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Biomedical subjects

Q Chen

Publications and source records attributed to Q Chen.

At least 523 records · Page 29Linked to original sources
PubMed1996

Establishment, characterization and drug sensitivity of four new human soft tissue sarcoma cell lines.

Four new cell lines were established from patients with soft tissue sarcomas. Drug sensitivity as well as genotypic characterization, which may be related to drug sensitivity in these cell lines, was determined. Karyotype, H-ras, c-myc and mutant p53 gene expression, Rb, G1- and S-phase cyclins, E2F and major cyclin/CDK inhibitors such as p16 and p21 and p-glycoprotein were analyzed using cytogenetic, Northern blot and immunological methods. Drug sensitivity was determined using growth inhibition tests. These cell lines differed in their morphology and growth rates, forming colonies in soft agar with a cloning efficiency of 4.3-13.4%, and 3 of the 4 cell lines grew in nude mice. Cytogenetic analysis of cell lines revealed highly aneuploid karyotypes. Deletion and/or translocation of chromosome 17 was seen in HS-16, HS-18 and HS-30 cells, and both copies of chromosome 13 were lost or re-arranged in the HS-18 cell line. Mutant p53 protein was present in all 4 cell lines. HS-18 cells showed no expression of the Rb protein and high levels of expression of E2F, cyclin A, cyclin E and CDK2. HS-16 expressed a higher level of cyclin D than the other 3 cell lines. p21WAF1 expression was seen in all cell lines, but p16ink4 was expressed only in HS-30 and HS-42 cell lines. These cell lines were sensitive to taxol and relatively resistant to methotrexate, vinblastine and 5-fluorouracil when compared with the fibrosarcoma cell line HT-1080. These new cell lines should provide a useful model for the study of soft tissue sarcomas and for evaluating new drugs or treatments.

Animals↗
PubMed1996

Identification of tyrosine 187 as a protein kinase C-delta phosphorylation site.

Protein kinase C-delta (PKC-delta) has been demonstrated to be phosphorylated on tyrosine residue(s) in many different biological systems (Li, W., Yu, J.-C., Michieli, P., Beeler, J. F., Ellmore, N., Heidaran, M. A., and Pierce, J. H. (1994) Mol. Cell. Biol. 14, 6727-6735; Li, W., Mischak, H., Yu, J.-C., Wang, L.-M., Mushinski, J. F., Heidaran, M. A., and Pierce, J. H. (1994) J. Biol. Chem. 269, 2349-2352; Denning, M. F., Dlugosz, A. A., Howett, M. A., and Yuspa, S. H. (1993) J. Biol. Chem. 268, 26079-26081). Tyrosine phosphorylation of PKC-delta has also been shown to occur in vitro when purified PKC-delta is coincubated with different tyrosine kinase sources. However, the tyrosine phosphorylation site(s) is currently unknown and the exact effect of this phosphorylation on its serine/threonine kinase activity and biological functions is still controversial. To directly investigate the potential role of PKC-delta tyrosine phosphorylation, tyrosine 187 was converted to phenylalanine (PKC-deltaY187F) by site-directed mutagenesis, and expression vectors containing PKC-deltaY187F cDNAs were transfected into both 32D myeloid progenitor cells and NIH 3T3 fibroblasts. The results showed that tyrosine 187 of PKC-delta became phosphorylated in vivo in response to 12-O-tetradecanoylphorbol-13-acetate stimulation or platelet-derived growth factor receptor activation. In vivo labeling and subsequent two-dimensional phosphopeptide analysis demonstrated that one phosphopeptide was absent in PKC-deltaY187F when compared to wild type PKC-delta, further substantiating that tyrosine 187 of PKC-delta is phosphorylated in vivo. Although the phosphotyrosine content of PKC-deltaY187F was reduced compared with PKC-deltaWT, the kinase activity of PKC-deltaY187F toward a PKC-delta substrate was not altered. Moreover, 12-O-tetradecanoylphorbol-13-acetate-mediated monocytic differentiation of 32D cells was not affected by expression of the PKC-deltaY187F mutant. Taken together, these results suggest that tyrosine phosphorylation of PKC-delta on 187 may not influence PKC-delta activation and known functions.

