The development of human endometrial carcinoma(HEC) is a complex pathologic process involves several oncogenes and tumor suppressor genes.The full-length paired-box gene 2(pax2),a recently discovered oncogene,promotes...The development of human endometrial carcinoma(HEC) is a complex pathologic process involves several oncogenes and tumor suppressor genes.The full-length paired-box gene 2(pax2),a recently discovered oncogene,promotes cell proliferation and growth and inhibits apoptosis of HEC cells.Here,we examined the effect of pax2 small interfering RNA(siRNA) on the growth of transplanted HEC cells in nude mice.The expression of Pax2 in 21 cases of normal endometrium and 38 cases of HEC was examined by immohistochemistry(IHC).HEC models were developed by subcutaneously transferring HEC cells into nude mice,followed by treatment with empty lentivirus vector,lentivirus vector-based pax2 siRNA,and phosphate buffered saline,respectively.Four weeks later,tumor size was measured,tumor inhibition rate was calculated,and histological analyses were conducted after staining with hematoxylin and eosin.The expression of Pax2 and Bcl-2 was detected by Western blot;proliferating cell nuclear antigen(PCNA) was detected by IHC.Significant differences were observed in the positive rate of Pax2 between normal endometrium and HEC(14.2% vs.60.5%,P<0.01).The expression index of Pax2 in well differentiated tumors was 1.88±1.68,much lower than that in tumors of moderate(3.07±1.96,P<0.05) or poor differentiation(5.45±2.76,P<0.01).Tumor necrosis increased,nuclear basophilia stain decreased,tumor growth was inhibited,and PCNA,Pax2,and Bcl-2 expression was reduced in HEC models treated with pax2 siRNA.These results indicate that Pax2 expression is related to HEC tumor biology with the increased expression of Pax2 correlated to malignancy.pax2 siRNA down-regulates Pax2 expression and inhibits tumorigenesis of HEC in nude mice,possibly due to cell apoptosis and the inhibition of tumor proliferation induced by down-regulation of Bcl-2.展开更多
AIM: Epidermal growth factor (EGF) plays an important role in the regulation of gastrointestinal tissue growth and development, and it can stimulate epithelial proliferation, cell differentiation and growth. It has be...AIM: Epidermal growth factor (EGF) plays an important role in the regulation of gastrointestinal tissue growth and development, and it can stimulate epithelial proliferation, cell differentiation and growth. It has been established that the EGF can promote gastric cytoprotection and ulcer healing. But the potential ability of EGF to regulate the gastric cancer growth is unknown. This study is to investigate the influence of EGF on human gastric cancer cell and the implanted tumor growth of nude mice. METHODS: The cell growth rates of human gastric adenocarcinoma cell lines MKN-28, MKN-45, SGC-7901 and normal human gastric epithelial cells 3T3 were assessed when incubated with recombinant human EGF (rhEGF, 0.05, 0.1, 0.5, 1.0, 10, 50, 100 mg.L(-1)) using MTT method. The cells of MKN-28, MKN-45, SGC-7901 (gastric cancer tissue 1.5mm(3)) were implanted in the BALB/cA nude mice for 10 days.The EGF was given intraperitoneally (15, 30, 60 microg.kg(-1)) for 3 weeks. The body weights of the tumor-bearing animals and their tumor mass were measured afterwards to assess the mitogenic effect of rhEGF in the nude mice. RESULTS: Within the concentration range of 0.05-100mg.L(-1), rhEGF could increase the cell growth of normal 3T3 cells (cell growth rate 100% vs 102.8%, P【0.05), but partially restrain the gastric cancer cell growth. The latter effect was related to cell differentiation. In 15-60 microg/kg rhEGF groups, the mean implanted tumor mass of MKN-28 cell were 1.75 g, 1.91 g, 2.08 g/NS group 1.97 g (P】0.05), the mean tumor mass of SGC-7901 cell were 1.53 g, 1.07 g, 1.20 g/NS group 1.07 g (P】0.05), and for MKN-45 cell, the tumor mass were respectively 1.92 g, 1.29 g, 1.77 /NS group 1.82 g (P】0.05). So rhEGF had no obvious effect on implanted MKN-28, SGC-7901 and MKN-45 tumor growth. CONCLUSION: EGF has no stimulating effect on the human gastric cancer cell growth neither in vitro nor in vivo.展开更多
AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carc...AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.展开更多
AIM: To characterize the anticancer function of cytokine-induced killer cells (CIK) and develop an adoptive immunotherapy for the patients with primary hepatocellular carcinoma (HCC), we evaluated the proliferation ra...AIM: To characterize the anticancer function of cytokine-induced killer cells (CIK) and develop an adoptive immunotherapy for the patients with primary hepatocellular carcinoma (HCC), we evaluated the proliferation rate, phenotype and the antitumor activity of human CIK cells from healthy donors and HCC patients in vitro and in vivo. METHODS: Peripheral blood mononuclear cells (PBMC) from healthy donors and patients with primary HCC were incubated in vitro and induced into CIK cells in the presence of various cytokines such as interferon-gamma (IFN-gamma), interleukin-1 (IL-1), IL-2 and monoclonal antibody (mAb) against CD3. The phenotype and characterization of CIK cells were identified by flow cytometric analysis. The cytotoxicity of CIK cells was determined by (51)Cr release assay. RESULTS: The CIK cells were shown to be a heterogeneous population with different cellular phenotypes. The percentage of CD3+/CD56+ positive cells, the dominant effector cells, in total CIK cells from healthy donors and HCC patients, significantly increased from 0.1-0.13% at day 0 to 19.0-20.5% at day 21 incubation, which suggested that the CD3+ CD56+ positive cells proliferated faster than other cell populations of CIK cells in the protocol used in this study. After 28 day in vitro incubation, the CIK cells from patients with HCC and healthy donors increased by more than 300-fold and 500-fold in proliferation cell number, respectively. CIK cells originated from HCC patients possessed a higher in vitro antitumor cytotoxic activity on autologous HCC cells than the autologous lymphokine-activated killer (LAK) cells and PBMC cells. In in vivo animal experiment, CIK cells had stronger effects on the inhibition of tumor growth in Balb/c nude mice bearing BEL-7402-producing tumor than LAK cells (mean inhibitory rate, 84.7% vs 52.8%, P【0.05) or PBMC (mean inhibitory rate, 84.7% vs 37.1%, P【0.01). CONCLUSION: Autologous CIK cells are of highly efficient cytotoxic effector cells against primary hepatocellular carcinoma cells and might serve as an alternative adoptive therapeutic strategy for HCC patients.展开更多
AIM： To establish nude mouse human gastric cancer orthotopic transplantation models using OB glue paste technique. METHODS： Using OB glue paste technique, orthtopic transplantation models were established by implant...AIM： To establish nude mouse human gastric cancer orthotopic transplantation models using OB glue paste technique. METHODS： Using OB glue paste technique, orthtopic transplantation models were established by implanting SGC-7901 and NKN-45 human gastric cancer cell strains into the gastric wall of nude mice. Biological features, growth of the implanted tumors, the success rate of transplantation and the rate of auto-metastasis of the two models were observed. RESULTS： The success rates of orthotopic transplanration of the two models were 94.20% and 96%. The rates of hepatic metastasis, pulmonary metastasis, peritoneal metastasis, lymphocytic metastasis and