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Biomedical subjects

Y Lee

Publications and source records attributed to Y Lee.

At least 73 records · Page 4Linked to original sources
PubMed2001

Ataxia telangiectasia mutated-dependent apoptosis after genotoxic stress in the developing nervous system is determined by cellular differentiation status.

Ataxia-telangiectasia (A-T) is a neurodegenerative syndrome resulting from dysfunction of ATM (ataxia telangiectasia mutated). The molecular details of ATM function in the nervous system are unclear, although the neurological lesions in A-T are probably developmental because they appear during childhood. The nervous systems of Atm-null mice show a pronounced defect in apoptosis that is induced by DNA damage, suggesting that ATM may function to eliminate DNA-damaged neurons. Here we show that Atm-dependent apoptosis occurs at discrete stages of neurogenesis. Analysis of gamma-irradiated mouse embryos showed that Atm-dependent apoptosis occurred only in the postmitotic populations that were present in the neuroepithelial subventricular zone of the developing nervous system. Notably, Atm deficiency did not prevent radiation-induced apoptosis in multipotent precursor cells residing in the proliferating ventricular zone. Atm-dependent apoptosis required p53 and coincided with the specific phosphorylation of p53 and caspase-3 activation. Thus, these data show that Atm functions early in neurogenesis and underscore the selective requirement for Atm in eliminating damaged postmitotic neural cells. Furthermore, these data demonstrate that the differentiation status of neural cells is a critical determinant in the activation of certain apoptotic pathways.

Animals↗
PubMed2001

Pertussis toxin-induced hyperacute autoimmune encephalomyelitis in Lewis rats is correlated with increased expression of inducible nitric oxide synthase and tumor necrosis factor alpha.

The involvement of inducible nitric oxide synthase (iNOS) and tumor necrosis factor alpha (TNF-alpha), which have diverse roles in the progression of autoimmune disease models, was studied in pertussis toxin (PT)-induced hyperacute experimental autoimmune encephalomyelitis (EAE) in Lewis rats. The expression of TNF-alpha mRNA (increased 5-fold, P<0.01) and iNOS protein (3-fold, P<0.01) was much greater in the spinal cords with PT(+) EAE at the peak stage of EAE than in those with PT(-) EAE, as shown by competitive PCR and Western blot analysis, respectively. Immunohistochemistry showed that the majority of ED1-positive macrophages in EAE lesions contained iNOS, and that there were many more iNOS-positive cells in the CNS lesions of PT(+) rats than in those of PT(-) rats. These findings suggest that PT-induced hyperacute EAE is partly mediated by the enhanced expression of iNOS and TNF-alpha in the early stages of rat EAE.

Animals↗
PubMed2001

Fas aggregation does not correlate with Fas-mediated apoptosis.

Cross-linking of cell surface Fas molecules by Fas ligand or by agonistic anti-Fas Abs induces cell death by apoptosis. We found that a serine protease inhibitor, N-tosyl-L-lysine chloromethyl ketone (TLCK), dramatically enhances Fas-mediated apoptosis in the human T cell line Jurkat and in various B cell lines resistant to Fas-mediated apoptosis. The enhancing effect of TLCK is specific to Fas-induced cell death, with no effect seen on TNF-alpha or TNF-related apoptosis-inducing ligand-induced apoptosis. TLCK treatment had no effect on Fas expression levels on the cell surface, and neither promoted death-inducing signaling complex formation nor decreased expression levels of cellular inhibitors of apoptosis (FLICE inhibitory protein, X chromosome-linked inhibitor of apoptosis, and Bcl-2). Activation of the Fas-mediated apoptotic pathway by anti-Fas Ab is accompanied by aggregation of Fas molecules to form oligomers that are stable to boiling in SDS and beta-ME. Fas aggregation is often considered to be required for Fas-mediated apoptosis. However, sensitization of cells to Fas-mediated apoptosis by TLCK or other agents (cycloheximide, protein kinase C inhibitors) causes less Fas aggregation during the apoptotic process compared with that in nonsensitized cells. These results show that Fas aggregation and Fas-mediated apoptosis are not directly correlated and may even be inversely correlated.

Adjuvants, Immunologic↗
PubMed2001

Testis-specific expression of an intronless gene encoding a human poly(A) polymerase.

