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Biomedical subjects

Z Chen

Publications and source records attributed to Z Chen.

At least 595 records · Page 33Linked to original sources
PubMed1999

ASK1 mediates apoptotic cell death induced by genotoxic stress.

ASKI mediates apoptotic cell death induced by genotoxic stress Genotoxic stress-induced apoptosis is mediated by caspase family proteases as triggered by other stimuli. In this study, we found that the DNA-damaging agent cisplatin (cDDP) activated MAP kinase kinase kinase ASK1 and subsequent downstream subgroups of MAP kinase kinase, SEK1 (or MKK4) and MKK3/MKK6, which in turn activated c-Jun N-terminal kinase 1/stress-activated protein kinase (JNK1/SAPK) and p38 MAP kinase prior to caspase family protease activation and the onset of apoptosis in human ovarian carcinoma (OVCAR-3) and human kidney (293T) cells. As reported previously, benzyloxy carbonyl-Asp-CH2OC(O)-2, 6-dichlorobenzene (Z-Asp), a preferential inhibitor of caspase family proteases, blocked the apoptosis of OVCAR-3 cells induced by the genotoxic stress cDDP. Z-Asp, however, did not inhibit ASKI activation and the subsequent kinase cascades. Overexpression of kinase-negative ASK1 (K709R), which inhibited ASK1 activation and the downstream MKK3-p38 and MKK4-JNK1 pathways, also suppressed the caspase protease activation and apoptosis induced by cDDP. These results indicate that the ASK1 pathway is involved in genotoxic stress-induced apoptosis and mediates apoptosis at a step upstream of caspase protease activation.

Apoptosis↗
PubMed1999

Simultaneous determination by capillary gas chromatography of organic acids, sugars, and sugar alcohols in plant tissue extracts as their trimethylsilyl derivatives.

A capillary gas chromatographic (GC) method for the simultaneous determination of organic acids, sugars, and sugar alcohols extracted from plant tissues is described. Plant leaves were extracted in 5% (w/v) perchloric acid and neutralized extracts were purified using C18 cartridges. Organic acids, sugars, and sugar alcohols in purified extracts were converted to their trimethylsilyl (TMS)/TMS-oxime derivatives prior to separation and detection by capillary GC with flame ionization detection (FID). Derivatization procedures were investigated in detail and the compounds of interest were readily converted to their TMS/TMS-oxime derivatives using hexamethyldisiazane reagent in acetonitrile solvent (1:6 v/v) at 100 degreesC for 60 min. The derivatives were sufficiently volatile and stable. The FID response to derivatized compounds was generally linear in the concentration range 30-300 microg ml-1, with detection limits in the order of 3-76 ng. The proposed method was demonstrated for the determination of organic acids, sugars, and sugar alcohols in leaf extracts of two native Australian plants.

Biochemistry↗
PubMed1999

Chromosomal aberrations during progression of chronic myeloid leukemia identified by cytogenetic and molecular cytogenetic tools: implication of 1q12-21.

To study the genomic abnormality underlying the acute transformation of chronic myeloid leukemia (CML), 15 CML patients in blast crisis (BC), 3 in accelerated phase (AP), and 20 in chronic phase (CP) were analyzed by conventional cytogenetics, comparative genomic hybridization (CGH), and dual-color chromosomal painting. Philadelphia (Ph) chromosome was identified in every case studied. Only 5 among 20 CP patients had additional abnormalities while 13 of 18 patients with disease progression (BC + AP) showed extra numerical and/or structural chromosomal aberrations. Cytogenetically, the most common chromosome gains during BC and AP were double or triple Ph chromosomes (5 of 14 cases) and trisomy 8 (5 of 14 cases). Trisomies 7 and 17 (1 of 14 cases each) were also observed. CGH analysis detected genetic imbalances in eight cases. Gains of chromosome 20 (3 cases) and 17q (2 cases) were observed, respectively. The recurrent chromosome loss was the deletion of the short arm of chromosome 17, seen in one case with i(17)(q10) and one case with an unbalanced translocation (1;17). In one case, a very complex chromosomal rearrangement, del(3),del(6),der(6)t(17;3;6),der(17)t(6;17), was seen. A novel finding of this work is the involvement of chromosome 1(q12-21qter) in CML disease progression. Overrepresentation of 1(q12-21qter) region was detected by CGH in one case which had a derivative chromosome 17. This abnormal chromosome was later confirmed by fluorescence in situ hybridization (FISH) painting to be a fusion between chromosome 1 and 17 to form the der(17)t(1;17) (q12-21;p11). Two other cases showed the same region being involved in translocations, t(1;10)(q12-21;q26) and t(1;11)(q12-21;p15). It is possible that one or more genes residing on chromosome 1q12-21 may be important in the acute transformation of CML. In conclusion, we find that the combined use of CGH, chromosome painting, and classic cytogenetic analysis allows a better evaluation of the genomic aberration involved in CML blastic transformation, and offers new directions for its further molecular investigations.

