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Biomedical subjects

S Lin

Publications and source records attributed to S Lin.

At least 109 records · Page 6Linked to original sources
PubMed2000

Down-regulation of cyclin D1 expression by prostaglandin A(2) is mediated by enhanced cyclin D1 mRNA turnover.

Prostaglandin A(2) (PGA(2)), an experimental chemotherapeutic agent, causes growth arrest associated with decreased cyclin D1 expression in several cancer cell lines. Here, using human non-small-cell lung carcinoma H1299 cells, we investigated the mechanisms whereby PGA(2) down-regulates cyclin D1 expression. Transcription rates of the cyclin D1 gene, studied using a cyclin D1 promoter-luciferase construct and nuclear run-on assays, were not affected by PGA(2) treatment. Instead, the cyclin D1 mRNA was rendered unstable after exposure to PGA(2). Since the stability of labile mRNA is modulated through binding of proteins to specific mRNA sequences, we sought to identify protein(s) recognizing the cyclin D1 mRNA. In electrophoretic mobility-shift assays using radiolabeled RNA probes derived from different regions of cyclin D1 mRNA, we observed that (i) lysates prepared from PGA(2)-treated cells exhibited enhanced protein-cyclin D1 RNA complex formation; (ii) the kinetics of complex formation correlated closely with that of cyclin D1 mRNA loss; and (iii) binding occurred within a 390-base cyclin D1 3' untranslated region (UTR) (K12). This binding activity could be cross-linked, revealing proteins ranging from 30 to 47 kDa. The RNA-binding protein AUF1, previously associated with the degradation of target mRNAs, bound cyclin D1 mRNA, because anti-AUF1 antibodies were capable of supershifting or immunoprecipitating cyclin D1 mRNA-protein complexes. Finally, insertion of K12 in the 3'UTR of reporter genes markedly reduced the expression and half-life of the resulting chimeric mRNAs in transfected, PGA(2)-treated cells. Our data demonstrate that PGA(2) down-regulates cyclin D1 expression by decreasing cyclin D1 mRNA stability and implicates a 390-base element in the 3'UTR in this regulation.

3' Untranslated Regions↗
PubMed2000

Comparative polymerase chain reaction analysis of c-myc amplification on archival breast fine-needle aspiration materials.

The oncogene c-myc is a key regulator of cell cycle progression (from G1 to S phase). The amplification of c-myc can either induce cell proliferation or apoptosis. As a part of our ongoing effort to develop methods for multiple tumor marker analysis, this study was carried out to determine whether biomarkers such as c-myc amplification could be analyzed on genetic materials collected from archival fine-needle aspiration (FNA) smears. A novel comparative PCR analysis was used to analyze c-myc amplification semiquantitatively. Genomic DNA was prepared using cells obtained from archival FNA materials that had undergone quantitative fluorescence image analysis (QFIA) for other biomarkers. Of the 72 cases selected from 1995 for this study, 53 had an adequate amount of DNA for analysis. A novel comparative PCR analysis was used to analyze c-myc amplification quantitatively. For each batch of experiments, DNA from the high c-myc expressing cells, HL-60, and DNA from the low expressing cells, K562, were served as positive and negative controls, respectively. c-myc amplification was observed in 16 (94.1%) of 17 malignant lesions, 5 (41.7%) of 12 proliferative breast diseases with nuclear atypia, and 4 (16.7%) of 24 other benign lesions (fibroadenoma or fibrocystic disease). The overall difference of c-myc expression among these groups was highly significant by chi2 analysis (P = 0.0002). We conclude that multiple phenotypic markers and genotypic markers may be combined in a risk assessment biomarker profile on small FNA samples that can be obtained on multiple occasions relatively noninvasively from the patient. The results of this study suggest that c-myc amplification may be a biomarker of breast cancer risk. However, additional large, prospective studies are needed to confirm the current observation.

Adult↗
PubMed2000

Androgenetic alopecia in heterozygous carriers of a mutation in the human hairless gene.

