Expression of RECK gene in the osteosarcoma cell line and its correlation with MMP-2

Objective： To detect the expression of RECK gene in the highly and low metastatic cell sublines of human osteosarcoma cell HOS and explore its possible roles on the occurrence and metastasis of osteosarcoma. Methods： RT-PCR, gelatin zymography, and matrigel invasion assay were respectively used to evaluate the endogenous expression of RECK mRNA, MMP-2 activation ratio and invasive capacity in the two osteosarcoma cell sublines. Results： The highly metastatic cell group expressed significantly lower mRNA level of RECK than the low metastatic cell group （P 〈 0.05）, but showed higher MMP-2 activation ratio and invasive capacity （P 〈 0.05 and P 〈 0.01, respectively）. Conclusion： The abnormal low expression of RECK may participate in osteosarcoma invasion and metastasis, and may be a new therapeutic target for osteosarcoma.     

Construction of eukaryotic expression plasmid pEGFP-N1-WWOX and its transient expression in SMMC-7721 cells

Objective： To construct eukaryotic expression plasmid pEGFP-NI-WWOX and transiently express it in SMMC-7721 cells. Methods： Total mRNA was extracted from normal human liver tissue. RT-PCR was used to amplify the aimed segments WWOX cDNA which was then digested with Hindlll and BamHI and inserted into a eukaryotic expression plasmid pEGFP-N 1 to construct pEGFP-N 1-WMVOX. The constructed plasmid was transfected into SMMC-7721 cells by lipofectamine 2000 - mediated transfer method. The expression of WWOX in transfected SMMC-7721 cells was detected 24, 36 and 48 h post-transfection with fluorescence microscope and the expression level of WWOX mRNA in transfected SMMC-7721 cells was assay by using RT-PCR. The change of MMWOX expression and cell proliferation rates were detected by immunocyto- chemistry and MTT methods respectively. Results： The results showed pEGFP-N1-WWOX was successfully constructed and expressed transiently in SMMC-7721 cells. At 48th hour post-transfection, the number of positive cells was increased significantly and much brighter green fluorescence could be detected, while no green fluorescence was detected in the control group. In SMMC-7721 cells transfected with pEGFP-NI-WWOX a high level of porcine WWOX was detected. WWOX ex- pressed by transfected cells could significantly inhibit the proliferation of SMMC-7721 cells. Conclusion： pEGFP-N1-WWOX was expressed successfully in SMMC-7721 cells, which suggested that might be used as a new therapeutic method for liver cancer.     

Expression of human yrdC gene promotes proliferation of gastric carcinoma cells

Objective： The aim of the research was to study the function of human yrdC gene in the gastric carcinoma cells. Methods： Human yrdC gene was isolated from human spleen tissue by RT-PCR. Anti-human yrdC monoclonal antibody was prepared by hybridoma cell technique. Recombinant adenovirus Ad.yrdC carrying yrdC gene was constructed by using the AdEasy adenoviral vector system. Recombinant adenovirus Ad.yrdCshRNA mediated yrdCshRNA was prepared by RNA interference technology. Gastric adenocarcinoma BGC-823 cells of moderate differentiation were transfected and absorbance of the transfected cells was calculated at 490 nm by methyl thiazolyl tetrazolium （MTT） method. Results： A value of the transfected Ad.yrdC group was significantly greater than that of the non-transfected and transfected Ad.Null groups, and A value of Ad.yrdCshRNA group was significantly lower than that of the non-transfected and transfected Ad.Null groups. Conclusion： Expression of yrdC gene has a function of promoting the proliferation of gastric carcinoma cells.     

