Abstract:Objective: To explore the roles of thrombospondin-1 (TSP-1) and transforming growth factor-β1 (TGF-β1) in mediating the secretion of extracellular matrix (ECM),such as fibronectin (FN) and collagen Ⅳ,from rat glomerular mesangial cell (GMC) induced by sublytic C5b-9 complexes. Methods: Rat GMC was cultured and divided into different groups. The expression levels of TSP-1 and TGF-β1 as well as FN and collagen Ⅳ in GMC with sublytic C5b-9 stimulation were measured. In addition,the synthetic blocking peptide of TSP-1(GGWSHW) and TGF-β1 neutralizing antibody were utilized to explore the role of TSP-1 and TGF-β1 in ECM secretion from GMC induced by sublytic C5b-9,as well as the relationship between TSP-1 and TGF-β1. Results: The protein levels of FN and collagen Ⅳ as well as TSP-1 and TGF-β1 including the activated TGF-β1 were significantly increased in rat GMC at 18 h after sublytic C5b-9 stimulation. The blocking peptides of TSP-1 (GGWSHW) could significantly inhibit TGF-β1 activation and reduce the production of FN and collagen Ⅳ. In addition,TGF-β1 neutralizing antibody could also obviously inhibit the expression of FN and collagen Ⅳ in GMC upon sublytic C5b-9 attack. Conclusion: Sublytic C5b-9 can promote ECM secretion and TSP-1 and TGF-β1 production as well as TGF-β1 activation in rat GMC in vitro,while the secretion of ECM stimulated by sublytic C5b-9 is possibly mediated in part by TSP-1-regulated TGF-β1 activation.

Abstract:Objective:To investigate the mechanism of deoxypodophyllotoxin(DOP) against the peripheral nervous system. Methods: The effects of DOP on voltage-dependant K+ channels were studied by using whole-cell patch-clamp recordings on acute isolated rat dorsal root ganglion(DRG) neurons. Results: DOP can reversibly block voltage-dependent potassium current(IK) and delayed-rectifier potassium current(IKDR) and make the curve of the current-voltage relation shift to positive potentials at the concentration of 10,20,40 and 80 μmol/L. This study thus demonstrated that DOP is a dose-dependant and reversibly blocker of IK and IKDR in DRG neurons. The inhibition rate of DOP on IKDR was higher than on IK. Conclusion: It can be concluded that the effect of DOP on the current of potassium channel of DRG neurons may be a possible contributing mechanism for the toxin on peripheral nervous system.

Abstract:Objective: To construct prokaryotic expression recombination plasmids containing GST-GSK3β1,GST-GSK3β2 and dominant-negative mutant GST-GSK3β1 K85M,then express,purify and identify these three fusion proteins. Methods: Using PCR and recombinant DNA technology,the target genes were amplified from templates Flag-GSK3β1,YFP-GSK3β2 and Flag-GSK3β1 K85M respectively and inserted into the prokaryotic expression vector pGEX-6P-1 with glutathione-S-transferase(GST) tag to construct the expression plasmids expressing GST-GSK3β1,GST-GSK3β2 and GST-GSK3β1 K85M fusion proteins. An effective mean of expression and purification was established by a series of expression,gentle lysis and purification experiments. In the end,Western blot was used to identify the purified fusion protein. Results: We succeed in constructing wild type and dominant-negative mutant plasmids of GST-GSK3β,expressing and purifying these fusion proteins. Conclusion: The successful construction,expression and purification of those target proteins facilitate us to further study the biology function of GSK3β.

Abstract:Objective: To investigate the potential influence of RANTES on the expression of membranous and cytoplasmic protease activated receptors (PARs) in P815 mast cells. Methods: P815 mast cells were challenged with different concentrations (0.1,1.0,10.0,100.0 ng/ml) of RANTES. The expression of PAR1,PAR2,PAR3 and PAR4 in the time of 2,6 and 16 h were determined by flow cytometry. Results: RANTES significantly down-regulated the expression of membranous rather than cytoplasmic PAR1,and the potentiated effect was abolished by monoclonal anti-RANTES. Nevertheless,RANTES had no effect on the expression of PAR2,PAR3 and PAR4. Conclusion: RANTES can selectively up-regulate membranous PAR1 expression in P815 mast cells,which reveals a novel immunobiologic profile of RANTES as a potential regulator of mast cells function.