BACKGROUND: Androgenetic alopecia is considered to be genetically determined. Recently, a rare autosomal recessive form of hereditary alopecia, termed atrichia with papular lesions (APL), was found to result from mutations in the human hairless gene. OBJECTIVE: Our aim was to assess the pattern of androgenetic alopecia in heterozygous carriers of a deleterious mutation in the human hairless gene. METHODS: Healthy male second-degree relatives (n = 31) of patients affected with APL and belonging to a large consanguineous kindred were interviewed and given a Hamilton score of baldness. DNA was obtained from each subject and analyzed for the presence of a mutation in the human hairless gene known to affect this family. The age at onset and extent of baldness were compared in healthy homozygotes and heterozygous carriers of the mutation. RESULTS: Statistical analysis of the results revealed no differences in age at onset and extent of androgenetic alopecia between the two groups of subjects. CONCLUSION: The present study reports the first attempt to characterize the phenotype of heterozygous carriers of a mutation in the human hairless gene. It indicates that the presence of a deleterious mutation in one allele of the hairless gene does not affect the pattern of androgenetic hair loss.

Adult↗
PubMed2000

Plasma CPU-86017 concentrations regarding suppression of ouabain-induced cardiac arrhythmias and decrease of heart rate in guinea pigs.

AIM: To determine the effective plasma levels of CPU-86017 which could suppress the cardiac arrhythmias induced by i.v. ouabain in guinea pigs. METHODS: The cardiac arrhythmias and the heart rate were monitored by ECG traces. Blood samples were collected to determine plasma levels using HPLC assay. TXB2 and 6-keto-PGF1 alpha were measured in plasma. RESULTS: The plasma concentrations of CPU-86017 which were effective to suppress ventricular fibrillation (VF) and heart rate were 0.13-0.23 mg/L and 0.13-0.31 mg/L, respectively. A reduction of TXB2 levels and an elevation of 6-keto-PGF1 alpha levels were observed after CPU-86017 i.v. administration. CONCLUSION: The arrhythmia-suppressing and heart rate-slowing effect of CPU-86017 followed a linear relationship with its concentrations in plasma.

6-Ketoprostaglandin F1 alpha↗
PubMed1999

Synthesis and structure-activity relationships of new second-generation taxoids.

A series of second-generation taxoids bearing a substituent on the C-2-benzoyl group and modifications at C-3'/C-10 positions was synthesized. These taxoids exhibited 2-3 orders of magnitude higher potency than that of paclitaxel against drug-resistant human breast cancer cell lines. It is also noteworthy that three taxoids showed almost no difference in activity against drug-resistant and drug-sensitive cell lines, which are categorized as "advanced second generation taxoids".

Alkaloids↗
PubMed1999

Human acyl-CoA:cholesterol acyltransferase-1 is a homotetrameric enzyme in intact cells and in vitro.

Acyl-CoA:cholesterol acyltransferase (ACAT) is a key enzyme in cellular cholesterol homeostasis and in atherosclerosis. ACAT-1 may function as an allosteric enzyme. We took a multifaceted approach to investigate the subunit composition of ACAT-1. When ACAT-1 with two different tags were co-expressed in the same Chinese hamster ovary cells, antibody specific to one tag caused co-immunoprecipitation of both types of ACAT-1 proteins. Radioimmunoprecipitations of cells expressing the untagged ACAT-1 or the 6-histidine-tagged ACAT-1 yielded a single radiolabeled band of predicted size on SDS-polyacrylamide gel electrophoresis. These results show that ACAT-1 exists as homo-oligomers in intact Chinese hamster ovary cells. We solubilized HisACAT-1 with the detergent deoxycholate or CHAPS (3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonic acid), performed gel filtration chromatography and sucrose density gradient centrifugations in H(2)O and D(2)O, and determined the Stokes radii and sedimentation coefficients of the HisACAT1-detergent complexes. The estimated molecular mass of HisACAT-1 is 263 kDa, which is 4 times that of the HisACAT-1 monomer (69 kDa). Finally, cross-linking experiments in intact cells and in vitro show that the increase in cross-linker concentrations causes an increase in size of the HisACAT-1-positive signals, forming material(s) 4 times the size of the monomer, supporting the conclusion that ACAT-1 is a homotetrameric enzyme.

Animals↗
PubMed1999

Studies of the DXS7 polymorphism at the MAO loci in unipolar depression.

