Objective To investigate the efficacy of sorafenib alone or combination with transarterial chemoembolization(TACE)and percutaneous local cryotherapy(PLCT)for advanced hepatocellular carcinoma patients without operation opportunity. Methods Sixty-four advanced hepatocellular carcinoma patients were selected as our subjects,who were underwent treatment of sorafenib alone or combination with TACE and PLCT. Thirty-two cases with sorafenib therapy were served as sorafenib group and another 32 cases with sorafenib in combination with transarterial chemoembolization and PLCT were served as combination group. All patients were followed up for 6 - 32 months. The treatment efficacy and tumor development were recorded. Results All surgeries of the patients were succeed and no death or serious operation complications occurred. Of 64 patients, 11 were achieved a complete remission( CR),31 cases with partial remission( PR),14 cases with stable development(SD),and 8 cases with progressive disease(PD). In the sorafenib group,3 cases were with CR,11 patients with PR,12 with SD,and 6 patients with PD. In the combination group,8 patients were with CR,20 patients with PR,2 patients with SD and 2 patients with PD,and the difference was significant between the two groups(χ2 = 14. 028,P = 0. 003). The median periods to tumor progression were 20 and 53 weeks in the sorafenib group and the combination group,and the difference was significant( χ2 = 14. 773,P = 0. 000). Conclusion For hepatocellular carcinoma patients without operation opportunity,sorafenib combined with TACE and PLCT can increase the tumor remission rate and prolong the periods to tumor progression in patients with hepatocellular carcinoma.

0.05).Compared with control group,contents of sialic acid in the high and moderate dose groups,epididymal coefficient and contents of carnitine in the high dose group reduced significantly(P

BACKGROUND: Amnesic shellfish poisoning(ASP) is caused by consumption of seafood contaminated by marine neurotoxins, mainly domoic acid (DA).OBJECTIVE: To ensure the safety of shellfish consumption by determining DA in shellfish by means of liquid chromatography(LC) combined with quadrupole time-of-flight mass spectrometry(Q-TOF MS).DESIGN: A self-controlled prospective study SETTING: The experiment was carried out in the Center for Animal, Plant and Food Inspection and Quarantine, Shanghai Exit-Entry Inspection and Quarantine Bureau.MATERIALS: MUS-1B, a nominal sample of homogenized mussel tissue (containing 36 ± 1μg DA per gram, Batch No. MUS-1B 0159) and the standard DA solution (100 mg/L, Batch No. DACS-1C 352) were purchased from National Research Council of Canada (Halifax, Canada). Positive musscl samples were collected from the waters during the occurrence of red tide in China East Sea, and blank mussel samples were obtained from local seafood market. LC system (Waters 2695) combined with Q-TOF MS (Qultima, Micromass) and ultravilet (UV) detection (Waters 2487 ) was employed for analysis of ASP toxins.METHODS: The shellfish tissue samples were extracted with methanol-water (1: 1, V/V) and cleansed with a SAX solid-phase extraction cartridge. Separation was carried out on a Zorbax Eclipse XDB-18 column (250×2.1 mm, 3.5μm) with gradient elution of CAN-H20 containing 0.05% formic acid at a flow rate of 0.20 mL per minute. In a positive electrospray ionization DA was ionized effectively, and collision-induced dissociation(CID) produced predominant characteristic ions[M + H] +allowing sensitive TOF MS detection.MAIN OUTCOME MEASURES: ① Limit of quantification(LOQ); ②Accuracy and precision of LC/Q-TOF MS method.RESULTS: The average recoveries of DA spiked to tissue homogenates through the complete cleanse procedure ranged from 87.8% to 94.6%, and the relative standard deviations(RSD) ranged from 8.4% to 11.9%. TheLOQ in the shellfish tissue was 0.1 μg/g.CONCLUSION: LC/Q-TOF MS suits well the purpose of detecting and identifying ASP toxins in shellfish with high sensitivity and specificity, and may provide reference values for monitoring the safety of consumption of shellfish.

