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Biomedical subjects

X Chen

Publications and source records attributed to X Chen.

At least 325 records · Page 18Linked to original sources
PubMed2001

Multicolor fluorescent differential display.

Differential display and DNA microarray have emerged as the two most popular methods for gene expression profiling. Here, we developed a multicolor fluorescent differential display (FDD) method that combines the virtues of both differential display in signal amplification and DNA microarray in signal analysis. As in DNA microarray, RNA samples being compared can be labeled with either a red or green fluorescent dye and displayed in a single lane, allowing convenient scoring and quantification of the differentially expressed messages. In addition, the multicolor FDD has a built-in signal proofreading capability that is achieved by labeling each RNA sample from a comparative study with both red and green fluorescent dyes followed by their reciprocal mixings in color. Thus, the multicolor FDD provides a platform upon which a sensitive and accurate gene expression profiling by differential display can be automated and digitally analyzed. It is envisioned that cDNAs generated by the multicolor FDD may also be used directly as probes for DNA microarray, allowing an integration of the two most widely used technologies for comprehensive analysis of gene expression.

Animals↗
PubMed2001

Universal SNP genotyping assay with fluorescence polarization detection.

The degree of fluorescence polarization (FP) of a fluorescent molecule is a reflection of its molecular weight (Mr). FP is therefore a useful detection methodfor homogeneous assays in which the starting reagents and products differ significantly in Mr. We have previously shown that FP is a good detection method for the single-base extension and the 5'-nuclease assays. In this report, we describe a universal, optimized single-base extension assay for genotyping single nucleotide polymorphisms (SNPs). This assay, which we named the template-directed dye-terminator incorporation assay with fluorescence polarization detection (FP-TDI), uses four spectrally distinct dye terminators to achieve universal assay conditions. Even without optimization, approximately 70% of all SNP markers tested yielded robust assays. The addition of an E. coli ssDNA-binding protein just before the FP reading significantly increased FP values of the products and brought the success rate of FP-TDI assays up to 90%. Increasing the amount of dye terminators and reducing the number of thermal cycles in the single-base extension step of the assay increased the separation of the FP values benveen the products corresponding to different genotypes and improved the success rate of the assay to 100%. In this study the genomic DNA samples of 90 individuals were typed for a total of 38 FP-TDI assays (using both the sense and antisense TDI primers for 19 SNP markers). With the previously described modifications, the FP-TDI assay gave unambiguous genotyping data for all the samples tested in the 38 FP-TDI assays. When the genotypes determined by the FP-TDI and 5'-nuclease assays were compared, they were in 100% concordance for all experiments (a total of 3420 genotypes). The four-dye-terminator master mixture described here can be used for assaying any SNP marker and greatly simplifies the SNP genotyping assay design.

Alkaline Phosphatase↗
PubMed2001

BindingDB: a web-accessible molecular recognition database.

This paper presents an initial description of the BindingDB, a public web-accessible database of measured binding affinities for various molecular types (http://www.bindingdb.org). The BindingDB allows queries based upon a range of criteria, including chemical similarity or substructure, sequence homology, numerical criteria (e.g. delta G(o) < 5 kcal/mol) and reactant names (e.g. "lysozyme"). Principles of Human-Computer Interactions are being employed in creating the query interface and user-feedback is being solicited. The data specification includes significant experimental detail. A full dictionary has been created for isothermal titration calorimetry data in consultation with experimentalists and data dictionaries for enzyme-inhibition and other measurement techniques are being developed. Currently, the BindingDB contains several data sets of broad interest, such as antigen-antibody binding and cyclodextrin/small molecule binding. However, it is anticipated that online deposition by experimentalists will ultimately contribute to a larger flow of data. We are actively developing software and file specifications to facilitate such deposition.

Binding Sites↗
PubMed2001

The relationship between new stroke onset and serum thiocyanate as an indicator to cigarette smoking.