Abstract:Objective: To investigate the possible mechanism of Smurf2 in the progression of mice renal interstitial fibrosis induced by unilateral ureteral obstruction(UUO). Methods: The UUO model was established through ligating left ureters of male CD-1 mice (UUO group). At the same time,the control group were set up by sham-operation of left kidney of CD-1 male mice(sham group). The animals were sacrified at day 3,7,14 after operation. The changes of kidney structure and the lesions of renal interstitial fibrosis were observed using HE,PAS and Masson’s trichrome staining assays. Furthermore,the expressions and distributions of Smurf2,E-cadherin,alpha-smooth muscle actin (-琢-SMA) and fibronectin (FN) in obstructed and normal kidneys were detected by Western blot and immunohistochemical staining assays. Results: Compared with the sham group,at day 7 after UUO operation,the atrophy and dilution of renal tubules,renal interstitial fibrogenesis were found in the obstructed kidneys. After UUO operation,the expression of Smurf2--琢-SMA or FN protein was upregulated in an time-dependent manner,respectively;while the expression of E-cadherin was remarkably decreased. Moreover,the Smurf2 expression was mainly distributed in the nucelus of the renal tubular epithelial cells;while the distribution of -琢-SMA protein was obviously posited in renal tubular epithelial cells and renal interstitium,and the distribution of FN was in renal interstitium of the obstructed kidneys. By regression analyses,the upregulation of Smurf2 expression had positive relationship with -琢-SMA(r = 0.765,P < 0.001) and FN (r = 0.773,P < 0.01),and negative relationship with E-cadherin (r = -0.656,P < 0.001)-Conclusion: These results suggest that Smurf2 maybe act as a key in the procession of renal interstitial fibrosis of obstructive nephropathy mice through inducing renal tubular epithelial-mesenchymal transition.

Abstract:Objective: To study the changes of phosphorylated eukaryotic translation initiation factor 2α[eIF2α(p)]and expression of activing transcription factor 4(ATF4) in neonatal rats with hypoxic-ischemic brain damage(HIBD). Methods: Eighty 7-day-old Sprague-Dawley rats were randomly divided into 2 groups:sham operation group(n = 40) and HIBD group(n = 40). HIBD models were established by Rice method. At 6,12,24 and 72 h after hypoxia-ischemia (HI),the brain tissue of rats in HIBD group was obtained to prepare section. The changes of phosphorylation of eIF2α and expression of ATF4 in the lesion side cerebral cortex in rats were detected by immunohistochemical staining. The sham operation group was dealt with the same method without HI. Results: The expression level of eIF2α (p) and ATF4 increased at 6 h after HIBD,peaked at 12 h,decreased significantly at 72 h,but was still higher than that of sham operation group(P < 0.01). The differences in the expression level of eIF2α(p) and ATF4 in HIBD group among different time points were significant(P < 0.01). Conclusion: eIF2α(p) and ATF4 participate in the pathophysiological process of HIBD. HIBD may induce endoplasmic reticulum stress reaction,which initiate PKR-like endoplasmic reticulum kinase(PERK) mediated unfolded-protein response(UPR) signal pathway.

Abstract:Objective: To construct pRetro-On/LTβR vector and obtain LTβR-expressing Jurkat cells,and explore the role of LTβR in T cells apoptosis. Methods: LTβR cDNA was obtained by PCR amplification and subcloned into pRetro-On plasmid. Then E.coli was transformed by the recombinant plasmid and positive clones under LB-Amp medium screen were used for recombinant plasmid extraction and purification. The recombinant plasmid pRetro-On/LTβR was identified by NotⅠ and BamHⅠ double enzyme digestion and DNA sequencing. Jurkat cells,a kind of T cell line,was transfected by the recombinant plasmid with Lipofectamine 2000. LTβR expression in Jurkat cells was induced by doxycycline(DOX) and identified by Western blot analysis and flow cytometry. The apoptosis of Jurkat cells induced by interaction of LTβR with the ligand,LIGHT,was detected by the method of annexin-V-PI double-stain flow cytometry. Results: Double enzyme digestion and DNA sequencing results confirmed that LTβR cDNA was inserted correctly into pRetro-On plasmid. The results of Western blot analysis and flow cytometry confirmed the LTβR expression in Jurkat cells. Apoptosis of transfected Jurkat cells significantly increased after LIGHT stimulation. Conclusion: The pRetro-On/LTβR was successfully constructed,and LTβR could be expressed in Jurkat cells using the recombinant plasmid. The apoptosis of LTβR-expressing Jurkat cells can be induced through LIGHT-LTβR pathway.

Abstract:Objective: To clone the hemagglutinin(HA) gene from H5N1 highly pathogenic avian influenza virus,and get eukaryotically expression. Methods: Total RNA was extracted from virus A/Jiangsu/08-6(H5N1),and used to reversely transcribe cDNA. HA gene was amplified with HA gene specific primers and then cloned into the pFastBac-GP67B plasmids with digestion by restriction enzyme BamHⅠ and XhoⅠ. The recombinant plasmid pFastBac-GP67B-HA was identified by restriction enzyme and gene sequencing. Following the transformation of E. coli DH10Bac competent cells with pFastBac-GP67B-HA,the recombinant bacmid rBacmid-HA was identified by blue/white selection and confirmed by PCR analysis. Then rBacmid-HA was transfected into sf9 cells using the Cellfectin Ⅱ reagent to eukaryotically express HA protein. The recombinant HA protein was characterized by SDS-PAGE,Western-blot and hemagglutination test and mass spectrographic analysis. Results: HA protein was efficiently expressed in sf9 cells with a molecular weight of about 70 000. In hemagglutination test,the recombinant protein HA can agglutinate chicken erythrocytes significantly. The mass spectrographic analysis confirmed the protein identity. Conclusion: The recombinant HA protein has the same biological activity as the natural HA protein,and the protein could be used to develop influenza specific subunit vaccine and functional antibody in future research.