Objective To establish human pancreatic cancer xenografts in nude mice, and to investigate the antitumor efficacy of human endostatin expressed by replication-competent adenovirus AdTPHre-hE in vivo. Methods Pancreatic cancer cells AsPC-1 were injected subcutaneously in BALB/c nude mice to establish the xenografts. Tumor growth was observed and measured after AdTPHre-hE treatment. Expression of endostatin was detected by ELISA assay. The tumors were harvested for pathologic examination and immunohistochemical staining. Results Tumors grew more slowly in the AdTPHre-hE group and their sizes were markedly smaller than those of the Ad-hE group (P＜0.01)and control group(P＜0. 01). Endostatin levels were detected in the sera of nude mice in all treated groups, and endostatin expression in AdTPHre-hE group increased with time. The endostatin level in the AdTPHre-hE treated group was much higher(P＜0. 01)and increased faster than that in the Ad-hE treated group. Immunohistochemical staining for Hexon of adenovirus capsid showed more positive tumor cells in the tumor tissues treated with AdTPHre-hE. Immunohistochemical staining for FⅧ revealed a decreased microvessel density in the tumor tissues treated with AdTPHre-hE. Conclusion The replication-competent adenovirus efficiently expressed high-level endostatin and significantly inhibited tumor growth in vivo.

Objective To develop a double-regulated replicative adenovirus carrying the Human endostatin gene(hEndo). Methods The plasmid pTPHre-hEndo was constructed by gene engineering technique, carrying human endostatin gene, in which El A gene and E1B gene were driven by human telomerase reverse transcriptase (hTERT) promoter and hypoxia response element (HRE) promoter,respectively. The pTPHre-hEndo was co-transfected with pBHGE3 in 293 cells to generate recombinant adenovirus AdTPHre-hEndo. Virus titer was measured by the TCID50 method. Virus replication assay was performed to evaluate the selective replication ability of AdTPHre-hEndo. The transgene expression of endostatin was detected by ELISA assay. Results A novel gene-viral therapeutic system AdTPHre-hEndo was constructed by gene engineering technique and its titer was 3. 25 X 1010 pfu/ml.Proliferative test revealed that AdTPHre-hEndo could proliferate selectively in telomerase positive tumors. Furthermore, in comparison with non-replicative adenovirus Ad-hEndo, the transgene expression of endostatin mediated with AdTPHre-hEndo was significantly increased (P < 0. 01).Conclusion The novel gene-viral therapeutic system AdTPHre-hEndo has the capacity to replicate in pancreatic cancer cells and expresses the endostatin efficiently, and may provide a new strategy for pancreatic cancer gene therapy.

Objective To construct the adenovirus vector containing mice CD40- IgG2aFc-IRS-GFP fusion gene and detect the expression of the fusion protein in transfeete 293cells.Methods The fragments of CD40, IgG2aFc were obtained and inserted into plasmid PDC316-IgG2aFc-GFP.After being verified by sequencing, Ad5-PDC316-CD40-IgG2aFc-GFP was contransfored with the bckbone vector into 293 cells.The virus titer was detected after replicating and purifing and the expression of the fusion protein was analyzed.Results A virus and plasmid were constructed successfully.The vitro infection showed that the virus can infect cells of which the fusion protein was confirmed by fluo-rescence.The expression of the fusion protein increased with the increased time and virus concentra-tion.The protein expression stopped increasing after the virus concentration came to a certain level.Conclusion CD40, IgG2aFc fragments are correctly ligated with plasmid PDC316-IgG2aFc-GFP, and the fusion protein can be expressed in 293cells.This might lay a foundation for further studies of the expresstion of virus, immune tolerance and mechanism of liver transplantation model in rats.

Objective To investigate the effect of a dual regulation of replicating adenovirus vector carrying human endostatin gene (AdTPHre-hEndo) on pancreatic cancer. Methods Human endostatin (hEndo) gene was cloned into the genome of replicating adenovirus specific for the tumor cells by virus recombination technology. The virus titer was 3.25 × 1010pfu/ml. A Balb/c nude mouse model carring sw1990 cells pancreatic cancer was established, the expression of human endostain and inhibition of tumor cells in vivo were detected. Results We successfully constructed AdTPHre-hEndo. The inhibition on pancreatic cancer cell line SW-1990 of AdTPHre-hEndo is better than Ad-hEndo (P ＜0. 01 ), and ONYX-015 ( P ＜ 0. 05 ). The endostatin expression of AdTPHre-hEndo group was significantly higher than Ad-hEndo group and the control group (P ＜ 0. 01 ). The intratumoral MVD also decreased significantly in the treated tumors(6. 8 ±2. 5 vs. 16. 0 ±4. 6、47. 2 ± 10. 0, P ＜0. 01 ). Conclusions The recombination adenovirus can express biologically active hEndo effectively, which results in inhibiting the growth of micro-blood vessels and proliferation slowly.

