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Volume 16,Issue 2,2009 Table of Contents
2009, 16(2):101-105.
DOI: 10.3872/j.issn.1007-385X.2009.2.001
Abstract:
Biochemotherapy refers to the combination of biotherapy and chemotherapy, and it has been rapidly developed since its emergence. Biochemotherapy has brought both new therapies and new therapeutic ideas for the treatment of malignant tumors. In this article, we reviewed the recent advances in tumor biochemotherapies, including expansion of the indications, retaining of therapy even when tumor progressed (unique to biochemotherapy), and reversal of chemotherapy resistance by targeted therapy, the predicting response to tumor biochemotherapy, recent clinical trials of immunotherapybased chemotherapy, and evaluation criteria for assessment of biochemotherapy response, and so on.
Biochemotherapy refers to the combination of biotherapy and chemotherapy, and it has been rapidly developed since its emergence. Biochemotherapy has brought both new therapies and new therapeutic ideas for the treatment of malignant tumors. In this article, we reviewed the recent advances in tumor biochemotherapies, including expansion of the indications, retaining of therapy even when tumor progressed (unique to biochemotherapy), and reversal of chemotherapy resistance by targeted therapy, the predicting response to tumor biochemotherapy, recent clinical trials of immunotherapybased chemotherapy, and evaluation criteria for assessment of biochemotherapy response, and so on.
2009, 16(2):106-112.
DOI: 10.3872/j.issn.1007-385X.2009.2.002
Abstract:
Objective: To compare the different protocols for amplifying human peripheral blood γδ T cells and establish the suitable expansion and storage conditions for clinical application.Methods: Human peripheral blood mononuclear cells were isolated and stimulated by immobilized or soluble antiTCR γδ, and further cultured in RPMI 1640 supplemented with newborn bovine serum, RPMI 1640 supplemented with human AB plasma, or serum free AIM V medium separately. Cell numbers and survival rates were detected by trypan blue staining; purities and phenotypes of the expanded γδ T cells were measured by immunofluorescence staining; cytotoxicities of γδ T cells against different tumor cell lines were detected by lactate dehydrogenase (LDH) method. Results: RPMI 1640 supplemented with 15% newborn bovine serum or 5% human AB plasma had similar effects on the proliferation of γδ T cells but better than that of AIM V serum free medium. Soluble antibody stimulated the expansion of peripheral blood γδ T cells as effectively as immobilized antibody did. The expanded γδ T cells had complete T cell receptor (TCR) pool, containing TCR Vδ1 and Vδ2 subtypes, and displayed cytotoxic activities aganist lung cancer, hepatocellular carcinoma, and ovarian carcinoma cells in vitro. These γδ T cells, when suspended in normal saline supplemented with 0.25% human albumin or 1% human AB plasma, could maintain 95% survival for 12 hours at 4 ℃. Conclusion:Human peripheral blood γδ T cells can be effectively expanded in vitro by soluble antiTCR γδ in RPMI 1640 culture medium containing human AB plasma, and these expanded γδ T cells can be stably preserved in normal saline supplemented with plasma or albumin.
Objective: To compare the different protocols for amplifying human peripheral blood γδ T cells and establish the suitable expansion and storage conditions for clinical application.Methods: Human peripheral blood mononuclear cells were isolated and stimulated by immobilized or soluble antiTCR γδ, and further cultured in RPMI 1640 supplemented with newborn bovine serum, RPMI 1640 supplemented with human AB plasma, or serum free AIM V medium separately. Cell numbers and survival rates were detected by trypan blue staining; purities and phenotypes of the expanded γδ T cells were measured by immunofluorescence staining; cytotoxicities of γδ T cells against different tumor cell lines were detected by lactate dehydrogenase (LDH) method. Results: RPMI 1640 supplemented with 15% newborn bovine serum or 5% human AB plasma had similar effects on the proliferation of γδ T cells but better than that of AIM V serum free medium. Soluble antibody stimulated the expansion of peripheral blood γδ T cells as effectively as immobilized antibody did. The expanded γδ T cells had complete T cell receptor (TCR) pool, containing TCR Vδ1 and Vδ2 subtypes, and displayed cytotoxic activities aganist lung cancer, hepatocellular carcinoma, and ovarian carcinoma cells in vitro. These γδ T cells, when suspended in normal saline supplemented with 0.25% human albumin or 1% human AB plasma, could maintain 95% survival for 12 hours at 4 ℃. Conclusion:Human peripheral blood γδ T cells can be effectively expanded in vitro by soluble antiTCR γδ in RPMI 1640 culture medium containing human AB plasma, and these expanded γδ T cells can be stably preserved in normal saline supplemented with plasma or albumin.