Abstract:Objective: To construct the recombinant eukaryote expression vector containing Arf6-T27N gene and detect the protein expression in its target cell line,MDA-MB-231. Methods: The fragment of Arf6-T27N gene from expression vector pXS-Arf6-T27N was cloned into the eukaryote expression vector pEGFP-N1,then the recombinant vector pEGFP-N1/Arf6-T27N were transfected into the MDA-MB-231 cells. After transfection of the recombinant vector,the protein expression of Arf6-T27N in MDA-MB-231 cells were detected by Western blot. The effect of Arf6-T27N on cell migration was investigated by Boyden chamber assay. Results: The recombinant eukaryote expression vector carrying Arf6-T27N was constructed successfully. The expression of Arf6-T27N in MDA-MB-231 cells could be detected by Western blot. In addition,Arf6-T27N protein could inhibit the migration of MDA-MB-231 cells. Conclusion: The new recombinant expression vector pEGFP-N1/Arf6-T27N was constructed successfully and the MDA-MB-231 cells which could express Arf6-T27N stably were also successfully made. Furthermore,our study suggested that Arf6-T27N protein could inhibit the migration of MDA-MB-231 cells.

Abstract:Objective:To investigate the influence of 32P on the biodegradation and radioactivity release rate of 32P-chromic phosphate-poly (L-lactide)(32P-CP-PLLA) seed. Methods:32P-CP-PLLA seeds with increased radioactivity and specific radioactivity were prepared(noted as S1-5) and their biodegradation,weight loss,radioactivity release rate and ultrastructure changes were investigated in different releasing medium such as fetal bovine serum,saline and dextran(noted as X,Y,Z),and after intratumor implantion. Results:32P-CP-PLLA seeds with increasing radioactivity were demonstrated increased radiation release rate at same time points,especially since day 32 and the increase seems corresponding with the change of the radioactivity of seed. And the increase of biodegradational weight loss matched with the increase of radiation release rate after intratumor implantation. The radiation release rate was much faster in the fetal bovine serum(sterile environment) than that in other medium and in vivo,and bacteria were found in the seeds by electron microscope. Conclusion: 32P may serve as one important modulator of the biodegradation and radiation release rate of 32P-CP-PLLA seed,and bacteria with possible secreting matters may also influence the biodegradation of PLLA.

Abstract:Objective: To study the effect of hypoxia on extracellular release of high mobility group box 1(HMGB1) in hypoxia-induced alveolar macrophages and its potential signaling pathway. Methods: The changes of HMGB1 mRNA expression and HMGB1 concentration in the culture medium were investigated after rat alveolar macrophages (NR8383) were subjected to hypoxia (1% O2) for different hours. The effect of JAK/STAT pathway inhibitors (tyrphostin AG 490 and fludarabine) with various concentrations on the HMGB1 release was observed. HMGB1 concentration was determined by enzyme-linked immunosorbent assay (ELISA). Results: HMGB1 concentration in the culture medium and HMGB1 mRNA expression significantly increased in a time-dependent manner after NR8383 cells were subjected to hypoxia culture. Tyrphostin AG 490 and fludarabine partly inhibited extracellular release of HMGB1 in hypoxia-induced NR8383 cells in a dose-dependent manner. Conclusion: Hypoxia induces alveolar macrophages to release HMGB1,the mechanism may be involved with intracellular JAK/STAT signaling pathway.

Abstract:Objective: To investigate the change and potential significance of adipokine in the rat of acute pancreatitis. Methods:Forty Sprague-Dawley rats were randomly divided into four groups: control group,sham operation group,acute edematous pancreatitis (AEP) group and acute necrotizing pancreatitis (ANP) group. AEP model was induced by intraperitoneal injection of cerulein and ANP model was induced by intraperitoneal injection of L-arginine while the control group didn’t receive anything,the sham operation group received same volume of 0.9% normal saline injected. The level of amylase,resistin,leptin,adiponectin,TNF--琢,IL-1β in rats blood serum were determined by ELISA,and pancreas/body weight ratio was evaluated and pancreatic pathology was observed. Results: The levels of amylase,pancreas/body weight ratio,pancreatic pathology,resistin,leptin,TNF--琢 and IL-1β in AEP group and ANP group were significantly higher than those in the control group and the sham operation group(P < 0.01),which in ANP group were higher than those in AEP group (P < 0.01 or P < 0.05). The levels of adiponectin in both AEP and ANP group were (3.26 ± 0.68),(2.27 ± 0.45) mg/L,respectively,which were significantly lower than those in the control and sham operation group(P < 0.01),it was lower in ANP group than in AEP group (P < 0.05). The level of resistin was correlated with the increase of TNF--琢,IL-1-茁 and the pathologic severity of pancreatitis(r = 0.711,0.794,0.812,respectively,P < 0.01). The level of leptin was correlated with the increase of TNF--琢,IL-1β and the pathologic severity of pancreatitis(r = 0.736,0.812,0.785,respectively,P < 0.01 or P < 0.05). But the level of adiponectin was correlated with the decrease of TNF--琢,IL-1β and the pathologic severity of pancreatitis(r = -0.796,-0.809,-0.788,respectively, P < 0.01 or P < 0.05). Conclusion: The adipokine was correlated with the pathogenesis and development of acute pancreatitis. The adipokine may be useful to predict the severity and prognosis of acute pancreatitis. The increased levels of resistin and leptin may act to aggravate the severity of acute pancreatitis. But the increased levels of adiponectin may act to protect the severity of acute pancreatitis.