Adult↗
PubMed1999

Improved free R factors for cross-validation of macromolecular structure - importance for real-space refinement.

Improvements in free R cross-validation are based on changed scaling procedures and the use, in map calculation, of estimates of the validation amplitudes which are independent of the actual observed values. The deleterious effects of the omitted test data are mitigated by reduction of the test-set size, which is made possible by constraining test and working sets to share the same scaling coefficients, thereby reducing the degrees of freedom and the dependence of free R on data selection. Further improvements come with use of a modified free R factor, R freeTA. Instead of omitting the validation reflections from map calculation, their amplitudes are replaced by the average of resolution peers that is (nearly) independent of the actual cross-validation amplitudes. The improvements are relevant to model building, phase refinement by density modification and especially to real-space refinement. Although for real data at about 3 A resolution, free R factors of about 0.25 are affected little, the precision of the structure is improved by about 0.1 A. Tests with simulated data show that with good agreement between observed and calculated amplitudes (as in very high resolution studies or simulated refinement tests), free R factors can be improved by factors greater than two.

Crystallography, X-Ray↗
PubMed1999

A minimax entropy registration framework for patient setup verification in radiotherapy.

In external beam radiotherapy (EBRT), patient setup verification over the entire course of fractionated treatment is necessary for accurate delivery of a specified dose to the tumor. We are working on the development of a minimax entropy registration framework for patient setup verification using dual portal images and the treatment planning 3D CT dataset. In this paper, we present an overview of our registration framework, where an iteratively and automatically estimated segmentation of the portal image is utilized to more accurately and robustly register the portal image to the 3D treatment-planning CT data. In addition, we describe initial testing of this approach. We note that, due to low resolution and low contrast of the portal images, this registration presents a difficult problem. We also note that the registration of the images in our proposed method is guided by the bony structure visible in the portal and the 3D CT images. However, since the prostate can move with respect to the pelvic bone, we propose using ultrasound images to quantify this movement.

Algorithms↗
PubMed1999

Constitutive activation of transcription factors NF-(kappa)B, AP-1, and NF-IL6 in human head and neck squamous cell carcinoma cell lines that express pro-inflammatory and pro-angiogenic cytokines.