2009, 16(2):113-119.
DOI: 10.3872/j.issn.1007-385X.2009.2.003
Abstract:
Objective: To construct a novel conditional duplicate adenovirus (CRA) regulated by AFP enhancer, AFP promoter and TK suicide gene, and to observe its duplicate abilities, cytopathic effects and cytotoxity against hepatocarcinoma cells when combined with predrug GCV.Methods: AFP gene promoter and enhancer were amplified using PCR from the genome DNA of HepG2 cells; pAFPpEGFPluc and pAFPepEGFPluc plasmids were constructed; pDC311AFPepE1A/CMVTK shuttle plasmid expression E1A gene was then constructed under the control of AFP promoter, AFP enhancer (AFPep) and TK gene. Ad.AFPepE1A/CMVTK vector was constructed by AdMax system. ElA expression, cytopathic effects and cytotoxity against hepatocarcinoma cells of Ad.AFPepE1A/CMVTK were evaluated by Western blotting, flow cytometry, CCK8 assay, respectively. Results: Ad.AFPepE1A/CMVTK was successfully constructed, which could selectively replicate in AFP positive HepG2 cells and specifically inhibit proliferation of AFP positive cells. Survival rates of AFP positive HepG2 and Hep3B cells after treated with combination of Ad.AFPepE1A/CMVTK and predrug GCV were (10.35±1.07)% and (15.49±5.80)%, respectively, which were significantly lower than those of AFP negative Chang liver cells and lung cancer NCIH460 cells (\[73.55±4.36\]% and \[74.54±9.89\]%, respectively, P<0.01). Conclusion: The constructed Ad.AFPepE1A/CMVTK, when combines with TK/GCV suicide gene system, can specifically kill AFP positive hepatocarcinoma cells, which may have a future in genebased therapy against hepatocarcinoma.
Objective: To construct a novel conditional duplicate adenovirus (CRA) regulated by AFP enhancer, AFP promoter and TK suicide gene, and to observe its duplicate abilities, cytopathic effects and cytotoxity against hepatocarcinoma cells when combined with predrug GCV.Methods: AFP gene promoter and enhancer were amplified using PCR from the genome DNA of HepG2 cells; pAFPpEGFPluc and pAFPepEGFPluc plasmids were constructed; pDC311AFPepE1A/CMVTK shuttle plasmid expression E1A gene was then constructed under the control of AFP promoter, AFP enhancer (AFPep) and TK gene. Ad.AFPepE1A/CMVTK vector was constructed by AdMax system. ElA expression, cytopathic effects and cytotoxity against hepatocarcinoma cells of Ad.AFPepE1A/CMVTK were evaluated by Western blotting, flow cytometry, CCK8 assay, respectively. Results: Ad.AFPepE1A/CMVTK was successfully constructed, which could selectively replicate in AFP positive HepG2 cells and specifically inhibit proliferation of AFP positive cells. Survival rates of AFP positive HepG2 and Hep3B cells after treated with combination of Ad.AFPepE1A/CMVTK and predrug GCV were (10.35±1.07)% and (15.49±5.80)%, respectively, which were significantly lower than those of AFP negative Chang liver cells and lung cancer NCIH460 cells (\[73.55±4.36\]% and \[74.54±9.89\]%, respectively, P<0.01). Conclusion: The constructed Ad.AFPepE1A/CMVTK, when combines with TK/GCV suicide gene system, can specifically kill AFP positive hepatocarcinoma cells, which may have a future in genebased therapy against hepatocarcinoma.
2009, 16(2):120-124.
