 28 normal persons by means of radioimmunoassay (RIA). The results showed that triiodothyronine reduced and reverse triiodothyronine elevated significantly in the patients compared with the normal (P < 0.05). The more severe heart failure developed, the more obvious changes of the hormones were observed. The results suggest that the alterations of serum thyroid level may be used as an index of the severity of heart failure.

Adult↗
PubMed1999

[Effect of IL-6 on the multiplication of Toxoplasma gondii].

AIM: To observe the effect of IL-6 on the multiplication of Toxoplasma gondii. METHODS: Peritoneal macrophages from C57BL/6 mouse were incubated with 3H-uracil-labelled T. gondii in vitro. RESULTS: Pretreatment (but not post-treatment) of Mphi with IL-6 enhanced T. gondii multiplication in a dose-dependent manner. Pretreatment with IFN gamma resulted in active killing of parasites whereas the addition of IL-6 resulted in a partial reversal of IFN gamma-mediated toxoplasmacidal activity. Combining TNF alpha with IL-6 and IFN gamma pretreatment resulted in restoration of toxoplasmacidal activity. Addition of polyclonal anti-TNF alpha antibodies to IL-6 and IFN gamma pretreatment resulted in enhancement in the IL-6-mediated impairment of IFN gamma function. CONCLUSION: IL-6 could enhance intraperitoneal multiplication of T. gondii and reverse IFN gamma-mediated toxoplasmacidal activity.

Animals↗
PubMed1999

[Noso-orbital related disease: an analysis of 39 cases].

OBJECTIVE: To enhance the Diagnostic and therapeutic levels of naso-orbital diseases. METHOD: Thirty-nine cases of naso-orbital diseases were analyzed retrospectively. RESULT: Among the 39 cases, 7 had no nasal symptoms. 9 had their symptoms mainly confined to eyes and were initially consulted with ophtalmologists. 1 case was consulted with neurologist at first. The main pathologies were as following: bony absorption of orbit wall in inflammatory disorders; compression, displacement and bony absorption of orbit wall in benign tumors and mucoceles; bony destruction in malignant tumors. Tranmatic fractures might involve multiple bony structures. CONCLUSION: Nosal and orbital organs are closely related anatomically. Imaging investigations are important in diagnosis of naso-orbital related diseases. CT is helpful for early diagnosis. Treatment for benign tumors is mainly by operation, while malignant tumors should be treated by operation and radiotherapy together.

Adolescent↗
PubMed1999

[Operative approach choosing to treat cholesteatoma otitis media (with clinical analysis of 860 cases)].

OBJECTIVE: In order to improve operative effect to cholesteatoma otitis media. METHOD: The clinical data of 860 cases of patients with cholesteatoma otitis media were treated by surgery through three approachs (through epitympanum, cribriform area or combined approach) were analyzed. RESULT: The results showed that the recurrent rates of them were 10%, 11% and 6% respectively; The rates of improvement of hearing were 56%, 40% and 69% five years after operation respectively, and after 5 to 15 years were 45%, 35% and 62%; The rates of different abnormal shapes of tympanic membrane (including retraction, adherence and perforation) were 43%, 47% and 12%. CONCLUSION: According to the location of foci, we regard that various and appropriate operative approach should be selected to treat the cases with cholesteatoma otitis media, and the combined approach tympanoplasty has advantage over the other two.

Adolescent↗