Objective: To investigate the feasibility of marking the human tumor cells with enhanced green fluorescent protein (EGFP) in vitro. Methods: The retroviral vector LGSN encoding EGFP was constructed and three human tum...Objective: To investigate the feasibility of marking the human tumor cells with enhanced green fluorescent protein (EGFP) in vitro. Methods: The retroviral vector LGSN encoding EGFP was constructed and three human tumor cell lines were infected with LGSN amphotropic virus. Tumor cell lines that stably express EGFP were selected with G418. The integration and expression of EGFP gene were analyzed by polymerase chain reaction, and flow cytometry (FCM). Results: After gene transfection and ping-pong transduction, amphotropic producer line Am12/LGSN was generated with a stable green fluorescence signal readily detectable by FCM in up to 97% of examined cells. The viral titer in the supernatants was up to 8.2×105CFU/ml. After transduction and selection, G418-resistant leukemia K562, mammary carcinoma MCF-7, and bladder cancer 5637 cells were developed, in which the integration of both EGFP and neomycin resistance gene was confirmed by DNA amplification. In comparison with uninfected cells, FCM analysis revealed EGFP expression in up to 90% (range 85.5%–90.0%) of tumor cells containing LGSN provirus. Conclusion: The retroviral vector LGSN can effectively mark the human tumor cells with a stably EGFP expression which may be in studying tumor growth, metastasis and angiogenesis.展开更多
Since taking part as leading actors in driving the metastatic process,circulating tumor cells(CTCs)have displayed a wide range of potential applications in the cancer-related research field.Besides their well-proved p...Since taking part as leading actors in driving the metastatic process,circulating tumor cells(CTCs)have displayed a wide range of potential applications in the cancer-related research field.Besides their well-proved prognostic value,the role of CTCs in both predictive and diagnostics terms might be extremely informative about cancer properties and therefore highly helpful in the clinical decision-making process.Unfortunately,CTCs are scarcely released in the blood circulation and their counts vary a lot among different types of cancer,therefore CTC detection and consequent characterization are still highly challenging.In this context,in vitro CTC cultures could potentially offer a great opportunity to expand the number of tumor cells isolated at different stages of the disease and thus simplify the analysis of their biological and molecular features,allowing a deeper comprehension of the nature of neoplastic diseases.The aim of this review is to highlight the main attempts to establish in vitro CTC cultures from patients harboring different tumor types in order to highlight how powerful this practice could be,especially in optimizing the therapeutic strategies available in clinical practice and potentially preventing or contrasting the development of treatment resistance.展开更多
OBJECTIVE: To investigate the response of multiple myeloma (MM) cells to arsenic trioxide (As2O3) and their possible mechanisms. METHODS: Two MM-derived cell lines RPMI8226 and U266 cells were used as in vitro models....OBJECTIVE: To investigate the response of multiple myeloma (MM) cells to arsenic trioxide (As2O3) and their possible mechanisms. METHODS: Two MM-derived cell lines RPMI8226 and U266 cells were used as in vitro models. Cell apoptosis was assessed by morphology, flow cytometry, and DNA gel electrophoresis. Mitochondrial transmembrane potentials (delta psi m) were evaluated by measuring cellular Rhodamine 123 staining intensity. Protein expression was analyzed using Western blot. RESULTS: Zero point one to 0.5 mumol/L As2O3 inhibited cell proliferation and 2.0 mumol/L As2O3 induced cell apoptosis, while 1.0 mumol/L As2O3 inhibited proliferation with a weak degree of apoptosis induction in RPMI8226 and U266 cell lines. As2O3-induced apoptosis was accompanied by mitochondrial transmembrane potentials (delta psi m) collapse and caspase-3 activation in the presence of intact membrane. Glutathione depleter buthionine sulfoximine enhanced, while disulfide bond-reducing agent dithiothreitol partially antagonized As2O3-induced delta psi m collapse and apoptosis in MM cells. All-trans retinoic acid (ATRA) could also induce apoptosis in RPMI8226 cells, but it did not show any cooperative effects with As2O3. CONCLUSION: As2O3 exerts apoptosis-inducing and growth-inhibiting effects on MM cells, and mitochondrium is a pivotal and common target of As2O3 for apoptosis induction.展开更多