The case subjects were 67 stroke patients (27 males and 40 females, mean age 65.7+/-7.1), who suffered from stroke attack and had participated in annual medical check-up between 1987 and 1988 at Oyabe Public Health Center. The controls, which were adjusted for sex, age and blood pressure level with the case subjects, were selected from participants attending their annual medical check-up in 1987-1988. The serum thiocyanate level of case subjects were significantly higher than that of controls (p<0.05). The usefulness of measurement of serum thiocyanate level as an indicator of smoking status was assessed by using multiple logistic regression analysis adjusting for body mass index, mean blood pressure, and serum cholesterol was found. An odds ratio of 3.00 (95% confidence interval: 1.06-8.48, p<0.05) in cerebral infarction. It is considered that serum thiocyanate may be useful as an indicator of smoking status to assess the relationship with stroke onset, especially for cerebral infarction.

Age Distribution↗
PubMed2001

Effects of free fatty acids on gluconeogenesis and autoregulation of glucose production in type 2 diabetes.

Effects of endogenously derived free fatty acids (FFAs) on rates of gluconeogenesis (GNG) (determined with 2H2O), glycogenolysis (GL), and endogenous glucose production (EGP) were studied in 18 type 2 diabetic patients and in 7 nondiabetic control subjects under three experimental conditions: 1) during an 8-h fast (from 16-24 h after the last meal), when plasma FFA levels increased slowly; 2) during 4 h (from 16-20 h) of nicotinic acid (NA) administration (fasting plus NA), when plasma FFAs decreased acutely; and 3) during 4 h (from 20-24 h) after discontinuation of NA (FFA rebound), when plasma FFAs increased acutely. During fasting, FFAs increased from 636 to 711 micromol/l in type 2 diabetic patients and from 462 to 573 micromol/l in control subjects (P < 0.04), but GNG did not change in diabetic patients (6.9 vs. 6.5 micromol x kg(-1) x min(-1), P > 0.05) or in control subjects (5.1 vs. 5.4 micromol x kg(-1) x min(-1), P > 0.05). During fasting plus NA, FFAs decreased in diabetic patients and control subjects (from 593 to 193 and from 460 to 162 micromol/l, respectively); GNG decreased (from 6.1 to 4.2 and from 4.7 to 3.5 micromol x kg(-1) x min(-1)), whereas GL decreased in diabetic patients (from 5.3 to 4.4 micromol x kg(-1) x min(-1)) but increased in control subjects (from 5.4 to 7.2 micromol x kg(-1) min(-1)). During the FFA rebound, FFAs increased in diabetic patients and control subjects (from 193 to 1,239 and from 162 to 1,491 micromol/l, respectively); GNG increased (from 4.2 to 5.4 and from 3.4 to 5.3 micromol x kg(-1) x min(-1) respectively), and GL decreased (from 4.4 to 3.4 and from 7.3 to 4.3 micromol x kg(-1) x min(-1), respectively). In summary, during an extended overnight fast, increasing plasma FFA levels stimulated GNG, whereas decreasing FFA levels inhibited GNG in both diabetic and control subjects; 20 h after the last meal, approximately one-third of GNG in both diabetic and control subjects was dependent on FFAs; and autoregulation of EGP by GL in response to decreasing GNG was impaired in diabetic patients.

Diabetes Mellitus, Type 2↗
PubMed2001

Internalisation of the protease-activated receptor 1: role of the third intracellular loop and of the cytoplasmic tail.

To analyse the mechanisms of PAR-1 internalisation, we constructed several PAR-1 mutants and stably expressed them in CHO cells. Our study shows that the Ser(306)-->Ala mutation (S306A), which eliminates a potential site of phosphorylation by PKC in the third intracellular loop of PAR-1, did not change the rate of phosphorylation but reduced the rate of thrombin-induced internalisation of the PAR-1 mutant (58 versus 78% of membrane PAR-1 in 15 min, p<0.005). Deletion of the last 43 amino acid residues of the PAR-1 cytoplasmic tail completely suppressed the thrombin phosphorylation of the mutated receptor and significantly reduced its internalisation upon activation. This deletion also inhibited the PMA-induced and the agonist-independent internalisation of the receptor. The Tyr(371)--> Ala mutation (Y371A), in a NPXXY motif of the seventh transmembrane domain of the receptor had no effect on the receptor behaviour. Our results indicate that both the C-tail and the third intracellular loop are involved in PAR-1 internalisation induced by thrombin while only the C-tail plays a role in the PMA-induced and in the agonist-independent PAR-1 internalisation.