Abstract:Objective: To investigate the effect of bufalin against proteinuria in adriamycin (ADR)-induced nephritic rats and its possible mechanisms. Methods: Forty SD rats were randomly divided into three groups: control group,model group and bufalin group. At the beginning,7.5 mg/kg ADR was singly administered through the tail vein except for the control rats. After 7 days,the rats in bufalin group were injected intraperitoneally with 0.1 mg/kg bufalin everyday unitl day 28. Total 24-h urine samples of all groups were collected weekly. At the end,biological parameters and oxidative stress in blood serum were measured,the histopathology of the renal cortex were detected by light microscopy and transmission electron microscopy,the expression of integrin-linked kinase(ILK) was qualified by Western blot. Results: Bufalin could reduce the 24-h urinary protein and blood lipid,alleviate the podocyte damage,inhibit the oxidative stress and down-regulate ILK in the protein level in ADR rats. Conclusion: Bufalin may against proteinuria by protecting podocyte,and its possible mechanisms against podocyte damage are related to inhibit the oxidative stress and down-regulate ILK in the protein level.

Abstract:Objective: To observe the effects of silencing of Livin gene expression on the apoptosis of MG-63 osteosarcoma cells. Methods: Double stranded RNA targeting the Livin gene was chemically synthesized in vitro and transfected into MG-63 osteosarcoma cells by LipofectamineTM 2000. The transfection efficiency was observed by fluorescence confocal microscopy. Reverse transcription-polymerase chain reaction(RT-PCR) and Western blot were used to detect the expression of Livin at mRNA and protein levels. Apoptosis analysis was performed by annexin V staining. Results: The Livin mRNA expression of the Livin-siRNA transfected group was 0.195 ± 0.019,significantly lower than those of blank control(0.539 ± 0.031) and the negative control group(0.438 ± 0.029). The Livin protein expression of the Livin-siRNA transfected group was 0.165 ± 0.022,significantly lower than those of blank control(0.632 ± 0.034) and the negative control group (0.625 ± 0.035). Increased apoptotic cells were observed by flow cytometry. Conclusion: siRNA can inhibit Livin expression of MG-63 osteosarcoma cells and induce cell apoptosis. Livin might serve as a target for apoptosis-inducing therapy of osteosarcoma.

Abstract:Objective: To investigate the correlative factors of hyperuricemia and metabolic syndrome (MS) and study the role of uric acid (UA) in the development of MS. Methods: Data of 12 249 persons undergoing health examination were collected. The disease history was investigated,the biochemical indices were detected,and the general physical data were measured. Multiple stepwise by multivariate Logistic regression model were used to analysis the relationship between UA and MS,at the same time,age,sex,low density lipoprotein cholesterol,total cholesterol were adjusted,the relationship of UA and other factors related to metabolism was also analyzed. Results: Age,BMI,blood pressure,lipids,glucose and UA were higher in MS group than those in non MS group(P < 0.01). The UA level was higher in men than that in women,and increased with accompany the costitution of MS. This phenomenon was obviously in woman. The UA level was increased in man under different glucose levels. Similarly,the UA level was related to the abnormal lipid metabolism,but not related to ages. After adjusting age,sex,total cholesterol,low density lipoprotein cholesterol,the rate of MS were higher in the hyperuricemia group when the UA was analyzed as continuous variable,but when uricemia was divided into two groups,the rate of MS in hyperuricemia group was 1.197 times of that in no hyperuricemia group,but non significant difference was found between them. When uricemia was divided into four groups,there were no differences among them. Conclusion: Age,BMI,lipids,glucose,uricemia were the risk factors of MS,hyperuricemia was found in MS group,and hyperuricemia group has higher rate of MS than the non hyperuricemia group,but no direct relationship between MS and hyperurcemia was found when the uricemia was divided into two groups or four groups

Abstract:Objective: To investigate the independent risk factors related to the development of diabetic retinopathy (DR) for type 2 diabetic patients. Methods: A case-control study containing 82 DR patients and 86 diabetics patients without DR (NDR) was performed from January,2009 to December,2010. The clinical parameter and biochemical indicators were compared between the two groups. Conditional Logistic regression analysis was further performed to assess variables independently associated with the DR developing. Results: In the univariate analysis,duration of diabetes,HbA1c,diastolic blood pressure and retinol-binding protein 4 (RBP4) were significant between DR and NDR. Logistic regression analysis showed that duration of diabetes(OR=2.41;95%CI:1.12~5.18),lipoprotein (a)(OR=2.22;95% CI:1.15~4.31) and RBP4 (OR=2.95;95% CI:1.54~5.65) were significant and independent predictors of the DR developing. Conclusion: These data suggest that duration of diabetes,lipoprotein (a) and RBP4 level are independent risk factors for the development of DR in type 2 diabetics.

