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Biomedical subjects

G Chen

Publications and source records attributed to G Chen.

At least 523 records · Page 29Linked to original sources
PubMed1998

GB virus C/hepatitis G virus infection among patients with hepatocellular carcinoma in the inshore area of the Yangtze River, China.

To investigate the association between GB virus C/hepatitis G virus (GBV-C/HGV) infection and the development of hepatocellular carcinoma (HCC) in H city, in the inshore area of the Yangtze River, where high prevalence of HCC has been reported, we determined hepatitis B virus (HBV) and hepatitis C virus (HCV) markers, GBV-C/HGV-RNA and GBV-C/HGV E2 antibody (anti-HG E2) among 114 HCC patients and the same number of age- and sex-matched controls. There were no significant differences in the clinical and demographic characteristics between them, except for serum alanine aminotransferase level and history of liver diseases. There was a significant difference of hepatitis B virus surface antigen (HBsAg) prevalence between the HCC patients (75.4%) and the controls (20.2%; P<0.01). Hepatitis C virus antibody was detected in 4.4% of the HCC patients, compared with 1.7% of the controls. GB virus-C/HGV-RNA and anti-HG E2 were detected in 14.9 and 1.7% of the HCC patients, respectively, compared with 7.0 and 1.7% of the controls, respectively. Nucleotide sequences and molecular evolutionary analysis showed the strains of GBV-C/HGV-RNA were classified into genotype 2 and 3 (HG and ASIA type). An effect analysis showed an odds ratio (OR) for developing HCC from GBV-C/HGV infection among HBsAg-positive subjects was 14.9, with a 95% CI of 4.9-45.4. HBsAg infection alone was 13.83 (95% CI 7.4-25.9) and GBV-C/HGV infection alone, 3.74 (95% CI 1.1-13.1), respectively. These data indicate that HBV infection is considered to be one of the major risk factors in patients with HCC and although GBV-C/HGV infection was observed in both the HCC and the control groups, it might not play an important role in the development of HCC in this area.

Adult↗
PubMed1998

Biochemical characterization of in vivo alkylating agent resistance of a murine EMT-6 mammary carcinoma. Implication for systemic involvement in the resistance phenotype.

While cancer drug resistance has been extensively studied in cell culture, little is known about more clinically relevant in vivo resistance. The in vivo resistance of a murine mammary carcinoma EMT-6 to alkylating agents was demonstrated in the present study to be associated with multiple biochemical changes. These included an up to 1.5-fold increase in activity of phase II drug metabolizing enzymes (DMEs), such as glutathione (GSH), glutathione reductase (GR), glutathione S-transferase (GST), glutathione peroxidase (GPX) and aldehyde dehydrogenase (ALDH), and an up to 88% decrease of phase I DME activity [7-ethoxycumarin O-deethylase (ECOD), P450 reductase (PR)] in the resistant tumors compared with the parental tumor. Transplant of either parental or resistant tumors to mice was accompanied by a decrease of both phase I and phase II DME activity in the livers of female Balb/C mice compared with the non-tumor mice. Moreover, at the protein level, while cytochrome P450 (CYP) IIB1/2 in the liver of mouse bearing both the sensitive and the resistant tumor was significantly diminished compared to that in the liver of non-tumor control mouse in Western analysis, there was actually an increase of this protein in the liver of the host bearing either of the two resistant tumors compared to that of the sensitive tumor-bearing animal. Although this in vivo resistance phenotype is not expressed in cell culture, the profile of most of the enzyme changes in the resistant tumors remained similar in in vitro culture of the isolated tumor cells. Collectively, these results demonstrate that this in vivo alkylating agent resistance is associated with multiple changes of both phase I and phase II DMEs in the resistant tumors, and some of these, such as CYP IIB1/2 protein are further altered in the resistant tumor-bearing mouse liver, suggesting a potential role of systemic factors in this resistance phenotype.

7-Alkoxycoumarin O-Dealkylase↗
PubMed1997

Cloning and functional characterization of mouse IkappaBepsilon.

