Objective: Clone, express and purify human recombinant calreticulin (CRT). Methods: Human CRT cDNA was amplified from total RNA of human lung cancer cell line A549 cells by RT-PCR. Then, PCR product was subcloned into prokaryotic expression vector pET-15b. After sequencing, this recombinant plasmid was transformed into E.coli. Rossetta. Recombinant CRT was expressed in host cells by IPTG induction. Resulted protein was purified by Ni-NTA resin under denature condition and dialyzed to recover its native structure. SDS-PAGE and Western blot method were used to identify the expression and purification of reconbinant CRT. Results: Human CRT cDNA was cloned from total RNA of A549 cells. CRT prokaryotic expression vector pET-15b-crt was constructed. Reconbinant CRT was induced to express in E.coli and purified by Ni-NTA affinity chromatograph. Conclusion: A method for prokaryotic expression and purification of human recombinant CRT was successfully established. This method laid a foundation for the subsequent CRT research.

Objective To evaluate the impact of sub-stage on the clinical prognosis of T 1 G3 and evaluate the feasibility of the T1me system in pT1G3 sub-staging.Methods The clinical data,pathological specimen and follow-up data were collected from 56 patients out of 87 patients diagnosed with initial high-grade T1 bladder urothelial carcinoma .The patients were divided into Group A [ T1-microinvasive (T1m),17 cases] and group B [T1-extensive-invasive (T1e),39 cases] according to pathological evaluation after transurethral resection.Clinical parameters were analyzed with Chi-square test,and recurrence-free and progression free survival were obtained by Kaplan-Meier analysis and Log-rank test.Results Age,tumor size,number and intravesical instilled medication showed no significant differences between the 2 groups ( P>0.05 ) .There were significant differences of 5-year recurrence-free rate ( P =0.037 ) and progression-free survival rate ( P =0.045 ) between the 2 groups, and the prognosis of group A was significantly better than that of group B .Conclusion The pathological sub-stage is an important predictor in initial high-grade T1 bladder urothelial carcinoma patients , and the T1me system is objective and feasible.

0.05).Conclusion The increasing of IL-15 in peripheral blood after MP infection may play a role in bronchitic asthma pathogenesis.

Objective To explore the effect of sealing tube with regular urokinase for the malfunction of central venous catheter(CVC) in patients with hemodialysis (HD).Methods This was a prospective randomized controlled study,132 HD patients with cuff CVC were studied.In the early stage,52 H D patients without 80 patients with regular catheter-locking solution of urokinase.To analyze effect of cuff CVC malfunction,blood flow(BF),dialysis adequacy,anemia and serum albumin (ALB) with regular catheter-locking solution of urokinase.The incidence of catheter malfunction was calculated based on the catheter dysfunction in the first 3 months.Clinical index was measured,recorded BF and calculated urea clearance rate (Kt/V).Results HD patients catheter-locking solution with regular urokinase could reduce the incidence of dialysis catheter malfunction ( 26.3％ vs 32.7％,x2 =32.727,P ＜0.01 ),increase the BF,Kt/V,levels of hemoglobin(HGB) and ALB(P＜0.05 or P ＜0.01).Conclusion Regular catheter-locking solution with urokinase is effective in reducing HD patients incidence of dialysis CVC malfunction,increasing patients BF,dialysis adequacy and the level of ALB,improving anemia.

Objective To explore the effect and safety of preoperative impact chemotherapy in the treatment of advanced gastric cancer patients.Methods 104 patients with advanced gastric cancer were randomly divided into two groups,including 54 cases of observation group,54 cases of control group.The observation group was given modified FOLFOX7 regimen for 2 courses before operation,and 6 courses FOLFOX7 regimen after operation.The control group was given routine operation and FOLFOX7 regimen for 8 coures.The effective rate,l-year,2-year,3-year survival rate and toxic effects after operation were observed.Results In observation group,the effective rate was 59.3％,the curative resection rate was 81.5％,and the overall resectability rate was 90.7％,and those was 35.2％,59.3％,75.9％ in control group,all the difference was statistically significant( x2 =8.55,6.39,4.27,all P ＜ 0.05 ).The 1 -year,2-year survival rates after operation were not significantly different ( x2 =0.38,2.06,all P ＞0.05 ),while the difference was significant at 3-year after operation( x2 =4.06,P ＜ 0.05 ).There was no significant difference on the toxic effects between the two groups ( P ＞ 0.05 ).Conclusion Modified FOLFOX7 regimen is effective and well-tolerable for patients with advanced gastric cancer,and it could contribute to improve the overall resectability rate and survival rate after operation.