splenic metastasis were 42.13% and 94.20%, 48.43% and 57.97%, 30.83% and 36.96%, 67.30% and 84.06%, and 59.75% and 10.53%, respectively. The occurrence of ascites was 47.80% and 36.96%. CONCLUSION： OB glue paste technique is easy to follow. The biological behaviors of the nude mouse human gastric cancer orthotopic transplantation models established with this technique are similar to the natural processes of growth and metastasis of human gastric cancer, and, therefore, can be used as an ideal model for experimental research of proliferative metastasis of tumors.展开更多
AIM: To evaluate antihepatoma effect of antisense phosphorothioate oligodeoxyribonucleotides (S-ODNs) targeted to alpha-fetoprotein (AFP) genes in vitro and in nude mice. METHODS: AFP gene expression was examined by i...AIM: To evaluate antihepatoma effect of antisense phosphorothioate oligodeoxyribonucleotides (S-ODNs) targeted to alpha-fetoprotein (AFP) genes in vitro and in nude mice. METHODS: AFP gene expression was examined by immunocytochemical method or enzyme-linked immunosorbent assay. Effect of S-ODNs on SMMC-7721 human hepatoma cell growth in vitro was determined using microculture tetrazolium assay. In vitro antitumor activities of S-ODNs were monitored by measuring tumor weight differences in treated and control mice bearing SMMC-7721 xenografts. Induction of cell apoptosis was evaluated by fluorescence-activated cell sorter (FACS) analysis. RESULTS: Antisense S-ODN treatment led to reduced AFP gene expression. Specific antisense S-ODNs, but not control S-ODNs, inhibited the growth of hepatoma cells in vitro. In vitro, only antisense S-ODNs exhibited obvious antitumor activities. FACS analysis revealed that the growth inhibition by antisense S-ODNs was associated with their cell apoptosis induction. CONCLUSION: Antisense S-ODNs targeted to AFP genes inhibit the growth of human hepatoma cells and solid hepatoma, which is related to their cell apoptosis induction.展开更多
AIM: The transcription factor EGR-1 (early growth response gene-1) plays an important role in cell growth, differentiation and development. It has identified that EGR-1 has significant transformation suppression activ...AIM: The transcription factor EGR-1 (early growth response gene-1) plays an important role in cell growth, differentiation and development. It has identified that EGR-1 has significant transformation suppression activity in some neoplasms, such as fibrosarcoma, breast carcinoma. This experiment was designed to investigate the role of egr-1 in the cancerous process of hepatocellular carcinoma (HCC) and esophageal carcinoma (EC), and then to appraise the effects of EGR-1 on the growth of these tumor cells. METHODS: Firstly, the transcription and expression of egr-1 in HCC and EC, paracancerous tissues and their normal counterpart parts were detected by in situ hybridization and immunohistochemistry, with normal human breast and mouse brain tissues as positive controls. Egr-1 gene was then transfected into HCC (HHCC, SMMC7721) and EC (ECa109) cell lines in which no egr-1 transcription and expression were present. The cell growth speed, FCM cell cycle, plate clone formation and tumorigenicity in nude mice were observed and the controls were the cell lines transfected with vector only. RESULTS: Little or no egr-1 transcription and expression were detected in HCC, EC and normal liver tissues. The expression of egr-1 were found higher in hepatocellular paracancerous tissue (transcription level P=0.000; expression level P=0.143, probably because fewer in number of cases) and dysplastic tissue of esophageal cancer (transcription level P=0.000; expression level P=0.001). The growth rate of egr-1-transfected HHCC (HCC cell line) cells and ECa109 (EC cell