Abstract:Objective: To evaluate the association of retinol binding protein (RBP4) with large vascular disease in type 2 diabetes mellitus(T2DM). Methods: There were 198 T2DM patients and 100 controls in study. The patients were divided into two groups based on with or without large vascular disease,and their clinical features,metabolic indices were compared. The T2DM patients were also divided into three groups based on the concentrations of RBP4 and the occurrence of atherosclerosis(AS) were compared. The relevance of RBP4 and other parameters were analyzed by Pearson correlation analysis. The factors that can influence AS in T2DM patients were analyzed by Logistic regression analysis. Results: Serum RBP4 level was higher in T2DM than that in the controls(P < 0.05). From RBP4 low and mediam tertile group to RBP4 high tertile group,the prevalence of AS were increased from 14.5%,21.4% to 48.2%,respectively. The AS prevalence of RBP4 high tertile group was significantly higher than the other two groups. It was showed by Pearson correlation analysis that the concentrationg of RBP4 in T2DM patients was positively correlated with body mass index(BMI),systolic blood pressure(SBP),fasting blood sugar(FBS),postprandial blood sugar(PBS),fasting serum insulin(FINS) and HOMA-IR. It was showed by mutiple stepwise Logistic regression analysis that AS in T2DM patients was related to plasma RBP4,age,course of disease and LDL-C. Conclusion: AS in T2DM patients is positively correlated with plasma RBP4 levels. High plasma RBP4 may be one of independent risk factors of AS in newly diagnosed T2DM patients.

Abstract:Objective: To investigate the potential relationship between amnestic mild cognitive impairment (aMCI) and metabolic disorders(MD). Methods: Totally,136 persons were included and divided into aMCI group(male/female=44/49,average age=73.01 ± 7.12 years) and control group (male/female=25/18,average age=73.77 ± 7.37 years). The histories of MD were all recorded. Fasting blood glucose(FBG) and glycated hemoglobin(HbA1c) were detected by dry chemistry method. Interleukin 6(IL-6) was detected by enzyme-linked immunosorbent assay. Results: IL-6 was highly expressed in the aMCI group[(78.57 ± 2.37) pg/ml],compared with control group[(72.48 ± 4.74)pg/ml,P < 0.05]. There was no significant difference between the two groups in FBG and HbA1c (P > 0.05). Furthermore,the diabetes and hypertension were proved as the common risk factors(P < 0.05). Interestingly,smoking is a risk factor only in male aMCI patients. Conclusion: The present study demonstrated that the history of diabetes,hypertension and male smoking placed an important part in the generation of aMCI. The reason might partly be ascribed to the inflammation state caused by the risk factors.

Abstract:Objective: To investigate the influence of inhaling different concentration of sevoflurane on post-operative cognitive dysfunction (POCD) in the elderly patients and evaluate the correlation between serum of homocysteine(Hcy) and POCD. Methods: Sixty ASA Ⅰ~Ⅱ patients undergoing lower gastrectomy were randomly assigned to two groups,group A inhaling low concentration of sevoflurane (n = 30) and group B inhaling high concentration of sevoflurane (n = 30). After the induction of anesthesia,1.5% sevoflurane (group A) or 3.0% sevoflurane (group B) were given for anesthetic maintenance. The mimi-mental state examination (MMSE) was used to assess the cognitive function before and 6,24,72 h after the operation. The serum Hcy before and 2 h in the operation and 24,72 h after the operation were determined. Results: The difference of MMSE scores between two groups before the operation were insignificant (P > 0.05). Compared with the preoperative values,MMSE scores were significantly reduced at 6 h in two groups(P < 0.05),which is lower in group B than group A(P < 0.05). Compared with the preoperative values,the serum Hcy at 2 h in the operation and 24,72 h after the operation in group A increased insignificantly(P > 0.05), while the serum Hcy at 2 h in the operation in group B increased significantly (P < 0.05),and returned to baseline at 24,72 h after the operation(P > 0.05). Conclusion: Inhaling high concentration of sevoflurane was easier to result in POCD than inhaling low concentration of sevoflurane. Serum Hcy level was related to the concentration of inhaling sevoflurane.

Abstract:Objective: To investigate the differencial expression of miRNA-26a in allergic asthma children and murine asthma model. Methods: Expressions of miRNA-26a in allergic asthma children and ovalbumin-induced mouse asthma model were analyzed by real-time PCR. The number of cells in the bronchoalveolar lavage fluid (BALF) of murine asthma model was acessed. The expression of GSK-3β in murine asthma model was acessed by Western blot. Results: miRNA-26a was up-regulated in allergic asthma children and murine asthma model compared with normal controls. The total cell number and acidocyte number in BALF from murine asthma model were significant higher than control. Enhanced eosinophilic inflammation in lung was observed in murine asthma model. Protein level of GSK-3β in murine asthma model was lower than control. Conculsion: miRNA-26a expression level was higher in allergic asthma children and murine asthma model than controls.

