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Biomedical subjects

Xiao-Jun Yan

Publications and source records attributed to Xiao-Jun Yan.

15 recordsLinked to original sources
PubMed2006

Detection of H. pylori antibody profile in serum by protein array.

AIM: To detect multiple H. pylori antibodies in serum samples of individuals who carry H. pylori by protein array. METHODS: Recombinant H. pylori antigens, urease B subunit (UreB), vacuolating toxin A (VacA) and cytotoxin associated gene A protein (CagA), were prepared and immobilized in matrixes on nitrocellulose membrane by robotics to bind the specific immunoglobulin G (IgG) antibodies in serum. Staphylococcus protein A (SPA) labeled by colloid gold was used to integrate the immuno-complex and gave red color signal. The scanner based on charge-coupled device (CCD) could collect the image signal and convert it into digital signal. RESULTS: When human IgG was printed on the membrane in increasing concentrations and incubated with immunogold, a linear dose response curve was obtained and the detection limit for IgG was about 0.025 ng. The cutoff values, which were defined as the mean grey level plus 3 times of standard deviation, were 27.183, 28.546 and 27.402, for anti-UreB IgG, anti-CagA IgG and anti-VacA IgG, respectively, as 400 human serum samples with negative H. pylori antibodies were detected by the protein array. When 180 serum samples from patients in hospital were employed for detection of IgG against UreB, CagA and VacA, the sensitivity of the protein array was 93.4%, 95.4%, 96.0%, and the specificity was 94.8%, 94.4% and 97.5%, respectively, as compared with the results obtained by ELISA. The assay also showed high reproducibility, uniformity and stability, and the results were available within 30 min. CONCLUSION: The protein array is a very practical method for rapid detection of multiple antibodies in serum samples. It is especially useful for large scale epidemiological investigation of the infection of H. pylori.

Antibodies, Bacterial↗
PubMed2006

Fascaplysin, a selective CDK4 inhibitor, exhibit anti-angiogenic activity in vitro and in vivo.

PURPOSE: This study was to evaluate the correlation of two important strategies, namely, cell cycle proliferation arrest and anti-angiogenesis. We chose fascaplysin, a marine natural product with selective CDK4 selective inhibition activity, to study its potential anti-angiogenesis effects in vivo and in vitro. METHODS: Chorioallantoic membrane (CAM) assay was initially used as an in vivo approach to evaluate anti-angiogenic activity of fascaplysin. In addition, human umbilical vein endothelial cell (HUVEC) line was used to further confirm the anti-angiogenic activity of fascaplysin in vitro. To explore the mechanism of anti-angiogenesis, we examined the effect of fascaplysin on vascular endothelial growth factor (VEGF) expression and secretion by hepatocarcinoma cells BeL-7402. RESULTS: The results of CAM assay suggested fascaplysin inhibited capillary plexus formation in a dose-dependent manner and suppressed VEGF in cross section. Moreover, the in vitro assay also confirmed that fascaplysin provided selective inhibition of endothelial cells proliferation towards tumor cells in low concentration. The immunocytochemical staining and ELISA verified fascaplysin could inhibit VEGF expression and secretion by BeL-7402. CONCLUSIONS: These findings strongly suggest that fascaplysin is a natural angiogenesis inhibitor.

Angiogenesis Inhibitors↗
PubMed2006

A simple parallel analytical method of prenatal screening.

Protein microarray has progressed rapidly in the past few years, but it is still hard to popularize it in many developing countries or small hospitals owing to the technical expertise required in practice. We developed a cheap and easy-to-use protein microarray based on dot immunogold filtration assay for parallel analysis of ToRCH-related antibodies including Toxoplasma gondii, rubella virus, cytomegalovirus and herpes simplex virus type 1 and 2 in sera of pregnant women. It does not require any expensive instruments and the assay results can be clearly recognized by the naked eye. We analyzed 186 random sera of outpatients at the gynecological department with our microarray and commercial ELISA kit, and the results showed there was no significant difference between the two detection methods. Validated by clinical application, the microarray is easy to use and has a unique advantage in cost and time. It is more suitable for mass prenatal screening or epidemiological screening than the ELISA format.

Dose-Response Relationship, Drug↗
PubMed2006

Corrosion resistance of a laser spot-welded joint of NiTi wire in simulated human body fluids.

The purpose of this study was to investigate corrosion resistance of a laser spot-welded joint of NiTi alloy wires using potentiodynamic tests in Hank's solution at different PH values and the PH 7.4 NaCl solution for different Cl- concentrations. Scanning electron microscope observations were carried out before and after potentiodynamic tests. The composition of a laser spot-welded joint and base metal were characterized by using an electron probe microanalyzer. The results of potentiodynamic tests showed that corrosion resistance of a laser spot-welded joint of NiTi alloy wire was better than that of base metal, which exhibited a little higher breakdown potential and passive range, and a little lower passive current density. Corrosion resistances of a laser spot-welded joint and base metal decreased with increasing of the Cl- concentration and PH value. The improvement of corrosion resistance of the laser spot-welded joint was due to the decrease of the surface defects and the increase of the Ti/Ni ratio.