S100A6 gene have a positive influence on the growth and proliferation of gastric cancer cell MKN45

Objective： S100A6 （a.k.a., calcyclin） is over-expressed in several human tumors, including gastric carcinoma, human melanoma, pancreatic carcinoma, squamous cell carcinoma, malignant fibrous histiocytoma （MFH）, and carcinomas of the thyroid, breast, and colon. However, little is known about the role S100A6 plays in gastric adenocarcinoma. In the present study, we intended to investigate the influence of S100A6 on the growth, proliferation, apoptosis, invasion and cell cycle of the gastric cancer cell MKN45. Methods： As an important member of S100 family, S100A6 cDNAwas subcloned into a constitutive vector pcDNA3.1 followed by transfection in gastric cancer cell line MKN45 by using liposome. Then stable transfectants were selected and appraised. The apoptosis and cell cycles of these clones were analyzed by using flow cytometric assay. The growth and proliferation were analyzed by cell growth curves and colony-forming assay respectively. The S100A6 stable expression clones （MKN-S100A6） were detected and compared with their control groups respectively. Results： MKN- S100A6 grew faster than MKN45 and MKN-PC （MKN45 transfected with pcDNA3.1 vector）. The cell counts of MKN-SI00A6 in the fifth, sixth and seventh days were significantly more than those of control groups （P 〈 0.05）. Cell cycle analysis showed that proportions of MKN-S100A6 in G0-G1 and G2-M were different significantly with those of its control groups respectively （P 〈 0.05）. The apoptosis rate of MKN-S100A6 was significantly lower than those of control groups （P 〈 0.05）. Results of colony-forming assay showed that the colon formation rate of MKN-S100A6 was higher than those of control groups （P 〈 0.05）. Conclusion： S100A6 can promote the growth and proliferation of gastric cancer cells. It can help tumor cell maintain malignant phenotype. In gastric cancer, S100A6 could be thought as a tumor-enhancing gene in some distance, but its role could be complicated.     

Influence of human mutant p27 gene on the biological behaviors of colon cancer cells

Objective： To observe the influence of human mutant p27 gene （p27mt） on the growth and apoptosis of colon cancer cells so as to investigate the function mechanism of p27mt in gene therapy for colon cancer. Methods： Colon cancer cell line SW480 was infected with recombinant replication defective adenovirus Ad-p27mt, and expression of p27mt protein was detected by Western blot; the inhibition effect of p27mt on SW480 cells was detected with cytometry. Cell cycle was decided with flow cytometer, and DNA fragment analytic process identified the occurrence of apoptosis. Results： After transfected SW480 cells with Ad-p27mt, high expression of p27 protein was identified with immunoblotting assay. PI staining and flow cytometer assay showed 77.96% colon cancer cells was blocked in phase G0/G1, while in Ad-LacZ group and blank control group, 27.57% and 25.29% cells were blocked in the same phase, respectively. Growth curve showed Ad-p27mt has an obvious inhibition effect on the growth of SW480 cells, DNA fragment assay demonstrated that p27mt was able to induce the apoptosis of colon cancer cells. Conclusion： p27mt has an obvious blocking effect on colon cancer cell cycle, and most cells were blocked in phase G0/G1. This blockage is related with the growth inhibition and apoptosis induction effect of p27mt.     

Biological effect of NK4 gene mediated by attenuated Salmonella typhimurium on hepatocellular carcinoma cell HepG2

Objective： The aim of the study was to construct a stable strain of recombined attenuated Salmonella typhimurium expressing NK4 gene, and observe the effect of the strain on the metastatic potentiality of HepG2 cells. Methods： The NK4 cDNA was isolated from PCAGGS/hNK4 plasmid by PCR, and subcloned into eukaryotic expression vector pcDNA4. The recombinant plasmid was electro-transferred into attenuated Salmonella typhimurium Ty21a to obtain the recombinant strain encoding NK4 gene （TPN）. Simultaneously, the recombinant attenuated Salmonella typhimurium carrying GFP gene （TPG） was also constructed. After the TPG and TPN were transferred into HepG2 cells, the transfection rate and the expression level of NK4 protein were detected by flow cytometry and ELISA, and the effects of expression product on the proliferation and migration of HepG2 and angiogenesis were observed. Results： The TPN and TPG were successfully constructed. Fortyeight hours after transfection with TPG, the infection rate was 82.58% ± 1.74%, and the expression level of NK4 protein in supernatant was （181.5 ± 11.7） ng/6 × 10^5 cells. The supematant had obviously depressant effect on the proliferative activity of HepG2 cells （P 〈 0.05）, and could obviously restrain the hepatocyte growth factor-mediated migration of tumor cells （P 〈 0.01）. The inhibitory effect of the expression product on the tumor angiopoiesis was obviously observed （P 〈 0.05）, without a dosage-effect relation. Conclusion： The TPN could effectively transfer tumor cells in vitro and express interest NK4 protein. The expression product could effectively inhibit the proliferation and migration of hepatocellular carcinoma cells and the tumor angiopoiesis.     