3T3 Cells↗
PubMed1996

Secretagogues increase the expression of surfactant protein A receptors on lung type II cells.

Since secretagogues have been shown to increase the internalization of surfactant phospholipid and protein by lung cells, we postulated that their action occurred through a mechanism involving increased surfactant protein A (SP-A) receptor density. Therefore, we evaluated the influence of secretagogues on the binding of iodinated SP-A to alveolar type II cells. Type II cells were isolated from rat lung and maintained in primary culture for 18 h on Transwell membranes. Upon exposure to 8-bromo-cyclic AMP (cAMP, 0.1 mM), phorbol 12-myristate 13-acetate (PMA, 10 nM), terbutaline (0.1 mM), or ATP (1 mM), the binding of SP-A increased 1.5-2-fold. This stimulation was cell substrate-dependent since type II cells plated on plastic dishes did not show this effect. A time course of the stimulation of SP-A binding due to secretagogues showed that both cAMP and PMA increased SP-A binding by 2-fold after 20 min. With cAMP, binding remained elevated for 2 h, while binding in the presence of PMA had returned to control values. The effects of submaximal concentrations of cAMP and PMA on binding were additive. Inhibition of cellular protein synthesis with cycloheximide did not alter the increase of SP-A binding stimulated by the secretagogues. Type II cells pretreated with PMA responded to subsequent treatment with cAMP by increasing SP-A binding, while these cells were refractory to subsequent treatment with PMA. Both constitutive and regulated binding of SP-A to type II cells were sensitive to trypsin. The binding of SP-A to type II cells showed saturation at a concentration of 1 microg/ml SP-A under control and secretagogue-stimulated conditions, with both total and calcium-dependent binding showing a 2-fold increase upon secretagogue exposure. The data are consistent with the hypothesis that secretagogues stimulate surfactant uptake, at least in part, through recruitment of SP-A receptors to the type II cell surface, resulting in an increase in the number of SP-A binding sites.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1996

Polymerization of three hemoglobin A2 variants containing Val6 and inhibition of hemoglobin S polymerization by hemoglobin A2.

To understand determinants for hemoglobin (Hb) stability and Hb A2 inhibition of Hb S polymerization, three Valdelta6 Hb A2 variants (Hb A2 deltaE6V, Hb A2 deltaE6V,deltaQ87T, and Hb A2 deltaE6V, deltaA22E,deltaQ87T) were expressed in yeast, and stability to mechanical agitation and polymerization properties were assessed. Oxy forms of Hb A2 deltaE6V and Hb A2 deltaE6V,deltaQ87T were 2- and 1.6-fold, respectively, less stable than oxy-Hb S, while the stability of Hb A2 deltaE6V,deltaA22E,deltaQ87T was similar to that of Hb S, suggesting that Aladelta22 and Glndelta87 contribute to the surface hydrophobicity of Hb A2. Deoxy Hb A2 deltaE6V polymerized without a delay time, like deoxy Hb F gammaE6V, while deoxy Hb A2 deltaE6V,deltaQ87T and deoxy Hb A2 deltaE6V,deltaA22E,deltaQ87T polymerized after a delay time, like deoxy Hb S, suggesting that beta87 Thr is required for the formation of nuclei. Deoxy Hb F gammaE6V,gammaQ87T showed no delay time and required a 3.5-fold higher concentration than deoxy Hb S for polymerization, suggesting that Thr effects on Valdelta6 Hb A2 and Valgamma6 Hb F variants are different. Mixtures of deoxy Hb S/Hb A2 deltaE6V,deltaQ87T polymerized, like deoxy Hb S, while polymerization of Hb S/Hb A2 deltaE6V mixtures was inhibited, like Hb S/Hb F gammaE6V mixtures. These results suggest alpha2betaSdelta6 Val, 87 Thr hybrids and Hb A2 deltaE6V,deltaQ87T participate in Hb S nucleation, while only 50% of alpha2betaSdelta6 Val hybrids and none of the Hb A2 deltaE6V participate. These findings are in contrast to those of mixtures of Hb S with Hb F gammaE6V or Hb F gammaE6V,Q87T, which both inhibit Hb S polymerization. Our results also suggest participation in nucleation of some alpha2betaSdelta hybrids in A2S mixtures but not alpha2betaSgamma hybrids in FS mixtures.

