Objectives:To observe the stress response and hemodynamic change during laparoscopic cholecystectomy(LC) under general anesthesia and general-epidural combined anesthesia.Methods:28 patients of LC were divided into general anesthesia group(G group n=14) and general-epidural anesthesia group(GE group n=14) randomly.The HR、SBP、MAP,plasma concentration of epinephrine(E),norepinepherine(NE),blood glucose、cortisol,PaCO2 and pH were measured before and after peritoneal insufflation of CO2.Results:The HR,SBP and MAP were increased significantly in group G than those in group GE(P

AIM: To establish the monoclonal antibody against human B lymphocyte stimulator (hBLyS) by DNA immunization and analyse its characterization. METHODS: The 858 bp DNA fragment of hBLyS was cloned into pcDNA3 plasmids. The cloned insert was identified by both sequence analysis and double digestion of the recombinant plasmid with restriction enzymes Xho I and EcoR I . After the splenocytes from BALB/c mice immunized with the recombinant plasmid of pcDNA3/hBLyS were fused with myeloma cells SP2/0,the hybridoma which can produce monoclonal antibodies against hBLyS were obtained. The specificity of anti-BLyS monoclonal antibody from hybridoma was verified by ELISA, Western blot and flow cytometry. RESULTS: The recombinant mammalian cell expression vector of pcDNA3/hBLyS was constructed, the sequence of the insert gene was identified to be the sequence encoding hBLy S antigen. The culture supernatants of hybridoma 9c10 were tested to be the monoclonal antibody with specificity against hBLyS on human peripheral blood CD3 + T cell activated by hIFN-? by ELISA, Western blot and flow cytometry. CONCLUSION: The monoclonal antibodies against hBLyS with high activity and specificity have been established successfully, and will be an useful tool in the studies of relationship between hBLyS and human autoimmunity diseases.

Objective To investigate the anti-proliferation effect of peroxisome proliferator-activated receptor γ(PPARγ)agonist troglitazone(TGZ)on leukemic Raji cells and its mechanisms.Methods Raji cells in culture medium in vitro were given different concentrations of TGZ(0～60 μmol/L)for 24,48 and 72 h.The inhibitory rates of the cells were measured by MTT assay,cell apoptotic rate was detected by flow cytometry(FCM),agarose gel electrophoresis was used to observe the DNA ladder,and western blotting was used to analyzed the variation of apoptosis related proteins bcl-2,Bax and Survivin.Results TGZ(over 20 μmol/L)could inhibit the growth of Raji cells and cause apoptosis remarkably,the suppression was both in time-and dose-dependent manner.DNA ladder was observed after the cells treated by TGZ for 72 h,and western blotting analysis revealed that anti-apoptotie proteins Survivin and bcl-2 were decreased remarkably while pro-apoptotic protein Bax increased significantly after the cells were treated by TGZ for 48 h.Conclusion PPARγ agonist TGZ can inhibit the growth and induce apoptosis on Raji cells significantly,downregnlating the expression of Survivin and bcl-2 as well as upregulating of Bax expression of Raji cells may be one of its most important mechanisms.

A novel chitosan derivant, N-octyl-N-arginine chitosan (OACS) with a mimetic structure of cell-penetrating peptides was synthesized by introducing hydrophilic arginine groups and hydrophobic octyl groups to the amino-group on chitosan's side chain. Structure of the obtained polymer was characterized by FT-IR and 1H NMR. The substitution degree of octyl and arginine groups was calculated through element analysis and spectrophotometric method, separately. The critical micelle concentration of OACS was 0.12 - 0.27 mgmL(-1) tested by fluorescence spectrometry. The solubility test showed OACS could easily dissolve in pH 1 - 12 solutions and self-assemble to form a micelle solution with light blue opalescence. The OACS micelles have a mean size of 158.4 - 224.6 nm, polydisperse index of 0.038 - 0.309 and a zeta potential of +19.16 - +30.80 mV determined by malvern zetasizer. AFM images confirmed free OACS micelle has a regular sphere form with a uniform particle size. MTT test confirmed that OACS was safe in 50 - 1 000 micromol-L(-1). The result of HepG2 cell experiment showed that the cell internalization of OACS micelles enhanced with increased substitution degree of arginine by 40 folds compared to chitosan. Thus, OACS micelles were a promising nano vehicle with permeation enhancement and drug carrier capability.