Expressions of genes related to genome stability and DNA repair in nasopharyngeal carcinoma clustering families

Objective： The aim of the study was to observe the expressions of genes related to genome stability and DNA repair in the members of nasopharyngeal carcinoma （NPC） clustedng families. Methods： In the Zhongshan City where there is highly incidence rate of NPC, we chose the members of the NPC clustering families as objects, and the patients of nasopharyngitis and NPC as the control group. We isolated the RNA from the nasopharyngeal tissue, and synthesized its cRNA, the genome stability and DNA repair genes chip technique, chemiluminescent detection and real-time fluorescence quantita- tive technique were used to examine the genome stability and DNA repair genes in the nasopharyngeal tissue. Results： More genome stability and DNA repair genes were up-regulated in the members of the NPC clustering families than the NPC patients, and the range of up-regulated was high, with the over up-regulated 100 times genes including TEP1, MSH4, PMS2LI. Fewer genome stability and DNA repair genes were down-regulated in the members of the NPC clustering families than the NPC patients, the ubiquitin genes almost were down-regulated, the results also could be confirmed by real-time fluorescence quantitative PCR. Conclusion： There are specially expression character of genome stability and DNA repair genes in the members of NPC clustering families.     

The latest advances of experimental research on targeted gene therapy for prostate cancer

The absence of effective therapies for castration-resistant prostate cancer （CRPC） establishes the need to de- velop novel therapeutic modality, such as targeted gene therapy, which is ideal for the treatment of CRPC. But its application has been limited due to lack of favorable gene vector and the reduction of ＂bystander effect＂. Consequently, scientists all over the world focus their main experimental research on the following four aspects： targeted gene, vector, transfer means and comprehensive therapy. In this paper, we reviewed the latest advances of experimental research on targeted gene therapy for prostate cancer.     

The difference between multi-drug resistant cell line A549/Gem and its parental cell A549

Objective： To discuss the difference between multi-drug resistant cell line A549/Gem and its parental cell A549 on the basis of establishment of human gemcitabine-resistant cell line A549/Gem so as to elaborate the possible mechanisms of gemcitabine resistance. Methods： Human gemcitabine-resistant non-small cell lung cancer cell line A549/Gem was established by the method of repeated clinical serous peak concentration plus gradually increasing concentration of gemcitabine from its parental cell human lung adenocarcinoma cell line A549 which was sensitive to gemcitabine. During the course of inducement, we had monitored their morphology, checked their resistance indexes and resistant pedigree by MTT method, gathered their growth curves and calculated their doubling time, examined their DNA contents and cell cycles by FCM; at the same time, we had measured their expressions of P53, EGFR, Cerb-B-2, PTEN, PCNA, c-myc, VEGF, MDR-1, Bcl-2, nm23, MMP-9, TIMP-1, and CD44v6 proteins via immunocytochemistry staining, RRM1 and ERCC1 mRNA by real-time fluorescent quantitative-PCR. Results： The resistance index of A549/Gem＇ cells （the deputy of cells in the process of inducement） to gemcitabine was 163.228, and the cell line also exhibited cross-resistance to vinorelbine, taxotere, fluorouraci, etoposide and cisplatin, but kept sensitivity to paclitaxol and oxaliplatin. The doubling time of A549/Gem＇ was shorter and figures in G0-G1 phases were increased than A549 cells. Compared with A549 cells, A549/Gem＇ cells achieved EGFR and c-myc proteins expressions, nm23 protein expression enhanced, P53, Cerb-B-2 and Bcl-2 proteins expressions reduced, PTEN, PCNA and MDR-1 proteins expressions vanished, but those of MMP-9, VEGF, CD44v6 and TIMP-1 proteins changed trivially. Meanwhile, expressions of RRM1 and ERCC1 mRNA were augmented markedly. The resistance index of A549/Gem cells to gemcitabine was 129.783, and the cell line also held cross-resistance to vinorelbine, taxotere, etoposide, cisplatin and sensitivity to     