Animals↗
PubMed1996

Properties of low density lipoproteins relevant to oxidative modifications change paradoxically during aging.

Atherosclerosis is a common problem among the elderly. Because lipid peroxidation is considered a contributor to the development of atherosclerosis, we compared oxidative properties of lipoproteins in an otherwise healthy (SENIEUR-classified) aged population (65-74 years) with young controls (18-30 years). Relative amounts of oxidatively altered low density lipoprotein (LDL), estimated by means of an antibody against LDL modified by 4-hydroxynonenal, a product of lipid peroxidation, were increased marginally in serum from the elderly (9.8 vs. 7.4%, P = 0.07). In contrast, isolated LDL from the elderly revealed a decreased susceptibility to in vitro oxidation: the lag time was increased (2.34 vs. 2.10 h, P < 0.01), and the maximal rate of LDL oxidation decreased (0.88 vs. 1.01 O.D./h, P = 0.001). However, there were no age-related changes in lipid composition of native LDL and consumption of fatty acids during in vitro oxidation. The serum concentrations of ascorbic acid and most lipophilic anti-oxidants (the latter expressed per g serum lipids) were significantly decreased in the elderly except tocopherols which tended to be higher. In conclusion, our data reveal paradox age-related alterations of LDL as to its behaviour in oxidation in vivo vs. in vitro.

Adolescent↗
PubMed1996

A functional phosphatidylinositol 3,4,5-trisphosphate/phosphoinositide binding domain in the clathrin adaptor AP-2 alpha subunit. Implications for the endocytic pathway.

Clathrin-coated pits are sites of concentration of ligand-bound signaling receptors. Several such receptors are known to recruit, bind, and activate the heterodimeric phosphatidylinositol-3-kinase, resulting in the generation of phosphatidylinositol 3,4, 5-trisphosphate. We report here that dioctanoyl-phosphatidylinositol-3,4,5-P3 binds specifically and saturably to soluble AP-2 and with greater affinity to AP-2 within assembled coat structures. Soluble -myo-inositol hexakisphosphate shows converse behavior. Binding to bovine brain clathrin-coated vesicles is evident only after detergent extraction. These observations and evidence for recognition of the diacylglyceryl backbone as well as the inositol phosphate headgroup are consistent with AP-2 interaction with membrane phosphoinositides in coated vesicles and with soluble inositol phosphates in cytoplasm. A discrete binding domain is identified near the N terminus of the AP-2 alpha subunit, and an expressed fusion protein containing this sequence exhibits specific, high affinity binding that is virtually identical to the parent protein. This region of the AP-2 alpha sequence also shows the greatest conservation between a Caenorhabditis elegans homolog and mammalian alpha, consistent with a function in recognition of an evolutionarily unchanging low molecular weight ligand. Binding of phosphatidylinositol 3,4, 5-trisphosphate to AP-2 inhibits the protein's clathrin binding and assembly activities. These findings are discussed in the context of the potential roles of phosphoinositides and AP-2 in the internalization and trafficking of cell surface receptors.

Adaptor Proteins, Vesicular Transport↗
PubMed1996

Antisense RNA, fur, iron, and the regulation of iron transport genes in Vibrio anguillarum.