DOI: 10.3872/j.issn.1007-385X.2009.2.004
Abstract:
Objective:To investigated the in vivo efficacy of altered CEA610D peptide vaccine against tumor, so as to provide a basis for its clinical use.Methods:Altered CEA peptide CEA610D, natural CEA peptide CEA605613 and MAGE3 peptide were synthesized by polypeptide solidphase synthesis system. The cytotoxicity T lymphocytes (CTLs) were induced by autologous mixed lymphocytes reaction implused with above peptides; proliferation and cytotoxicity of different CTLs were measured by MTT; phenotypes of the CTLs were detected by flow cytometry; expression of perforin in different CTLs were assayed by RTPCR; IFNγ levels in the supernatants of different CTLs were detected by ELISA.Results: CEA610D peptide more efficiently induced CTL than CEA605613 and MAGE3 peptide did (P<0.05). The number of CD8+T cells in CEA610D group was significantly larger than those in CEA605613 and MAGE3 groups (P<005). When the E∶T was 40∶1, the cytotoxicity of CTLs induced by CEA610D and CEA605613 against CEA+HLAA2+ T84 cells were (56.7±373)% and (51.2±1.86)%, respectively. But the CTL cytotoxicities induced by the three peptides against CEA+HLAA2Lovo cells were all at the background level. Perforin expression and IFNγ level of CTLs in CEA610D group were significantly higher than those in the other two groups (P<0.01).Conclusion:Compared with natural CEA605613 peptide, altered CEA610D peptide can effectively break the immune tolerance to self peptide, and thus has a stronger antitumor activity in vitro.
Objective:To investigated the in vivo efficacy of altered CEA610D peptide vaccine against tumor, so as to provide a basis for its clinical use.Methods:Altered CEA peptide CEA610D, natural CEA peptide CEA605613 and MAGE3 peptide were synthesized by polypeptide solidphase synthesis system. The cytotoxicity T lymphocytes (CTLs) were induced by autologous mixed lymphocytes reaction implused with above peptides; proliferation and cytotoxicity of different CTLs were measured by MTT; phenotypes of the CTLs were detected by flow cytometry; expression of perforin in different CTLs were assayed by RTPCR; IFNγ levels in the supernatants of different CTLs were detected by ELISA.Results: CEA610D peptide more efficiently induced CTL than CEA605613 and MAGE3 peptide did (P<0.05). The number of CD8+T cells in CEA610D group was significantly larger than those in CEA605613 and MAGE3 groups (P<005). When the E∶T was 40∶1, the cytotoxicity of CTLs induced by CEA610D and CEA605613 against CEA+HLAA2+ T84 cells were (56.7±373)% and (51.2±1.86)%, respectively. But the CTL cytotoxicities induced by the three peptides against CEA+HLAA2Lovo cells were all at the background level. Perforin expression and IFNγ level of CTLs in CEA610D group were significantly higher than those in the other two groups (P<0.01).Conclusion:Compared with natural CEA605613 peptide, altered CEA610D peptide can effectively break the immune tolerance to self peptide, and thus has a stronger antitumor activity in vitro.
2009, 16(2):125-129.
DOI: 10.3872/j.issn.1007-385X.2009.2.005
Abstract:
Objective:To evaluate the inhibitory effect of Nitroreductase/CB1954 (NTR/CB1954) suicide gene system，which contains 5 copies of hypoxiaresponsive element (5HRE) and promoter of alphafetoprotein gene (AFPp), against human hepatocellular carcinoma cell line HepG2 under hypoxia condition in vitro. Methods: 5HRE and AFPpregulated nitroreductase (NTR) gene eukaryotic expression vector was transfected into AFPpositive human hepatocellular carcinoma cell line HepG2 and AFPnegative human gastric carcinoma cell line MKN45 by Lipofectamine 2000. Stably transfected cell lines were selected by G418. RTPCR and Western blotting were employed to examine the expression of NTR mRNA and NTR protein, respectively. Prodrug CB1954 was added into stablytransfected cell lines; inhibitory activity of its derivative product (4hydroxylamine) was observed by MTT. Results: Monoclonal HepG2 cells stably expressing NTR were successfully obtained. Expression of NTR mRNA and NTR protein in monoclonal HepG2 cells under hypoxia condition was significantly higher than those under normoxia condition as confirmed by RTPCR and Western blotting, respectively. Monoclonal MKN45 cells did not express NTR protein under either hypoxia condition or normoxia condition. NTR effectively activated CB1954 under hypoxia condition, resulting in dosedependent inhibition on the proliferation of HepG2 cells as shown by MTT assay (P<0.05). But CB1954 did not inhibit proliferation of MKN45 cells and wild type HepG2 cells. Conclusion: 5HRE and AFPpregulated NTR/CB1954 suicide gene system can specifically kill human hepatocellular carcinoma cell line HepG2 under hypoxia condition.