The biological activity of the transcription factor NF-kappaB is mainly controlled by the IkappaB proteins IkappaBalpha and IkappaBbeta, which restrict NF-kappaB in the cytoplasm and enter the nucleus where they terminate NF-kappaB-dependent transcription. In this paper we describe the cloning and functional characterization of mouse IkappaBepsilon. Mouse IkappaBepsilon contains 6 ankyrin repeats required for its interaction with the Rel proteins and is expressed in different cell types where we found that it is up-regulated by NF-kappaB inducers, as is the case for IkappaBalpha and human IkappaBepsilon. IkappaBepsilon functions as a bona fide IkappaB protein by restricting Rel proteins in the cytoplasm and inhibiting their in vitro DNA binding activity. Surprisingly, IkappaBepsilon did not inhibit transcription of genes regulated by the p50/p65 heterodimer efficiently, such as the human interferon-beta gene. However, IkappaBepsilon was a strong inhibitor of interleukin-8 expression, a gene known to be regulated by p65 homodimers. In addition, IkappaBepsilon appears to function predominantly in the cytoplasm to sequester p65 homodimers, in contrast with the other two members of the family, IkappaBalpha and IkappaBbeta, which also function in the nucleus to terminate NF-kappaB-dependent transcriptional activation.

Amino Acid Sequence↗
PubMed1997

The E1B 19K/Bcl-2-binding protein Nip3 is a dimeric mitochondrial protein that activates apoptosis.

Nip3 (nineteen kD interacting protein-3) is an E1B 19K and Bcl-2 binding protein of unknown function. Nip3 is detected as both a 60- and 30-kD protein in vivo and in vitro and exhibits strong homologous interaction in a yeast two-hybrid system indicating that it can homodimerize. Nip3 is expressed in mitochondria and a mutant (Nip3(163)) lacking the putative transmembrane domain and COOH terminus does not dimerize or localize to mitochondria. Transient transfection of epitope-tagged Nip3 in Rat-1 fibroblasts and MCF-7 breast carcinoma induces apoptosis within 12 h while cells transfected with the Nip3(163) mutant have a normal phenotype, suggesting that mitochondrial localization is necessary for induction of cell death. Nip3 overexpression increases the sensitivity to apoptosis induced by granzyme B and topoisomerase I and II inhibitors. After transfection, both Nip3 and Nip3(163) protein levels decrease steadily over 48 h indicating that the protein is rapidly degraded and this occurs in the absence of cell death. Bcl-2 overexpression initially delays the onset of apoptosis induced by Nip3 but the resistance is completely overcome in longer periods of incubation. Nip3 protein levels are much higher and persist longer in Bcl-2 expressing cells. In conclusion, Nip3 is an apoptosis-inducing dimeric mitochondrial protein that can overcome Bcl-2 suppression.

Adenovirus E1B Proteins↗
PubMed1997

Conversion of dorsal from an activator to a repressor by the global corepressor Groucho.

The Dorsal morphogen acts as both an activator and a repressor of transcription in the Drosophila embryo to regulate the expression of dorsal/ventral patterning genes. Circumstantial evidence has suggested that Dorsal is an intrinsic activator and that additional factors (corepressors) convert it into a repressor. These corepressors, however, have previously eluded definitive identification. We show here, via the analysis of embryos lacking the maternally encoded Groucho corepressor and via protein-binding assays, that recruitment of Groucho to the template by protein:protein interactions is required for the conversion of Dorsal from an activator to a repressor. Groucho is therefore a critical component of the dorsal/ventral patterning system.

Animals↗
PubMed1997

Apoptosis in gastric epithelial cells is induced by Helicobacter pylori and accompanied by increased expression of BAK.

Carriage of the bacterium H. pylori in the human stomach is associated with evidence of increased epithelial cell apoptosis. This may be of significance in the etiology of gastritis, peptic ulcers, and neoplasia. The ability of H. pylori to directly induce epithelial apoptosis was examined in vitro by fluorescence and electron microscopy, flow cytometry, and DNA fragmentation ELISA. The induction of apoptosis by H. pylori was time and concentration-dependent and inhibited by preventing direct bacterial-epithelial cell contact. Apoptosis was accompanied by increased expression of Bak, with little change in expression of other Bcl-2 family proteins. The expression of Bak was also increased in gastric biopsies from patients colonized by H. pylori. Thus, H. pylori induces gastric epithelial cell apoptosis, by a Bak-dependent pathway.