Gene therapy targeted to telomerase in HCC by AF-hTERT-TK/GCV

Objective： The aim of this study was to observe the affection of targeted therapy to plasmid AF-pGL3-hTERT-TK in HCC cell line HepG2. Methods： We constructed therapeutic plasmid pGL3-hTERT-TK （containing suicide gene TK that promoted by promoter of hTERT） and was conjugated with AF-liposome （a protein that can combine with the receptor ASPGR on HCC cell surface）. Then we transfected HCC cell line HepG2 and hepatic cell L02 with AF-pGL3-hTERT-TK, observed the effects of therapeutic plasmid AF-pGL3-hTERT-TK for HCC cell line growth and apoptosis in vitro by Flow cytometry and Tun- nel method. Results： Our results showed that TK gene was 1100 bp in plasmid pGL3-hTERT-TK. Plasmid pGL3-hTERT-TK can effectively transfect HCC cell HepG2 and the transfection rate was 8.91%. By recognizing and combining effects of recep- tor protein ASPGR on HCC cell surface the therapeutic plasmid AF-pGL3-hTERT-TK could easily enter into HCC cell HepG2 and induce its apeptosis. The apoptosis rate was 85.87% while only 8.65% in L02 cell. Four days after AF-pGL3-hTERT-TK transfected HepG2 was intervention by ganciclovir （GCV）, a lot of apeptotic bodies were found by Tunnel analysis, while little apoptotic body was found in hepatic cell L02. Conclusion： AF-pGL3-hTERT-TK can target to HCC cell line and induce it to apoptesis, almost has no influence on hepatic cell L02. AF-pGL3-hTERT-TK has the potential therapeutic effects for HCC.     

Detection of PRL-2 gene expression in hepatocellular carcinoma by real-time fluorescence quantitative PCR

Objective： To quantitatively detect the expression level of PRL-2 in primary hepatocellular carcinoma using realtime fluorescence quantitative PCR. Methods： Total RNA isolated from human HCC and liver tissue adjacent to the tumor was reversely transcribed into cDNA. Real-time fluorescence quantitative PCR （Q-PCR） method was used to analyze the expression level of PRL-2 gene. Results： The Q-PCR method was performed successfully to precisely detect RNA level. PRL-2 was expressed in all portal vein tumor thrombosis （PVTT） and HCC, but only in some paratumor tissue. The highest expression level of PRL-2 gene was recorded in PVTT; meanwhile expression level of PRL-2 was higher than that in paratumor liver tissues and in HCC （P 〈 0.01）, and it was higher in HCC than that in paratumor liver tissues. Conclusion： The Q-PCR may be the most precise method to quantitatively detect RNA level and can be used in gene expression changes. The PRL- 2 gene has higher expression in PVTT than that in HCC and in paratumor liver tissue cells, indicating that it plays an important role in the development and metastasis of the HCC.     

Expression of coxsackie and adenovirus receptor in small cell lung cancer

Objective： We explored the expression of coxsackie and adenovirus receptor （CAR） in small cell lung cancer （SCLC） tissue. Methods： CAR expression in 31 SCLC was assessed in formaldehyde-fixed, paraffin-embedded tissue according to the EnVision immunohistochemistry procedure, while 3 samples of surgical specimens of non-malignant lung disease were taken as the negative control. Results： We observed that the expression of CAR was detectable positive in all the 31 cases from the small cell lung cancer tissue, in contrasting that non-malignant lung tissue control. Conclusion： The high expression of CAR appeared in SCLC tissue indicates that it play an important role in of adenovirus vector-based gene therapy in SCLC.     

