To investigate the potential role of hepatitis B virus(HBV) preS1 protein in mediating HBV adhesion to liver cell, we prepared recombinant proteins of HBV preS1 in yeast. PCR was performed to amplify the gene of HBV preS1 from the plasmid pCP10/HBV ayw subtype containing the whole fragment of HBV and the PCR product was cloned into pGEM T vector. The gene of HBV preS1 was cut from pGEM T vector and cloned into yeast expression plasmid pGBKT7, the pGBKT7 plamids containing preSl were transformed into yeast cell AH109. The yeast protein was isolated and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and Western blotting. The results showed that the presence of HBV presl proteins in yeast cells was confirmed by Western slot analysis. the molecular weight of the expressed product was about 30000 Da. The findings indicated that HBV preS1 was successfully expressed in yeast system.

Protein protein binding is the basis of virus and host cell interactions. With the application of technology of studying of protein interactions, more knowledge of replication and pathogenesis of hepatitis C virus (HCV) could be acquired. Non structure protein NS5A is one of the important regulatory factors in virus replication , transcription and signal transduction, but there are controversy in effect on HCV pathogenesis and resistance to interferon ?(IFN?). In order to describe the relationship between NS5A and host proteins, we use yeast two hybrid system 3 to screen the gene encoding proteins that could interact with NS5A from human hepatocyte library. Thirty five clones were obtained including apo A1, apo A2, apo B100, haplotype mitochondrion complete genome, phosphatidic acid phosphatase type 2B, albumin similar to tumor endothelial marker 5 precursor, matrix metalloproteinase 14, and three of the Homo sapiens hypothetical proteins. The study paved a way for further studies on the pathogenesis of HCV NS5A

To clone the genes of hepatocyte protein interacting with hepatitis C virus NS3 protein, "bait" plasmids of hepatitis C virus NS3 were constructed. After verifying that hepatitis C virus NS3 protein could be steadily expressed in AH109 yeast strain, yeast two hybrid assay was berformed by mating AH109 with Y187 that pre transformed with liver cDNA library plasmids pACT2, and the diploidy yeast cells were plated on quadruple dropout (QDO) medium and assayed for X ? gal activity. Nineteen yeast colonies that could grow on QDO and had ? gal activity were obtained, then the library plasmids were extracted and sequenced. The gene sequences from the 19 positive colonies were aligned with the genes deposited in GenBank. It was found hepatitis C virus NS3 protein could interact with some proteins which have different functions.

Humans ; Hypopharyngeal Neoplasms ; Hypopharynx ; pathology ; Male ; Middle Aged ; Neoplasms, Muscle Tissue

