AIM: To establish the role of vascular endothelial growth factor (VEGF) in the oncogenesis of human gastric carcinoma more directly. METHODS: The expression of VEGF and its receptor kinase-domain insert containing rec...AIM: To establish the role of vascular endothelial growth factor (VEGF) in the oncogenesis of human gastric carcinoma more directly. METHODS: The expression of VEGF and its receptor kinase-domain insert containing receptor (KDR) in human gastric cancer tissue were observed by immunohistochemical staining. VEGF levels were manipulated in human gastric cancer cell using eukaryotic expression constructs designed to express the complete VEGF(165) complimentary DNA in either the sense or antisense orientation. The biological changes of the cells were observed in which VEGF was up-regulated or down-regulated. RESULTS: VEGF-positive rate was 50%, and VEGF was mainly localized in the cytoplasm and membrane of the tumor cells, while KDR was mainly located in the membrane of vascular endothelial cells in gastric cancer tissues and peri-cancerous tissue. In 2 cases of 50 specimens, the gastric cancer cells expressed KDR, localized in both the cytoplasm and membrane. Introduction of VEGF(165) antisense into human gastric cancer cells (SGC-7901, immunofluorescence intensity, 31.6%)) resulted in a significant reduction in VEGF-specific messenger RNA and total and cell surface VEGF protein (immunofluorescence intensity, 8.9%) (P【0.05). Conversely, stable integration of VEGF(165) in the sense orientation resulted in an increase in cellular and cell surface VEGF (immunofluorescence intensity, 75.4%) (P【0.05). Lowered VEGF levels were associated with a marked decrease in the growth of nude mouse xenografted tumor (at 33 days postimplantation, tumor volume: 345.40 +/- 136.31 mm3)(P【0.05 vs control SGC-7901 group: 1534.40 +/- 362.88 mm3), whereas up-regulation of VEGF resulted in increased xenografted tumor size (at 33 days postimplantation, tumor volume: 2350.50 +/- 637.70 mm3) (P【0.05 vs control SGC-7901 group). CONCLUSION: This study provides direct evidence that VEGF plays an important role in the oncogenesis of human gastric cancer.展开更多
AIM: To test the hypothesis to block VEGF expression of SMMC-7721 hepatoma cells may inhibit tumor growth using the rat hepatoma model. METHODS: Amplify the 200 VEGF cDNA fragment and insert it into human U6 gene cass...AIM: To test the hypothesis to block VEGF expression of SMMC-7721 hepatoma cells may inhibit tumor growth using the rat hepatoma model. METHODS: Amplify the 200 VEGF cDNA fragment and insert it into human U6 gene cassette in the reverse orientation transcribing small antisense RNA which could specifically interact with VEGF165, and VEGF121 mRNA. Construct the retroviral vector containing this antisense VEGF U6 cassette and package the replication-deficient recombinant retrovirus. SMMC-7721 cells were transduced with these virus and positive clones were selected with G418. PCR and Southern blot analysis were performed to determine if U6 cassette integrated into the genomic DNA of positive clone. Transfected tumor cells were evaluated for RNA expression by ribonuclease protection assays. The VEGF protein in the supernatant of parental tumor cells and genetically modified tumor cells was determined with ELISA. In vitro and in vivo growth properties of antisense VEGF cell clone in nude mice were analyzed. RESULTS: Restriction enzyme digestion and PCR sequencing verified that the antisense VEGF RNA retroviral vector was successfully constructed.After G418 selection, resistant SMMC-7721 cell clone was picked up. PCR and Southern blot analysis suggested that U6 cassette was integrated into the cell genomic DNA. Stable SMMC-7721 cell clone transduced with U6 antisense RNA cassette could express 200 bp small antisense VEGF RNA and secrete reduced levels of VEGF in culture condition. Production of VEGF by antisense transgene-expressing cells was 65+/-10 ng/L per 10(6) cells, 42045 ng/L per 10(6) cells in sense group and 485+/-30 ng/L per 10(6) cells in the negative control group, (P【 0.05). The antisense-VEGF cell clone appeared phenotypically indistinguishable from SMMC-7721 cells and SMMC-7721 cells transfected sense VEGF. The growth rate of the antisense-VEGF cell clone was the same as the control cells. When S.C. was implanted into nude mice, growth of antisense-VEGF cell lines was greatly inhibited compared with control cells. CONCLUSION: Expression of antisense VEGF RNA in SMMC-7721 cells could decrease the tumorigenicity, and antisense-VEGF gene therapy may be an adjuvant treatment for hepatoma.展开更多