From the fact that DXS7 polymorphism is closely related to monoamine oxidase (MAO) genes and MAO inhibitors are widely used in the treatment of unipolar depression, it is of particular interest to study the relationship between the DXS7 polymorphism and unipolar depression. Thus, this study examined the possible association between DXS7 polymorphism and unipolar depression in 66 cases versus 85 controls from Shanghai. Polymerase chain reaction and amplification fragment length polymorphism techniques were used for genotyping of the DXS7 locus in this study. Four alleles at the DXS7 locus were detected with length generated by polymerase chain reaction amplification ranging from 157 to 167 bp. Comparison of allele frequency in the DXS7 locus showed no difference between unipolar depression cases and normal controls in the total population set. When subclassified by age, a significant difference of allele frequency distribution was observed between early onset (before age 40) and late onset (after age 40) patients. The frequency of the 157-bp allele was decreased, whereas the frequency of the 165 allele was increased in late onset patients (0.3810 for the 157-bp allele and 0.5238 for the 165-bp allele) compared with that of early onset patients (0.6304 for the 157-bp allele and 0. 3261 for the 165-bp allele). There was also a difference of allele frequency between patients and normal controls with age over 40 years. The frequency of 165-bp allele increased significantly in late onset patients (0.5238) compared with that of controls within the same age range (0.3454). Association studies suggested that in the population with age over 40 years, presence of the 165-bp allele of DXS7 locus was significantly associated with unipolar depression (relative risk = 2.08, P < 0.05), whereas in the total population set, this association did not exist. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 88:598-600, 1999.

Adolescent↗
PubMed1999

Suppression of activin-induced apoptosis by novel antisense strategy in human prostate cancer cells.

Apoptosin, a novel gene encoding a mitotic kinase-motif protein, is stimulated by activin, a member of TGF-beta family, in human LNCaP prostate cancer cells and in patient tissues. We employed a gene knockout methodology based on the covalent bonding of chemically modified antisense probes to apoptosin mRNAs in LNCaP cells. The mRNA-antisense hybrid duplexes were neither translated nor post-transcriptionally modified, resulting in no protein synthesis. Introducing antisense apoptosin into activin-induced apoptotic LNCaP cells prevented apoptosis, interfered with genomic DNA fragmentation and released cell cycle checkpoint. These findings suggest that the apoptosin, in addition to p53, is important in apoptotic regulation of human prostate cancers.

Activins↗
PubMed1999

Picosecond transient absorption spectroscopy in the blue spectral region of photosystem I.

Picosecond transient absorption difference spectroscopy in the blue wavelength region (380-500 nm) was used to study the early electron acceptors in photosystem I. Samples were photosystem I core particles with about 100 chlorophylls per reaction center isolated from the cyanobacterium Synechocystis sp. PCC 6803. After excitation at 590 nm at room temperature, decay-associated spectra (DAS) were determined from global analysis in the blue region, yielding two transient components and one nondecaying component. A 3 ps decay phase is interpreted as primarily due to antenna excited-state redistribution. A 28 ps decay phase is interpreted as due to overall excited-state decay by electron transfer. The nondecaying component is ascribed to the difference spectrum of P(700) and the quinone or A(1) electron acceptor (P(700)(+)A(1)(-) - P(700)A(1)). Decay curves on the millisecond time scale at different wavelengths were measured with an autoxidizable artificial electron acceptor, benzyl viologen, and the (P(700)(+) - P(700)) difference spectrum was constructed. The (A(1)(-) - A(1)) difference spectrum was obtained by taking the difference between the above two difference spectra. A parallel picosecond experiment under strongly reducing conditions was also done as a control experiment. These conditions stabilize the electron on an earlier acceptor, A(0). The nondecaying component of the DAS at low potential was assigned to (P(700)(+)A(0)(-) - P(700)A(0)) since the electron-transfer pathway from A(0) to A(1) was blocked. The [(P(700)(+)A(0)(-) - P(700)A(0)) - (P(700)(+) - P(700))] subtraction gives a spectrum, interpreted as the (A(0)(-) - A(0)) difference spectrum of a chlorophyll a molecule, consistent with previous studies. The (A(1)(-) - A(1)) spectrum resolved on the picosecond time scale shows significant differences with similar spectra measured on longer time scales. These differences may be due to electrochromic effects and spectral evolution.

Benzyl Viologen↗
PubMed1999

Endothelin antagonists: substituted mesitylcarboxamides with high potency and selectivity for ET(A) receptors.