We previously reported that human head and neck squamous cell carcinomas (HNSCCs) express the pro-inflammatory and pro-angiogenic cytokines interleukin (IL)-1alpha, IL-6, IL-8, and granulocyte-macrophage colony-stimulating factor in vitro and in vivo. The promoter region of the genes encoding these cytokines include binding sites for the transcription factors nuclear factor (NF) kappaB/Rel A, activator protein-1 (AP-1), and CCAAT enhancer-binding protein beta (C/EBPbeta, or NF-IL6), which have been reported to contribute to activation of these cytokine genes. In the study presented here, we examined the activation, composition, and function of these transcription factors in HNSCC cell lines that express pro-inflammatory cytokines, by using electrophoretic mobility shift and reporter-gene assays. Constitutive activation of NF-kappaB, AP-1, and NF-IL6 DNA-binding proteins was detected. Supershift analysis with antibodies specific for NF-kappaB, AP-1, and NF-IL6 binding proteins showed that the NF-kappaB-binding protein included p65/Rel A and p50; AP-1 activity included c-jun, junB, junD, and Fra-1; and NF-IL6 included C/EBPbeta. Mutational analysis of the NF-kappaB, AP-1, and NF-IL6 sites in the IL-8 promoter region showed that NF-kappaB and AP-1 sites contributed to constitutive IL-8 reporter activity in HNSCC. HNSCC lines that exhibited increased IL-8 secretion relative to simian virus 40-immortalized and primary keratinocyte cell lines also demonstrated a concordant increase in NF-kappaB reporter activity relative to nonmalignant keratinocytes. We concluded that the early transcription factors NF-kappaB, AP-1, and NF-IL6 are constitutively activated in human HNSCC cell lines and that NF-kappaB and AP-1 promote expression of the pro-inflammatory and pro-angiogenic cytokine IL-8 in HNSCC. The demonstration of the activation of these transcription factors will be helpful in defining the identity and role of these and other early gene products that contribute to pathogenesis of the malignant phenotype in HNSCC and in defining potential targets for pharmacologic and molecular therapy of HNSCC. Mol. Carcinog. 26:119-129, 1999. Published 1999 Wiley-Liss, Inc.

CCAAT-Enhancer-Binding Proteins↗
PubMed1999

Long-term complications with prostate implants: iodine-125 vs. palladium-103.

The linear quadratic model predicts that the normal tissue biologically effective dose (BED) will be lower with palladium-103 (Pd-103) vs. iodine-125 (I-125) for the currently prescribed minimum tumor doses (MTD) used for I-125 (160 Gy) and Pd-103 (115 Gy) prostate cancer brachytherapy. The predicted BEDs for I-125 and Pd-103 suggest that the long-term complication rates should be lower with Pd-103 vs. I-125 in clinical practice. A review of 123 early stage T1c and T2 prostate cancer patients implanted at Yale University with I-125 (82 patients) or Pd-103 (41 patients) reveals a significantly lower overall complication rate with Pd-103 (0%) vs. I-125 (13%). Most important, the grade III-IV complication rate for Pd-103 was 0% vs. 6% for I-125. The 3-year actuarial probability of remaining free of a long-term complication was 100% for Pd-103 vs. 82% for I-125 (P<0.01). A review of the literature for 992 patients implanted with I-125 vs. 540 patients implanted with Pd-103 shows a consistently higher complication rate for I-125 vs. Pd-103. Assuming that the MTD for Pd-103 may be increased to produce an equivalent late-reacting normal tissue BED to that for I-125, then the radiobiology model predicts the log10 cell kill for Pd-103 implant will be greater than that of an I-125 implant for all tumor doubling times (high-grade tumors and low-grade tumors). The implications of these findings are discussed in terms of future research directions for prostate implants.

Actuarial Analysis↗
PubMed1999

Fast acquisition of quantitative T2 maps.

A rapid new technique called T2 fast acquisition relaxation mapping (T2 FARM) was developed to allow T2 maps to be reconstructed directly from k-space data acquired in 3 sec. A single acquisition measured two sets of k-space data, the first with T1 and T2 weighting independent of phase encode position, and the second with T2 weighting varying with phase encode position. These data were processed using an iterative least squares algorithm to produce a quantitative T2 map from the k-space data. T2 values of phantoms and an egg were determined from T2 FARM maps and spectroscopic Carr-Purcell-Meiboom-Gill (CPMG) measurements, demonstrating the validity of the new technique.

Algorithms↗
PubMed1999

Improved contrast agent bolus tracking using T1 FARM.