Apoptosis↗
PubMed1997

Oncogenic transformation potentiates apoptosis, S-phase arrest and stress-kinase activation by etoposide.

The exact mechanisms for the selective toxicity of chemotherapeutic drugs against tumor cells are not fully understood. We designed a series of experiments to test the possibility that the positive proliferative signal initiated by oncogenes might change the sensitivity for apoptosis induction by the anticancer drug etoposide (VP16), an inhibitor of topoisomerase II (Topo II). Treatment with VP16 induced significantly increased apoptosis in NIH3T3 cells transformed by oncogenic src, ras or raf, compared with the normal 3T3 cells. Apopototic changes involved nuclear DNA fragmentation, morphological alterations and decreased viability. Furthermore it was shown that stress-activated protein kinase (SAPK) was activated much more strongly in all three transformed lines compared to untransformed cells by VP16 treatment, while slight activation of extracellular signal-regulated kinase (ERK1) was observed in all four cell lines. In addition, the transformed cells displayed arrest in mid-S-phase following the treatment, whereas NIH3T3 cells were primarily arrested in late S and G2/M phase. Finally, the cyclin-dependent kinase inhibitor p21 WAF1 was induced in all four cell lines, although induction of p53 was not detected in any of these cell lines. Taken together our results demonstrated that oncogenic transformation can sensitize the cells to apoptosis induction, stress kinase activation and cell cycle arrest in response to VP16 treatment. These results may have important implications for understanding the selective toxicity of anti-cancer drugs in tumor cells.

3T3 Cells↗
PubMed1997

Photo-immobilization of epidermal growth factor enhances its mitogenic effect by artificial juxtacrine signaling.

Photo-reactive epidermal growth factor (EGF) was synthesized by coupling EGF with azidobenzoic acid and was immobilized onto the wells of a polystyrene culture plate by photo-irradiation. The photo-immobilized EGF enhanced the growth of anchorage-dependent cells more than native or azidobenzoyl derivatized EGF. A small amount of photo-immobilized EGF was sufficient to enhance the growth of cells and the maximal mitogenic effect was greater than that of native or derivatized EGF. On the other hand, the photo-immobilized EGF did not enhance growth of anchorage-independent cells. In addition, signal transduction in the cells adhered only on the EGF-immobilized surface was observed by staining of phosphotyrosine residues by anti-phosphotyrosine antibodies. These results showed that the enhanced cell growth was due to direct interaction between the cells and the immobilized EGF. Photo-immobilization could be a universal means of fixing growth factors onto an artificial matrix that is devoid of chemically functional groups scaffolding growth factors and could provide a new tool to elucidate signal transduction mechanism and could lead to the development of a new protein-free cell culture system or tissue engineering materials.

Animals↗
PubMed1997

Patterns of failure in children with medulloblastoma: effects of preirradiation chemotherapy.