line) cells was much slower than that of the controls. The proportion of S phase cell, clone formation and tumorigenicity were significantly lower than these of the controls' (decreased 45.5% in HHCC cells and 34.1% in ECa109 cells; 46.6% and 41.8%; 80.4% and 72.6% respectively). There were no obvious differences between SMMC7721 (HCC) egr-1-transfected cells and the controls with regard to the above items. CONCLUSION: The decreased expression of egr-1 might play a role in the dysregulation of normal growth in the cancerous process of HCC and EC. Egr-1 gene of transfected HHCC and ECa109 cells showed obvious suppression of the cell growth and malignant phenotypes, but no suppression in SMMC7721 (HCC cell line) cells.展开更多
目的探究细胞有丝分裂前期检查点(checkpoint with forkhead-associated and ring finge,CHFR)对鼻咽癌(NPC)5-8F细胞裸鼠移植瘤生长抑制作用,并探究可能的机制。方法将重组慢病毒LV-CHFR及空载体慢病毒-绿色荧光蛋白质粒(LV-GFP)作为...目的探究细胞有丝分裂前期检查点(checkpoint with forkhead-associated and ring finge,CHFR)对鼻咽癌(NPC)5-8F细胞裸鼠移植瘤生长抑制作用,并探究可能的机制。方法将重组慢病毒LV-CHFR及空载体慢病毒-绿色荧光蛋白质粒(LV-GFP)作为阴性对照导入5-8F细胞中,构建LV-CHFR/5-8F重组细胞,激光共聚焦显微镜观察细胞感染效率;将裸鼠随机分为3组(每组5只):空白对照组(接种0.2 ml未感染病毒的5-8F细胞悬液)、阴性对照组(接种0.2 ml稳转株LV-GFP/5-8F细胞悬液)、过表达CHFR组(接种0.2 ml稳转株LV-CHFR/5-8F细胞悬液),并绘制肿瘤生长曲线图;免疫组化法检测瘤体组织中丝氨酸/苏氨酸蛋白激酶Polo样激酶1(polo like kinase 1,PLK1)蛋白表达情况;原位末端转移酶标记(TUNEL)法检测各组瘤体组织中细胞凋亡情况;免疫印迹法检测各组瘤体组织中PLK1、CHFR、细胞周期素B(CyclinB1)、细胞周期蛋白(CyclinB1)、周期蛋白依赖性激酶2(CDC2)蛋白表达。结果随处理时间(9、12、15、18、21 d)延长,过表达CHFR组各时间点裸鼠瘤体体积均小于空白对照组、阴性对照组(F=7.872、5.778、6.717、70.020、11.782,P=0.007、0.011、0.010、0.001)。免疫组化检测显示,在空白对照组、阴性对照组裸鼠移植瘤组织中PLK1呈强阳性,在过表达CHFR组中,裸鼠移植瘤组织中PLK1表达呈弱阳性。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤细胞凋亡率升高(P<0.005)。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤细胞凋亡率升高(F=355.256,P=0.000)。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤组织中PLK1、Cyclin B1、CDC2蛋白水平均降低(F=5.201、2.0304、129.946,P=0.024、0.142、0.000),CHFR蛋白水平升高(F=38.919,P=0.000)。结论过表达CHFR对NPC 5-8F细胞裸鼠移植瘤生长有一定抑制作用,可能与抑制PLK1通路活化有关。展开更多
文摘The development of human endometrial carcinoma(HEC) is a complex pathologic process involves several oncogenes and tumor suppressor genes.The full-length paired-box gene 2(pax2),a recently discovered oncogene,promotes cell proliferation and growth and inhibits apoptosis of HEC cells.Here,we examined the effect of pax2 small interfering RNA(siRNA) on the growth of transplanted HEC cells in nude mice.The expression of Pax2 in 21 cases of normal endometrium and 38 cases of HEC was examined by immohistochemistry(IHC).HEC models were developed by subcutaneously transferring HEC cells into nude mice,followed by treatment with empty lentivirus vector,lentivirus vector-based pax2 siRNA,and phosphate buffered saline,respectively.Four weeks later,tumor size was measured,tumor inhibition rate was calculated,and histological analyses were conducted after staining with hematoxylin and eosin.The expression of Pax2 and Bcl-2 was detected by Western blot;proliferating cell nuclear antigen(PCNA) was detected by IHC.Significant differences were observed in the positive rate of Pax2 between normal endometrium and HEC(14.2% vs.60.5%,P<0.01).The expression index of Pax2 in well differentiated tumors was 1.88±1.68,much lower than that in tumors of moderate(3.07±1.96,P<0.05) or poor differentiation(5.45±2.76,P<0.01).Tumor necrosis increased,nuclear basophilia stain decreased,tumor growth was inhibited,and PCNA,Pax2,and Bcl-2 expression was reduced in HEC models treated with pax2 siRNA.These results indicate that Pax2 expression is related to HEC tumor biology with the increased expression of Pax2 correlated to malignancy.pax2 siRNA down-regulates Pax2 expression and inhibits tumorigenesis of HEC in nude mice,possibly due to cell apoptosis and the inhibition of tumor proliferation induced by down-regulation of Bcl-2.
