Objective To evaluate the influence of expression of pre S1 protein on the genomic expression of hepatitis B virus (HBV) infected hepatocyte with microarray. Methods The differentially expressed genes between the hepatoblastoma cell line HepG2 transfected by pcDNA3.1(-) and pcDNA3.1(-)-preS1, were respectively compared by cDNA microarray technique. The HBV pre-S1 coding DNA fragment was amplified with polymerase chain reaction (PCR) technique by using G376-7 plasmid DNA containing the full length of HBV genome as the template. The expressive vector of pcDNA3.1-preS1 was constructed by routine molecular biological methods. HepG2 cells were transfected by pcDNA3.1(-) and pcDNA3.1-preS1, respectively, using FuGENE6 Transfection Reagent. The total RNA was isolated and reversely transcribed. Results The cDNAs were subjected for microarray screening with 1152 cDNA probes. From the scanning results, it was found that 30 genes were up-regulated and 38 genes were down-regulated by pre-S1 protein of HBV. Conclusion The expression of pre-S1 protein affected the genomic expression spectrum of HBV infected hepatocyte(HepG2 cell line).

A set of specific primers was synthesized according to DNA sequence of HBV found in China, the reverse transcriptase (RT) region in polymerase gene was amplified by PCR method from the serum of 3 patients with chronic HBV infection, and then the PCR products were subcloned into pGEM Teasy vectors. 13 clones were sequenced. Sequence comparison of the selected clones was made to look for the difference. After being compared, 13 sequences of RT were found different. Besides 2 clones with long sequence deletion, the different rate of RT coding nucleic acid sequences, RT and HBsAg amino acid sequences of the 11 clones is 5 1%, 4 9% and 7 5%, respectively. Many mutation types, including point substitution and deletion mutation, were found in this region. There is quasispecies population and defective HBV genome in patients with chronic HBV infection.

Objective To construct plasmid pVR1012 M as nuclei acid vaccine for hepatitis B,was constructed to immunize mice with or without plasmid pcDNA 3.1 - IL 18 to identify the effect of Interleukin 18(IL 18). Methods Polymerase chain reaction method was used to amplify the PreS2 and S region of HBV and reconstruct plasmid pVR1012 M as nuclei acid vaccine. Plasmid pcDNA 3.1 - IL 18 was used as a co stimulator. Twenty five Balb/c mice were divided into 3 groups, group 1 immunized with 100 ?g plasmid pVR1012 group 2 pVR1012 M,Group 3,pVR1012 M with pcDNA 3.1 - IL 18, respectively, every 2 weeks for 3 times. Anti HBs were detected in serum 2 weeks after each injection. Lactated ehydrogenase (LDH) cytotoxicity assay was done to analyze the cytotoxic T lymphocytes funciton. Results The positive rate and the antibody titer of serum from mice injected pVR1012 M increasing gradually with the increasing frequency of inoculation, while those from mice injected pVR1012 M and pcDNA 3.1 - IL 18(joint group) were lower than those injected with pVR1012 M alone (Difference after 3rd inoculation was significant, P

It was found that pre-X region was present in five clones of hepatitis B virus (HBV) genome from 2 patients with chronic HBV infection. The open reading frame (ORF) consisted of 168 base pair (bp), and could be transslated with the X gene in frame. To investigate the activity of pre-X gene promoter, promoter DNA sequence was amplified from HBV-DNA by polymerase chain reaction (PCR), The amplified product was cloned into pCAT3 vector, The HepG2 cells were transfected by pCAT3-Pre-X-p. The CAT activity was detected by an enzyme-linked immunosorbent assay (ELISA) kit. It was found that pCAT3-Pre-X-p had higher activity of CAT, which was 3.5 fold over that of pCAT3-promoter. This result implicated that pCAT3-Pre-X-p possessed promoter activity.

To investigate the promoter activity of pre-pre-S gene, which was found in five clones of hepatitis B virus (HBV) genome from 2 patients with chronic HBV infection, promoter DNA sequence was amplified from HBV-DNA by polymerase chain reaction (PCR).The amplified product was cloned into pCAT3 reporter vector. The HepG2 cells were transfected by pCAT3-pre-pre-S-p. The CAT activity was detected by an enzyme-linked immunosorbent assay (ELISA) kit. It was found that pCAT3-pre-pre-S-p had higher activity of CAT which was 10 fold over that of pCAT3-basic. This result implicated that pCAT3-pre-pre-S-p had promoter activity.

