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.

The nonstructural protein 5A (NS5A) of the hepatitis C virus (HCV) has been shown to interact with a variety of cellular proteins and implicated in the regulation of cell growth, interferon resistance, and other cellular signaling pathways. Using the yeast-two hybrid method, we have isolated a clone that encodes a novel NS5A--associated binding protein: NS5ABP37. Reverse transcription polymerase chain reaction (RT-PCR) method was employed to amplify the full fragment,and the plasmid pGADT7-NS5ABP37 with the Saccharomyces cerevisiae vector pGADT7 was constructed. To prove the interaction, yeast cell Y187 transformed with pGADT7-NS5ABP37 was mated with yeast cell AH109 containing pGBKT7-NS5A to verify the interaction between the novel protein coded by the new gene NS5ABP37 and NS5A.

Objective HCV NS3 protein plays an important role in disease caused by HCV. We investigate the gene expression of HCV NS3 in yeast for future study of the function of the protein. Methods PCR was performed to amplify the gene of HCV NS3 from the plasmid pBRTM/HCV containing the whole fragment of HCV and the gene was cloned into pGEM T vector. Thereafter, HCV NS3 gene was cut from pGEM T vector and cloned into yeast expression plasmid pGBKT7, and recombinant pGBKT7∶NS3 was transformed into yeast AH109. The yeast protein was isolated and analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and Western blotting. Results HCV NS3 gene was successfully cloned into pGBKT7. The results of SDS PAGE and Western blotting assay showed that the molecular weight of the expressed product was about 22000 Da and HCV NS3 protein was existed within yeast cells.Conclusions HCV NS3 was successfully expressed in yeast expression system.

Objective To study the effect of new barrel theory in improving evaluation of hospitalized patients. Methods Eight hundred hospitalized patients from September 2012 to August 2013 were randomized equally into the control group and the observation group.The last one or two items affecting patient satisfaction from the control group were used as objectives to be improved.Causal effect analysis was done pertinent to the items and the worksheet was bettered and improved and then enforced.The two groups were compared after intervention with new barrel theory in terms of satisfaction of patients during admission and discharge. Result The satisfaction of patients in the observation group during admission and discharge was significantly better than that in the control group(P<0.05).Conclusion The new barrel theory used to detect the flaws in nursing service and improve the workflow can improve assessment from the patients so that the management quality can be enhanced.

Objective To observe the clinical efficacy and safety of propafenone on paroxysmal supraventricular tachycardia (PSVT).Methods 37 patients with PSVT were injected with 70 mg propafenone intravenously in 5 minutes.The unrecovered patients in 20 minutes were injected with 70 mg propafenone intravenously again,who were given 70 mg propafenone intravenously once again if not controlled in late 30 minuted.Blood pressure,heart beat,12 lead electrocardiogram were recorded before and after use of drug.Results The significant effective rate was 77.5% and effective rate was 21.6% with overall effective rate of 97.3% and effective time of 1～55 (7.1?2.8) minutes.The average accumulated dose of propafenone was 105 mg.Some side effects were observed in part of patients,including slight reduction of blood pressure,P R interval.QRS wave and Q T interval prolonged,temporary type sino atrial block,vertigo and nausea,etc whice could vanish gradually if not treated.Conclusion Propafenone by intravenous injection can quickly contro PSVT with saftey and effectiveness.

This study investigated the expression of hemeoxygenase-1 (HO-1) in rats with acute lung rejection and its implication. A valid rat orthotopic left lung transplantation model (SD rat-->Wistar rat) was established by using an improved three-cuff anastomosis technique. The rats were divided into control group, CoPP (HO-1 inducer)-treated group and ZnPP (HO-1 inhibitor)- treated group. The severity of acute rejection was graded on the basis of the morphologic changes of the lung samples stained with HE. The expression of HO-1 protein in lung tissue was detected by using immunohistochemistry and Western blot, and HO-1 mRNA activity was assayed by RT-PCR. The results showed that the expression of HO-1 protein was significantly increased with the acute rejection grading in rats (P<0.01). As compared with control and ZnPP-treated groups, the severity of acute rejection was not alleviated and the grade not reduced significantly in CoPP-treated group (P>0.05). It was concluded that HO-1 protein might be involved in the pathological process of post-graft acute rejection. The expression of HO-1 protein was increased gradually with aggravation of acute rejection, and HO-1 protein might be used as an index to monitor acute rejection after lung transplantation.
Graft Rejection/*enzymology ; Heme Oxygenase (Decyclizing)/genetics ; Heme Oxygenase (Decyclizing)/*metabolism ; Lung Transplantation ; RNA, Messenger/genetics ; RNA, Messenger/metabolism ; Rats, Sprague-Dawley ; Rats, Wistar