The negative regulation of the expression of iron transport genes fatA and fatB in Vibrio anguillarum is mediated by a chromosome-encoded Fur protein and a plasmid pJM1-derived antisense RNA (RNAalpha), which is preferentially expressed under iron-rich conditions. In this work, we characterized the RNAalpha promoter region, and by using promoter fusion and rifampicin experiments we were able to demonstrate that iron regulates RNAalpha synthesis posttranscriptionally by stabilizing RNAalpha half-life rather than enhancing transcription initiation. The Fur protein is also essential for RNAalpha synthesis at the transcription initiation level, independently of the iron status of the cell. From experiments assessing the relative contribution of Fur and RNAalpha, we were able to show that RNAalpha may indeed play an important role on the negative regulation of the expression of the iron transport genes under physiological conditions.

Bacterial Outer Membrane Proteins↗
PubMed1996

Integrin-mediated activation of MEK and mitogen-activated protein kinase is independent of Ras [corrected].

The integrins are a family of cell surface receptors that mediate adhesive interactions with the extracellular matrix and also generate signals that influence cell growth and differentiation. Ligation and clustering of integrins causes activation and autophosphorylation of focal adhesion kinase (FAK), a cytoplasmic tyrosine kinase, and results in the transient activation of p42 and p44 mitogen-activated protein (MAP) kinases. Initial evidence has suggested that the integrin signaling pathway may share common elements with the canonical Ras signal transduction cascade activated by peptide mitogens such as epidermal growth factor (EGF). In this report we demonstrate that Raf-1 and MAP or extracellular signal-related kinase kinase (MEK), key cytoplasmic kinases of the Ras cascade, are activated subsequent to integrin-mediated adhesion of mouse NIH 3T3 fibroblasts. We also show that MAP kinase is downstream of MEK in the integrin signaling pathway. However, in contrast to the receptor tyrosine kinase signaling cascade, integrin-mediated signal transduction seems to be largely independent of Ras. Dominant negative inhibitors of Ras-dependent signaling failed to block integrin-mediated activation of MEK. In addition, while treatment with the peptide mitogen EGF clearly increased GTP-loading of Ras, little effect was observed in response to integrin-dependent cell adhesion. Thus, integrin-mediated activation of MEK and MAP kinase in 3T3 cells occurs primarily by a mechanism that is distinct from the Ras signal transduction cascade.

3T3 Cells↗
PubMed1996

Oct-4 regulates alternative platelet-derived growth factor alpha receptor gene promoter in human embryonal carcinoma cells.

Expression of the platelet-derived growth factor alpha-receptor (PDGFalphaR) gene is tightly controlled in mammalian embryogenesis. A well established model system to study human embryogenesis is the embryonal carcinoma cell line Tera2. We have shown previously that retinoic acid-differentiated Tera2 cells express two PDGFalphaR transcripts of 6.4 kilobase pairs (kb) (encoding the full-length receptor) and 3.0 kb, respectively, whereas in contrast, undifferentiated Tera2 cells express PDFGalphaR transcripts of 1.5 kb and 5.0 kb. Here we show that this switch in PDGFalphaR expression pattern during differentiation of Tera2 cells results from alternative promoter use. In undifferentiated cells, a second promoter is used, which is located in intron 12 of the PDGFalphaR gene. Functional analysis shows that this promoter contains a consensus octamer motif, which can be bound by the POU domain transcription factor Oct-4. Oct-4 is expressed in undifferentiated Tera2 cells but not in retinoic acid-induced differentiated cells. Mutation of the octamer motif decreases promoter activity, while ectopic expression of Oct-4 in differentiated Tera2 cells specifically enhances the activity of this PDGFalphaR promoter. Therefore, we suggest that an important aspect in the maintenance of the undifferentiated state of human embryonal carcinoma cells results from Oct-4 expression, which thereupon activates this PDGFalphaR promoter.

Alternative Splicing↗
PubMed1996

Spectroscopic recognition of guanine dimeric hairpin quadruplexes by a carbocyanine dye.