Influence of CDK1 and CDK2 siRNA interference on tumor cell cycle and cell apoptosis

Objective： We investigated the influence of CDK1 and CDK2 expression inhibited by cotransfection of CDK1 and CDK2 siRNA on cell cycle and apoptosis, explored the exact role of cell cycle master regulator in tumor cell apoptosis process. Methods： The siRNA targeting the CDK1 and CDK2 genes were synthesized and simultaneously cotransfected into Hela cells by lipofectamine 2000.48 or 60 h after the cotransfection, CDK1 and CDK2 protein expressions were examined by Western blot. Cell cycle distribution was analyzed by flow cytometry. Cell apoptosis was detected by the Annexin V/PI method. The changes of the transfected cell morphological under a microscope after Wright-Giemsa Staining were studied. Results： CDK1 and CDK2 protein expression was decreased at 48 or 60 h after cotransfection. The accumulation of the G2/M and S phase population in cell cycle of the cotransfected cells at 48 or 60 h after transfection was enhanced obviously compared with control. The ratio of apoptotic cell of cotransfected cells at 48 or 60 h after transfection was increased significantly compared with control. More binucleate or multinucleate ceJls among cotransfected cells were observed under the microscope. Conclusion： The decreased expression of CDK1 and CDK2 by cotransfection of CDK1 and CDK2 siRNA not only leads to tumor cell cycle arrest in S phase and G2/M phase, but also induces tumor cell apoptosis.     

The expression of cyclooxygenase-2 （COX-2） and p16 in non-Hodgkin＇s lymphomas and its clinical significance

Objective： To investigate the expression of cyclooxygenase-2 （COX-2） and p16 proteins in non-Hodgkin＇s lymphomas （NHL） and their relationship with the genesis and progress of it. Methods： The expression of COX-2 and p16 protein were studied in the lymph nodes tissue from 60 NHL patients and 10 control patients with non-malignant diseases by flow cytometry. Results： Positive rate of COX-2 protein expression in NHL tissues （63.3%, 38/60） was higher than that in normal lymphaden tissues （0, 0/10）. The difference was significant between the two groups （P 〈 0.01）. Expression of COX-2 protein was related with the clinical stage of NHL. In stage Ⅰ ＋ Ⅱ patients, it was significantly lower （35.0% ± 54.6%） than that in stage Ⅲ ＋ Ⅳ patients （84.6% ±87.5%） （P 〈 0.01）. In different sex, age, tumor malignant degree, IPI grade, extranodal involvement and B symptoms groups, the differences of COX-2 expression were not statistically significant （P 〉 0.05）. Positive rate of p16 protein expression （41.7%, 25/60） in NHL＇ was statistically lower than that in normal lymphomas （100%, 10/10） （P 〈 0.01）. Expression of p16 protein was related to malignant degree of NHL. The positive rates of p16 protein in low malignant degree tissues （64.7%, 11/17） was higher than that in high malignant degree tissues （14.3%, 2/14） （P 〈 0.05）. Positive rates of p16 protein of NHL tissues in different sex, age, IPI grade, extranodal involvement, clinical stages and B symptoms were not statistically significant （P 〉 0.05）. The p16 protein expression in COX-2 positive patients was 47.4% （18/38）, and in negative patients it was 31.8% （7/22）. There was no statistically difference between them （P 〉 0.05）. Correlation analysis revealed there was no correlation between expression of COX-2 and p16 protein. Conclusion： Both COX-2 and p16 protein may all have relationship with the genesis and progress of NHL. The expression of COX-2 protein in NHL may     

Efficacy of percutaneous ethanol injection in the adjuvant treatment of hepatocellular carcinoma after TACE