Abstract:Objective: To detect the expression of CK-19 mRNA in peripheral blood of non-small cell lung cancer(NSCLC) patients by real-time fluorescence quantitative polymerase chain reaction(RT-QPCR),and discuss the value of the experiment. Methods: The expression of CK-19 mRNA in peripheral blood of 113 NSCLC patients(NSCLC group),50 non-cancer patients and 50 healthy person(control group) was detected using RT- PCR and found out the positive cases. Then,RT-QPCR was used to detect the expression of relative levels of CK-19 mRNA in positive peripheral blood. The accuracy of this trial was analyzed by receiver operating characteristic curve(ROC curve) and the area under ROC curve(AUC). Results: ① The expression rates of CK-19 mRNA in the NSCLC group and the control group were 38.94%(44/113) and 1.00%(1/100),respectively,there was significant difference between them(P < 0.01),and the expression of CK-19 was related to TNM classification and differentiation of cancer. ② The analysis of ROC curve showed that when the point of transition was 0.236,its sensitivity and specificity were 95.45% and 77.27% respectively,and the AUC was 0.896. Conclusion: CK-19 mRNA might be the specific and sensitive marker to detect circulating tumor cells in peripheral blood of NSCLC patients,but it still needs to be proved by further follow-up observation in a large volume sample.

Abstract:Objective: To evaluate the value of serum soluble triggering receptor expressed on myeloid cells-1(sTREM-1) on antibacterial agents useage in treatment of acute exacerbations of chronic obstructive pulmonary disease(AECOPD). Method: From August 2007 to February 2009,a total of 246 patients requiring hospitalization for AECOPD were randomly assigned into two groups:standard therapy group(n = 123) and sTREM-1-guided group(n = 123). Serum sTREM-1 levels of all patients were measured after hospital admission by enzme-linked immunosorbent assay. On the base of similarly normal treatment,standard therapy group received antibacterial agents according to guideline for antimicrobial drug useage,and sTREM-1-guided group were treated with antibacterial agents according to serum sTREM-1 levels:antibacterial agents treatment was applied when sTREM-1 level≥20ng/ml and was discouraged when sTREM-1 level < 20 ng/ml and CRP< 5 mg/L. Length of hospitalization,clinical efficacy,costs of hospitalization and antibacterial agents,hospital mortality,rate of exacerbation and rehospitalization within 1 year were observed. Results: Clinical efficacy,hospital mortality,length of hospitalization,rate of exacerbation and rehospitalization within 1 year were similar in two groups(P = 0.621-0.734,0.571,0.793 and 0.883);costs of antibacterial agents and hospitalization of sTREM-1-guided group were lower than that of standard therapy group(P = 0.000和0.026). Conclusion: sTREM-1 could be detected as a reference for guiding the use of antibacterial agents for AECOPD treatment which may reduce the costs of antibacterial agents and hospitalization.

Abstract:Objective: To observe the clinical effects in patients with biliary tract cancer treated with OLF (L-OHP,LV,5-Fu) combined with hydroxycamptothecin(HCPT) which was administrated as intraperitoneal perfusion hyperthermia. Methods: Twenty-four patients were given chemotherapy as well as intraperitoneal chemotherapy combined with hyperthermia. The curative effects were estimated according the criterea of RECIST and the clinical benefit response rate(CBR) was evaluated. The CBR refers to pain,physical status and body weight. Results: There was no complete response among all the patients. Partial response was observed in 7 patients(29.2%) and stable disease was 33.3%. As to CBR,16 patients of total(66.7%) achieved the clinical benefit response and their quality of life was improved. The toxicities were marrow depression and alimentary tract reactions. Conclusion: This thermo-chemotherapy comprehensive regimen was an effective and safty methods for patients with advanced biliary tract cancer.

Abstract:Objective: To detect plasma vascular endothelial growth factor (VEGF) in gastric carcinoma patients with liver metastasis and investigate its significance in chemotherapy. Methods: Plasma levels of VEGF in 43 gastric cancer patients and 20 superficial gastritis patients were detected by ELISA test. Results: Plasma levels of VEGF in gastric carcinoma patients were found to be higher than those in gastritis patients. The plasma levels of VEGF in complete response (CR) or partial response (PR) gastric cancer patients decreased significantly after chemotherapy(P < 0.05). However,the plasma VEGF did not differ significantly in stable disease(SD) gastric cancer patients although they were given the same treatment. In progressive disease(PD) patients,the plasma VEGF increased after the treatment. Conclusion: The plasma VEGF level determined by ELISA could be a reference for evaluating the efficacy of treatment and progression of illness for gastric cancer patients with liver metastasis.

Abstract:Objective: To investigate the clinical efficacy of neoadjuvant chemotherapy used for locally advanced cervical cancer patients. Methods: Total 52 cases of Ⅰb2~ Ⅱb cervical cancer were included. According to pathological types of the cancer cells,2 to 3 cycles of combined chemotherapy were used before surgery,then extensive hysterectomy and pelvic lymph node dissection would be done. Results: After preoperative chemotherapy,symptoms and signs of all patients were significantly improved,it created favorable conditions for the radical surgery,total clinical effective rate was 92.5%,and surgery rate was 88.50%(46/52). Conclusion: The use of neoadjuvant chemotherapy for the locally advanced cervical cancer before surgery can alleviate clinical symptoms significantly,reduce size as well as the invaded area of the tumor,lower clinical stage and improve the surgery rate,reduce risk factors and the relapse rate,and it can be expected to extend 5-year survival.