Objective:To evaluate the inhibitory effect of Nitroreductase/CB1954 (NTR/CB1954) suicide gene system，which contains 5 copies of hypoxiaresponsive element (5HRE) and promoter of alphafetoprotein gene (AFPp), against human hepatocellular carcinoma cell line HepG2 under hypoxia condition in vitro. Methods: 5HRE and AFPpregulated nitroreductase (NTR) gene eukaryotic expression vector was transfected into AFPpositive human hepatocellular carcinoma cell line HepG2 and AFPnegative human gastric carcinoma cell line MKN45 by Lipofectamine 2000. Stably transfected cell lines were selected by G418. RTPCR and Western blotting were employed to examine the expression of NTR mRNA and NTR protein, respectively. Prodrug CB1954 was added into stablytransfected cell lines; inhibitory activity of its derivative product (4hydroxylamine) was observed by MTT. Results: Monoclonal HepG2 cells stably expressing NTR were successfully obtained. Expression of NTR mRNA and NTR protein in monoclonal HepG2 cells under hypoxia condition was significantly higher than those under normoxia condition as confirmed by RTPCR and Western blotting, respectively. Monoclonal MKN45 cells did not express NTR protein under either hypoxia condition or normoxia condition. NTR effectively activated CB1954 under hypoxia condition, resulting in dosedependent inhibition on the proliferation of HepG2 cells as shown by MTT assay (P<0.05). But CB1954 did not inhibit proliferation of MKN45 cells and wild type HepG2 cells. Conclusion: 5HRE and AFPpregulated NTR/CB1954 suicide gene system can specifically kill human hepatocellular carcinoma cell line HepG2 under hypoxia condition.
2009, 16(2):130-135.
DOI: 10.3872/j.issn.1007-385X.2009.2.006
Abstract:
Objective: To explore whether apoptotic ovarian cancer cells induced by chemotherapy drugs paclitaxel and cisplatin can be crosspresented by dendritic cells (DCs) and enhance immune responses. Methods: DCs were induced from peripheral blood monocytes cells by GMCSF/IL4 for 6 d, then they were stimulated with either apoptotic ovarian cancer HO8910 cells, frozenthawed HO8910 cells or control cells for 4 h. Their surface markers and phagocytotic ability were detected by flow cytometry and confocal microscopic scanning assay, respectively. DCs of different groups were cultured with CD8+ T cells isolated by magnetic cell sorting, and the ability of DCs to activate CD8+ T cells was evaluated by 3HTdR, the activity of CTL to kill tumor cells was evaluated by LDH. Production of IFNγ by CD8+ T cells was measured by ELISPOT. Results: Apoptotic ovarian cancer cells induced by chemotherapy drugs paclitaxel and cisplatin could be phagocytized by DCs, which subsequently promoted the maturation and antigen presenting ability of DCs. Apoptotic ovarian cancer cells implused DCs significantly promoted proliferation of CD8+ T cells compared with that of control cells (P＜0.05）; they also significantly increased the cytotocity of CTL to kill tumor cells compared with that of frozenthawed HO8910 cells and control cells as detected by LDH and ELISPOT（all P＜0.01). Conclusion: Apoptotic ovarian cancer cells induced by paclitaxel and cisplatin exhibit strong immunogenicity and enhanced ability to promote DCs maturation and antigen presentation, subsequently enhance CD8+ T cell proliferation and promote their ability to secrete IFNγ and kill tumor cells.