Objective To evaluate the effect of mechanical stretch preconditioning on pathological stretch-induced activation of γ-aminobutyric acid (GABA) signaling pathway in human type Ⅱ alveolar epithelial cells (AEC Ⅱ).Methods AEC Ⅱ cell line (A549 cells) culturedin vitro were divided into control group (group C), pathological stretch group (group P1) and mechanical stretch preconditioning group (group P2). In group C, A549 cells were cultured routinely. In group P1, A549 cells were exposed to 20% cyclic stretch for 6 hours. In group P2, A549 cells were exposed to 5% cyclic stretch for 60 minutes, and then exposed to 20% cyclic stretch for 6 hours. The cells were harvested for determination of the cell viability by methyl thiazolyl tetrazolium assay, lactate dehydrogeuase (LDH) release was determined by colorimetric method, the levels of interleukin (IL-1β and IL-6) and tumor necrosis factor-α (TNF-α) were determined by enzyme linked immunosorbent assay (ELISA), the mRNA expressions of IL-1β, IL-6 and TNF-α were determined by reverse transcription-polymerase chain reaction (RT-PCR), and the protein expressions of glutamic acid decarboxylase (GAD) and γ-aminobutyric acid A receptor (GABAAR) were determined by Western Blot.Results Compared with group C, the cell viability of group P1 was significantlydecreased (A value: 0.196± 0.071 vs. 0.886±0.107), the release rate of LDH was significantly increased [(12.3±2.4)% vs. (1.9±0.5)%]; the contents and mRNA expressions of IL-1β, IL-6 and TNF-α in cell culture medium were significantly increased [IL-1β (ng/L): 138.6±19.7 vs. 32.7±7.4, IL-6 (ng/L): 196.5±31.7 vs. 55.4±13.8, TNF-α (ng/L): 111.3±21.8 vs. 20.8±7.6; IL-1β mRNA (2-ΔΔCT): 2.79±0.44 vs. 0.83±0.12, IL-6 mRNA (2-ΔΔCT): 1.99±0.25 vs. 0.56±0.11, TNF-α mRNA (2-ΔΔCT): 2.54±0.37 vs. 0.72±0.09]; the protein expressions of GAD and GABAAR were significantly decreased [GAD (gray value): 0.38±0.12 vs. 1.75±0.45, GABAAR (gray value): 0.29±0.09 vs. 1.68±0.39; allP < 0.05]. Compared with group P1, the cell viability of group P2 was significantly increased (A value: 0.523±0.132 vs. 0.196±0.071),the release rate of LDH was significantly decreased [(6.9±1.7)% vs. (12.3±2.4)%]; the contents and mRNA expressions of IL-1β, IL-6 and TNF-α in cell culture medium were significantly decreased [IL-1β (ng/L): 79.2±11.6 vs. 138.6±19.7, IL-6 (ng/L): 89.6±15.6 vs. 196.5±31.7, TNF-α (ng/L): 55.9±11.4 vs. 111.3±21.8; IL-1β mRNA (2-ΔΔCT): 1.92±0.36 vs. 2.79±0.44, IL-6 mRNA (2-ΔΔCT): 1.09±0.18 vs. 1.99±0.25, TNF-α mRNA (2-ΔΔCT): 1.77±0.25 vs. 2.54±0.37]; the protein expressions of GAD and GABAAR were significantly increased [GAD (gray value): 1.26±0.33 vs. 0.38±0.12, GABAAR (gray value): 1.04±0.15 vs. 0.29±0.09; allP < 0.05]. Conclusion The mechanism by which mechanical stretch preconditioning attenuates pathological stretch-induced injury in human AECⅡ is related to the activation of GABA signaling pathway.

OBJECTIVE: To determine imperatorin as the active ingredient of Angelica dahuricae in Jiedu tongqiao pills by HPLC. METHODS: The separation was performed on Waters Symmetryshielb33 C18(250 mm?4.6 mm,5 ?m) column with detection wavelength of 248 nm. The mobile phase consisted of methanol-water (65 ∶ 35) with flow rate of 1.0 mL?min-1.RESULTS:The linear range of imperatirin were 0.15～1.35 ?g. The average recovery was 100.4%(RSD=1.89%,n=9).CONCLUSION: The method is simple, accurate and reproducible for quality control of Jiedu tongqiao pills.

Objective To establish and test the reliability and validity of the disease related psychological pressure scale of pregnant women with chronic HBV infection,so as to provide mental stress assessment tool for the pregnant women.Methods Item pool was developed after the interview and comprehensively retrieved literatures,then after expert review and rebuilt,the scale items were fixed.Totals of 368 pregnant women with chronic HBV infection disease were investigated and the reliability and validity of scale were test.Results The scale had 5 factors and 24 items,and the correlation coefficient between each factor score and total score was 0.712 - 0.894.The correlation coefficient among each factor was 0.409 - 0.631 which was significantly lower than that between each factor score and total score ( P ＜ 0.01 ).The cumulative contribution rate of 5 factors was 64.055％,and tes-retest reliability of 5 factors was 0.856,0.887,0.828,0.813.The Cronbach' s α of internal consistency of 5 factors was 0.788 - 0.865,and the Cronbach' s α of scale was 0.932.Conclusions The scale has good reliability and validity,and can be used for measuring the disease related pregnancy psychological pressure for chronic HBV infection pregnant women.