PURPOSE: To evaluate the effects of preirradiation chemotherapy on patterns of failure in children with medulloblastoma. METHODS AND MATERIALS: Fifty-three patients (pts) with medulloblastoma were given preirradiation chemotherapy as initial postoperative treatment at St. Jude Children's Research Hospital from November 1984 to September 1993. Patients < or = 3 years of age (n = 23) received chemotherapy (CH) with delayed craniospinal irradiation (CSI). Children > or = 3 years with more advanced disease (T3b-T4, M+ or measurable residual after resection) were given CH followed by CSI (30 patients). Chemotherapy regimen depended on protocol, but usually included cis- or carboplatin and etoposide, +/- cyclophosphamide and vincristine. RESULTS: Actuarial overall survival and event-free survival rates are 60% (95% confidence interval [41,79]) and 37% [19,55] at 5 years. Children < or = 3 at diagnosis: six of 23 pts completed CH without progression and received consolidative CSI; all six are alive with no evidence of disease (NED) at 2.4-9.1 years. Seventeen patients progressed during CH and were then given CSI. Sites of progression during CH were posterior fossa (PF) in 11 patients, neuraxis (NEUR) in 4, and PF+NEUR in 2. Following CSI, 7 patients are alive NED at 2.0-8.6 years; 10 patients died of progressive disease. Eleven patients had M0 disease at diagnosis; 8 (73%) progressed during CH, 3 in the neuraxis. Children > or = 3 at diagnosis: 20 of 30 patients completed pre-CSI CH without progression; 15 are alive NED at 1.3-9.2 years, and 5 showed post-CSI progression in the PF (n = 3), in the NEUR (n = 1) and in bone marrow (n = 1). Ten of the 30 (33%) patients progressed on CH (6 in NEUR, 4 in PF); 5 are alive and NED or with stable disease. Seventeen patients had M0 disease at diagnosis; 3 out of 17 (18%) progressed during CH, 2 in NEUR and 1 in an extraneural site. In the total group of 30 patients, 11 have had disease recurrence after completion of XRT. The actuarial rate of failure was 23 +/- 9% for the patients < or = 3 years of age and 21 +/- 8% for the older children when evaluated at 4 months after diagnosis (at the completion of chemotherapy in the older children but during the ongoing chemotherapy in the younger children). CONCLUSIONS: In patients presenting with M0 disease and receiving pre-CSI chemotherapy, the risk of neuraxis progression seems to increase with duration of chemotherapy. The sites of progression during preirradiation chemotherapy are nearly equally divided between posterior fossa and other neuraxis sites. CSI salvage of patients progressing on chemotherapy is possible in approximately 50% of patients. Following CSI, neuraxis progression is more frequent than posterior fossa relapse.

Adolescent↗
PubMed1997

Direct antidiabetic effect of leptin through triglyceride depletion of tissues.

Leptin is currently believed to control body composition largely, if not entirely, via hypothalamic receptors that regulate food intake and thermogenesis. Here we demonstrate direct extraneural effects of leptin to deplete fat content of both adipocytes and nonadipocytes to levels far below those of pairfed controls. In cultured pancreatic islets, leptin lowered triglyceride (TG) content by preventing TG formation from free fatty acids (FFA) and by increasing FFA oxidation. In vivo hyperleptinemia, induced in normal rats by adenovirus gene transfer, depleted TG content in liver, skeletal muscle, and pancreas without increasing plasma FFA or ketones, suggesting intracellular oxidation. In islets of obese Zucker Diabetic Fatty rats with leptin receptor mutations, leptin had no effect in vivo or in vitro. The TG content was approximately 20 times normal, and esterification capacity was increased 3- to 4-fold. Thus, in rats with normal leptin receptors but not in Zucker Diabetic Fatty rats, nonadipocytes and adipocytes esterify FFA, store them as TG, and later oxidize them intracellularly via an "indirect pathway" of intracellular fatty acid metabolism controlled by leptin. By maintaining insulin sensitivity and preventing islet lipotoxicity, this activity of leptin may prevent adipogenic diabetes.

3-Hydroxybutyric Acid↗
PubMed1997

Mechanistic studies on the aminopeptidase from Aeromonas proteolytica: a two-metal ion mechanism for peptide hydrolysis.