OBJECTIVE: To investigate genes involved in cancer metastasis, mRNA differential display was used to compare the levels of gene expression in two cell sublines derived from human giant cell carcinoma of lung (PG) whic...OBJECTIVE: To investigate genes involved in cancer metastasis, mRNA differential display was used to compare the levels of gene expression in two cell sublines derived from human giant cell carcinoma of lung (PG) which had different metastatic potentials. METHODS: Using in vivo tumorigenicity and a spontaneous metastasis assay in nude mice, two sublines (BE1, LH7) from human giant cell carcinoma of lung (PG) with different metastatic potentials were isolated and characterized. mRNA differential display was used to compare the levels of gene expression between them and the obtained results were confirmed by Northern hybridization. RESULTS: One differentially expressed band was nearly identical (99% homology) to Ras-GTPase-Activating protein SH3 domain binding protein (G3BP). G3BP displayed a strong expression in LH7 (non-metastatic in recipient nude mice) and a very weak expression in BE1 (100% metastatic frequency). The same different expression level of G3BP was detected in Northern hybridization with another panel of cell sublines with different metastatic potentials (established in our lab) derived from human prostate carcinoma cell line PC-3M. CONCLUSION: Our results indicate that G3BP was implicated in cancer metastasis because of its differential expressions in the two panels of cell sublines with different metastatic potentials.展开更多
OBJECTIVE: To study the apoptotic effects of influenza A virus on the Raji cell line. METHODS: Cultured Raji cells were infected with influenza A virus at a multiplicity of infection (m.o.i) of 20 and the effects of a...OBJECTIVE: To study the apoptotic effects of influenza A virus on the Raji cell line. METHODS: Cultured Raji cells were infected with influenza A virus at a multiplicity of infection (m.o.i) of 20 and the effects of apoptosis were detected at different time points post infection using the following methods: electron microscope, DNA agarose gel electrophoresis, PI stained flow cytometry (FCM) and Annexin-V FITC/PI stained FCM. RESULTS: Raji cells infected with influenza A virus showed changes of morphology apoptosis, DNA agarose electrophoresis also demonstrated a ladder-like pattern of DNA fragments in a time-dependent manner. PI stained FCM showed 'apoptosis peak' and FITC/PI stained FCM showed apoptotic cells. Quantitative analysis indicated that the percentage of apoptotic Raji cells increased after infection, and cycloheximide (CHX), an eukaryotic transcription inhibitor, could effectively inhibit the apoptotic effects of influenza A virus in vitro. CONCLUSIONS: Influenza A virus can induce apoptosis in Raji cell line suggesting that it may lead to a potential method for tumor therapy.展开更多
文摘Objective: To investigate the feasibility of marking the human tumor cells with enhanced green fluorescent protein (EGFP) in vitro. Methods: The retroviral vector LGSN encoding EGFP was constructed and three human tumor cell lines were infected with LGSN amphotropic virus. Tumor cell lines that stably express EGFP were selected with G418. The integration and expression of EGFP gene were analyzed by polymerase chain reaction, and flow cytometry (FCM). Results: After gene transfection and ping-pong transduction, amphotropic producer line Am12/LGSN was generated with a stable green fluorescence signal readily detectable by FCM in up to 97% of examined cells. The viral titer in the supernatants was up to 8.2×105CFU/ml. After transduction and selection, G418-resistant leukemia K562, mammary carcinoma MCF-7, and bladder cancer 5637 cells were developed, in which the integration of both EGFP and neomycin resistance gene was confirmed by DNA amplification. In comparison with uninfected cells, FCM analysis revealed EGFP expression in up to 90% (range 85.5%–90.0%) of tumor cells containing LGSN provirus. Conclusion: The retroviral vector LGSN can effectively mark the human tumor cells with a stably EGFP expression which may be in studying tumor growth, metastasis and angiogenesis.
文摘Since taking part as leading actors in driving the metastatic process,circulating tumor cells(CTCs)have displayed a wide range of potential applications in the cancer-related research field.Besides their well-proved prognostic value,the role of CTCs in both predictive and diagnostics terms might be extremely informative about cancer properties and therefore highly helpful in the clinical decision-making process.Unfortunately,CTCs are scarcely released in the blood circulation and their counts vary a lot among different types of cancer,therefore CTC detection and consequent characterization are still highly challenging.In this context,in vitro CTC cultures could potentially offer a great opportunity to expand the number of tumor cells isolated at different stages of the disease and thus simplify the analysis of their biological and molecular features,allowing a deeper comprehension of the nature of neoplastic diseases.The aim of this review is to highlight the main attempts to establish in vitro CTC cultures from patients harboring different tumor types in order to highlight how powerful this practice could be,especially in optimizing the therapeutic strategies available in clinical practice and potentially preventing or contrasting the development of treatment resistance.