Current MRI bolus tracking techniques, such as saturation recovery Turbo-FLASH (srTFL), suffer from signal saturation at high contrast concentrations. In this study T1 Fast Acquisition Relaxation Mapping (T1 FARM) was compared to srTFL. In phantoms, T1 FARM maps were linear with [Gd-DTPA] up to 7.0 mmol/L while srTFL images saturated above 2.0 mmol/L. In the canine left ventricle, blood concentration curves determined from T1 FARM saturated with bolus injections exceeding 0.075 mmol/kg, while srTFL curves saturated above 0.025 mmol/kg of Gd-DTPA. Also, T1 FARM improved contrast-to-noise ratio in tissue concentration curves since higher contrast concentrations could be measured without saturating.

Animals↗
PubMed1999

Correlation between IL-3 receptor expression and growth potential of human CD34+ hematopoietic cells from different tissues.

CD123 (alpha-subunit of IL-3 receptor) expression on primitive and committed human hematopoietic cells was studied by multicolor sorting and single-cell culture. The sources of cells included fetal liver (FLV), fetal bone marrow, umbilical cord blood, adult bone marrow and mobilized peripheral blood. Three subsets of CD34+ cells were defined by the levels of surface CD123: CD123negative, CD123low, and CD123bright. Coexpression of lineage markers showed that a majority of CD34+CD123bright cells were myeloid and B-lymphoid progenitors, while erythroid progenitors were mainly in the CD34+CD123negative subset. The CD34+CD123low subset contained a heterogeneous distribution of early and committed progenitor cells. Single CD34+ cells from the CD123 subsets were cultured in a cytokine cocktail of stem cell factor, interleukin 3 (IL-3), IL-6, GM-CSF, erythropoietin, insulin-like growth factor-1, and basic fibroblast growth factor. After 14 days of incubation, a higher cloning efficiency (CE) was observed in the CD34+CD123negative and CD34+CD123low fractions (37+/-23% and 44+/-23%, respectively) than in the CD34+CD123bright fraction (15+/-21%). Using previously published criteria that colonies containing dispersed, translucent cells (dispersed growth pattern, DGP) were derived from primitive cells and that colonies composed solely of clusters were from committed cells, early precursors were distributed evenly in the CD34+CD123negative and CD34+CD123low subsets. When CD38 and CD90 (Thy-1) were used for further characterization of CD34+ cells from FLV, CE increased from 37+/-23% in CD123negative to 70+/-19% in CD123negativeCD38- and from 44+/-23% in CD123low to 66+/-19% in CD123lowCD38-. No significant increase in CE or DGP progenitors was observed when CD34+ cells were sorted by CD90 and CD123. We concluded that: A) high levels of CD123 were expressed on B-lymphoid and myeloid progenitors; B) early erythroid progenitors had little or no surface CD123, and C) primitive hematopoietic cells are characterized by CD123negative/low expression.

ADP-ribosyl Cyclase↗
PubMed1999

The refolding, purification, and activity analysis of a rice Bowman-Birk inhibitor expressed in Escherichia coli.

A putative rice trypsin/chymotrypsin inhibitor of the Bowman-Birk family, RBBI-8 of about 20 kDa, was expressed in Escherichia coli as a fusion protein bearing an N-terminal (His)6 purification tag. The expressed recombinant protein, rRBBI-8, is insoluble and accumulates as inclusion bodies. The insoluble protein was solubilized in 8 M urea under reducing environment and then refolded into its active conformation under optimized redox conditions. Strategies used to optimize yield and efficiency include selecting the redox system, increasing protein concentration during refolding by adding the denatured protein in a stepwise way, utilizing additives to prevent aggregation, and selecting buffer-exchanging conditions. A Ni-chelate affinity column was then employed to purify the renatured protein. rRBBI-8 shows strong inhibitory activity against trypsin and it can slightly inhibit chymotrypsin. In this study, a refolding and purification system was set up for this cysteine-rich recombinant protein expressed in a prokaryotic system.

Amino Acid Sequence↗
PubMed1999

Successful design and development of genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose from cellulosic biomass to fuel ethanol.