Objective： To evaluate the efficacy of percutaneous ethanol injection （PEI） in the adjuvant treatment of hepatocellular carcinoma （HCC） after transcatheter arterial chemoembolization （TACE） by primary end points of time to progress （TTP）. Methods： The study population consisted of 73 consecutive patients with inoperable HCC （China Classification System IIN liB）. Among them, 22 patients were treated with TACE and PEI （experimental group）, and the rest 51 were treated only with TACE （control group）, and then the time to progress （TTP） and overall survival （OS） of these two groups were analyzed. Results： The median TTP was 10 months [95% confidence interval （CI）, 7.9-12.1 months] in experimental group and 6 months （95% CI, 4.7-7.3 months） in control group. The 3-month,6-month, and 1-year Progression Free Survival （PFS） rates were respectively 77.3%, 63.6%, and 48.1% in experimental group, and 76.5%, 42.15%, and 24.8% in control group. The TTP of experimental group was significantly longer than that of control group （P 〈 0.05）. The median survival period was 17 months [95% confidence interval （CI）, 11-23 months] of experimental group and 12 months （95% CI, 10-14 months） of control group （P 〉 0.05）. Conclusion： Compared with single TACE, the combination of TACE and PEI can obviously postpone disease progress and prolong survival of HCC patients.     

Hepatitis B virus envelope as a targeting gene transfer vector for hepatic cancer cells

Objective： The aim of the study was to observe the transfection efficacy of hepatitis B virus envelope （HBVE） and evaluate its ability as a gene transfer vector for liver cancer cells. Methods： To obtain HBVE, the supematant fluid of HepG 2.2.15 cells was mixed with a PEG8000 solution for concentration and was inactivated by β-propiolactone. The acquired HBVE was used to pack plRES2-EGFP to test its package ability. Then, we examined its quantity and quality with ELISA, PCR, SDS-PAGE and electron microscopy. The plRES2-EGFP was packed with HBVE and obtained the product HBVE-GFP. The plRES2-EGFP was packed with liposome and obtained the product liposome-GFP. HBVE-GFP and liposome-GFP were used to transfer HepG 2 cells to study the transfection efficiency. HBVE-GFP was used to transfer HepG 2, A549, HeLa and FB cells to study the targeting ability. The green fluorescent protein （GFP） expression was observed under a fluorescent microscope. The rate of GFP positive cells was determined by flow cytometry. Results： 1. The acquired HBVE could retain the surface protein HBsAg ＋ pre S1 ＋ pre S2 and had no virus DNA. It had good package ability for plRES2-EGFP. 2. Transfection efficiency： The GFP could be observed in both the liposome group and HBVE group under the fluorescent microscope. But the HBVE group had a higher fluorescent intensity than liposome group. The transfection rate of liposome group was 49.97% ＋ 2.37% while the HBVE group was 70.65% ＋ 3.15% and the fluorescent intensity of the HBVE group was 3-4 times （P = 0.000） for liposome group with the determination of flow cytometry. 3. Targeting ability： The GFP could be observed in the four groups under the fluorescent microscope. The HepG 2 group had the highest fluorescent intensity among the four groups. The transfection rate of HepG 2 group was 71.35% ＋ 0.03% which was highly expressed than other groups （P = 0.000） and the fluorescent intensity of the HepG 2 group was 2-3 times （P = 0.000） for the other 3 groups with t     

Effect of focal adhesion kinase on cytoskeletal arrangement of HepG2 cells induced by hypoxia