Objective: To explore whether apoptotic ovarian cancer cells induced by chemotherapy drugs paclitaxel and cisplatin can be crosspresented by dendritic cells (DCs) and enhance immune responses. Methods: DCs were induced from peripheral blood monocytes cells by GMCSF/IL4 for 6 d, then they were stimulated with either apoptotic ovarian cancer HO8910 cells, frozenthawed HO8910 cells or control cells for 4 h. Their surface markers and phagocytotic ability were detected by flow cytometry and confocal microscopic scanning assay, respectively. DCs of different groups were cultured with CD8+ T cells isolated by magnetic cell sorting, and the ability of DCs to activate CD8+ T cells was evaluated by 3HTdR, the activity of CTL to kill tumor cells was evaluated by LDH. Production of IFNγ by CD8+ T cells was measured by ELISPOT. Results: Apoptotic ovarian cancer cells induced by chemotherapy drugs paclitaxel and cisplatin could be phagocytized by DCs, which subsequently promoted the maturation and antigen presenting ability of DCs. Apoptotic ovarian cancer cells implused DCs significantly promoted proliferation of CD8+ T cells compared with that of control cells (P＜0.05）; they also significantly increased the cytotocity of CTL to kill tumor cells compared with that of frozenthawed HO8910 cells and control cells as detected by LDH and ELISPOT（all P＜0.01). Conclusion: Apoptotic ovarian cancer cells induced by paclitaxel and cisplatin exhibit strong immunogenicity and enhanced ability to promote DCs maturation and antigen presentation, subsequently enhance CD8+ T cell proliferation and promote their ability to secrete IFNγ and kill tumor cells.
2009, 16(2):136-139.
DOI: 10.3872/j.issn.1007-385X.2009.2.007
Abstract:
Objective:To study the inhibitory effect of GMCSF（granulocytemacrophage colony stimulating factor) membraneanchored B16.F10 (murine melanoma cell line) vaccine against implanted tumors in mice. Methods: GMCSF membraneanchored B16.F10 vaccine was prepared by fusing biotinavidinGMCSF together. BALB/c mice were randomly divided into 5 groups: GMCSF membraneanchored B16.F10, B16.F10 mixed with GMCSF, B16.F10 cells, GMCSF and 0.9% sodium chloride. At day 1 and 7, all BALB/c mice were immunized; at day 14, all the mice received subcutaneous injection of 0.2 ml wildtype B16.F10 cells (1×106 cells). Tumorfree rate and survival time of different groups were measured. INFγ levels in splenocytes were detected by ELISA at day 24 in different groups.Results: Sixty days and 90 d after subcutaneous injection with B16.F10 cells, tumorfree rate and survival rate of mice in GMCSF membraneanchored B16.F10 group, B16.F10 mixed with GMCSF group, B16.F10 cell group were 0% and 0%, 100% and 100%; 50% and 40%, 70% and 40%; 20% and 20%, and 80% and 20%, respectively. The tumorfree rate and survival rate of mice in GMCSF group and 0.9% sodium chloride group were both 100% and 0%, respectively. INFγ levels in splenocytes in GMCSF membraneanchored B16.F10 group were significantly higher than those of other groups（P<0.01）. Conclusion: GMCSF membraneanchored B16.F10 vaccine can induce specific antitumor immunity in mice and can prevent later B16.F10 tumor cells challenge.
Objective:To study the inhibitory effect of GMCSF（granulocytemacrophage colony stimulating factor) membraneanchored B16.F10 (murine melanoma cell line) vaccine against implanted tumors in mice. Methods: GMCSF membraneanchored B16.F10 vaccine was prepared by fusing biotinavidinGMCSF together. BALB/c mice were randomly divided into 5 groups: GMCSF membraneanchored B16.F10, B16.F10 mixed with GMCSF, B16.F10 cells, GMCSF and 0.9% sodium chloride. At day 1 and 7, all BALB/c mice were immunized; at day 14, all the mice received subcutaneous injection of 0.2 ml wildtype B16.F10 cells (1×106 cells). Tumorfree rate and survival time of different groups were measured. INFγ levels in splenocytes were detected by ELISA at day 24 in different groups.Results: Sixty days and 90 d after subcutaneous injection with B16.F10 cells, tumorfree rate and survival rate of mice in GMCSF membraneanchored B16.F10 group, B16.F10 mixed with GMCSF group, B16.F10 cell group were 0% and 0%, 100% and 100%; 50% and 40%, 70% and 40%; 20% and 20%, and 80% and 20%, respectively. The tumorfree rate and survival rate of mice in GMCSF group and 0.9% sodium chloride group were both 100% and 0%, respectively. INFγ levels in splenocytes in GMCSF membraneanchored B16.F10 group were significantly higher than those of other groups（P<0.01）. Conclusion: GMCSF membraneanchored B16.F10 vaccine can induce specific antitumor immunity in mice and can prevent later B16.F10 tumor cells challenge.