Alanine↗
PubMed2001

Role of the third intracellular loop and of the cytoplasmic tail in the mitogenic signaling of the protease-activated receptor 1.

The activation of the protease-activated receptor 1 (PAR-1) by thrombin has been shown to induce an activation of the MAP kinase cascade and to stimulate cell proliferation. To examine the mechanisms of signal transduction by PAR-1, we constructed several PAR-1 mutants which were stably expressed in CHO cells. When compared to wild-type PAR-1, mutation of Ser306-->Ala (S306A) in the third intracellular loop of PAR-1 inhibited MAP kinase activation and cell proliferation stimulated by thrombin. The thrombin activation of MAP kinase was inhibited by pertussis toxin, suggesting a role for a Gi-like protein. As shown by calcium signaling and inosotol trisphosphate generation, the Ser306-mutated PAR-1 induced a strong activation of phospholipase C after thrombin addition. Deletion of the cytoplasmic tail of PAR-1 also inhibited thrombin-induced DNA synthesis but the MAP kinase pathway was activated as with wild-type PAR-1. In contrast, the deletion of the C-tail of PAR-1 prevented almost completely the activation of the phospholipase C pathway. Taken together these results suggest that the C-tail of PAR-1 is a critical site for PAR-1 coupling to phospholipase C activation, while the third intracellular loop of PAR-1 is implicated in PAR-1 coupling to Gi and MAP kinase activation. In addition, these results also show that MAP kinase activation is necessary but not sufficient for thrombin to induce cell proliferation.

Animals↗
PubMed2001

[Genetic analysis and gene deduction of powdery mildew resistance in T. durum-Ae. squarrosa amphidiploids].

Eleven seedlings with resistance and two amphidiploids with disease resistance at all developmental stage were screened by using No. 15 isolate of E. graminis f. sp. tritici to inoculate 99 T. durum-Ae. squarrosa amphidiploids. The two amphidiploids with disease resistance at all developmental stage, M53 and M81, and their Ae. squarrosa parents were resistant to No. 15 isolate of powdery mildew payhogen, but their T. durum parents were susceptible. Therefore, the powdery mildew resistance of M53 and M81 were derived from Ae. squarrosa. Amphidiploid M74, which had the same durum parent as M53 but different Ae. squrrosa parent, was susceptible to No. 15 isolate in adult stage. M29 and M35, which had the same durum parent as M81 but different Ae. squarrosa parent were susceptible in all stage. These results were also proved that the powdery mildew resistance genes were derived from Ae. sqarrosa. Genetic analysis showed that powdery mildew resistance gene in M53 or M81 was a single dominant gene. A series of lines with known powdery mildew resistance genes and two unknown lines were used to test response patterns with 14 differential isolates of E. graminis f. sp. tritici. The response patterns in M53 and M81 were different from Pm1 to Pm21 and indicated that M53 and M81 conferred a new powdery mildew resistance gene respectively.

Genes, Plant↗
PubMed2001

[Comparative survey of cardiac arrests during anesthesia and surgery in the 1980 s and 1999 s].

A survey of unexpected cardiac arrests, excluding those associated with cardiac surgery, that had occurred during anesthesia and surgery in the period 1980-1999 was conducted. There was no significant difference between the number of such cardiac arrests that occurred in the 1980 s (29/36,159, 0.080%) and the number in the 1990 s (33/37,643, 0.088%). According to the classification by Keenan et al., there were 3 cases (0.0083%) in the 80 s and 4 cases (0.0106%) in the 90 s that occurred due to anesthetic management. Cardiac arrests due to surgery itself significantly decreased from 21 cases in the 80 s to 6 cases in the 90 s, probably due to proper treatment for massive bleeding, a decrease in the number of cases of anaphylactic shock, and the development of novel anesthetics. Conversely, cardiac arrests due to preoperative conditions of the patients significantly increased from 5 cases in the 80 s to 23 cases in the 90 s. This increase seems to be due to an increase in the number of severe and multiple injuries and an increase in complicated major surgery. The increase in number of cases due to preoperative conditions also depends on coronary spasm and cardiac conduction insufficiency. Taking into consideration the improvement in intraoperative monitoring and the development of novel anesthetics in the 90 s, greater efforts should be made by anesthesiologists to reduce the incidence of cardiac arrest due to anesthetic management, and preoperative evaluation of surgical patients needs to be reconsidered.