Hepatocellular carcinoma (HCC) is one of the most common malignancies in China. To date, surgery is still the best solution to it. However, metastatic recurrences after curative hepatic resections are very common. Tan...Hepatocellular carcinoma (HCC) is one of the most common malignancies in China. To date, surgery is still the best solution to it. However, metastatic recurrences after curative hepatic resections are very common. Tang et al have reported that recurrence rate within 5 years of curative hepatic resection is 61.5% [1]. As curative hepatic resection has a high tendency for metastatic recurrence, therapeutic interventions such as transarterial embolization and antiangiogenesis have been tried to further improve prognosis of HCC patients. Therefore, establishing a dependable, sensitive, easy, and economical method to predict metastatic recurrence following curative hepatic resection is of clinical urgency.展开更多
BACKGROUND： This study was undertaken to investigate the expression of hypoxia-inducible factor-1α （HIF-1α） in rat cerebral cortex and the effects of β-sodium aescinate （SA） administration after return of spon...BACKGROUND： This study was undertaken to investigate the expression of hypoxia-inducible factor-1α （HIF-1α） in rat cerebral cortex and the effects of β-sodium aescinate （SA） administration after return of spontaneous circulation （ROSC）.METHODS： Sixty rats were divided into three groups： SA group, injected intraperitoneally with SA instantly after ROSC; control group, injected intraperitoneally with normal saline; and sham-operated group, without cardiac arrest or SA. The cardiac arrest model was established using asphyxiation and intravenous potassium chloride. Blood was sampled 1, 6, 12, and 24 hours after ROSC. Protein and mRNA levels of HIF-1α, VEGF and EPO were detected in the cerebral cortex by immunohistochemistry and real-time RT-PCR; serum levels of NSE and S100β were determined by enzyme-linked immunosorbent assays.RESULTS： Serum S100β and NSE were signi？ cantly increased in the control group versus the sham-operated group 1, 6, 12 and 24 hours after ROSC （P〈0.05）. Protein and mRNA levels of HIF-1α, VEGF and EPO were signi？ cantly increased in the control rats （P〈0.05）. Serum NSE and S100β were significantly decreased in the SA group versus the control group 1, 6, 12 and 24 hours after ROSC （P〈0.05）. Protein and mRNA levels of HIF-1α, VEGF and EPO were signi？ cantly increased in the SA group （P〈0.05）.CONCLUSIONS： The expression of HIF-1α is increased in rat cerebral cortex after ROSC, and SA up-regulates the expression of HIF-1α. The up-regulation of HIF-1α improves the resistance of the cortex to ischemia and hypoxia and contributes to neuroprotection, possibly because of up-regulation of EPO and VEGF expression.展开更多
OBJECTIVE: To investigate the feasibility of endothelialization of bioprosthesis by transfer of vascular endothelial growth factor (VEGF) gene. METHODS: Bovine pericardium treated with glutaraldehyde and L-glutamic ac...OBJECTIVE: To investigate the feasibility of endothelialization of bioprosthesis by transfer of vascular endothelial growth factor (VEGF) gene. METHODS: Bovine pericardium treated with glutaraldehyde and L-glutamic acid was positioned into the pig right atrium. pcD(2)/hVEGF(121) gene (1 mg) was transferred into the right ventricular myocardium using surgical sutures Reverse transcri ption polymerase chain reaction (RT PCR) was employed to evaluate the expression of myocardial VEGF mRNA. The determination of concentrations of VEGF protein in blood from both the right atrium and peripheral vein, and histological and ultrastructural analysis of implanted bovine pericardium were completed simultaneously. RESULTS: The concentration of VEGF derived from the right atrium in pcD(2)/hVEGF(121) group was significantly higher than that in the pcD(2) group 10 days after VEGF gene transfer (P展开更多