Alloys↗
PubMed2006

[Expression, purification, and serum antibody detection of HPV16 E6 in cervical cancer patients].

BACKGROUND & OBJECTIVE: Human papillomavirus type 16 (HPV16) is the predominant high-risk type of HPV in cervical cancer tissues. Serum antibody responded to HPV16-related proteins is associated with the development of cervical cancer. This study was to construct and purify recombinant HPV16 E6 protein, detect its corresponding serum antibody among different populations, and explore the correlation of HPV16 E6 serum antibody reaction to cervical cancer. METHODS: HPV16 E6 early gene was constructed into pRSET-A expression vector. The plasmids were transfected into BL21 (DE3) cells, which were induced to express HPV16 E6 protein by isopropylthio-beta-D-galactoside (IPTG). E6 inclusions were denatured, purified through Ni column, and renatured. After the activity of HPV16 E6 protein being identified, the antibodies against HPV16 E6 in serum samples from 80 healthy women, 46 chronic cervicitis patients, and 32 cervical cancer patients were determined by ELISA using the fusion protein as antigen. HPV DNA genotype was estimated in cervical cancer tissues by fluorescence polarization. RESULTS: HPV16 E6 fusion protein of Mr 24x10(3) was expressed in pRSET-16E6 after induction of IPTG. The fusion protein was accounted for 22.3% of total bacterial proteins, and expressed as inclusive body. After purification with Ni-NTA agarose resin, the purity of the recombinant protein was over 95%, and its activity was identified by ELISA. The antibody-positive rate was significantly higher in cervical cancer patients than in healthy women and chronic cervicitis patients (31.2% vs. 5.0% and 6.5%, P<0.01). In the 32 cervical cancer patients, the positive rates of HPVs DNA and HPV16 DNA in cancer tissues were 90.61% and 46.88%. The antibody-positive rate of HPV16 E6 was higher in HPV16 DNA-positive cervical cancer patients than in HPV16 DNA-negative patients (46.7% vs. 17.6%), but the difference was not significant. CONCLUSIONS: HPV16 E6 fusion protein obtained from pRSET-A/BL21 can be used in serologic studies on cervical cancer-related HPV infection. Serum antibody against HPV16 E6 is more common in cervical cancer patients than in healthy women and chronic cervicitis patients.

Adult↗
PubMed2005

A microarray-based gastric carcinoma prewarning system.

AIM: To develop a microarray-based prewarning system consisting of gastric cancer chip, prewarning data and analysis software for early detection of gastric cancer and pre-cancerous lesions. METHODS: Two high-density chips with 8,464 human cDNA sites were used to primarily identify potential genes specific for normal gastric mucosa, pre-cancerous lesion and gastric cancer. The low-density chips, composed of selected genes associated with normal gastric mucosa, precancerous lesion and gastric cancer, were fabricated and used to screen 150 specimens including 60 specimens of gastric cancer, 60 of pre-cancerous tissues and 30 of normal gastric mucosa. CAD software was used to screen out the relevant genes and their critical threshold values of expression levels distinguishing normal mucosa from pre-cancerous lesion and cancer. All data were stored in a computer database to establish a prewarning data library for gastric cancer. Two potential markers brcaa1 and ndr1 were identified by Western blot and immunohistochemistry. RESULTS: A total of 412 genes associated with three stages of gastric cancer development were identified. There were 216 genes displaying higher expression in gastric cancer, 85 genes displaying higher expression in pre-cancerous lesion and 88 genes displaying higher expression in normal gastric mucosa. Also 15 genes associated with metastasis of gastric cancer and 8 genes associated with risk factors were screened out for target genes of diagnosis chip of early gastric cancer. The threshold values of 412 selected genes to distinguish gastric cancer, pre-cancerous lesion from normal gastric mucosa were defined as 6.01+/-2.40, 4.86+/-1.94 and 5.42+/-2.17, respectively. These selected 412 genes and critical threshold values were compiled into an analysis software, which can automatically provide reports by analyzing the results of 412 genes obtained by examining gastric tissues. All data were compiled into a prewarning database for gastric cancer by CGO software. Northern blot and immunohistochemistry analysis confirmed that gene and protein of brcaa1 displayed lower expression in normal gastric mucosa and higher expression in gastric cancer tissues, conversely, ndr1 displayed lower expression in gastric cancer and higher expression in normal gastric mucosa. CONCLUSION: The microarray-based prewarning system for gastric cancer was developed. This system consisted of gastric cancer-associated gene chip, prewarning data and analysis software, which has a high potential for applications in the early detection of gastric cancer. The two potential markers brcaa1 and ndr1 identified may be used to distinguish cancer status fand non-cancer status.