BACKGROUND:In an additional changing magnetic field, magnetosomes can invade the tumor cells and inhibit their proliferation.OBJECTIVE: To study the effect of targeted delivery of pHSP70-shPLK1/DOX compounds with magnetosome of magnetotactic bacteria on the treatment of osteosarcoma.METHODS: Human osteosarcoma cell U2OS was divided into four groups: group A: magnetosomes of magnetotactic bacteria+pHSP70-shPLK1/DOX complex; group B: magnetosomes of magnetotactic bacteria; group C: magnetosomes of magnetotactic bacteria+pHSP70-shPLK1/DOX complex, combined with magnetic intervention; group D: magnetosomes of magnetotactic bacteria, combined with magnetic intervention. After 12, 24, 48 and 72 hours of culture, the uptake rate of osteosarcoma cells was observed; cell cycle was evaluated by flow cytometry; PLK1 mRNA and protein expression levels were detected by RT-PCR and western blot, respectively; cell proliferation was explored by MTT assay; cell adhesion and invasion were checked by adhesion assay and Transwel chamber assay, respectively; and the apoptosis rate was measured by apoptosis detection Kit.RESULTS AND CONCLUSION: (1) After culture of 12 and 24 hours, the uptake rate was the highest in group D; meanwhile, the uptake rate was the highest in group B after 48 and 72 hours of culture. (2) The ratio of G2/M phase decreased and the ratio of G0/G1 increased gradualy in group A, group C and group D; the G2/M phase ratio of C group was the lowest at each time point, followed by group D, and that of group A was the highest; and the G2/M phase ratio increased gradualy in group B. (3) Folowing 12 and 24 hours of culture, the expression of PLK1 in group B and group D was higher than that in group A and group C (P < 0.05); in addition, the expression of PLK1 in group B was higher than that in the other three groups after 48 and 72 hours of culture (P < 0.05), and the PLK1 level in group D was higher than that in group A and group C (P < 0.05). (4) The proliferation, adhesion and invasion ability in group B were the highest at different time points, but the apoptosis rate was the lowest. Furthermore, the proliferation, adhesion and invasion abilities in group C were the lowest at different time points, and the apoptosis rate was the highest. To conclude, our experimental results show that the targeted delivery of pHSP70-shPLK1/DOX complex by magnetosomes of magnetotactic bacteria can enhance the apoptosis of osteosarcoma cells and inhibit their proliferation and invasion.

Objective To analyse risk factors for bile leakage after liver resection.Methods Between January 2011 and December 2012,469 patients underwent elective hepatectomy.We prospectively collected and retrospectively analyzed demographic data,pathological variables,and perioperative variables.Univariate analysis screened the related factors of bile leakage after liver resection.Multivariate analysis identified the independent risk factors of postoperative bile leakage.Results 469 patients were included in the analysis.The prevalence of bile leakage was 22.6％ (n =106).Univariate analysis identified the following risk factors as male gender,portal hypertension,steatosis,cirrhosis,ChildPugh grade,ascites,operative time,intraoperative transfusion,intraoperative blood loss,portal triad clamping,microwave solidification,lymphadenectomy,number of tumor,tumor margin,tumor capsular,diameter of tumor,portal vein invasion or portal branch thrombosis,number of abdominal drains.Multivariate analysis identified 4 independent risk factors for postoperative bile leakage:Cirrhosis [OR =13.2 (2.3,76.9),P =0.004],steatosis [OR =73.1 (17.7,301.5),P ＜ 0.001],infusion volume of the surgery day [OR=1.0 (1.0,1.0),P=0.019] and diameter of tumor [OR=1.2 (1.1,1.3),P=0.003].Conclusions Cirrhosis,steatosis,transfusion volume of the surgery day,and tumor size were risk factors for bile leakage after major liver resection.