文摘AIM: To establish the role of vascular endothelial growth factor (VEGF) in the oncogenesis of human gastric carcinoma more directly. METHODS: The expression of VEGF and its receptor kinase-domain insert containing receptor (KDR) in human gastric cancer tissue were observed by immunohistochemical staining. VEGF levels were manipulated in human gastric cancer cell using eukaryotic expression constructs designed to express the complete VEGF(165) complimentary DNA in either the sense or antisense orientation. The biological changes of the cells were observed in which VEGF was up-regulated or down-regulated. RESULTS: VEGF-positive rate was 50%, and VEGF was mainly localized in the cytoplasm and membrane of the tumor cells, while KDR was mainly located in the membrane of vascular endothelial cells in gastric cancer tissues and peri-cancerous tissue. In 2 cases of 50 specimens, the gastric cancer cells expressed KDR, localized in both the cytoplasm and membrane. Introduction of VEGF(165) antisense into human gastric cancer cells (SGC-7901, immunofluorescence intensity, 31.6%)) resulted in a significant reduction in VEGF-specific messenger RNA and total and cell surface VEGF protein (immunofluorescence intensity, 8.9%) (P【0.05). Conversely, stable integration of VEGF(165) in the sense orientation resulted in an increase in cellular and cell surface VEGF (immunofluorescence intensity, 75.4%) (P【0.05). Lowered VEGF levels were associated with a marked decrease in the growth of nude mouse xenografted tumor (at 33 days postimplantation, tumor volume: 345.40 +/- 136.31 mm3)(P【0.05 vs control SGC-7901 group: 1534.40 +/- 362.88 mm3), whereas up-regulation of VEGF resulted in increased xenografted tumor size (at 33 days postimplantation, tumor volume: 2350.50 +/- 637.70 mm3) (P【0.05 vs control SGC-7901 group). CONCLUSION: This study provides direct evidence that VEGF plays an important role in the oncogenesis of human gastric cancer.
基金Project supported by National Natural Science Foundation of China,No.863 Z2001-04
文摘AIM: To test the hypothesis to block VEGF expression of SMMC-7721 hepatoma cells may inhibit tumor growth using the rat hepatoma model. METHODS: Amplify the 200 VEGF cDNA fragment and insert it into human U6 gene cassette in the reverse orientation transcribing small antisense RNA which could specifically interact with VEGF165, and VEGF121 mRNA. Construct the retroviral vector containing this antisense VEGF U6 cassette and package the replication-deficient recombinant retrovirus. SMMC-7721 cells were transduced with these virus and positive clones were selected with G418. PCR and Southern blot analysis were performed to determine if U6 cassette integrated into the genomic DNA of positive clone. Transfected tumor cells were evaluated for RNA expression by ribonuclease protection assays. The VEGF protein in the supernatant of parental tumor cells and genetically modified tumor cells was determined with ELISA. In vitro and in vivo growth properties of antisense VEGF cell clone in nude mice were analyzed. RESULTS: Restriction enzyme digestion and PCR sequencing verified that the antisense VEGF RNA retroviral vector was successfully constructed.After G418 selection, resistant SMMC-7721 cell clone was picked up. PCR and Southern blot analysis suggested that U6 cassette was integrated into the cell genomic DNA. Stable SMMC-7721 cell clone transduced with U6 antisense RNA cassette could express 200 bp small antisense VEGF RNA and secrete reduced levels of VEGF in culture condition. Production of VEGF by antisense transgene-expressing cells was 65+/-10 ng/L per 10(6) cells, 42045 ng/L per 10(6) cells in sense group and 485+/-30 ng/L per 10(6) cells in the negative control group, (P【 0.05). The antisense-VEGF cell clone appeared phenotypically indistinguishable from SMMC-7721 cells and SMMC-7721 cells transfected sense VEGF. The growth rate of the antisense-VEGF cell clone was the same as the control cells. When S.C. was implanted into nude mice, growth of antisense-VEGF cell lines was greatly inhibited compared with control cells. CONCLUSION: Expression of antisense VEGF RNA in SMMC-7721 cells could decrease the tumorigenicity, and antisense-VEGF gene therapy may be an adjuvant treatment for hepatoma.