2009, 16(2):140-144.
DOI: 10.3872/j.issn.1007-385X.2009.2.008
Abstract:
Objective: To evaluate the role of proteasome inhibitor MG132 in reversing the acquired TRAIL resistance of human colon cancer cell line DLD1TRAIL/R and the related mechanisms. Methods:Colon cancer cell line DLD1TRAIL/Rwas treated with MG132 combined with TRAIL protein. The viability of DLD1TRAIL/R cells was determined by MTT assay; the apoptotic rate was detected by flow cytometry, and the expression of apoptosisrelated proteins was examined by Western blotting analysis. Results: The viability of DLD1TRAIL/R cells was dramatically decreased after combined treatment with MG132 and TRAIL protein(P<0.01) and the apoptotic rate was significantly increased (P<0.01). Western blotting analysis showed that MG132 dramatically enhanced the cleavage of apoptotic molecules, including caspases8, 9, 3, Bid, and PARP in DLD1TRAIL/R cells after combined treatment and increased the release of cytochrome C and Smac from mitochondria. Further study demonstrated that MG132 upregulated DR5 and Bik proteins, but had no detectable effects on DR4, Bax, Bak, BclXL, XIAP or survivin. Moreover, we found MG132 induced phosphorylation of kinase JNK, and the inhibitor of JNK (SP600125) blocked MG132induced expression of DR5, but not the expression of Bik. Furthermore, SP600125 did not attenuate the apoptosis of DLD1TRAIL/R cells induced by MG132 in the presence of TRAIL protein (P<0.05). Conclusion: Proteasome inhibitor MG132 can reverse the acquired drug resistance to TRAIL and induce upregulation of DR5 and Bik protein in DLD1TRAIL/R cells. The underlying mechanism may involve the initiation of mitochondrionrelated apoptosis caused by Bik protein expression, not by activation of JNK pathway.
Objective: To evaluate the role of proteasome inhibitor MG132 in reversing the acquired TRAIL resistance of human colon cancer cell line DLD1TRAIL/R and the related mechanisms. Methods:Colon cancer cell line DLD1TRAIL/Rwas treated with MG132 combined with TRAIL protein. The viability of DLD1TRAIL/R cells was determined by MTT assay; the apoptotic rate was detected by flow cytometry, and the expression of apoptosisrelated proteins was examined by Western blotting analysis. Results: The viability of DLD1TRAIL/R cells was dramatically decreased after combined treatment with MG132 and TRAIL protein(P<0.01) and the apoptotic rate was significantly increased (P<0.01). Western blotting analysis showed that MG132 dramatically enhanced the cleavage of apoptotic molecules, including caspases8, 9, 3, Bid, and PARP in DLD1TRAIL/R cells after combined treatment and increased the release of cytochrome C and Smac from mitochondria. Further study demonstrated that MG132 upregulated DR5 and Bik proteins, but had no detectable effects on DR4, Bax, Bak, BclXL, XIAP or survivin. Moreover, we found MG132 induced phosphorylation of kinase JNK, and the inhibitor of JNK (SP600125) blocked MG132induced expression of DR5, but not the expression of Bik. Furthermore, SP600125 did not attenuate the apoptosis of DLD1TRAIL/R cells induced by MG132 in the presence of TRAIL protein (P<0.05). Conclusion: Proteasome inhibitor MG132 can reverse the acquired drug resistance to TRAIL and induce upregulation of DR5 and Bik protein in DLD1TRAIL/R cells. The underlying mechanism may involve the initiation of mitochondrionrelated apoptosis caused by Bik protein expression, not by activation of JNK pathway.
2009, 16(2):145-150.