The aminopeptidase from Aeromonas proteolytica (AAP) is uncompetitively inhibited by fluoride ion at pH 8.0 with an inhibition constant (Ki) of 30 mM. Thus, fluoride inactivates AAP only after substrate binding, and only a single fluoride ion binds to AAP. On the other hand, chloride ion does not inhibit AAP up to concentrations of 2 M at pH 8.0. The pH dependence of fluoride inhibition of AAP was measured over the pH range 6.0-9.5. Between pH values of 6.0 and 9.0, fluoride ion acts as a pure uncompetitive inhibitor of AAP, and the Ki increases from 1.2 to 370 mM. From a plot of pKi vs pH, a pKa value of 7.0 +/- 0.3 was extracted which corresponds to a single deprotonation process. At pH values higher than 9.0, the fluoride inhibition pattern changes to competitive. This change in inhibition pattern was attributed to a change in ionic strength or perhaps pH of the solution since fluoride ion was also found to become a competitive inhibitor of AAP at pH 8.0 in the presence of 2 M NaCl. These data, taken together with previous kinetic studies of mono- and dinuclear hydrolases with fluoride ion, suggest that a Zn(II)-bound water/hydroxide exists at the dimetal active site of AAP with a pKa of 7.0 and that this water/hydroxide acts as the active site nucleophile. The hydrolysis of L-leucine-p-nitroanilide was measured spectrophotometrically in triplicate between 25 and 85 degrees C at eight substrate concentrations ranging from 5 to 800 microM. From these data, Km values were derived at each temperature studied and were found to increase exponentially with increasing temperature. Moreover, the calculated Vmax values were also found to increase over this temperature range, mimicking the Km values. An Arrhenius plot was constructed from k(cat) values and was found to be linear over the temperature range 25-85 degrees C, indicating that the rate-limiting step in AAP peptide hydrolysis is product formation and does not change as a function of temperature. From the slope of the line, the activation energy (Ea) was calculated to be 36.5 kJ/mol. The enthalpy and entropy of activation at 25 degrees C calculated over the temperature range 298-358 K were found to be 34.0 kJ/mol and -94.2 J/(mol x K), respectively. The free energy of activation at 25 degrees C was found to be 62.1 kJ/mol. Combination of the available X-ray crystallographic data with the present kinetic and thermodynamic results, as well as the previously reported kinetic and spectroscopic data, has allowed a detailed catalytic mechanism for AAP to be proposed.

Aeromonas↗
PubMed1997

Multidrug-resistant human sarcoma cells with a mutant P-glycoprotein, altered phenotype, and resistance to cyclosporins.

A variant of the multidrug-resistant human sarcoma cell line Dx5 was derived by co-selection with doxorubicin and the cyclosporin D analogue PSC 833, a potent inhibitor of the multidrug transporter P-glycoprotein. The variant DxP cells manifest an altered phenotype compared with Dx5, with decreased cross-resistance to Vinca alkaloids and no resistance to dactinomycin. Resistance to doxorubicin and paclitaxel is retained. The multidrug resistance phenotype of DxP cells is not modulated by 2 microM PSC 833 or cyclosporine. DxP cells manifest a decreased ability to transport [3H]cyclosporine. DNA heteroduplex analysis and sequencing reveal a mutant mdr1 gene (deletion of a phenylalanine at amino acid residue 335) in the DxP cell line. The mutant P-glycoprotein has a decreased affinity for PSC 833 and vinblastine and a decreased ability to transport rhodamine 123. Transfection of the mutant mdr1 gene into drug-sensitive MES-SA sarcoma cells confers resistance to both doxorubicin and PSC 833. Our study demonstrates that survival of cells exposed to doxorubicin and PSC 833 in a multistep selection occurred as a result of a P-glycoprotein mutation in transmembrane region 6. These data suggest that Phe335 is an important binding site on P-glycoprotein for substrates such as dactinomycin and vinblastine and for inhibitors such as cyclosporine and PSC 833.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Patterned artificial juxtacrine stimulation of cells by covalently immobilized insulin.

Insulin was immobilized on prescribed areas of poly(ethylene terephthalate) film by photolithography. Chinese hamster ovary cells overexpressing human insulin receptors were cultured on the pattern-immobilized film. Pattern-immobilized insulin did not enhance cell adhesion but transduced a signal to the cells through phosphorylation of tyrosine residues of signal proteins. In addition, only cells on immobilized insulin grew in medium containing no serum. The enhancement of cell growth was considered to be a consequence of signal transduction. This microprocessing technique is useful for investigating the effect of immobilized biosignal molecules and for constructing new tissue engineering materials.

Animals↗
PubMed1997

Adsorption Enthalpy and Adsorption Isotherm of Tetradecylpyridinium Bromide on Na-Montmorillonite

The adsorption enthalpy and the adsorption isotherm of tetradecylpyridinium bromide (TPB) onto a Na-montmorillonite suspension and the dilution enthalpy of the TPB solution have been determined at 298.15 K. The dilution process is endothermic and the adsorption process is exothermic. For ion exchange adsorption, the interaction between TP+ and the montmorillonite is much stronger than that between Na+ and the montmorillonite because the cumulative adsorption enthalpy increases dramatically with adsorption amount. For molecular adsorption, however, the interaction between TPB and the clay is much weaker.