Ethanol is an effective, environmentally friendly, nonfossil, transportation biofuel that produces far less pollution than gasoline. Furthermore, ethanol can be produced from plentiful, domestically available, renewable, cellulosic biomass. However, cellulosic biomass contains two major sugars, glucose and xylose, and a major obstacle in this process is that Saccharomyces yeasts, traditionally used and still the only microorganisms currently used for large scale industrial production of ethanol from glucose, are unable to ferment xylose to ethanol. This makes the use of these safest, most effective Saccharomyces yeasts for conversion of biomass to ethanol economically unfeasible. Since 1980, scientists worldwide have actively been trying to develop genetically engineered Saccharomyces yeasts to ferment xylose. In 1993, we achieved a historic breakthrough to succeed in the development of the first genetically engineered Saccharomyces yeasts that can effectively ferment both glucose and xylose to ethanol. This was accomplished by carefully redesigning the yeast metabolic pathway for fermenting xylose to ethanol, including cloning three xylose-metabolizing genes, modifying the genetic systems controlling gene expression, changing the dynamics of the carbon flow, etc. As a result, our recombinant yeasts not only can effectively ferment both glucose and xylose to ethanol when these sugars are present separately in the medium, but also can effectively coferment both glucose and xylose present in the same medium simultaneously to ethanol. This has made it possible because we have genetically engineered the Saccharomyces yeasts as such that they are able to overcome some of the natural barrier present in all microorganisms, such as the synthesis of the xylose metabolizing enzymes not to be affected by the presence of glucose and by the absence of xylose in the medium. This first generation of genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts relies on the presence of a high-copy-number 2 mu-based plasmid that contains the three cloned genetically modified xylose-metabolizing genes to provide the xylose-metabolizing capability. In 1995, we achieved another breakthrough by creating the super-stable genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts which contain multiple copies of the same three xylose-metabolizing genes stably integrated on the yeast chromosome. This is another critical development which has made it possible for the genetically engineered yeasts to be effective for cofermenting glucose and xylose by continuous fermentation. It is widely believed that the successful development of the stable glucose-xylose-cofermenting Saccharomyces yeasts has made the biomass-to-ethanol technology a step much closer to commercialization. In this paper, we present an overview of our rationales and strategies as well as our methods and approaches that led to the ingenious design and successful development of our genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose to biofuel ethanol.

Biomass↗
PubMed1999

Diagnosis of disorders of platelet function by flow cytometry.

The platelet membrane glycoprotein (GP) IIb/IIIa, GPIX were detected by immunogold assay and flow cytometry, respectively, in two cases of Glannzmann's thrombasthenia (GT). The immunogold assay showed that the GP IIb/IIIa gold granules in GT were decreased obviously, compared with that in normal controls. Flow cytometry results showed that GP IIb/IIIa in two cases of GT was 0.1% and 0.5% of normal control, respectively. While GPIX showed no difference between the GT and normal control. The patients' father showed no bleeding symptoms, whose GP IIb/IIIa accounted for 36.99% of normal control. The experiment results suggested that flow cytometry is a quick, simple and sensitive method for the diagnosis of GP disorders.

Adult↗
PubMed1999

Multidrug resistance P-glycoprotein function of bone marrow hematopoietic cells and the reversal agent effect.

The multidrug resistance P-glycoprotein (P-gp) expression and function in hematopoietic stem/progenitor cells were studied to investigate whether the inhibition of hematopoietic cell P-gp function by multidrug resistance reversal agent increases the cytotoxicity of chemotherapy drugs on the hematopoietic cells. The expression of P-gp on the surface of CD34+ cells from healthy human marrow was examined by flow cytometry. The multidrug resistance reversal agent MS-209 was used to measure the effects of MS-209 on the Rhodamin-123 uptaking of CD34+ hematopoietic cells. By using methylcellulose semi-solid culture, normal human granulocyte-macrophage clonal formation unit (CFU-GM) was cultured. The changes in CFU-GM inhibitory rate caused by daunorubicin were determined in the presence or absence of MS-209. The results showed that the P-gp expression rate of bone marrow CD34+ cells was 13.3%. MS-209 obviously increased the Rhodamin-123 uptake of CD34+ positive cells. The mean inhibitory rate of daunorubicin for CFU-GM was 29.6%, but it was increased to 43.3% in the presence of MS-209 with the difference being significant (P < 0.05). It was concluded that hematopoietic cells expressed P-gp protein and possessed active function. MS-209 could inhibit the membrane efflux pump and increase the cytotoxicity of chemotherapy drugs to the clonal growth of hematopoetic stem cells, suggesting the side effects of these drugs on the hematopoietic system should be taken into consideration in the clinical use.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed1999

Measurement of the activation of IL-2 on bone marrow by flow cytometry.