Objective To explore the feasibility of lithium heparin anticoagulant plasma instead of serum in biochemical test . Methods 54 biochemical indexes were comparatively detected in 100 samples of lithium heparin anticoagulant plasma and serum . Results The detection results of 45 biochemical indexes in 100 samples of lithium heparin anticoagulant blood plasma and serum showed no statistically significant differences (P>0 .05) .There were statistically significant differences in the indexes of total pro-tein(TP) ,potassium ion(K+ ) ,lactic dehydrogenase(LDH) ,glucose(GLU) ,creatine kinase-MB(CK-MB) and lipase(LPS) between lithium heparin anticoagulant plasma and serum(P<0 .05) ,while results showed good correlation(the maximum r value was 0 .998 , the minimum r value was 0 .887);3 indexes of transferrin(TRF) ,α-L-fucosidase(AFU) and leucine aminopeptidase (LAP) had statistically significant differences (P<0 .05) and showed no correlation(r<0 .6) .Conclusion Lithium heparin has the strongly an-ticoagulant ability with the advantages of non-influence on cell volume and no hemolysis ,which can be used for routine biochemical test ,especially suitable for the outpatient service ,emergency and the patients with blood coagulation dysfunction ,but the detection of TRF ,AFU and LAP can not be suitable .

Objective Studies show that the ERCC1 gene may be involved in secondary cisplatin resistance.This article aims to investigate the effects of shRNA targeting silencing excision repair cross-complementation group 1 (ERCC1-shRNA) on the proliferation and apoptosis of lung cancer A549/DDP cells treated with different concentrations of cisplatin.Methods Lung cancer A549/DDP cells were divided into a negative control, a blank control, an ERCC1-shRNA1, and an ERCC1-shRNA2 group.Human interfering RNA (RNAi) targeting the human ERCC1 gene was constructed and transfected into the A549/DDP cells using Lipofectamine 2000.The mRNA and protein expressions of ERCC1 in the A549/DDP cells were detected by real-time PCR and Western blot respectively, the proliferation-inhibition rate was assessed by MTT, and their cell cycle and apoptosis were determined by flow cytometry.Results ERCC1-shRNA was successfully constructed and transfected into the A549/DDP cells.Both the mRNA and protein expressions of ERCC1 were significantly lower in the ERCC1-shRNA1 (0.20±0.04 and 0.24±0.10) and ERCC1-shRNA2 (0.47±0.28 and 0.37±0.11) than in the negative control (0.96±0.12 and 1.32±0.13) and blank control groups (0.84±0.07 and 1.45±0.23) (P<0.01).Compared with the negative and blank control groups, the ERCC1-shRNA1 group showed a significantly decreased IC50 value (16.71±2.33 and 16.69±1.69 vs 7.78±0.54, P<0.01) and an increased proportion of G0/G1 phase cells ([72.87±3.23] and [71.75±4.56] vs [82.99±4.23]%, P<0.05), with the cell cycle arrested in the G0/G1 phase.The apoptosis rate of the cells in the ERCC1-shRNA1 group was remarkably lower after treated with cisplatin at the concentrations of 6.25 and 12.5 μg/mL than at 0 μg/mL ([8.17±0.65] and [11.91±1.41] vs [29.97±3.14]%, P<0.05).Conclusion ERCC1-shRNA can inhibit the proliferation and enhance the apoptosis of A549/DDP cells by silencing the expression of the ERCC1 gene.

Objective To evaluate the application of “expert-guided network tracking system” in training community general practitioner for hypertension management.Methods Total 855 general practitioners (GPs) from 20 community health service centers in Hangzhou,Jiaxing and Shaoxing of Zhejiang province were divided into two groups: 430 GPs from 10 community health centers received training for hypertension management using “ expert-guided networking tracking system” (intervention group) in December 2008 and 425 GPs from another 10 health centers received conventional training (control group).Questionnaire surveys about the knowledge and skill of hypertension management were conducted before and 3 y after training.Results The awareness rates of knowledge about hypertension prevention and control,non-drug therapy and drug therapy increased from 37.0％ (159/430),45.6％ (196/430),42.8％ (184/430) to 66.8％ (284/425),81.4％ (346/425),77.6％ (330/425),respectively and the accuracy of case analysis increased from 38.4％ (490/1290) to 73.3％ (946/1290) (P ＜0.01) after training in intervention group.Meanwhile the knowledge and case management abilities in control group were also improved after conventional training,but the degree was significantly lower than that in the intervention group.Conclusions “Expert-guided networking tracking system” can significantly improve the knowledge and skills for hypertension management in community general practitioners.