OBJECTIVE:To study the effects of Anzi mixture on Toll like receptor 4(TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear facter-κB(NF-κB)signaling pathway of antiphospholipid antibodies(APA)positive abortive mice,and to inves-tigate the mechanism of anti-APA positive abortion. METHODS:BALB/c mice(female)were randomly divided into blank control group,model group,aspirin group (positive control,0.0195 g/kg) and Anzi mixture low-dose,medium-dose and high-dose groups (37.7,75.4,150.8 g/kg,calculated by crude drug),with 10 mice in each group. Except for blank control group,other groups were given human β2-glycoprotein Ⅰ as derivant to establish APA positive abortion model. From the first day of pregnancy, treatment groups were given relevant medicine intragastrically,and blank control group and model group were given constant vol-ume of normal saline intragastrically,once a day,for consecutive 9 d. mRNA and protein levels of TLR4,myeloid differentiation 2 (MD2),MyD88 and NF-κB in placental tissue of mice were determined by RT-PCR and immunohistochemical method. RE-SULTS:Compared with blank control group,mRNA and protein expression of TLR4,MD2,MyD88 and NF-κB in placental tis-sue were increased markedly in the model group(P<0.01). Compared with model group,mRNA and protein expression of TLR4, MD2 and MyD88 in aspirin group and Anzi mixture low-dose and medium-dose groups were decreased significantly as well as the protein expression of TLR4 in Anzi mixture high-dose group and the protein expression of NF-κB in all medicine groups(P<0.05 or P<0.01). mRNA expression of TLR4 and MD2 and the protein expression of MD2 and MyD88 in Anzi mixture low-dose groups were lower than those in aspirin group (P<0.05 or P<0.01). CONCLUSIONS:Anzi mixture can inhibit TLR4/MyD88/NF-κB signaling pathway of APA positive abortive mice,which may be one of anti-APA positive abortion mechanisms.

Anterior approach refers to a method of hepatectomy which is first to resect the hepatic parenchyma and then to free the liver; hanging maneuver refers to placing a tape before the inferior vena cava for hanging the liver during hepatectomy.In October 2011,anatomical trisectionectomy was performed on a 54-year-old male patient with large hepatocellular carcinoma in the left medical lobe and right lobe with anterior approach and hanging maneuver.The diameter of the tumor was 16 cm,and was in the ⅢA/T3NOM0 stage.The indocyanine green retention at fifteen minutes was 5.4％,and the ratio of hepatic left lateral lobe volume over the standard total liver volume was 44％.The left bile duct was slightly dilated because of the compress of the tumor.The operation started with the isolation and dissection of the inflow vessels,including the right hepatic artery,the right portal vein,the middle hepatic artery,the portal vein branches of left internal lobe.The hepatic parenchyma transection was performed along the fight side of the falciform ligament.A tape was passed between the anterior surface of inferior vena cava and liver,and the liver was suspended during the transection.The left bile duct was cut at the right side of round ligament,and then the middle hepatic vein and the right hepatic vein were resected.The ligaments around the liver were dissected and the right hepatic lobe was removed.Finally,the end-toend anastomosis between the left hepatic duct and the common hepatic duct was performed.The operation lasted for 4 hours and the intra-operative blood loss was 350 mL.The patient was recovered well.At the end of 4 months after surgery,magnetic resonance cholangiopancreatography showed that the anastomosis of the bile duct was unobstructed,and there was no recurrence of tumor inside the liver.

A new magnetic affinity immunoassay (MAIA) strategy based on enzyme-labeled phage displayed antibody was developed. The assay consisted of a sandwich format in which immobilized polyclonal antibody (pcAb) on magnetic microparticle was used for capture probe, and enzyme-labeled phage displayed antibody for specific detection probe to increase enzyme amount and enhance detection signal. By the proposed method,β-bungarotoxin (β-BGT) was successfully detected. A linear relationship between absorbance value and the concentration of β-BGT in the range of 0. 016-62. 5 μg / L was obtained. The linear regression equation was Y=0. 641X+1. 355 (R =0. 9925, n = 13, p<0. 0001) with a detection limit of 0. 016 μg / L. In comparison with the traditional ELISA, this method gave a 10-fold better sensitivity in β-BGT detection. This strategy also gave a 4-fold better sensitivity comparing with the MAIA based on enzyme labeled monoclonal antibody (mcAb). Due to low detection limit, acceptable reproducibility and high specificity, this method holds great promise in toxin trace detection.