Objective To evaluate the effect of dexmedetomidine on microRNA (miRNA)-155-hypoxia-inducible factor-1α (HIF-1α)-heme oxygenase-1 (HO-1) signaling pathway in a rat model of endotoxin-induced acute lung injury.Methods Forty adult male Wistar rats,weighing 220-250 g,were equally and randomly divided into 4 groups using a random number table:control group (group C),dexmedetomidine group (group D),endotoxin-induced acute lung injury group (group L),and endotoxin-induced acute lung injury+dexmedetomidine group (group LD).Acute lung injury was induced by intraperitoneal lipopolysaccharide (LPS) 5 mg/kg in L and LD groups.In D and LD groups,dexmedetomidine was infused in a loading dose of 1 μg · kg-1 · h-1 for 10 min starting before intraperitoneal injection of normal saline or LPS followed by an infusion of 5 μg · kg-1 · h-1 throughout the operation.At 6 h after normal saline or LPS injection,blood samples were taken from the carotid artery for detection of arterial oxygen partial pressure (PaO2).The left lung was lavaged,and broncho-alveolar lavage fluid (BALF) was collected for determination of concentrations of total protein,interleukin-1β (IL-1β),tumor necrosis factor-α (TNF-α),and intercellular adhesion molecule-1 (ICAM-1).The rats were then sacrificed,and lungs were removed for determination of the wet to dry lung weight ratio (W/D ratio),mRNA expression of miR-155,IL-1β,TNF-α and ICAM-1,and protein expression of HIF-1α and HO-1,and for examination of the pathological changes which were scored.Results Compared with group C,the PaO2 was significantly decreased,and the W/D ratio,lung injury score,concentrations of total protein,IL-1β,TNF-α and ICAM-1 in BALF,mRNA expression of miR-155,IL-1β,TNF-α and ICAM-1,and protein expression of HIF-1α and HO-1 were significantly increased in L and LD groups (P ＜ 0.05).Compared with group L,the PaO2 and protein expression of HIF-1α and HO-1 were significantly increased,and the W/D ratio,lung injury score,concentrations of total protein,IL-1β,TNF-α and ICAM-1 in BALF,and mRNA expression of miR-155,IL-1β,TNF-α and ICAM-1 were significantly decreased in group LD (P＜0.05).Conclusion Dexmedetomidine reduces endotoxin-induced acute lung injury through activating miR-155-HIF-1α-HO-1 signaling pathway in rats.

Objective This study was to assess the three-dimensional gross tumor volume(GTV)motion of lung cancer caused by respiration using four-dimensional computed tomography(4DCT),and to analyze the influenee factors.Methotis Four-DCT scans of 22 lung focuses in 21 patients with lung cancer were analyzed.The gross tumor volume was contoured in all 10 respiration phases of 4DCT scans.The changes in volume of GTV,the 3D motion of the centroid,boundary of GTV and the 3D spatial motion vectors were calculated and the irdluenee factors were analyzed.Results The average change in volume of GTV was+14.3%(0.2%.42.5%)/-8.4%(0.4%-38.6%),the average movement amplitude of GTV centroid and GTV boundary were(0.18±0.12)cm,(0.20±0.16)cm,(0.53±0.59)cm and(0.42±0.23)cm,(0.41±0.22)cm,(0.57±0.70)cm in medio-lateral,vertro-dorsal,cranio-caudal(CC) direction,respectively.The CC movement was larger than other directions(Z=-2.12,P=0.034;Z:-2.10,P=0.035),and no significant difference was observed in 3D motion of GTV boundary(Z=-0.81.P=0.417;Z=-0.86,0.391).The CC motion of GTV eentroid in lower lobe was larger than that in upper lobe[(0.87±0.64)and(0.35±0.49)cm,(t=-2.12,P=0.047)],and no significant difference was found in other directions[(0.23±0.10)and(0.19±0.18)em(t=-0.49,P=0.629),(0.21±0.13)and(0.17±0.11)cm(t=0.76,P=0.460)].There was no correlation of the 3D movement and 3D spatial motion vector of GTV to the volume of GTV(r=-0.306,-0.062,-0.279,-0.300;P=0.189,0.796.0.234,0.199).Conclusions GTV motion of patients with lung cancer is individual,the CC movement is the moat obvious,using 4DCT to assess is comparatively accurate.The motion amplitude of lower lobe focuses is larger.No significant correlation of the GTV motion to the volume was observed.Larger sample study is needed to analyze the influence of adjacency to the GTV motion.

Female ; Gastroenteritis ; complications ; Humans ; Infant ; Male ; Nitric Oxide ; physiology ; Rotavirus Infections ; complications ; Seizures ; etiology