Antigens, Neoplasm↗
PubMed2005

Detection of hepatitis B virus DNA by real-time PCR using TaqMan-MGB probe technology.

AIM: To develop a real-time PCR for detecting hepatitis B virus (HBV) DNA based on TaqMan technology using a new MGB probe. METHODS: Plasmid containing the sequence of X gene (1414-1744 nt) was constructed as HBV-DNA standard for quantitative analysis. A TaqMan-MGB probe between primers for amplification was designed to detect PCR products. The interested sequence contained in the plasmid and in clinical specimens was quantitatively measured. RESULTS: The detection limit of the assay for HBV DNA was 1 genome equivalent per reaction. A linear standard curve was obtained between 10(0) and 10(9) DNA copies/reaction (r>0.990). None of the negative control samples showed false-positive reactions in duplicate. HBV DNA was detected in 100% (50/50) of HBV patients with HbeAg, and in 72.0% (36/50) with HBsAg, HBeAb and HBcAb. The coefficient of variation for both intra- and inter-experimental variability demonstrated high reproducibility and accuracy. CONCLUSION: Real-time PCR based on TaqMan-MGB probe technology is an excellent method for detection of HBV DNA.

Base Sequence↗
PubMed2005

[Inhibitory effect of agarohexaose on antimycin A induced generation of reactive oxygen species].

AIM: To evaluate the hepatocyte protective effect of agarohexaose against indirect oxidative stress injury induced by antimycin A. METHODS: Antimycin A was used to induce oxidative injury of human hepatocyte L-02. The oxidative degree in cells was detected by dichlorofluorescin diacetate (DCFH-DA) and the fluorescence generation was recorded by flow cytometer and fluorescent microscope. The apoptosis of L-02 cells induced by oxidation was identified by TUNEL test, and the morphologic features of cells were also observed. RESULTS: Agarohexaose at concentration of 1 mg x mL(-1) inhibited the oxidation of DCFH into DCF significantly. The fluorescence intensity and oxidized cell number decreased after the incubation with agarohexaose. The photomicrographs of antimycin A and agarohexaose treated cells revealed that agarohexaose could reduce the apoptotic morphologic features. The TUNEL results also indicated that the number of apoptotic cells decreased significantly after the treatment of agarohexaose. CONCLUSION: Agarohexaose could inhibit the sudden increase reactive oxygen species (ROS) in cells significantly, and it also protected cells against oxidative stress injury in vitro.

Antimycin A↗
PubMed2004

Antioxidant activities of agaro-oligosaccharides with different degrees of polymerization in cell-based system.

The aim of the present study was to evaluate the antioxidant activity of agaro-oligosaccharides with different degrees of polymerizations (DPs) and establish a relationship between the activity and DPs. The attenuate effect of oligosaccharides on 1,1-diphenyl-2-picryl-hydrazyl (DPPH(*)) was initially assessed, and the result indicated that agarohexaose showed the highest scavenging DPPH(*) capability (IC(50)=1.85 mg/ml). Following that, the intracellular antioxidant ability of agaro-oligosacharides was investigated by using the dichlorofluorescein (DCF) assay in human liver cell L-02 system. Different levels of antioxidant activities of agaro-oligosaccharides with various DPs were observed, and their scavenging reactive oxygen species (ROS) capability was associated with the improvement of the cell viability. In these oligosaccharides, agarohexaose possessed the highest scavenging capability, which could reduce 50% of oxidants generated by H(2)O(2) at 1 mg/ml. Furthermore, the antioxidant effect of agarohexaose on the indirect oxidation of cells induced by antimycin A (AA) was also tested. The results showed that agarohexaose could scavenge ROS generated by electron leakage and protect cells against apoptosis induced by ROS. It is concluded that agaro-oligosaccharides are generally considered as novel antioxidants which could protect cell damage caused by reactive oxygen species, especially agarohexaose exhibiting most desirable effects.

Antimycin A↗
PubMed2003

Rapid and high throughput detection of HBV YMDD mutants with fluorescence polarization.