文摘AIM: Epidermal growth factor (EGF) plays an important role in the regulation of gastrointestinal tissue growth and development, and it can stimulate epithelial proliferation, cell differentiation and growth. It has been established that the EGF can promote gastric cytoprotection and ulcer healing. But the potential ability of EGF to regulate the gastric cancer growth is unknown. This study is to investigate the influence of EGF on human gastric cancer cell and the implanted tumor growth of nude mice. METHODS: The cell growth rates of human gastric adenocarcinoma cell lines MKN-28, MKN-45, SGC-7901 and normal human gastric epithelial cells 3T3 were assessed when incubated with recombinant human EGF (rhEGF, 0.05, 0.1, 0.5, 1.0, 10, 50, 100 mg.L(-1)) using MTT method. The cells of MKN-28, MKN-45, SGC-7901 (gastric cancer tissue 1.5mm(3)) were implanted in the BALB/cA nude mice for 10 days.The EGF was given intraperitoneally (15, 30, 60 microg.kg(-1)) for 3 weeks. The body weights of the tumor-bearing animals and their tumor mass were measured afterwards to assess the mitogenic effect of rhEGF in the nude mice. RESULTS: Within the concentration range of 0.05-100mg.L(-1), rhEGF could increase the cell growth of normal 3T3 cells (cell growth rate 100% vs 102.8%, P【0.05), but partially restrain the gastric cancer cell growth. The latter effect was related to cell differentiation. In 15-60 microg/kg rhEGF groups, the mean implanted tumor mass of MKN-28 cell were 1.75 g, 1.91 g, 2.08 g/NS group 1.97 g (P】0.05), the mean tumor mass of SGC-7901 cell were 1.53 g, 1.07 g, 1.20 g/NS group 1.07 g (P】0.05), and for MKN-45 cell, the tumor mass were respectively 1.92 g, 1.29 g, 1.77 /NS group 1.82 g (P】0.05). So rhEGF had no obvious effect on implanted MKN-28, SGC-7901 and MKN-45 tumor growth. CONCLUSION: EGF has no stimulating effect on the human gastric cancer cell growth neither in vitro nor in vivo.
基金Supported by National Natural Science Foundation of China, No.81201963Inner Mongolia Natural Science Foundation of China,No.2010MS1123
文摘AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.
基金Science and Technology Development Foundation of Beijing Institute of Infectious Diseases,No.01 Z094
文摘AIM: To characterize the anticancer function of cytokine-induced killer cells (CIK) and develop an adoptive immunotherapy for the patients with primary hepatocellular carcinoma (HCC), we evaluated the proliferation rate, phenotype and the antitumor activity of human CIK cells from healthy donors and HCC patients in vitro and in vivo. METHODS: Peripheral blood mononuclear cells (PBMC) from healthy donors and patients with primary HCC were incubated in vitro and induced into CIK cells in the presence of various cytokines such as interferon-gamma (IFN-gamma), interleukin-1 (IL-1), IL-2 and monoclonal antibody (mAb) against CD3. The phenotype and characterization of CIK cells were identified by flow cytometric analysis. The cytotoxicity of CIK cells was determined by (51)Cr release assay. RESULTS: The CIK cells were shown to be a heterogeneous population with different cellular phenotypes. The percentage of CD3+/CD56+ positive cells, the dominant effector cells, in total CIK cells from healthy donors and HCC patients, significantly increased from 0.1-0.13% at day 0 to 19.0-20.5% at day 21 incubation, which suggested that the CD3+ CD56+ positive cells proliferated faster than other cell populations of CIK cells in the protocol used in this study. After 28 day in vitro incubation, the CIK cells from patients with HCC and healthy donors increased by more than 300-fold and 500-fold in proliferation cell number, respectively. CIK cells originated from HCC patients possessed a higher in vitro antitumor cytotoxic activity on autologous HCC cells than the autologous lymphokine-activated killer (LAK) cells and PBMC cells. In in vivo animal experiment, CIK cells had stronger effects on the inhibition of tumor growth in Balb/c nude mice bearing BEL-7402-producing tumor than LAK cells (mean inhibitory rate, 84.7% vs 52.8%, P【0.05) or PBMC (mean inhibitory rate, 84.7% vs 37.1%, P【0.01). CONCLUSION: Autologous CIK cells are of highly efficient cytotoxic effector cells against primary hepatocellular carcinoma cells and might serve as an alternative adoptive therapeutic strategy for HCC patients.