A mouse intronless gene, encoding a testis-specific poly(A) polymerase (mPAPT), was previously identified. mPAPT may play a role as a putative enzyme that is responsible for polyadenylation regulation during mouse spermatogenesis. In order to understand how PAPT genes are conserved in mammals, we isolated a human cDNA homolog encoding a human PAPT (hPAPT), which was specifically expressed in the testis. The structure of hPAPT was very similar to that of mPAPT. The about 100 residues at the C-terminal region of a nuclear poly(A) polymerase, PAP II, were missing in both PAPT proteins. An analysis of the genomic DNA showed that the hPAPT gene is an intronless gene that is similar to the mPAPT gene. Interestingly, the sequence homology between hPAPT and mPAPT was much lower than the homology between hPAP II and mPAP II. The phylogenetic analysis suggests that PAPTs arose through retrotransposition after the amphibian-amniote split during evolution.

Animals↗
PubMed2001

Biomedical applications of collagen.

Collagen is regarded as one of the most useful biomaterials. The excellent biocompatibility and safety due to its biological characteristics, such as biodegradability and weak antigenecity, made collagen the primary resource in medical applications. The main applications of collagen as drug delivery systems are collagen shields in ophthalmology, sponges for burns/wounds, mini-pellets and tablets for protein delivery, gel formulation in combination with liposomes for sustained drug delivery, as controlling material for transdermal delivery, and nanoparticles for gene delivery and basic matrices for cell culture systems. It was also used for tissue engineering including skin replacement, bone substitutes, and artificial blood vessels and valves. This article reviews biomedical applications of collagen including the collagen film, which we have developed as a matrix system for evaluation of tissue calcification and for the embedding of a single cell suspension for tumorigenic study. The advantages and disadvantages of each system are also discussed.

Animals↗
PubMed2001

Definition of an optimal cytotoxic T lymphocyte epitope in the latently expressed Kaposi's sarcoma-associated herpesvirus kaposin protein.

Cytotoxic T lymphocytes (CTL) recognize and kill virus-infected cells and contribute to immunologic control of viral replication. For many herpesviruses (e.g., Epstein-Barr and cytomegalovirus), virus-specific CTL responses can be readily detected in infected persons, but CTL responses against Kaposi's sarcoma-associated herpesvirus (KSHV) appear to be weak and remain poorly characterized. Using a human leukocyte antigen (HLA) binding motif-based epitope prediction algorithm, we identified 37 HLA-A*0201 binding peptides from 8 KSHV open-reading frames (ORFs). After in vitro stimulation of peripheral blood mononuclear cells from KSHV-infected persons, CTL responses against 1 peptide in the KSHV kaposin protein (ORF K12) were detected in 2 HLA-A*0201-positive subjects. The optimal CTL epitope was identified by HLA restriction analysis and peptide titration assays. These data describe a latent phase viral gene product targeted by CTL that may be relevant for KSHV immunopathogenesis.

Acquired Immunodeficiency Syndrome↗
PubMed2001

Glycogen synthase kinase 3beta regulates GATA4 in cardiac myocytes.

Inactivation of glycogen synthase kinase 3beta (GSK3beta) is critical for transcription of atrial natriuretic factor (ANF) by beta-adrenergic receptors in cardiac myocytes. We examined the mechanism by which GSK3beta regulates ANF transcription. Stimulation of beta-adrenergic receptors induced nuclear accumulation of GATA4, whereas beta-adrenergic ANF transcription was suppressed by dominant negative GATA4, suggesting that GATA4 plays an important role in beta-adrenergic ANF transcription. Interestingly, GATA4-mediated transcription was markedly attenuated by GSK3beta. GSK3beta physically associates with GATA4 and phosphorylates GATA4 in vitro. Overexpression of GSK3beta suppressed both basal and beta-adrenergic increases in nuclear expression of GATA4, whereas inhibition of GSK3beta by LiCl caused nuclear accumulation of GATA4, suggesting that GSK3beta negatively regulates nuclear expression of GATA4. The nuclear exportin Crm1 reduced nuclear expression of GATA4, and the reduction was enhanced by GSK3beta but not by kinase-inactive GSK3beta. Leptomycin B, an inhibitor for Crm1, increased basal nuclear GATA4 and suppressed GSK3beta-induced decreases in nuclear GATA4. These results suggest that GSK3beta negatively regulates nuclear expression of GATA4 by stimulating Crm1-dependent nuclear export. Inhibition of GSK3beta by beta-adrenergic stimulation abrogates GSK3beta-induced nuclear export of GATA4, causing nuclear accumulation of GATA4, which may represent an important signaling mechanism mediating cardiac hypertrophy.

Adenoviridae↗
PubMed2001

Intracardiac echocardiographic guidance and monitoring during percutaneous endomyocardial gene injection in porcine heart.