Objective T7 cDNA phage display system and bioinformatics methods were employed to find the binding protein to the PreS1 protein of hepatitis B virus (HBV). Methods PreS1 protein was coated in ELISA plate as the target protein, and then T7 cDNA library phage display system was used to scan the binding protein or peptide. A piece of cDNA was found to have the function to bind the PreS1 protein, and the product was named as PreS1 binding protein (PreS1BP). Using BLAST in GenBank, the amino acid sequence of PreS1BP was compared in the protein sequence database. Results The amino acid sequence of PreS1BP was identified as a piece of glioma tumor suppressor candidate region gene 2 (GLTSCR2), and the length of cDNA of PreS1BP was proved to be 1436 nt. The gene was located at chromosome 19q arm (19q13.3) with a length of 11445 base pair between 10403483 and 10414989, containing 13 exons and 12 introns. Conclusion HBV PreS1BP gene could be obtained by T7 cDNA phage display system in combination with bioinformatics methods.

0.05),the incidence of adverse drug reactions were 3.5% and 2.4%,respectively and the total treatment costs were(4 146.45?282.15)yuan and(4 807.20?318.15)yuan,respectively;and the cost-minimization analysis showed that manicol was the preferred therapy as compared with Glycerol Fructose.In the treatment of the patients with cerebral infarction complicating renal dysfunction,the total effective rates of the two drugs were 80.0% and 93.3%,respectively(P

Objective To investigate the clinical value of serum homocysteine (Hcy),Cystatin C (CysC) detection in coronary heart disease prevention,diagnosis and treatment.Methods A total of 200 cases of coronary heart disease patients including 50 cases of acute myocardiac infarction (AMI) (AMI group),85 cases of unstable angina pectoris (UAP)(UAP group.) and 65 cases of stable angina pectoris (SAP) (SAP group) were selected,and 120 cases of healthy controls were selected as control group.The serum levels of Hcy and CysC were detected and compared.Results The serum levels of Hcy and CysC in AMI group,UAP group and SAP group were significandy higher than those in control group[Hcy:(22.35 ± 8.18),(16.54±7.56),(14.52±6.38) μmol/Lvs.(9.35±3.23) μmol/L; CysC:(1.32±0.27),(1.88± 0.66),(1.19 ± 0.46) mg/L vs.(0.80 ± 0.33) mg/L],and there were significant differences between two groups (P＜ 0.01).The serum level of Hcy in AMI group was higher than that in UAP group and SAP group [(22.35 ± 8.18) μ mol/L vs.(16.54 ± 7.56),(14.52 ± 6.38) μ mol/L],and there was significant difference (P ＜ 0.01).Conclusions The relationship between Hcy,CysC level and coronary heart disease is close,and Hcy levels increase with increasing degree of coronary heart disease.Detection of Hcy and CysC has some clinical value on prevention,diagnosis,treatment of coronary heart disease.

Objective To study the low dose hyper-radiosensitivity in human lung cancer cell line A549,and its possible mechanisms.Methods Exponentially growing A549 cells were irradiated with 60Co γ-rays at doses of 0-2 Gy.Together with flow cytometry for precise cell sorting,cell survival fraction was measured by mean of conventional colony-formation assay.ATM1981 Ser-P protein expression was examined by Western blot.Apoptosis was identified by Hoechst 33258 fluorescent staining,and Annexin V-FITC and propidium iodide staining flow cytometry.Cell cycle distribution was observed by flow cytometry.Results There was an excessive cell killing per unit dose when the doses were below about 0.3 Gy,and the cells exhibited more resistant response at the doses between 0.3 and 0.5 Gy,the cell survival fraction was decreased as the doses over 0.5 Gy.The expression of ATM1981Ser-P protein was first observed at 0.2 Gy,followed by an increase over 0.2 Gy,and reached the peak at 0.5 Gy(compared with 0.2 Gy group,t=7.96,P＜0.05),with no further increase as the doses at 1.0 and 2.0 Gy(t=0.69,0.55,P＞0.05).24 hours after irradiation,part cells presented the characteristic morpholos4cal change of apoptosis,and the apoptosis curve was coincident with the dose-survival curve.Compared with the control group,the cell cycle had no change post-irradiation to 0.1 and 0.2 Gy.G2/M phase arrest was manifested at 6 and 12 hours post-irradiation to 0.3,0.4 and 0.5 Gy(t=2.87,2.88,4.92 and 3.70,3.12,8.11,P＜0.05),and the ratio of G2/M phase was decreased at 24 hours post-irradiation(t=3.87,4.77,3.01,P＜0.05).Conclusions A549 cells displays the phenomenon of hyper-radiosensitivity(HRS)/induced radioresistance(IRR).The model of cell death induced by low dose irradiation is mainly apoptosis.The activity of ATM and cell cycle change might play an important role in HRS/IRR.

Acute Kidney Injury ; etiology ; Hemolytic-Uremic Syndrome ; complications ; etiology ; therapy ; Humans ; Shiga Toxin ; toxicity