The changes of cell surface markers before and after activation by IL-2 were detected by flow cytometry (FCM) to establish a more convenient and precise criterion for the judgment of the activation of bone marrow. By using the measurement of the release of lactate dehydrogenase (LDH) the cytotoxicity of mononuclear cells (MNCs) from bone marrow, activated or inactivated, on tumor cell line K562 was evaluated, and at the same time the changes of surface markers on MNCs before and after activation were examined by using FCM. The results showed that the cytotoxicity of MNCs from bone marrow activated by IL-2 on tumor cell line K562 was increased obviously and the number of CD25+ and CD70+ positive cells in bone marrow MNCs was higher than before activation. The enhanced cytotoxicity of MNCs on tumor cell line K562 was synchronous with the increase of the number of CD25+ and CD70+ positive cells in 48 to 72 h. It is more direct, simple and precise to demonstrate the activation of IL-2 on bone marrow by detecting the changes of the amount of the CD25+ and CD70+ positive cells in bone marrow by FCM.

Bone Marrow Cells↗
PubMed1999

Clonal growth of human acute leukemia cells in serum-free methylcellulose medium.

The clonal growth of human acute leukemia cell line (K562) and acute myeloid leukemia cells in the serum-free culture (SFC) was studied in order to establish a SFC system which could replace the effects of serum by using semi-solid methylcellulose culture technique. Our results showed that the clonal growth of K562 cells in semi-solid culture was dependent on exogenous serum. The K562 could be grown in SFC supplemented with 4 major replacing substances. The multifactor and multilevel orthogonal experiment demonstrated that the colony formation was statistically influenced by the 4 replacing substances at various concentrations (P < 0.01). Among them, bovine serum albumin had greatest effect on clonal growth of K562 cells with the optimal concentration being 15 mg/L, followed by transferring, cholesterol and insulin with their optimal concentrations being of 150 mg/L, 7.8 mg/L and 7.0 mg/L respectively. SFC system was formed with the 4 substances at their optimal concentrations. Colony formation of the blast cells in 10 patients with acute myeloid leukemia was observed in this SFC system. There was a heterogeneity of acute myeloid leukemia cells among the 10 patients in response to the growth substances. In SFC system, there was a linear relationship between the number of the clonal formation and the count of the added cells, indicating the colony growth of the cells. Primary acute leukemia cells maintained in SFC system in 10 cases could completely form clones. The colony formation number in some cases in SFC system was more than that of the serum-containing culture. The SFC system could partially replace the serum for study of the clonal formation of human leukemia cells.

Cell Division↗
PubMed1999

Microdissection and microcloning of rye (Secale cereale L.) chromosome 1R.

Chromosome 1R was microdissected and collected from mitotic metaphase spreads of rye (Secale cereale L.) by using glass needles. The isolated chromosomes were amplified in vitro by Sau3A linker adaptor-mediated polymerase chain reaction (PCR). After amplification, the presence of rye-specific DNA was verified by Southern hybridization. The second-round PCR products from five 1R chromosomes were cloned into a plasmid vector to create a chromosome-specific library, which produced approximately 220,000 recombinant clones. Characterization of the microclone library showed that the 172 clones evaluated ranged in size from 300-1800 bp with an average size of 950 bp, of which approximately 42% were medium/high copy and 58% were low/unique copy clones. Chromosome in situ hybridization confirmed that the PCR products from microdissected chromosomes originated from chromosome 1R, indicating that many chromosome 1R-specific sequences were present in the library.

Base Sequence↗