Objective To investigate the effects of lipnsomal prostnglandin E1 (Lipo-PGE1) on the renal function in patients undergoing cardiopulmonary bypass (CPB) .Methods Twenty ASA Ⅱ or Ⅲ patients of both sexes aged 34-56 yr weighing 48-81 kg scheduled for elective cardiac valve replacement with CPB were randomly divided into 2 groups (n = 10 each): normal saline group (group NS) and Lipo-PGE1 group. In group Lipo-PGE1, Lipo-PGE1 was infused at 3 ng·kg-1·min-1 from the initiation of CPB to the end of CPB, and then added into the priming solution of the extracorporeal circulation machine with its concentration set at 5 ng/ml. In group NS, the equal volume of normal saline was administered instead of Lipo-PGE1 . The concentrations of thromboxane B2 ( TXB2 ), 6-keto-prostnglandin F1α ( 6-Keto-PGF1α ), and free hemoglobin ( F-Hb ) in plasma, and α1-microgipbulin (α1-MG), β2-micmglobulin (β2-MG) and Cystatin C (Cys C) in serum were measured after heparinization and before CPB (T1), at 30 min of CPB (T2) and at 0, 1 and 24 h after termination of CPB (T3-5). Results Compared to T1, the concentrations of TXB2, 6-Keto-PGF1α, α1-MG and β2-MG and TXB2/6-Keto-PGF1α ratio at T2-4, and the concentrations of F-Hb and Cystatin C at T2-5 were significantly increased in both groups ( P < 0.05). The concentration of 6-Keto-PGF1α at T2-4 was significantly higher, and concentrations of TXB2, α1-MG and β2-MG and TXB2/6-Keto-PGF1α ratio at T2-4 and concentrations of F-Hb and Cys C at T2-5 were significantly lower in group Lipo-PGE1 than in group NS ( P < 0.05). Conclusion Lipo-PGE1 can obviously improve the renal function in patients undergoing CPB.

Objective To establish a method for determining oxalate and citrate in urine simultaneously by capillary electrophoresis.The components,the concentration and pH of the buffer solution,the separation voltage and the injection time on theseparation were studied in detail.Methods The separations were carried out using potassium dihydrogen phosphatebuffer ina fused-silica capillary tubeby capillary zone electrophoresis (CZE) and the detection were monitored by UV.24 h-urine samples from patients (n =5) and health control (n =5) were collected from Zhongshan Hospital of Fudan University for systematically validating the method developed.Results The optimized separations were carried out using a 50 mmol/L potassium dihydrogen phosphatebuffer solution (pH 6.5) in a fused-silica capillary tube of 50 cm × 50 μm I.D.Injections were made by using the pressure mode for 10 s at 34 mbar.The detections were monitored by a UV at 200 nm after samples were separated at avohage of 30 kV.Under the seconditions,urinary oxalate and citrate were separated completely within 5 min.The relative standard deviations of migration time and peak area within-run foroxalate and citrate were less than 1％ and 3.0％ and the betweenrun relative standard deviations were less than 2.0％ and 4.0％,respectively.The detection limits were 1 mg/L for both oxalate and citrate.The linearity ranges of oxalate and citrate were both 0-500 mg/L with the correlation coefficient between 0.999 5 and 0.995 4 (P ＜ 0.05),respectively.The average recoveries were 102.38％ for oxalate and 92.74％ for citrate.Conclusion This method is proved to be simple,sensitive and accurate,and also applied to determine oxalate and citrate in urine samples with satisfactory results.