文摘AIM： To establish nude mouse human gastric cancer orthotopic transplantation models using OB glue paste technique. METHODS： Using OB glue paste technique, orthtopic transplantation models were established by implanting SGC-7901 and NKN-45 human gastric cancer cell strains into the gastric wall of nude mice. Biological features, growth of the implanted tumors, the success rate of transplantation and the rate of auto-metastasis of the two models were observed. RESULTS： The success rates of orthotopic transplanration of the two models were 94.20% and 96%. The rates of hepatic metastasis, pulmonary metastasis, peritoneal metastasis, lymphocytic metastasis and splenic metastasis were 42.13% and 94.20%, 48.43% and 57.97%, 30.83% and 36.96%, 67.30% and 84.06%, and 59.75% and 10.53%, respectively. The occurrence of ascites was 47.80% and 36.96%. CONCLUSION： OB glue paste technique is easy to follow. The biological behaviors of the nude mouse human gastric cancer orthotopic transplantation models established with this technique are similar to the natural processes of growth and metastasis of human gastric cancer, and, therefore, can be used as an ideal model for experimental research of proliferative metastasis of tumors.
基金Supported by the National Postdoctoral Science Foundation of China,No.199711.
文摘AIM: To evaluate antihepatoma effect of antisense phosphorothioate oligodeoxyribonucleotides (S-ODNs) targeted to alpha-fetoprotein (AFP) genes in vitro and in nude mice. METHODS: AFP gene expression was examined by immunocytochemical method or enzyme-linked immunosorbent assay. Effect of S-ODNs on SMMC-7721 human hepatoma cell growth in vitro was determined using microculture tetrazolium assay. In vitro antitumor activities of S-ODNs were monitored by measuring tumor weight differences in treated and control mice bearing SMMC-7721 xenografts. Induction of cell apoptosis was evaluated by fluorescence-activated cell sorter (FACS) analysis. RESULTS: Antisense S-ODN treatment led to reduced AFP gene expression. Specific antisense S-ODNs, but not control S-ODNs, inhibited the growth of hepatoma cells in vitro. In vitro, only antisense S-ODNs exhibited obvious antitumor activities. FACS analysis revealed that the growth inhibition by antisense S-ODNs was associated with their cell apoptosis induction. CONCLUSION: Antisense S-ODNs targeted to AFP genes inhibit the growth of human hepatoma cells and solid hepatoma, which is related to their cell apoptosis induction.