Isolated guanine quadruplex structures have been described at high resolution both in solution and in the solid state. The existence of this unusual DNA structure in vivo and its biological significance remain to be determined. We describe the binding of 3,3'-diethyloxadicarbocyanine to dimeric hairpin guanine quadruplexes. This interaction results in a set of unique spectrophotometric signatures, none of which arises from binding to single strands or Watson-Crick duplexes. These unique signatures include a new absorbance peak (lambda max = 534 nm), an induced circular dichroism (lambda = 534-626 nm), a quenching of the dye fluorescence upon excitation with visible light, and strong energy transfer from DNA. This last effect provides the basis for detecting hairpin quadruplex structures in the presence of excess amounts of nonquadruplex DNA structures, such as single strands and Watson-Crick duplexes. The mechanism of quadruplex recognition by this dye is discussed, along with the possibility of using this dye as a probe for hairpin quadruplex structures in vitro and in vivo.

Base Sequence↗
PubMed1996

Cortical activation in confrontation naming.

Functional magnetic resonance imaging was used to detect cortical activation in the right and left perisylvian cortex of seven young adult right-handed volunteers in response to a letter fluency task and to a visual naming task using standardized line drawings. Both letter fluency and visual naming activated left dorsolateral prefrontal cortex (Brodmann's areas 6, 9, 44 and 45). Only visual naming activated area 37 (a cortical region with strong connections to visual association areas), visual association area 19, and areas 39 and 21 previously shown to activate with auditory semantic tasks. This study supports a role for area 37 as participant in a visual lexicosemantic processing network which may otherwise overlap the auditorysemantic network.

Adult↗
PubMed1996

Laminoplasty with foraminotomy for coexisting cervical myelopathy and unilateral radiculopathy: a preliminary report.

STUDY DESIGN: An assessment was made of the efficacy of a combined laminoplasty and foraminotomy operation for patients with coexisting myelopathy and unilateral radiculopathy. The procedure was done in 17 patients. OBJECTIVES: The patients were followed with lateral flexion and extension radiographs, computed tomography scans, and an assessment system specially designed to qualitatively evaluate the patients' neurologic status. Follow-up period averaged 4 years (range, 2.1-9.3 years). SUMMARY OF BACKGROUND DATA: Excellent-to-good results were obtained for 76% (13 of 17) of the patients without any significant functional compromise based on the radiographs. Sixteen nerve roots were decompressed with a less than 25% foraminotomy, whereas eight were decompressed by a 25%-50% foraminotomy without serious neurologic damage, except for one patient. The neurologic results appeared unrelated to the extent of foraminotomy. METHODS: A refined procedure for combined laminoplasty and foraminotomy was reviewed retrospectively in terms of neurologic outcome and radiographic data. RESULTS: The present series is small, and results are not comparable directly with other methods. The procedure appears effective for myelopathy and radiculopathy. This procedure is applicable to patients with myelopathy and coexisting nerve root impingement anterolaterally or in the neural foramen. CONCLUSION: The combined laminoplasty and foraminotomy operation may provide greater neurologic improvement in patients with coexisting myelopathy and unilateral radiculopathy, while maintaining cervical spine stability after surgery.

Adult↗
PubMed1996

Lumbosacral dumb-bell neurilemmoma approached by microsurgical interlaminar foraminotomy. A case report.

The case history of a patient with a dumb-bell neurilemmoma arising from the L5 nerve root is described. The tumour extended into the outlet of the neural foramen at L5-S1 on the left and was also compressing the dural sac and the S1 nerve root. A limited laminotomy at L4-5 and L5-S1, with preservation of the neural arch, was followed by microsurgical medial foraminotomy at L5-S1 which allowed visualisation of the nerve root and enucleation of the tumour. The technique maintains stability of the posterior elements and spinal movement, and is recommended for the removal of this type of tumour.

Adult↗
PubMed1996

Cellular expression of ionotropic glutamate receptor subunits on specific striatal neuron types and its implication for striatal vulnerability in glutamate receptor-mediated excitotoxicity.