We have previously disclosed the discovery of 2,4-disubstituted anilinothiophenesulfonamides with potent ET(A)-selective endothelin receptor antagonism and the subsequent identification of sitaxsentan (TBC11251, 1) as a clinical development compound (Wu et al. J. Med. Chem. 1997, 40, 1682 and 1690). The orally active 1 has demonstrated efficacy in a phase II clinical trial of congestive heart failure (Givertz et al. Circulation 1998, 98, Abstr. #3044) and was active in rat models of myocardial infarction (Podesser et al. Circulation 1998, 98, Abstr. #2896) and acute hypoxia-induced pulmonary hypertension (Chen et al. FASEB J. 1996, 10 (3), A104). We now report that an additional substituent at the 6-position of the anilino ring further increases the potency of this series of compounds. It was also found that a wide range of functionalities at the 3-position of the 2,4,6-trisubstituted ring increased ET(A) selectivity by approximately 10-fold while maintaining in vitro potency, therefore rendering the compounds amenable to fine-tuning of pharmacological and toxicological profiles with enhanced selectivity. The optimal compound in this series was found to be TBC2576 (7u), which has approximately 10-fold higher ET(A) binding affinity than 1, high ET(A)/ET(B) selectivity, and a serum half-life of 7.3 h in rats, as well as in vivo activity.

Administration, Oral↗
PubMed1999

A Haemophilus influenzae gene that encodes a membrane bound 3-deoxy-D-manno-octulosonic acid (Kdo) kinase. Possible involvement of kdo phosphorylation in bacterial virulence.

The lipopolysaccharide of Haemophilus influenzae contains a single 3-deoxy-D-manno-octulosonic acid (Kdo) residue derivatized with either a phosphate or an ethanolamine pyrophosphate moiety at the 4-OH position. In previous studies, we identified a kinase unique to H. influenzae extracts that phosphorylates Kdo-lipid IV(A), a key precursor of lipopolysaccharide in this organism. We have now identified the gene encoding the Kdo kinase by using an expression cloning approach. A cosmid library containing random DNA fragments from H. influenzae strain Rd was constructed in Escherichia coli. Extracts of 472 colonies containing individual hybrid cosmids were assayed for Kdo kinase activity. A single hybrid cosmid directing expression of the kinase was found. The kinase gene was identified by activity assays, sub-cloning, and DNA sequencing. When the putative kinase gene was expressed in E. coli behind a T7 promoter, massive overproduction of kinase activity was achieved ( approximately 8000-fold higher than in H. influenzae membranes). The catalytic properties and the product generated by the overexpressed kinase, assayed with Kdo-lipid IV(A) as the substrate, were the same as observed with H. influenzae membranes. Unexpectedly, the kinase gene was identical to a previously characterized open reading frame (orfZ), which had been shown to be important for establishing bacteremia in an infant rat model (Hood, D. W., Deadman, M. E., Allen, T., Masoud, H., Martin, A., Brisson, J. R., Fleischmann, R., Venter, J. C., Richards, J. C., and Moxon, E. R. (1996) Mol. Microbiol. 22, 951-965). However, based solely on the genome sequence of H. influenzae Rd, no biochemical function had been assigned to the product of orfZ, which we now designate kdkA ("Kdo kinase A"). Although Kdo phosphorylation may be critical for bacterial virulence of H. influenzae, it does not appear to be required for growth.

Amino Acid Sequence↗
PubMed1999

Potentiation of the action of anandamide on hippocampal slices by the fatty acid amide hydrolase inhibitor, palmitylsulphonyl fluoride (AM 374).

The electrically-evoked release of [3H]acetylcholine from hippocampal brain slices is inhibited by cannabinoid receptor agonists. The effect of palmitylsulphonyl fluoride (AM 374), a recently developed inhibitor of fatty acid amide hydrolase, in influencing the potency of exogenously added anandamide in this preparation was examined. Anandamide alone had relatively little effect on [3H]acetylcholine release. By contrast, in the presence of AM 374 (0.1 microM), anandamide produced a significant inhibition of [3H]acetylcholine release at all concentrations tested (0.1-10 microM). In addition to experiments with AM 374 the effects of N-(4-hydroxyphenyl)arachidonamide (AM 404), a putative anandamide uptake inhibitor, was also examined. However, AM 404 at concentrations up to 10 microM, was not found to significantly enhance the effect of anandamide on electrically-evoked [3H]acetylcholine release. These results indicate that AM 374 potently inhibits endogenous amidase activity and thus facilitates access of exogenous anandamide to cannabinoid receptors in the hippocampal tissue.