基金theNationalNaturalScienceFoundationofChina (No 39970 312andNo 39730 2 70 ) NationalOutstandingYoungScientificFoundationofC
文摘OBJECTIVE: To investigate the response of multiple myeloma (MM) cells to arsenic trioxide (As2O3) and their possible mechanisms. METHODS: Two MM-derived cell lines RPMI8226 and U266 cells were used as in vitro models. Cell apoptosis was assessed by morphology, flow cytometry, and DNA gel electrophoresis. Mitochondrial transmembrane potentials (delta psi m) were evaluated by measuring cellular Rhodamine 123 staining intensity. Protein expression was analyzed using Western blot. RESULTS: Zero point one to 0.5 mumol/L As2O3 inhibited cell proliferation and 2.0 mumol/L As2O3 induced cell apoptosis, while 1.0 mumol/L As2O3 inhibited proliferation with a weak degree of apoptosis induction in RPMI8226 and U266 cell lines. As2O3-induced apoptosis was accompanied by mitochondrial transmembrane potentials (delta psi m) collapse and caspase-3 activation in the presence of intact membrane. Glutathione depleter buthionine sulfoximine enhanced, while disulfide bond-reducing agent dithiothreitol partially antagonized As2O3-induced delta psi m collapse and apoptosis in MM cells. All-trans retinoic acid (ATRA) could also induce apoptosis in RPMI8226 cells, but it did not show any cooperative effects with As2O3. CONCLUSION: As2O3 exerts apoptosis-inducing and growth-inhibiting effects on MM cells, and mitochondrium is a pivotal and common target of As2O3 for apoptosis induction.
基金86 3HighTechnologyProject (No # 10 2 10 0 1 0 9)andSpecialFoundationforPh DTrainingProgramofEducationMinistry (No 19990 0
文摘OBJECTIVE: To investigate genes involved in cancer metastasis, mRNA differential display was used to compare the levels of gene expression in two cell sublines derived from human giant cell carcinoma of lung (PG) which had different metastatic potentials. METHODS: Using in vivo tumorigenicity and a spontaneous metastasis assay in nude mice, two sublines (BE1, LH7) from human giant cell carcinoma of lung (PG) with different metastatic potentials were isolated and characterized. mRNA differential display was used to compare the levels of gene expression between them and the obtained results were confirmed by Northern hybridization. RESULTS: One differentially expressed band was nearly identical (99% homology) to Ras-GTPase-Activating protein SH3 domain binding protein (G3BP). G3BP displayed a strong expression in LH7 (non-metastatic in recipient nude mice) and a very weak expression in BE1 (100% metastatic frequency). The same different expression level of G3BP was detected in Northern hybridization with another panel of cell sublines with different metastatic potentials (established in our lab) derived from human prostate carcinoma cell line PC-3M. CONCLUSION: Our results indicate that G3BP was implicated in cancer metastasis because of its differential expressions in the two panels of cell sublines with different metastatic potentials.
基金This work was supported by a grant from the Natural Science Foundation of China (No.39970830).
文摘OBJECTIVE: To study the apoptotic effects of influenza A virus on the Raji cell line. METHODS: Cultured Raji cells were infected with influenza A virus at a multiplicity of infection (m.o.i) of 20 and the effects of apoptosis were detected at different time points post infection using the following methods: electron microscope, DNA agarose gel electrophoresis, PI stained flow cytometry (FCM) and Annexin-V FITC/PI stained FCM. RESULTS: Raji cells infected with influenza A virus showed changes of morphology apoptosis, DNA agarose electrophoresis also demonstrated a ladder-like pattern of DNA fragments in a time-dependent manner. PI stained FCM showed 'apoptosis peak' and FITC/PI stained FCM showed apoptotic cells. Quantitative analysis indicated that the percentage of apoptotic Raji cells increased after infection, and cycloheximide (CHX), an eukaryotic transcription inhibitor, could effectively inhibit the apoptotic effects of influenza A virus in vitro. CONCLUSIONS: Influenza A virus can induce apoptosis in Raji cell line suggesting that it may lead to a potential method for tumor therapy.