In an effort to develop a guiding and monitoring tool for transmyocardial gene transfer, we have evaluated the feasibility of intracardiac echocardiography (ICE) to guide percutaneous endomyocardial gene transfer (PEGT), and monitor complications, in a pig model. ICE (5.5-10 MHz), complemented by fluoroscopy, was utilized to guide a needle injection into the heart in 19 normal pigs. Using this system, we injected Evans blue dye into eight pigs (group I), a mixture of pCK-CAT plasmid and India ink into seven pigs (group II), and pCK-LacZ plasmid into four pigs (group III). In all pigs, ICE contributed to the injection procedure by guiding the catheter to anatomically distinct sites, and by assisting stabilization of the catheter-endocardial contact. ICE predicted the injection sites correctly in 56 of 64 sites (87.5%) in group I, and in 42 of 42 sites (100%) in group II. Leakage of injectate into the left ventricular cavity could be detected by the microbubbles generated. The sites of injections appeared as foci of bright myocardial echodensity, which persisted until the end of the procedure. The procedures were not associated with significant morbidity or mortality. The expression of the chloramphenicol acetyltransferase (CAT) gene was identified in 40 sites from 42 injections (95.2%) in group II. In group III, histology showed positive beta-galactosidase staining of myocytes limited around the needle track with low transfection efficiency (<1%). These results suggest that real-time ICE monitoring proves safe and useful during PEGT for guiding needle injection, monitoring leakage, ensuring delivery of injectate into the myocardium, and instantly diagnosing cardiac complications, resulting in successful gene transfer.

Animals↗
PubMed2001

Scanning electrochemical microscopy. 41. Theory and characterization of ring electrodes.

Ring ultramicroelectrodes, which are of particular interest as probes for scanning electrochemical microscopy (SECM), combined with near-field scanning optical microscopy, were investigated. Theoretical SECM tip current-distance (approach) curves for a ring electrode were calculated by numerical (finite element) analysis. The SECM curves obtained were a function of the geometry of the tips including the thickness of the ring and the insulating sheath. Theoretical approach curves over conductive substrates showed a strong dependence on the ratio of inner to outer radii of ring microelectrodes (a/b) and were relatively insensitive to the thickness of the insulating sheath (r(g)). For insulating substrates, however, the approach curves varied significantly with r(g), but much less with the a/b ratio. Comparison of experimental and theoretical SECM curves provided a good method of evaluating the size and shape of ring electrodes. Good agreement of the experimental and theoretical curves was found with a ring microelectrode with a nominal 200-nm ring thickness, yielding values of 1.7, 1.9, and 5.7 microm for the inner (a) and outer (b) radii of a ring and the outermost radius of insulating sheath (r(g)), respectively.

Journal Article↗
PubMed2001

Scanning optical microscopy with an electrogenerated chemiluminescent light source at a nanometer tip.

Electrogenerated chemiluminescence at electrodes with effective diameters down to 155 nm was used as a stable light source for near-field scanning optical microscopy imaging of an interdigitated array and a submicrometer size test substrate. Light was generated in a thin (approximately 500 microm) layer of an aqueous solution of 15 mM Ru(bpy)3(2+) and 100 mM tri-n-propylamine in a pH 7.5 buffer. The resolution obtained was compared to that found with a micrometer size electrode. The shear force from the tip attached to a quartz tuning fork was used to monitor and control the tip-to-substrate separation within the near field regime.

Journal Article↗
PubMed2001

Evaluation on the adsorption capabilities of new chemically modified polymeric adsorbents with protoporphyrin IX.

A chemically modified polymeric adsorbent was synthesized to evaluate the availability as an adsorbent for solid-phase extraction (SPE) of phenol and chlorophenols. Commercially available Amberlite XAD-2 and XAD-4 resins were modified with macrocyclic protoporphyrin IX (PPIX) through the ketone linkage. Adsorption isotherms were obtained by batch experiments and the data were fitted to the Freundlich equation to calculate the adsorption parameters. Breakthrough volumes were measured by column experiments. Physical properties such as surface area, average pore diameter and micropore volume of resins were measured to correlate with the adsorption characteristics. As a result, adsorption capacity was increased for the chemically modified resins and it can be concluded that the increase of pi-pi interaction due to the introduction of the porphyrin molecule is the major factor for the increase of the adsorption capacity.

Adsorption↗
PubMed2001

Induction of alpha1-acid glycoprotein mRNA by cytokines and differentiation in human colon carcinoma cell.