Objective： To study focal adhesion kinase （FAK） expression in hypoxic HepG2 cells and the effect of FAK siRNA on cytoskeletal arrangement of HepG2 cells induced by hypoxia. Methods： HepG2 cells were cultured in 21% O2 and 1% O2. Morphological changes were observed after hypoxia treatment. Western blot was used to measure FAK expression. The siRNA expression vector pshRNA-FAK targeting the mRNA of FAK and vector pGensil-2 （as a control） were constructed, and then transfected into HepG2 cells. Western blot was used to detect FAK. The cytoskeletal arrangement of HepG2 cells transfected with pshRNA-FAK induced by hypoxia was analyzed by phalloidin. The migratory ability of HepG2 cells transfected with pshRNA-FAK induced by hypoxia was analyzed by cell migration assay. Results： Hypoxia-treated cells displayed a more elongated shape with a large degree of cell detachment. FAK expression increased in hypoxic HepG2 cells. FAK protein level was decreased by 75.64% ± 3.12% （P 〈 0.01） after the pshRNA-FAK transfection. Hypoxia induced cytoskeletal arrangement of HepG2 cells. However, cytoskeletal arrangement of HepG2 cells transfected with pshRNA-FAK induced by hypoxia was inhibited in 1% O2. As cell migration assay showed, the migrating number of HepG cells transfected with pshRNA-FAK was significantly lower than that of control （P 〈 0.05）. Conclusion： The expression of FAK in hypoxic HCC might have a close relationship to the cytoskeletal arrangement of HepG2 cells induced by hypoxia. Up-regulation of FAK expression may be one of mechanisms of cytoskeletal arrangement and invasion of hepatocellular carcinoma induced by hypoxia.     

Comparison of diagnostic value of PET using 18- fluoro-2-deoxyglucose,CT and MR1 in detecting skull base invasion of nasopharyngeal carcinomas

Objective： We compared positron emission tomography （PET） using 18-fluoro-2-deoxyglucose （FDG）, enhanced computed tomography （CT） and magnetic resonance imaging （MRI） in detecting skull base invasion of nasopharyngeal carcinomas （NPC） and to evaluate the value of these three methods in determining the existence of skull base invasion of nasopharyngeal carcinomas. Methods： The images of enhanced CT, MRI and PET-CT scans, performed at intervals -〈 20 days on 57 NPC patients from July 2004 to February 2007, were selected and reviewed. The endpoints of the comparison were sensitivity, specificity, accuracy, positive predictive value （PPV） and negative predictive value （NPV） of Enhanced CT, MRI and PET-CT, based on histopathologic findings or clinical imaging follow-up for at least 6 months. Results： For detecting skull base invasion of NPC, the sensitivity of enhanced CT, MRI and PET-CT were 68.18%, 84.09%, 97.67% respectively; speci- ficity were 76.92%, 69.23%, 57.14% respectively; accuracy were 70.18%, 80.7%, 87.72% respectively; PPV were 90.9%, 90.24%, 87.5% respectively; NPV were 41.67%, 56.25%, 88.89% respectively. Conclusion： PET-CT has obvious advantages in sensitivity over CT （P 〈 0.05） and MRI, better than the two methods in accuracy and NPV and may be more valuable for new patients in detecting skull base invasion of NPC patients.     

Inhibitory effect of metformin on the proliferation of human hepatoma HepG2 cells and its potential mechanism

Objective： This work aimed to study the inhibitory effect and the related mechanism of metformin （MET） on the proliferation of human hepatoma HepG2 cells. Methods： Human hepatoma HepG2 cells were treated with MET （0, 2, 10, and 50 mM）. The inhibitory effect of MET on the proliferation of HepG2 cells was determined by MTT method. The apoptosis of HepG2 cells was detected by flow cytornetry. The expression of cyclin D1 in HepG2 cells was examined by Western blot. ROS-DHE fluorescence probe was used to stain the reactive oxygen species （ROS） generated by HepG2 cells after treat- ment. Results： MET could inhibit the proliferation of HepG2 cells in a dose and time dependent manner. MET promoted the apoptosis of HepG2 cells. In addition, MET suppressed the expression of cell cycle protein cyclin D1 and induced the produc- tion of ROS in HepG2 cells. Conclusion： MET can inhibit the proliferation of human hepatoma HepG2 cells and induce cell apoptosis. Meanwhile, MET has the ability to decrease the expression of cyclin D1 and induce ROS generation, which may be involved in the mechanism of inhibiting hepatoma cells proliferation.