Acetylcholine↗
PubMed1999

A PSTTLRE-form of cdc2-like gene in the marine microalga Dunaliella tertiolecta.

To understand the genetic control of algal cell division cycle that pertains to phytoplankton bloom dynamics in the sea, we cloned and analyzed a gene coding for a cyclin-dependent kinase (CDK) for the chlorophyte Dunaliella tertiolecta. The cDNA cloned, 1061 bp long, contained an open reading frame of 314 amino acids. FASTA and GAP analyses showed that this sequence was most homologous to cdc2 out of all known cdks, with an identity of 54-68% and a similarity of 65-76% to cdc2 in higher plants, animals, and yeast. Several signature domains of cdc2 were identified from this sequence, although the PSTAIRE and GDSEID motifs were replaced with PSTTLRE and GDCELQ, respectively. Southern blot hybridization demonstrated that this gene occurred as a single copy in this species, and quantitative RT-PCR showed that the transcription of this gene was constitutive. The present results suggest that the universal cdc2 is conserved in the lower eukaryote with unique structural characteristics.

Algal Proteins↗
PubMed1999

Genetic organization of alpha-bungarotoxins from Bungarus multicinctus (Taiwan banded krait): evidence showing that the production of alpha-bungarotoxin isotoxins is not derived from edited mRNAs.

Two genomic DNAs with a size of approximately 2.8 kb, isolated from the liver of Bungarus multicinctus (Taiwan banded krait), encode the precursors of the long neurotoxins, alpha-Bgt(A31) and alpha-Bgt(V31), respectively. Both genes share virtually identical overall organization with three exons separated by two introns, which were inserted in the same positions in the coding regions of the genes. Moreover, their nucleotide sequences share approximately 98% identity. This result indicates that the two genes co-exist in the genome of B.multicinctus, and probably arose from gene duplication. The exon/intron structures of the alpha-Bgt genes were essentially the same as those reported for the short neurotoxins. This reflects that the long and short neurotoxins should share a common evolutionary origin. Comparative analyses on long neurotoxin and short neurotoxin genes showed that the protein coding regions of the exons were more diverse than the introns except for the signal peptide domain. This implies that the protein coding regions of the neurotoxins may have evolved via accelerated evolution. PCR amplification of venom gland cDNA mixtures revealed that only two amino acid sequences corresponding to alpha-Bgt(A31) and alpha-Bgt(V31) could be deduced from the cDNAs. The results of chromatographic analyses and protein sequencing again emphasized the view that, with the exception of alpha-Bgt(A31) and alpha-Bgt(V31), no other alpha-Bgt isotoxins with amino acid substitutions were present in B.multicinctus venom. In contrast to the proposition of Liu et al. ( Nucleic Acids Res., 1998,26, 5624-5629), our findings strongly suggest that each alpha-Bgt isotoxin is derived from the respective gene, and that alpha-Bgt RNA polymorphism does not originate from one single, intronless gene by the mechanism of RNA editing.

Animals↗
PubMed1999

Crucial role of the interleukin 1 receptor family member T1/ST2 in T helper cell type 2-mediated lung mucosal immune responses.

T1/ST2 is an orphan receptor of unknown function that is expressed on the surface of murine T helper cell type 2 (Th2), but not Th1 effector cells. In vitro blockade of T1/ST2 signaling with an immunoglobulin (Ig) fusion protein suppresses both differentiation to and activation of Th2, but not Th1 effector populations. In a nascent Th2-dominated response, anti-T1/ST2 monoclonal antibody (mAb) inhibited eosinophil infiltration, interleukin 5 secretion, and IgE production. To determine if these effects were mediated by a direct effect on Th2 cells, we next used a murine adoptive transfer model of Th1- and Th2-mediated lung mucosal immune responses. Administration of either T1/ST2 mAb or T1/ST2-Ig abrogated Th2 cytokine production in vivo and the induction of an eosinophilic inflammatory response, but failed to modify Th1-mediated inflammation. Taken together, our data demonstrate an important role of T1/ST2 in Th2-mediated inflammatory responses and suggest that T1/ST2 may prove to be a novel target for the selective suppression of Th2 immune responses.

Allergens↗