The mRNA for the alpha1-acid glycoprotein (AAG) was expressed not only in hepatoma cells, but also in non-hepatic cancer cells. The expression of the AAG mRNA in HT-29 human colon carcinoma cells is induced by cytokines, IL-6, IL-1, and TNF-alpha, in a manner characteristic of the acute phase response, and the expression of AAG mRNA was up-regulated in differentiated HT-29 cells.

Cell Differentiation↗
PubMed2001

Congestion of right liver graft in living donor liver transplantation.

BACKGROUND: Left liver graft from a small donor will not meet the metabolic demands of a larger adult recipient. One solution to this problem is to use a right liver graft without a middle hepatic vein (MHV). However, the need for drainage from the MHV tributaries has not yet been described. METHODS: Five right liver grafts without a MHV were transplanted in patients including two hepatitis B virus-cirrhosis, two fulminant hepatic failure and one secondary biliary cirrhosis. The graft weight ranged from 650 to 1,000 g, corresponding to 48 to 83% of the standard liver volume of the recipients. RESULTS: Two of five recipients were complicated with severe congestion of the right median sector immediately after reperfusion, followed by prolonged massive ascites and severe liver dysfunction. One of the patients died of sepsis with progressive hepatic dysfunction 20 days after the operation. CONCLUSIONS: Preservation and reconstruction of the MHV tributaries is recommended to prevent congestion of the right liver graft without MHV.

Body Weight↗
PubMed2001

Temporary inactivation of plasma amine oxidase by alkylhydrazines. A combined enzyme/model study implicates cofactor reduction/reoxidation but cofactor deoxygenation and subsequent reoxygenation in the case of hydrazine itself.

It has been known for some time that hydrazine and its methyl and 1,1-dimethyl analogues induce inactivation of the copper-containing quinone-dependent plasma amine oxidase but that the activity recovers over time, suggesting metabolism of all three inhibitors. However, the mechanism responsible for loss and regain of activity has not been investigated. In this study a combination of enzyme studies under a controlled atmosphere along with model studies using 5-tert-butyl-2-hydroxy-1,4-benzoquinone to mimic the 2,4,5-trihydroxyphenylalanine quinone (TPQ) cofactor of the enzyme suggest that regain of enzyme activity represents two different O(2)-dependent processes. In the case of methylhydrazine and 1,1-dimethylhydrazine, we propose that the inactive methylhydrazone/azo form of the enzyme slowly rehydrates and eliminates MeN=NH to give the triol cofactor form, which instantly reoxidizes to the catalytically active quinone form in the presence of O(2). Metabolism of methylhydrazine represents its conversion to CH(4) and N(2), and of 1,1-dimethylhydrazine to CH(2)=O, CH(4), and N(2). In the case of hydrazine itself, however, we propose that the inactive hydrazone/azo form of the enzyme instead undergoes a slow decomposition, probably facilitated by the active-site copper, to give N(2) and a novel 5-desoxy resorcinol form of the cofactor. The latter undergoes a rapid, but noninstantaneous reoxygenation at C5 to restore the active cofactor form, also probably mediated by the active-site copper.

Amine Oxidase (Copper-Containing)↗
PubMed2001

Characterization and regulation of Schizosaccharomyces pombe gene encoding thioredoxin.

A cDNA coding thioredoxin (TRX) was isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The 438 bp EcoRI fragment, which was detected by Southern hybridization, reveals an open reading frame which encodes a protein of 103 amino acids. The genomic DNA encoding TRX was also isolated from S. pombe chromosomal DNA using PCR. The cloned sequence contains 1795 bp and encodes a protein of 103 amino acids. However, the C-terminal region obtained from the cDNA clone is -Val-Arg-Leu-Asn-Arg-Ser-Leu, whereas the C-terminal region deduced from the genomic DNA appears to contain -Ala-Ser-Ile-Lys-Ala-Asn-Leu. This indicates that S. pombe cells contain two kinds of TRX genes which have dissimilar amino acid sequences only at the C-terminal regions. The heterologous TRX 1C produced from the cDNA clone could be used as a subunit of T7 DNA polymerase, while the TRX 1G from the genomic DNA could not. The upstream sequence and the region encoding the N-terminal 18 amino acids of the genomic DNA were fused into the promoterless beta-galactosidase gene of the shuttle vector YEp357 to generate the fusion plasmid pYKT24. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by hydrogen peroxide, menadione and aluminum chloride. It indicates that the expression of the cloned TRX gene is induced by oxidative stress.

Aluminum Chloride↗