Objective To investigate the association between carotid atherosclerosis(AS) and serum magnesium(Mg) in hemodialyzed (HD) patients.Methods Clinical index was measured,and intimamedia thickness (IMT) of extracranial common carotid artery and presence of atherosclerotic plaques were determined by high-resolution B-mode ultrasonography.The data were analyzed between plaque positive group and plaque negative group.Results The age,serum phosphate (P),total cholesterol (TCH),low density lipoprotein (LDL),serum C-reactive protein (CRP),serum albumin(Alb),and serum Mg all had significant difference between two groups(t =4.153,2.908,2.301,6.322,5.791,2.341,7.778,P ＜0.01 or P ＜ 0.05).The risk factors of HD patients with AS were serum Mg and CRP(P ＜ 0.01).Conclusions The occurrence of AS was related to low Mg and high CRP in HD patients.

Objective To regulate nutrition support nursing, facilitate the development of nursing specialty, cultivate nutrition support nurse. Methods Establish nursing specialist team on nutrition support. 30 nurses received theory training, skill training, clinical practice, and completion reporting. Conduct nutrition knowledge management survey. Establish standardized wards of nutrition support nursing and conduct special inspection. Results Nursing specialist team on nutrition support was established, and 29 nutrition support nurses were cultivated. Before training, nurses' knowledge of nutrition and nutrition nursing management score were (3.46 ± 0.47), (0.70 ± 0.24) points, and the score were (4.63±0.14), (0.90±0.27) points after training, results was statistically significant (t=12.848, 2.981, P<0.01). Before and after special inspection of enteral nutrition, enteral nutrition pump and special infusion tube cases were 106,60 and 106, 60, results were statistically significant (χ2=4.747, 4.471, P<0.05). Before and after special inspection of parenteral nutrition, correct infusion and speed cases were 90, 92 and 61, 54, results were statistically significant (χ2=4.642, 7.644, P<0.05 or 0.01). Conclusions The development of nursing specialty of nutrition support was promoted, nurses′ comprehensive ability and self-identity was improved, management quality of nutrition support was.

AIM: To prepare recombinant human spermine oxidase (SMO) and polyclonal antibody against human SMO by gene recombination techniques. METHODS: Human SMO cDNA was amplified from total RNA of A549 cells through reverse transcription PCR. The cDNA was then cloned into pET-15b to construct SMO prokaryotic expression vector. After transforming, the vector was induced to express recombinant SMO by IPTG in E. coli BL21 (DE_3). Recombinant SMO was purified by Ni-NTA resin under denaturing condition and then was dialyzed to renature. The enzyme activity of recombinant SMO was analyzed by chemical fluorescent method. SMO polyclonal antibody was prepared by using recombinant human SMO protein purified by polyacrylamide gel electrophoresis as antigen to inoculate rabbit intradermally. The titer and specificity of anti-sera were determined by ELISA, Western blot and Immune Cell Chemistry. RESULTS: Purified and dialyzed recombinant human SMO has the specificicity of oxidizing the spermine. The polyclonal antibody has high titer and specificity against human SMO. CONCLUSION: This research established a method for prokaryotic expression, purification and polyclonal antibody preparation of human SMO. The method lays a foundation for the future functional research of SMO.

Aim To study the effects of polyamine analogue CPENSpm on the human lung cancer line A549 in cell proliferation and apoptosis.Methods MTS was used to assay the cell proliferation,chemical analysis methods were used to determine the activities of enzymes in the polyamine metabolism,HPLC was performed to assay the intracellular concentration of polyamines,Sub-G1 and DNA fragmentation assays were used to determine the cell apoptosis.Results Treating A549 lung cancer cells by CPENSpm resulted in:①cell-growth inhibition and cell apoptosis;②inhibition of ODC(key enzyme in polyamine synthetic pathway)and activation of SSAT and SMO(key enzymes in polyamine catabolism);③great decrease of intracellular polyamine concentrations.MDL72527,the SMO inhibitor,can antagonize the effect of CPENSpm on inhibiting the proliferation of A549 cells.Conclusion CPENSpm inhibits proliferation and induces apoptosis of human A549 lung cancer cell line by interfering the polyamine metabolism,depleting intracellular polyamine contents that are need by quick-growth of cancer cells and inducing production of H2O2.

Objective:To probe into the status characteristics and the dynamic change trend of the psychological stress among medical staff at four time points after exposed to blood-borne pathogen.Methods:A longitudinal study was carried out among 78 medical personnel in this study,with 67 of hepatitis B virus,5 of hepatitis C virus,4 of HIV and 2 of treponema pallidum respectively.Perceived stress scale (PSS-4),Post-traumatic stress disorder Checklist-5 (PCL-5),Hospital anxiety and depression scale (HADS) were used to assess the psychological status,including perceived stress,post-traumatic stress symptom,anxiety and depression at the four times.Data analysis was performed using repeated measures analysis of variance (RMANOVA) and paired sample t test to explore time effect of the psychological stress reaction in the exposed population.Results:The scores of PSS-4,PCL-5,HADS-A,and HADS-D increased from exposure,and gained highest at one month after exposed,then gradually declined with time (Ps ＜ 0.05).Conclusion:It suggests that the levels of perceived stress,post-traumatic stress symptom,anxiety,depression may be the highest at one month after exposed.