Abstract:Objective: To analyze the risk factors of prolonged mechanical ventilation(PMV) after coronary artery bypass graft surgery(CABG) with the purpose of improving the management of these patients. Methods: The preoperative,operative and postoperative clinical data of 170 patients,who underwent CABG in 2010,were retrospectively analyzed for the risk factors of PMV. Binary Logistic regression models were used to assess the affection of factors. Results: Statistics show that the elder age,currently smoker,left ventricular ejection fraction(LVEF),duration of operation,volume of chest drainage and postoperative complications were risk factors of PMV,meanwhile LVEF,duration of operation,volume of chest drainage and postoperative had extremely affection. Conclusion:Greater importance should be attached to healthcare education,preoperative heart function regulation,lower duration of operation,reduce postoperative chest drainage and better preservation of organs during operation to decrease the rate of PMV and optimize the clinical quality.

Abstract:Objective: To explore the correlation between the occurrence of tubal pregnancy and the expression level of leukemia inhibitory factor (LIF) and LIF receptor (LIFR). Methods: Semi-quantitative analysis of LIF and LIFR by immunohistochemistry was performed in 11 fallopian tubal tissues of non-pregnant group,and 36 of tubal pregnancy group,respectively. Results: The LIFR expression level in fallopian tubal epithelium cells of tubal pregnancy group was significantly higher than that of non-pregnant group. While the LIFR abundance in interstitial cells of tubal pregnancy group was significantly lower than that of non-pregnant group. However,no significant difference of LIF abundance was found between two groups. Conclusion: Aberrant expression of LIFR in oviduct tissues might be involved in the ectopic implantation process of tubal pregnancy.

Abstract:Objective: To detect serum levels of soluble endoglin (sEng) during different periods in normal pregnancy and patients with preeclampsia and evaluate its role in the pathogenesis of preeclampsia. Methods: Women were enrolled in following groups:healthy pregnant control(n = 43) and preeclampsia patients,which contained mild preeclampsia(n = 31),severe preeclampsia (n = 26). The serum sEng levels were measured by enzyme linked immunoassay (ELISA). Results: The concentration of sEng in control group during 12~16,20~24,28~32 and 37~41 weeks were (7.7 ± 1.5),(8.2 ± 1.6),(9.3 ± 1.7) and (11.4 ± 2.6) ng/ml,respectively. They were(7.8 ± 1.5),(9.4 ± 1.8),(14.2 ± 2.9),(21.1 ± 3.7) ng/ml in mild preeclampsia group,and(7.9 ± 1.6),(12.5 ± 2.8),(25.2 ± 3.9),(33.6 ± 4.5) ng/ml in severe preeclampsia group,respectively. The levels of maternal plasma sEng were significantly higher in the preeclamptic women than in normal controls during the latter three periods(P < 0.01) except for the first period. Conclusion: Alterations in the levels of sEng in maternal serum were associated with the occurrence and development of preeclampsia.

Abstract:Objective:To summarize clinical characteristics of Kawasaki disease (KD) in infants younger than 6 months and explore indices for early diagnosis. Methods: Retrospective data from 43 cases of KD in infants younger than 6 months were collected in Nanjing Children’s Hospital from March 2005 to February 2009, and compared with 238 cases of KD in children older than 6 months in the same period. Results: Among 43 cases of KD in infants younger than 6 months, male-female ratio was 2.31∶1. Compared with 238 cases of KD in children older than 6 months, the incidence of some clinical features, such as BCG vaccination erythema, symptoms in digestive system and nervous system at acute period were increased significantly(P < 0.05), while cervical lymphadenopathy was absent in 43 cases of KD in infants younger than 6 months. The levels of neutrophils(N%), platelet(PLT), erythrocyte sedimentation rate(ESR) and lactate dehydrogenase(LDH) in infants were higher(P < 0.05) and the level of Hb was lower(P < 0.05) than children. The incidence of incomplete KD cases and coronary artery aneurysm in infants increased statistically(P < 0.05). Conclusion: Incomplete KD cases and coronary artery aneurysm are more common in infants younger than 6 months. Examination of coronary artery abnormalities, BCG vaccination erythema, symptoms in digestive system and nervous system and the levels of N%, PLT, ESR, LDH and Hb in time would be helpful for early diagnosis.

Abstract:Objective: To investigate cortical-lumbar-anorectal neural pathways in patients with functional constipation (FC) and to explore potential neural mechanisms in FC. Methods: Sixty cases of FC patients and twenty five healthy controls were enrolled in the study. FC group included 28 males and 32 females,mean age was (49.44 ± 15.38) years;normal control group included 11 males and 14 females,mean age was (24.67 ± 7.14) years. Anorectal motor evoked potentials(MEPs) were recorded following transcranial magnetic stimulation (TMS) over motor cortices and lumbar nerve root bilaterally. Results: ①TMS-MEP results:MEP amplitude in the FC group was significantly lower,and the differentiation of wave form and reproducibility were worse than those in the control group(P < 0.05),but latency and peak-interval changes were not changed significantly. The comparison of bilateral parameters showed no significant difference in both the FC group and the control group. ②MS-LMEP results:In the FC group,evoked potential latency was prolonged and amplitude was significantly lower than the control group(P < 0.05). Conclusion: Cortex-lumbar-anorectal nerve pathway conduction disturbances may be the potential onset FC neurophysiological mechanisms.