基金the National Natural Scientific Foundation of China,No.39670298
文摘AIM: The transcription factor EGR-1 (early growth response gene-1) plays an important role in cell growth, differentiation and development. It has identified that EGR-1 has significant transformation suppression activity in some neoplasms, such as fibrosarcoma, breast carcinoma. This experiment was designed to investigate the role of egr-1 in the cancerous process of hepatocellular carcinoma (HCC) and esophageal carcinoma (EC), and then to appraise the effects of EGR-1 on the growth of these tumor cells. METHODS: Firstly, the transcription and expression of egr-1 in HCC and EC, paracancerous tissues and their normal counterpart parts were detected by in situ hybridization and immunohistochemistry, with normal human breast and mouse brain tissues as positive controls. Egr-1 gene was then transfected into HCC (HHCC, SMMC7721) and EC (ECa109) cell lines in which no egr-1 transcription and expression were present. The cell growth speed, FCM cell cycle, plate clone formation and tumorigenicity in nude mice were observed and the controls were the cell lines transfected with vector only. RESULTS: Little or no egr-1 transcription and expression were detected in HCC, EC and normal liver tissues. The expression of egr-1 were found higher in hepatocellular paracancerous tissue (transcription level P=0.000; expression level P=0.143, probably because fewer in number of cases) and dysplastic tissue of esophageal cancer (transcription level P=0.000; expression level P=0.001). The growth rate of egr-1-transfected HHCC (HCC cell line) cells and ECa109 (EC cell line) cells was much slower than that of the controls. The proportion of S phase cell, clone formation and tumorigenicity were significantly lower than these of the controls' (decreased 45.5% in HHCC cells and 34.1% in ECa109 cells; 46.6% and 41.8%; 80.4% and 72.6% respectively). There were no obvious differences between SMMC7721 (HCC) egr-1-transfected cells and the controls with regard to the above items. CONCLUSION: The decreased expression of egr-1 might play a role in the dysregulation of normal growth in the cancerous process of HCC and EC. Egr-1 gene of transfected HHCC and ECa109 cells showed obvious suppression of the cell growth and malignant phenotypes, but no suppression in SMMC7721 (HCC cell line) cells.
文摘目的探究细胞有丝分裂前期检查点(checkpoint with forkhead-associated and ring finge,CHFR)对鼻咽癌(NPC)5-8F细胞裸鼠移植瘤生长抑制作用,并探究可能的机制。方法将重组慢病毒LV-CHFR及空载体慢病毒-绿色荧光蛋白质粒(LV-GFP)作为阴性对照导入5-8F细胞中,构建LV-CHFR/5-8F重组细胞,激光共聚焦显微镜观察细胞感染效率;将裸鼠随机分为3组(每组5只):空白对照组(接种0.2 ml未感染病毒的5-8F细胞悬液)、阴性对照组(接种0.2 ml稳转株LV-GFP/5-8F细胞悬液)、过表达CHFR组(接种0.2 ml稳转株LV-CHFR/5-8F细胞悬液),并绘制肿瘤生长曲线图;免疫组化法检测瘤体组织中丝氨酸/苏氨酸蛋白激酶Polo样激酶1(polo like kinase 1,PLK1)蛋白表达情况;原位末端转移酶标记(TUNEL)法检测各组瘤体组织中细胞凋亡情况;免疫印迹法检测各组瘤体组织中PLK1、CHFR、细胞周期素B(CyclinB1)、细胞周期蛋白(CyclinB1)、周期蛋白依赖性激酶2(CDC2)蛋白表达。结果随处理时间(9、12、15、18、21 d)延长,过表达CHFR组各时间点裸鼠瘤体体积均小于空白对照组、阴性对照组(F=7.872、5.778、6.717、70.020、11.782,P=0.007、0.011、0.010、0.001)。免疫组化检测显示,在空白对照组、阴性对照组裸鼠移植瘤组织中PLK1呈强阳性,在过表达CHFR组中,裸鼠移植瘤组织中PLK1表达呈弱阳性。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤细胞凋亡率升高(P<0.005)。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤细胞凋亡率升高(F=355.256,P=0.000)。与空白对照组、阴性对照组比较,过表达CHFR组移植瘤组织中PLK1、Cyclin B1、CDC2蛋白水平均降低(F=5.201、2.0304、129.946,P=0.024、0.142、0.000),CHFR蛋白水平升高(F=38.919,P=0.000)。结论过表达CHFR对NPC 5-8F细胞裸鼠移植瘤生长有一定抑制作用,可能与抑制PLK1通路活化有关。