Objective To evaluate the performance of sST 2 ELISA kit and investigate the clinical application of sST2.Methods This verification study validated the precision , linearity of sST2 ELISA kit according to the CLSI EP-15A, EP-6A protocols.300 healthy adults(aged from 20 to 85, 124 male and 176 female) from 5 different districts of Shanghai were used to establish serum sST 2 reference interval .The correlations between sST2, NT-ProBNP, LVEF and NYHA class were analyzed in 117 patients diagnosed with heart failure who were grouped according to the New York heart association ( NYHA).Receiver operating characteristic (ROC) curve was used to compare the ablity of sST2, NT-ProBNP, LVEF in distinguishing heart failure patients .Results The within-lot and between-lot variation of three level samples were below 4% and 10% respectively.There was a good linear correlation ( Y=0.995X+0.005, R2 =0.999) between theoretical value and actual detection result in the range of 0 to 200 μg/ml.The reference interval of sST2 was 10.2 to 41.0μg/ml for males and 8.9 to 28.1μg/ml for females.sST2 was positively correlated with NT-ProBNP and NYHA class but did not correlate with LVEF in heart failure patients . Patients with NYHA class>II (Median:28.3,IQR:19.5-39.2)had higher serum sST2 level than patients with NYHA class≤II (Median:45.1,IQR:34.1 -85.6), P<0.05.The AUC of sST2 in distinguishing heart failure patients from normal people was 0.815(sensitivity :51.2%,specificity:92.7%).The AUC of sST2 ,sST2+NT-ProBNP and sST2+NT-ProBNP+LVEF in distinguishing patients between NYHA class≤II and>II were 0.743, 0.810, 0.831 respectively and the sensitivity of sST 2 +NT-ProBNP+LVEF was 94.7%.Conclusions Experimental results show that this sST 2 ELISA kit has a good performance in the precision, linearity.sST2 correlates with NT-ProBNP and NYHA class but do not correlates with LVEF . Serum sST2 level is not influenced by age , BMI, renal function.sST2 could be a good supplement of NT-ProBNP and LVEF in distinguishing patients between NYHA class≤II and>II.

Objective To evaluate the performance of serum sialic acid detection kit using enzymic method and investigate the clinical diagnosis value of sialic acid.Methods one hundred and fifty healthy adults were enrolled in this case control study to establish serum SA reference interval.The analytical performance (accuracy,precision,linearity) of serum sialic acid detection kit using enzymic method was assessed.Two hundred and forty patients were classified into different malignant tumor groups according to their pathological types.Serum SA level of each tumor group was compared with that of normal control group.In tumor groups with statistical difference,benign disease groups were further collected.Receiver operating characteristic (ROC) curve and area under curve (AUC) were used to evaluate the diagnostic value of SA compared with other tumor markers.t test,one-way ANOVA,Mann-Whitney U test were used as statistical methods.Results The reference interval of SA was 479 to 715 mg/L.The detection result of 2 level controls was 584 and 1 482 mg/L respectively,which were both within the acceptable limits.The within-lot and between-lot variations of three level samples were both below 5％.There was a good linear correlation (Y =0.995X-0.177,R2 =0.999) between theoretical value and actual detection result in range of 0-1 052 mg/L.The serum level of SA was (757 ± 177),(514 ± 86) and (597 ± 60) mg/L in gastric cancer group,benign disease control group and normal control group respectively,which had statistically significant difference(F =55.2,P ＜ 0.01).The serum level of SA was(659 ± 127) and (545 ± 66) mg/L in colorectal cancer group and benign disease control group respectively,which had statistically significant difference(F =42.8,P ＜ 0.01).The serum level of SA was (738 ± 157) and (672 ± 161) mg/L in colorectal cancer group and benign disease control group respectively,which did not have statistically significant difference(F =26.3,P ＞ 0.05).The AUC of SA was 0.804,0.724,0.755 in gastric cancer group,colorectal cancer group and lung cancer group respectively,which was higher than that of CEA and CA72-4.In gastric cancer group,the sensitivity of SA was higher than that of CEA (59.5％,24.3％).The AUC of SA was 0.791,0.687,0.790 in gastric cancer,colorectal cancer and lung cancer patients with normal CEA serum level respectively.Conclusions Experimental results show that serum sialic acid detection kit using enzymic method has good performance in the precision and linearity.Sialic acid has some value in the diagnosis of gastric cancer and colorectal cancer and could be a good supplement of CEA in screening of cancer.