AIM: To develop a simple and rapid detection of HBV gene variants and prediction of lamivudine-resistance in patients. METHODS: Initially, plasmids harboring the wild-type or mutant HBV DNA fragments were used in a model system. The technique was then applied to clinical samples for an analysis of YMDD mutations. The sera were extracted from chronic hepatitis patients who had received lamivudine treatment for more than one year. P region gene of HBV was amplified by polymerase chain reaction. The excess primers and dNTPs in PCR products were removed by cleaning-up reagents. Template-directed dye-terminator incorporation reaction was performed and R110 or TAMRA labeled acyclo-terminator was added on the 3' end of TDI-primer specifically. Fluorescence polarization value was measured with Victor 2 multilabel counter and the genotypes of HBV were analyzed. RESULTS: The YMDD genotypes in recombined positive plasmid and 56 serum samples of HBV infected patients were analyzed by using our TDI-FP method and the specificity and sensitivity were confirmed by DNA sequencing. Five of 56 serum samples showed YVDD phenotype (9%), including 1 YMDD and YVDD mixed infection. Four of 56 showed YIDD phenotype (7.1%). CONCLUSION: This is a simple, rapid, low cost and high throughput assay to detect HBV polymerase gene variants and suitable for large-scale screening and prediction of the lamivudine-resistance in clinical samples.

Drug Resistance, Viral↗
PubMed2003

Detection and genotyping of human papillomavirus DNA in cervical cancer tissues with fluorescence polarization.

To evaluate the type-specific prevalence of eight common types of human papillomavirus (HPV) in patients with cervical cancer living in Shanxi, China, with fluorescence polarization detection, crude DNA extracted from 137 samples of early-stage cervical cancer (within stage IIa) and chronic cervicitis was subjected to HPV L1 consensus GP5+/GP6+ system. Then, the HPV-positive products identified by GP5 + /GP6+ PCR were genotyped based on template-directed dye-terminator incorporation assay with fluorescence polarization detection (TDI-FP): the PCR products were respectively hybridized with designed type-specific probes within the GP5+/GP6+ amplicons for eight common HPV types (HPV 6, 11, 18, 16, 31, 33, 35, and 58), and specific fluorescence-labeled ddNTPs (TAMRA-ddTTP or R110-ddGTP) were directly incorporated to the ends of the corresponding probes under directing of the corresponding template in PCR products, which was reflected and read by high FP values for TAMRA or R110. HPV DNA was detected in 38.89% (28/72) cases of chronic cervicitis, and 87.69% (57/65) cases of cervical cancer. There was a significant difference in HPV prevalence between these two groups. The four commonly identified types in patients with cervical cancer were HPV 16 (45.6%), HPV 18 (22.8%), HPV 58 (17.5%), and HPV 31 (7.02%), and in those with chronic cervicitis were HPV 16 (35.7%), HPV 11 (32.1%), HPV 6 (21.4%), and HPV 18 (10.7%). 57.14% of HPV types detected in patients with chronic cervicitis were high-risk types. HPV 16 was the most common viral type identified in both groups. Type-specific prevalence of HPV DNA has some characteristics in patients with chronic cervicitis and cervical cancer living in Shanxi, China and the worldwide uncommon type HPV 58 is relatively common in both kinds of cases. The high prevalence of HPV 58 in Chinese women should been considered in diagnosis and vaccine designs of HPV.

Adult↗
PubMed2000

Mechanism of exogenous nucleic acids and their precursors improving the repair of intestinal epithelium after gamma-irradiation in mice.

AIM:To clone expressed genes associated with repair of irradiation-damaged mice intestinal gland cells treated by small intestinal RNA, and to explore the molecular mechanism of exogenous nucleic acids improving repair of intestinal crypt.METHODS:The animal mode of test group and control group was established, forty-five mice being irradiated by gamma ray were treated with small intestinal RNA as test group, forty mice being irradiated by gamma ray were treated with physiological saline as control group,five mice without irradiation were used as normal control, their jejunal specimens were collected respectively at 6h, 12h,24h, 4d and 8d after irradiation. Then by using LD-PCR based on subtractive hybridization, these gene fragments differentially expressed between test group and control group were obtained, and then were cloned into T vectors as well as being sequenced. Obtained sequences were screened against. GeneBank, if being new sequences, they were submitted to GeneBank.RESULTS:Ninety clones were associated with repair of irradiation-damaged intestinal gland cells treated by intestinal RNA. These clones from test group of 6h, 12h, 24h, 4d and 8d were respectively 18, 22, 25, 13, 12. By screening against GeneBank, 18 of which were new sequences, the others were dramatically similar to the known sequences, mainly similar to hsp, Nmi,Dutt1, alkaline phosphatase, homeobox, anti-CEA ScFv antibody, arginine/serine kinase and BMP-4,repA. Eighteen gene fragments were new sequences,their accept numbers in GeneBank were respectively AF240164-AF240181.CONCLUSION:Ninety clones were obtained to be associated with repair of irradiation damaged mice intestinal gland cells treated by small intestinal RNA, which may be related to abnormal expression of genes and matched proteins of hsp, Nmi, Dutt1, Na, K-ATPase,alkalineph-osphatase, glkA, single stranded replicative centromeric gene as well as 18 new sequences.

Journal Article↗