Objective:To express recombinant protein mIL-21-hIgGFc in 293E cells,and investigate its effect on CD8+T cell.Methods:Total RNA was extracted from the mouse spleen cells ,and then IL-21 gene was amplified by RT-PCR and inserted into expression vector PTT3-hIgGFc.PTT3-mIL-21-hIgGFc were transfected into 293E cells by calcium phosphate method.The supernatants were collected at 48 hours and 72 hours and concentrated by MOLLIPORE Labscale TM TFF system ( 5 kD membrane ).The mIL-21-hIgGFc fusion protein was purified with HiTrap TM Protein G column.The protein was quantified by SDS-PAGE and ELISA.The biological activity of the protein was determined by detecting the change of the phenotypes of CD 8+ T cells treated with the protein.Results: The constructed recombinant plasmid PTT 3-mIL-21-Fc was confirmed by sequencing.PTT3-mIL-21-Fc was transfected into 293E cells,mIL-21-Fc protein in culture supernatant was collected after 48 hours and 72 hours.The protein in cell su-pernatant reached a concentration of 787 ng/ml which was determined by ELISA.The protein was purified by Protein G chromatography column.P1A-specific T cells were treated with mIL-21-hIgGFc, and found that the CD44low CD62Lhi CD8+ population increased compared to the control.Conclusion:We built PTT3-mIL-21-hFc recombinant plasmid, expressed mIL21-hFc fusion protein in 293E cells,and purified by Protein G column.By treating mIL-21-hFc ,the antigen-primed CD8+T cells prefer to differentiate into CD44low CD62Lhi CD8+T cells which had been reported as a memory stem phenotype .This protein may be used to improve the effectness of adoptive T cell cancer therapy.

Objective To explore the diagnostic value of four tumor markers analyzed with Logistic regression and receiver operator characteristic (ROC) curve in patients with lung cancer.Methods The serum levels of carcinoembryonic antigen (CEA) 、carbohydrate antigen 125 (CA125),cytokeratin 19 fragment (CYFRA21-1) and neuron specific enolase (NSE) were determined by radioimmunoassay in 112 patients with lung cancer and 74 patients with benign pulmonary disease.Four tumor markers were analyzed by Logistic regression and ROC curve.Results The serum levels of CEA,CA125,CYFRA21-1 and NSE in lung cancer patients were [4.53(2.22-11.53)ng/ml,28.97 (11.39-62.10) U/ml,4.05(2.29-8.18)ng/ml,14.11 (11.35-24.12) ng/ml],respectively,which were significantly higher than those in health adults[2.08 (1.45-2.52) ng/ml,12.90 (9.80-19.44) U/ml,1.53 (1.21-2.17) ng/ml,11.38 (9.07-12.80) ng/ml] (all P ＜ 0.01).According to regression equation Y =1/[1 + EXP (4.902-0.394X1-0.627X2-0.165X3)],the area under the ROC curve (AUC),sensitivity,specificity,and accuracy of new variable Y were 0.915 ± 0.020,79.46％,93.24％,and 84.95％,respectively.Conclusions Application of logistic regression and ROC curve increases diagnostic accuracy in lung cancer.

Objective To evaluate the role of nodal gene modulating malignancy of a hepatocellular carcinoma cell lines SMMC7721. Methods To silence the expression of nodal gene in human hepatocellular carcinoma cells by RNA interference ( RNAi),and to observe the effect on cells biological behaviour and vasculogenic mimicry.4 expression vectors of nodal gene targeting small interference RNA were constructed and transfected into SMMC-7721 cells.Real-time quantitive PCR and Western blot were used to examine nodal gene expression. The effects of nodal gene RNA interference on proliferation,apoptosis,infestation,migration and vasculogenic mimicry of SMMC-7721 were studed. Results The expression of nodal gene was suppressed in SMMC-7721 cells by RNA interference.In the first 4,5,6 days of proliferation experiment,the proliferation of interference group was significantly lower than the control group(separately F =17 098.922,18 135.107,32 641.075,all P ＜ 0.05 ); 48 h after transfection,the apoptosis rate of interference group was significantly higher than the control group (F =1136.452,P ＜0.05); In the infestation and migration experiments,the cells through the transwell chamber in the interference group were less than the control group( separately F =83.6,1126.857,all P ＜ 0.05 ) ; 24 h and 48 h after transfection,the vasculogenic mimicry in the interference group did not form which was significantly different from the control group. Conclusions Interfering the expression of nodal gene inhibits the malignant biological behaviour and the formation of vasculogenic mimicry in human hepatocellular carcinoma cells.