Glutamate receptors are composed of subtype-specific subunits. Variation in the precise subunit composition of a receptor may result in significant functional differences. Thus, a precise knowledge of subunit composition on striatal neurons is a prerequisite for understanding the selective vulnerability of striatal neurons to excitatory amino acids. In the present study, we used an immunohistochemical double-labelling approach to localize ionotropic glutamate receptor subunits (NMDAR1, GluR1, GluR2/3, GluR4 and GluR5/6/7) on specific striatal neuron populations. Our results showed that striatal cholinergic and somatostatin interneurons were not labelled for the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate, receptor subunits GluR1, GluR2/3 and GluR4. Most cholinergic and somatostatin interneurons (83.3% to 100%), however, were double-labelled for the N-methyl-D-aspartate receptor subunit NR1 and kainic acid receptor subunits GluR5/6/7. All parvalbumin interneurons were labelled for GluR1 and GluR4, and 96% GluR1 positive and 95% GluR4 positive neurons were also double-labelled as parvalbumin interneurons. About half of all parvalbumin interneurons co-localized with GluR2/3, and over 97% were labelled for NR1 and GluR5/6/7. Among striatal projection neurons, enkephalin-positive (mainly striatopallidal) neurons, striatonigral neurons (mainly substance P-positive) and calbindin-positive matrix neurons were not immunostained for GluR1 or GluR4. In contrast, 95% to 100% of each of these types of projection neurons were double-labelled for NR1, GluR2/3 and GluR5/6/7. Our results demonstrate that striatal neuron types differ in their expression of ionotropic glutamate receptor subunits and subtypes. The clear difference between striatal interneurons and projection neurons in ionotropic glutamate receptor subtypes/subunits supports the idea that differential glutamate receptor expression mechanism may account for the selective vulnerability of striatal projection neurons to excitotoxicity, and that glutamate receptor-mediated excitotoxicity may be involved in the striatal neurodegenerative diseases.

Animals↗
PubMed1996

Mapping drug-induced changes in cerebral R2* by Multiple Gradient Recalled Echo functional MRI.

A multiple Gradient Recalled Echo MRI sequence was used to map spatial and temporal changes in the rate of MR signal decay (R2*) in response to L-3,4-dihydroxyphenylalanine (levodopa) in the striatal dopaminergic system of a rhesus monkey unilaterally lesioned with 4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP). R2* decreased significantly in the right (dopamine depleted) putamen and caudate following levodopa. More focal areas of smaller R2* decline were also observed in these structures in the left hemisphere. The observed spatial and temporal patterns of R2* change support the view that the method is monitoring changes in neural activity.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1996

Time-resolved fluorescence studies of tomaymycin bonding to synthetic DNAs.

Tomaymycin reacts covalently with guanine in the DNA minor groove, exhibiting considerable specificity for the flanking bases. The sequence dependence of tomaymycin bonding to DNA was investigated in synthetic DNA oligomers and polymers. The maximum extent of bonding to DNA is greater for homopurine and natural DNA sequences than for alternating purine-pyrimidine sequences. Saturation of DNA with tomaymycin has little effect on the melting temperature in the absence of unbound drug. Fluorescence lifetimes were measured for DNA adducts at seven of the ten unique trinucleotide bonding sites. Most of the adducts had two fluorescence lifetimes, representing two of the four possible binding modes. The lifetimes cluster around 2-3 ns and 5-7 ns; the longer lifetime is the major component for most bonding sites. The two lifetime classes were assigned to R and S diastereomeric adducts by comparison with previous NMR results for oligomer adducts. The lifetime difference between binding modes is interpreted in terms of an anomeric effect on the excited-state proton transfer reaction that quenches tomaymycin fluorescence. Bonding kinetics of polymer adducts were monitored by fluorescence lifetime measurements. Rates of adduct formation vary by two orders of magnitude with poly(dA-dG).poly(dC-dT), reacting the fastest at 4 x 10(-2) M-1 s-1. The sequence specificity of tomaymycin is discussed in light of these findings and other reports in the literature.

Antibiotics, Antineoplastic↗