Anesthesia↗
PubMed2001

[Effects of homocysteine on post-implantation rat embryo cultured in vitro].

In order to investigate the effect of homocysteine(HCY) on the development of rat embryos, the post-implantation whole embryo culture(WEC) technique was used. Neural plate stage(GD9.5) rat embryos were explanted in rat serum medium(immediately centrifugal serum, ICS) with D,L-HCY(0,0.15, 1.5,2.0,4.0,6.0,8.0,10.0 mmol/L), and cultured for 48 hours. The results showed that HCY did cause damage to embryonic development and the damages of HCY on embryos were characterized in a significant dose-response pattern. The minimum teratogenic dose of D,L-HCY was 0.15 mmol/L, the incidence of dysmorphogenic embryos was 8.33%. Significant inhibiting effects of HCY on yolk sac(including reduced yolk sac diameter, shriveled surface, small or defective yolk sac vessels) and embryonic growth and morpholological differentiation were apparent with increased HCY, while HCY were 4 mmol/L or more (P < 0.05). The abnormalities included neural tube defects, delayed cardiac tube formation, pericardial effusion, incomplete flexion, small or missing forelimb buds and irregular somites, etc. These findings suggested that HCY might exert a direct effect on embryos and a "double effect" both on yolk sac and embryos.

Animals↗
PubMed2001

Prognostic significance of periodic acid-Schiff-positive patterns in primary cutaneous melanoma.

The patterns of periodic acid-Schiff (PAS) staining of extracellular matrix in histological sections of certain melanomas may be predictive of outcome. Recent in vitro and molecular genetic data suggest that the appearance of these patterns in both uveal and cutaneous melanoma is a function of aggressive tumor cells. We studied 96 patients with primary cutaneous melanomas treated at the University of Illinois at Chicago who were monitored for disease-free survival. Survival probabilities were determined by Kaplan-Meier estimates, and prognostic factors were evaluated by multivariate analysis. By univariate analysis, there was a significant decrease in disease-free survival among patients whose tumors contained parallel with cross-linking or network patterns (PXNs; P = 0.0070). Stepwise regression with Cox models that included the combinations of the PAS-positive patterns, tumor thickness, female gender, ulceration, and age yielded a model with thickness and the PAS-positive parallel with cross-linking or networks. Despite the relatively small sample size in this study, the detection of the PAS-positive parallel with cross-linking or networking in cutaneous melanoma was associated with a decrease in disease-free outcome. Additional studies of the prognostic significance of these patterns is warranted on larger data sets.

Adolescent↗
PubMed2001

[Extraction of MUAP from NEMG signal using self-organization competing NN].

The pattern classification of motor unit action potential (MUAP) of needle electrode electromyogram (NEMG) signal by means of self-organization competing Neural Network(NN) has been accomplished in this paper. The parameters and the power of excited white-noise of auto-regressive (AR) model are taken as the feature of MUAP. The results of simulated NEMG and real NEMG all show that this method of classification is very effective and correct, and thus it presents a new approach to the extraction of MUAP template in the study of decomposition of NEMG.

Action Potentials↗
PubMed2001

Determination of nucleic acids with crystal violet by a resonance light-scattering technique.