Journal Article↗
PubMed1997

In vitro scanning saturation mutagenesis of an antibody binding pocket.

We have combined PCR mutagenesis with in vitro transcription/translation and ELISA for the rapid generation and characterization of antibody mutants. The PCR products are used directly as the template for the in vitro transcription/translation reactions and because no cloning steps are required, the in vitro saturation mutagenesis of one residue can be completed in duplicate within a week by a single investigator. In vitro scanning saturation mutagenesis was used to analyze the role and plasticity of six key contact residues (H:Tyr-33, H:Asn-35, H:Tyr-50, H:Trp-100, L:Val-94, and L:Pro-96) in the binding pocket of a single chain Fv antibody derived from the 26-10 monoclonal antibody. A total of 114 mutant antibodies were produced; all 19 substitutions at each of the 6 chosen positions. The mutants were analyzed for binding to digoxin, digitoxin, digoxigenin, and ouabain resulting in the generation of a comprehensive data base of 456 relative affinity values. Excellent agreement between the relative affinity values obtained with in vitro synthesized mutant antibodies and equilibrium affinity data obtained with previously reported purified mutant monoclonal antibodies was observed. Approximately 75% of the single amino acid mutants exhibited significant binding to one or more of the digoxin analogs. Mutations that alter and, in some cases, reverse specificity for the different digoxin analogs were identified. In vitro scanning saturation mutagenesis represents a new tool for protein structure-function and engineering studies and can be interfaced with laboratory automation so that an even higher throughput of protein mutants can be constructed and analyzed.

Binding Sites, Antibody↗
PubMed1997

Guanine nucleotide binding proteins in opioid-dependent patients.

Guanine nucleotide binding (G) protein levels and functioning in the platelets of 19 methadone-maintained patients were compared to age and sex matched, normal controls. We found that in the methadone patients, G alpha s-levels were significantly higher, while the levels of G alpha i 1/2 and pertussis toxin catalyzed [32P]ADP ribosylation were significantly lower compared to control subjects in platelet membranes. We have further found that when all three of these biochemical indicators were combined in a discriminant function analysis, 79% of the methadone patients were correctly classified and 83% of the controls were correctly classified.

Adenosine Diphosphate Ribose↗
PubMed1997

Inhibition of lymphocyte function by head and neck carcinoma cell line soluble factors.

BACKGROUND: Immunosuppression in patients with head and neck cancer is well recognized. Previous investigations have demonstrated graded immunosuppression that becomes more pronounced as lymphocyte activity is measured closer to the primary neoplasm. In fresh tumors, a soluble factor has been identified that may partly account for the observed graded immunosuppression. OBJECTIVE: To examine the effect of soluble factors produced by head and neck sqamous cell carcinoma cell lines on the generation of lymphokine activated killer cell cytotoxicity and peripheral blood lymphocyte proliferation induced by interleukin 2, concanavalin A, and phytohemagglutinin. DESIGN: Conditioned supernatant fluids were generated in a 4-day incubation period, using 5 head and neck squamous cell carcinoma cell lines, and were assayed for the ability to inhibit the generation of lymphokine activated killer cell cytotoxicity and naive peripheral blood lymphocyte proliferation induced by interleukin 2, concanavalin A, and phytohemagglutinin. RESULTS: All conditioned supernatant fluids significantly inhibited the generation of lymphokine activated killer cell cytotoxicity relative to controls, and this inhibition was dose dependent. In contrast, supernatant fluids from a myelogenous leukemic tumor cell line, K562, and an ovarian epithelial cell line, SKOV-3, failed to inhibit cytotoxicity. Supernatant fluids conditioned with head and neck squamous cell carcinoma cell lines also profoundly inhibited the naive peripheral blood lymphocyte proliferation induced by interleukin 2, concanavalin A, and phytohemagglutinin. CONCLUSIONS: These studies demonstrate that the cell lines of head and neck squamous cell carcinoma produce soluble factors that inhibit lymphocyte function. Furthermore, these experiments suggest that the inhibition previously observed with enzymatically disaggregated fresh tumors is due to factors produced by the tumor cells rather than by other cells within the tumor matrix.

Biological Factors↗