Objective To investigate the effect of ulinastatin treatment on the inflammatory factor expression and prognosis in patients with ventilator-associated pneumonia (VAP). Methods One hundred patients with VAP were enrolled, and the patients were given the standardized treatment of VAP. The patents were divided into high dose group (33 cases, using the ulinastatin 20 000 U/d), normal dose group (34 cases, using the ulinastatin 10 000 U/d) and control group (33 cases, no using the ulinastatin) by random digits table method. The serum C-reactive protein (CRP), procalcitonin, interleukin (IL)-6 and tumor necrosis factor (TNF)-αlevels at the first, third, fifth and seventh day of diagnosis were detected. All the patients were followed up for 1 month, and the antibiotics treatment time, mechanical ventilation time, ICU stay time and mortality were recorded. Results The CRP, procalcitonin, TNF-αand IL-6 from the first day of diagnosis to the seventh day of diagnosis in 3 groups showed the downward trend, and there were statistical differences (P<0.05). There were no statistical differences in CRP and procalcitonin at the third, fifth and seventh day of diagnosis among 3 groups (P>0.05). At the third, fifth and seventh day of diagnosis, the TNF-α levels in high dose group were (46.02 ± 4.65), (23.88 ± 7.76) and (11.05 ± 2.56) ng/L, the IL-6 levels were (15.53 ± 4.54), (11.33 ± 3.45) and (6.62 ± 2.45) ng/L;the TNF-αlevels in normal dose group were (56.02 ± 6.42), (38.88 ± 9.34) and (27.05 ± 3.42) ng/L, the IL-6 levels were (18.23 ± 2.45), (15.33 ± 4.34) and (11.23 ± 3.34) ng/L; the TNF-α levels in control group were (68.13 ± 4.77), (52.88 ± 7.46) and (42.12 ± 3.76) ng/L, the IL-6 levels were (20.02 ± 3.23), (17.23 ± 2.34) and (15.33 ± 2.33) ng/L. The TNF-αand IL-6 levels at the third, fifth and seventh day of diagnosis in high dose group were significantly lower than those in normal dose group and control group, and those in the normal dose group were significantly lower than those in control group, and there were statistical differences (P<0.05). The mechanical ventilation time, antibiotics treatment time and ICU stay time in high dose group were significantly shorter than those in normal group and control group:(15.34 ± 5.67) d vs. (18.44 ± 6.32) and (22.34 ± 5.21) d, (7.45 ± 2.54) d vs. (10.45 ± 4.56) and (14.43 ± 6.24) d, (18.42 ± 7.45) d vs. (20.43 ± 4.98) and (26.35 ± 5.97) d, and those in normal group were significantly shorter than those in control group, and there were statistical differences (P<0.05). There was no statistical differences in the mortality among 3 groups (P>0.05). Conclusions Ulinastatin can inhibit the expression of IL-6 and TNF-α in patents with VAP, shorten the antibiotics treatment time, mechanical ventilation time, ICU stay time and mortality, and improve prognosis.

Objective To explore the role of toll like receptors-2 and toll like receptors-4 in the pathogenesis of rheumatoid arthritis(RA). Methods The expression rate and intensity of TLR2 and TLR4 on the CD14 positive monocytes in peripheral blood (PBMC) of 32 RA patients were determined by multi-colore immunofluorescence flow cytometry. Results ①The expression rate and intensity of TLR2 and TLR4 on the CD14 positive monocytes in peripheral blood of RA patients were higher than that of normal control. ② There was no difference in the expression pattern between the early stage RA group and the late stage RA group. ③The intensity of TLR2, but not TLR4 was correlated with serum CRP level. Conclusion The results indicate that innate immunomolecules such as TLR2 and TLR4 may be involved both in the initiation stage and perpetuating stage of RA.