For the first time, Crystal Violet (CV) was used to determine nucleic acid concentrations using the resonance light-scattering (RLS) technique. Based on the enhancement of the RLS of CV by nucleic acids, a new quantitative determination method for nucleic acids in aqueous solutions has been developed. At pH 5.03 and ionic strength 0.005 mol kg-1, the interaction of CV with nucleic acids results in three characteristic RLS peaks at 344.0, 483.0 and 666.0 nm. With 4.0 x 10(-5) mol l-1 of CV, linear relationships were found between the enhanced intensity of RLS at 666.0 nm and the concentration of nucleic acids in the range 0-2.5 micrograms ml-1 for herring sperm DNA, 0-4.0 micrograms ml-1 for calf thymus DNA and 0-4.5 micrograms ml-1 for yeast RNA. The limits of determination were 13.8 ng ml-1 for herring sperm DNA, 36.8 ng ml-1 for calf thymus DNA and 69.0 ng ml-1 for yeast RNA. The assay is convenient, rapid, inexpensive and simple.

Animals↗
PubMed2001

[Changes in sedative level during induction of anesthesia using a single volatile anesthetic].

We investigated the changes in sedative level during induction of anesthesia using a single volatile anesthetic, sevoflurane, compared with the changes when propofol and sevoflurane were used. We used a bispectral index (BIS) monitor (Aspect, Newton, USA) to monitor the sedative level. Thirty patients were randomly divided into one group receiving sevoflurane alone (S group, n = 15) and another group receiving propofol and sevoflurane (PS group, n = 15). The S group received 5% sevoflurane with nitrous oxide 6 l.min-1 and oxygen 3 l.min-1 via a face mask, while the PS group received propofol 1.5 mg.kg-1 i.v. followed by step-wise increasing sevoflurane up to 5%. Induction times, as assessed by loss of eyelash reflex, were 32 s in the PS group and 65 s in the S group. The decrease in the BIS value was smooth in the S group, whereas it increased transiently and varied greatly in the PS group. Two patients in the PS group showed body movement during induction of anesthesia, and the BIS values in both cases transiently exceeded 60. BIS monitor is useful for monitoring the sedative level during induction of anesthesia, and induction using a single volatile anesthetic such as sevoflurane is smooth.

Adult↗
PubMed2001

[The expression of tPA directed by the bovine BLG regulatory elements in the mammary gland of transgenic mice].

In order to get the regulatory elements which are essential for generating mammary gland bioreactors, the whole 8.4 kb bovine BLG gene was obtained by PCR amplification. The 1.6 kb chicken lysozyme matrix attachment region (MAR) was used to overcome position effects. The bovine BLG-tPA expression vector was constructed and the BLG-tPA fusion gene was introduced into fertilized eggs of mice by microinjection to generate transgenic mouse. 170 offsprings were obtained, of which 9 were proved to be transgenic mice based on PCR and Southern-blot analysis. The tPA expression level amounted to 12 micrograms/mL in the milk of mice. The bovine BLG-tPA fusion gene integrated in the founders was inheritable.

Animals↗
PubMed2001

[Establishment of ELISA with TT virus(TTV) and its application to patients with hepatitis and healthy people].

OBJECTIVE: To establish the EIA method for detection of antibody to TT virus (anti-TTV) and to investigate the distribution of anti-TTV in patients with hepatitis and in healthy people. METHODS: The ELISA method using the recombinant expressed product of truncated gene of TTV ORF1 as antigen was developed and was applied to detect anti-TTV in patients with hepatitis and in healthy control. The TTV DNA were detected by nested-PCR. RESULTS: The positive rates of anti-TTV antibodies in different populations were as follows: 1.3% in healthy people, 10.5% in patients with hepatitis A, 12.5% in patients with hepatitis B, 8.3% in patients with hepatitis C, 7.7% in patients with hepatitis D, 12.0% in patients with hepatitis E, 6.5% in patients with hepatitis G, 32.3% in patients with hepatitis non-A to G. The positive rate of TTV DNA was significantly related to that of anti-TTV (P < 0.05). CONCLUSION: The anti-TTV antibodies were detected positive in patients with hepatitis and healthy people, however, its positive rate in patients with hepatitis non-A to G was significantly higher than that in other groups. The fact that anti-TTV antibodies coexist with TTV DNA suggests that the antibody to TTV ORF1 antigen may be an infectious marker of TTV similar to that of anti-HCV.

Antibodies, Viral↗