Abstract:Objective: To develop the gentamicin sulfate/chitosan nanospheres loaded plates and evaluate its characteristics such as surface structure and the drug release behavior of the drug in vitro and its anti-Staph aureus efficacy in vitro. Methods: The solvent casting technology was adopted to prepare the gentamicin sulfate/chitosan nanospheres loaded plates. The surface appearance of the plates was observed by SEM,the drug release behavior in vitro was studied by immerse the plates into PBS. And the gentamicin sulfate/chitosan nanospheres loaded plates were placed in the centre of the Mueller-Hinton(MH) agar which were inoculated with the Staph aureus at the concentration of 1 × 108 CFU/ml. The bacteria were cultured on MH agar for 48 h and the diameter of the inhibition zone was measured,and this procedure was repeated until there was no inhibition zone. Result: The result of the SEM shows the gentamicin sulfate/chitosan nanospheres distributed on the surface of plates uniform,the drug release behavior in vitro could continue about 14 days,and the max cumulative release of the drug was about 52%. The anti-Staph aureus efficacy in vitro could continue about 9 days and the max diameter of the inhibition zone was (3.82 ± 0.03) cm. Conclusion: The gentamicin sulfate/chitosan nanospheres loaded plates show well surface appearance and the plates have a steady drug release behavior and significant anti-Staph aureus efficacy in vitro.

Abstract:Objective: To explore the effects of the laryngeal mask airway (LMA) and the traditional endotracheal intubation (TT) in general anesthesia children with congenital heart disease. Methods: Sirty children (ASA Ⅰ~Ⅱ) with congenital heart disorders,were randomly divided into two groups:laryngeal mask airway group (LMA group,30 cases) and endotracheal intubation group (TT group,30 cases). LMA was inserted following the intravenous infusion of propofol (1.5 mg/kg) and remifentanil (2 μg/kg) in LMA group,while in TT group endotracheal intubation was set up following the intravenous administration of propofol (1.5 mg/kg),remifentanil (2 μg/kg) and rocuronium (0.6 mg/kg). Anesthesia was maintained with propofol and remifentanil micro pump infusion,intermittent intravenous rocuronium. The length of time for operation and recovery from anaesthesia in two groups were recorded,and the values of MAP,HR,SpO2 at different time points(before anaesthesia induction,before insection of LMA or TT,immediately after insection of LMA or TT,before and after the removal of LMA or TT) were observed. Meanwhile,the successful rate of first placement was compared in two groups. Results: There were no significant differences between the two groups in the length of time for operation and recovery from anaesthesia(P > 0.05),as well as the values of MAP,HR,SpO2 before induction,before insection and before remove of LMA or TT(P > 0.05). However,the values of MAP,HR,SpO2 were significantly higher in TT group than those in LMA group immediately after insection and after removal of LMA or TT (P < 0.05). The incidence of postoperative respiratory complications in LMA group was significantly lower than that in TT group. Conclusion: Application of laryngeal mask airway for general anesthesia in the interventional therapy of children with congenital heart disease could not only achieve an effect of hemodynamic-stabilization,but also reduce postoperative respiratory complications because of its less stress response.

Abstract:Objective:To compare shear bond strengths and adhesion failure modes of three cements between titanium and noble alloy. Methods: Fifteen pairs of titanium and noble alloy samples were divided into three groups. Then three cements(Clearfil SA Cement,RelyX Unicem,3M glass ionomer cement) were applied on them respectively. The tensile bond strengths between titanium and alloy were determined after 24 h of water bath. Adhesion failure modes were observed under a stereomicroscope. Results: Clearfil SA Cement and RelyX Unicem had significantly higher shear bond strengths than 3M glass ionomer cement. There was no significant difference between Clearfil SA Cement and RelyX Unicem. Conclusion: Clearfil SA Cement and RelyX Unicem have higher shear bond strengths between titanium and noble alloy.

Abstract:Objective: To investigate the role of ultrasound in the differential diagnosis of Kikuchi disease (KD) and tuberculous lymphadenopathy (TLPN). Methods: An analysis included 75 cases of KD and 215 cases of TLPN with their grayscale and color doppler flow features was performed retrospectively. Results: Both KD and TLPN had yong female predominance,and mostly occurred at the neck area. On sonography KD had homogeneous appearances with intact and sharp capsules and clear hilum. The cortex in KD was homogeneous hypoechoic. The distribution of blood was hilum pattern and flow density had a predominance of grade Ⅱ~Ⅲ. In TLPN,peri-nodal edema was shown. And in most diseased nodes,the hilum disappeared,the nodes usually fused to form a mass,the capsule was thickened or even disappeared. Liquifications and calcifications were common findings in TLPN. The distribution of blood revealed peripheral type or bloodless type and flow density were mainly grade 0 ~Ⅰ. The resistive index(RI) was higher in TLPN(P < 0.001). Conclusion: The sonography apperances are distingishable between KD and TLPN of superficial lymph nodes and will supply as indications for biopsy.