基金the Shanghai Leading Medical Subjects Grant(№983001)State Key Basic Research Grant(№G1998051211)for financial supports.
文摘Hepatocellular carcinoma (HCC) is one of the most common malignancies in China. To date, surgery is still the best solution to it. However, metastatic recurrences after curative hepatic resections are very common. Tang et al have reported that recurrence rate within 5 years of curative hepatic resection is 61.5% [1]. As curative hepatic resection has a high tendency for metastatic recurrence, therapeutic interventions such as transarterial embolization and antiangiogenesis have been tried to further improve prognosis of HCC patients. Therefore, establishing a dependable, sensitive, easy, and economical method to predict metastatic recurrence following curative hepatic resection is of clinical urgency.
基金supported by the Liaoning Province Natural Science Foundation(20092162)
文摘BACKGROUND： This study was undertaken to investigate the expression of hypoxia-inducible factor-1α （HIF-1α） in rat cerebral cortex and the effects of β-sodium aescinate （SA） administration after return of spontaneous circulation （ROSC）.METHODS： Sixty rats were divided into three groups： SA group, injected intraperitoneally with SA instantly after ROSC; control group, injected intraperitoneally with normal saline; and sham-operated group, without cardiac arrest or SA. The cardiac arrest model was established using asphyxiation and intravenous potassium chloride. Blood was sampled 1, 6, 12, and 24 hours after ROSC. Protein and mRNA levels of HIF-1α, VEGF and EPO were detected in the cerebral cortex by immunohistochemistry and real-time RT-PCR; serum levels of NSE and S100β were determined by enzyme-linked immunosorbent assays.RESULTS： Serum S100β and NSE were signi？ cantly increased in the control group versus the sham-operated group 1, 6, 12 and 24 hours after ROSC （P〈0.05）. Protein and mRNA levels of HIF-1α, VEGF and EPO were signi？ cantly increased in the control rats （P〈0.05）. Serum NSE and S100β were significantly decreased in the SA group versus the control group 1, 6, 12 and 24 hours after ROSC （P〈0.05）. Protein and mRNA levels of HIF-1α, VEGF and EPO were signi？ cantly increased in the SA group （P〈0.05）.CONCLUSIONS： The expression of HIF-1α is increased in rat cerebral cortex after ROSC, and SA up-regulates the expression of HIF-1α. The up-regulation of HIF-1α improves the resistance of the cortex to ischemia and hypoxia and contributes to neuroprotection, possibly because of up-regulation of EPO and VEGF expression.
基金agrantfromtheEducationAssociationofJiangsuProvince ,China (No .98JKB32 0 0 0 8)
文摘OBJECTIVE: To investigate the feasibility of endothelialization of bioprosthesis by transfer of vascular endothelial growth factor (VEGF) gene. METHODS: Bovine pericardium treated with glutaraldehyde and L-glutamic acid was positioned into the pig right atrium. pcD(2)/hVEGF(121) gene (1 mg) was transferred into the right ventricular myocardium using surgical sutures Reverse transcri ption polymerase chain reaction (RT PCR) was employed to evaluate the expression of myocardial VEGF mRNA. The determination of concentrations of VEGF protein in blood from both the right atrium and peripheral vein, and histological and ultrastructural analysis of implanted bovine pericardium were completed simultaneously. RESULTS: The concentration of VEGF derived from the right atrium in pcD(2)/hVEGF(121) group was significantly higher than that in the pcD(2) group 10 days after VEGF gene transfer (P