Objective: To quantitatively study the morphological structure and parameter of right atrial appendage (RAA) by 256-slice spiral CT to provide RAA imaging reference for interventional or surgical therapy in clinical practice. Methods: A total of 200 patients with negative CTA results examined in our hospital were studied including 96 male and 104 female; by age division, Age≤40 years group,n=29, Age (40-60) years group,n=114 and Age>60 years group,n=57. The original scanning data was reconstructed for RAA establishment, indexes of volume, radial lines were measured and compared between different gender and age groups to obtain 95% normal imaging references of RAA. Results:①The volume, height, basilar part circumference, anteroposterior spread of RAA in male were greater than female, allP<0.05; with surface area standardization, the long and short diameter of basilar part, area, circumference in female were greater than male, allP<0.05.②Anteroposterior angle and spread of RAA in Age<40 years group were higher than both Age (40-60) years group and Age>60 years group,P<0.05, the above parameters were similar between Age (40-60) years group and Age>60 years group,P>0.05.③95% normal reference ranges of RAA were as following: volume (4.83-19.97) ml, height (21.46-41.80) mm; basilar part longest diameter (25.89-41.65) mm, short diameter (19.34-36.24) mm, area (459.30-1093.28) mm2, circumference (86.16-147.04) mm; anteroposterior angle (1.67-31.23) °, spread (8.07-34.37) mm. Conclusion: 256-slice CT may quantitatively study the morphological structure and parameter of RAA and establish its 95% normal imaging references for clinical practice.

Objective To explore the expression and relationship of PTEN and mTOR in the development of cholangiocarcinoma,determine the effect of the PI3K/PTEN/AKT/mTOR signal transduction on the(development) of cholangiocarcinoma.Methods The expression of mTOR and PTEN in the cholangiocarcinoma was detected by the immunohistochemistry and RT-PCR method.Results Compared to normal tissues,the(expression) of mTOR gene in cholangiocarcinoma significantly increased,but the expression of PTEN gene(decreased).There was a negative correlation between mTOR gene and PTEN gene expression in(cholangiocarcinoma). Conclusions The expression of mTOR gene in cholangiocarcinoma was increased and the expression of PTEN was decreased.It suggested that the mTOR gene and PTEN gene could play an important role in the process of development of cholangiocarcinoma.

AIM: To investigate the effects of the selective mitochondrial ATP-sensitive K+ channels opener diazoxide on mitochondrial respiratory function and enzyme activity in isolated rat myocardium under ischemia/reperfusion.METHODS: Observation was made on rat hearts perfused with Langendorff apparatus.72 Sprague-Dawley(SD) rats were randomly divided into 4 groups: normal group(NOR),ischemia reperfusion(IR),diazoxide group(DIA) and 5-hydroxydecanoate(5-HD) antagonized diazoxide group(5HD-DIA).Hearts isolated from SD rats were mounted on a Langendorff apparatus and started with a 20 min perfusion for equilibration.NOR went on perfusion for another 100 min after equilibration.IR underwent 40 min global ischemia and followed by 30 min reperfusion after 30 min stabilization.DIA was administered with K-H solution containing diazoxide at concentration of 50 ?mol/L for 10 min before ischemia and reperfusion.5HD-DIA was infused with 100 ?mol/L 5-HD(a specific mitochondrial ATP sensitive K+ channel blocker) and the same procedure was carried out as DIA group.Hearts were taken down to extract mitochondrial at the end-equation,before ischemia and at the end-reperfusion for determination of mitochondrial respiratory function and the enzyme activity of mitochondria.RESULTS: At the end of reperfusion,mitochondrial respiratory function(mitochondrial respiratory control rate,P/O ratio and state 3 respiration) and mitochondrial enzyme activity(NADH oxidase,succinate oxidase and cytochrome C oxidase) in DIA group were better than those in IR group and 5HD-DIA group(P0.05).CONCLUSION: Preconditioning with mitochondrial ATP sensitive potassium channel opener,diazoxide,protects rat heart mitochondria against ischemia-reperfusion injury.The mechanisms are involved in the safeguarding of respiratory function and activity of enzymes of respiratory chain.

Objective To evaluate the prevalence and risk factors of progressive liver fibrosis in patients with non-alcoholic fatty liver disease (NAFLD).Methods A total of 2 054 subjects who underwent health check up and were diagnosed as NAFLD in 9 institutions were included in the study.Blood routine and biochemical findings were collected to calculate aspartate aminotransferase-to-platelet ratio index (APRI).Subjects were divided into three groups according to diagnostic threshold of liver fibrosis:APRI ＜0.43 group,APRI 0.43-0.53 group and APRI ≥ 0.54 group.The correlation between APRI and biochemical variables was analyzed,and the risk factors of progressive fibrosis were also analyzed.Results Among 2 054 subjects (male/female 1 598/456) there were 61 cases with APRI ≥ 0.98 (2.97％,progressive fibrosis),318 with APRI ≥0.54 (15.48％),1 475 with APRI ＜ 0.43 (71.81％),261 with APRI 0.43-0.53 (12.71％).Logistic stepwise regression analysis showed that TG (P =0.002,OR =1.095,95％CI:1.033-1.161),2 hPG(P =0.000,OR =1.103,95％ CI:1.058-1.151,BUN(P =0.034,OR =1.215,95 ％ CI:1.014-1.454) were risk factors,and H DL-C (P =0.034,OR =0.353,95 ％ CI:O.135-0.924) was a protective factor for the progression of fibrosis.Conclusion The progressive fibrosis in patients with NAFLD is closely associated to blood glucose and lipid metabolism disorder.