DOI: 10.3872/j.issn.1007-385X.2009.2.009
Abstract:
Objective: To study the effects of lentivirusmediated bcr/abl RNAi on the biological characteristics of human leukemia cell line K562. Methods:Bcr/abl RNAi lentivirus vector pNLB/AEGFP was constructed and was used to transfect K562 cells, the stable tansfectants（B/AK562）were selected. RNAi efficiency was assessed by Realtime PCR and Western blotting. Cell proliferation was detected by trypan blue staining and colony formation assay, cell differentiation was investigated by benzidine staining, PTK activity was determined by ELISA, apoptosis was observed by AOEB staining, and caspase3 and caspase9 activation were measured by chromometry. K562 cells and mock transfected EGFPK562 cells were used as controls.Results: pNLB/AEGFP stably transfected K562 cells (B/AK562) was successfully constructed. Realtime PCR and Western blotting analysis confirmed that lentivirusmediated bcr/abl RNAi downregulated bcr/abl mRNA and P210bcr/abl protein expression in K562 cells. The doubling time of B/AK562 cells was obviously longer than those of K562 cells and EGFPK562 cells (37.1 vs 20.4, 23.3 h). Furthermore, B/AK562 cells showed decreased colony formation ability, strengthened differentiation toward erythrocytes, decreased activation of PTK, increased apoptosis and enhanced caspase3 and caspase9 activation (P<0.05 or P<0.01). Conclusion: Lentivirusmediated bcr/abl RNAi can result in long time silencing of bcr/abl gene, inhibit malignant proliferation and induce differentiation and apoptosis in K562 cells.
Objective: To study the effects of lentivirusmediated bcr/abl RNAi on the biological characteristics of human leukemia cell line K562. Methods:Bcr/abl RNAi lentivirus vector pNLB/AEGFP was constructed and was used to transfect K562 cells, the stable tansfectants（B/AK562）were selected. RNAi efficiency was assessed by Realtime PCR and Western blotting. Cell proliferation was detected by trypan blue staining and colony formation assay, cell differentiation was investigated by benzidine staining, PTK activity was determined by ELISA, apoptosis was observed by AOEB staining, and caspase3 and caspase9 activation were measured by chromometry. K562 cells and mock transfected EGFPK562 cells were used as controls.Results: pNLB/AEGFP stably transfected K562 cells (B/AK562) was successfully constructed. Realtime PCR and Western blotting analysis confirmed that lentivirusmediated bcr/abl RNAi downregulated bcr/abl mRNA and P210bcr/abl protein expression in K562 cells. The doubling time of B/AK562 cells was obviously longer than those of K562 cells and EGFPK562 cells (37.1 vs 20.4, 23.3 h). Furthermore, B/AK562 cells showed decreased colony formation ability, strengthened differentiation toward erythrocytes, decreased activation of PTK, increased apoptosis and enhanced caspase3 and caspase9 activation (P<0.05 or P<0.01). Conclusion: Lentivirusmediated bcr/abl RNAi can result in long time silencing of bcr/abl gene, inhibit malignant proliferation and induce differentiation and apoptosis in K562 cells.
2009, 16(2):151-155.
DOI: 10.3872/j.issn.1007-385X.2009.2.010
Abstract:
Objective: To observe the sensitivities of breast cancer cells and its implanted tumors to chemotherapeutic drug epirubicin after downregulation of HER2 expression by RNA interference (RNAi). Methods: HER2siRNApU6 vector containing HER2 RNAi was constructed and was used to transfect HER2 positive breast cancer cell line SKBR3. The expression of HER2 mRNA and protein were analyzed by RTPCR and Western blotting, respectively. Transfected SKBR3 cells were treated with different concentrations of epirubicin; the growth of SKBR3 cells and IC50 of epirubicin were observed by MTT. SKBR3 cells were injected into nude mouse to establish breast cancer model; the sensitivity of mouse model to epirubicin was observed after HER2shRNApU6 treatment. Results: Expression of HER2 mRNA and HER2 protein were downregulated in SKBR3 cells after transfection with HER2shRNApU6. Furthermore, the proliferation of SKBR3 cells transfected with HER2shRNApU6 was significantly decreased compared with mock tansfected group（P<0.05）. Chemosensitivity of SKBR3 cells to epirubicin was enhanced after treatment with HER2shRNApU6, with the IC50 values of HER2shRNApU6, mock, and negative tansfected group being 0.25, 3.46 and 3.69 μg/μl, respectively. The tumor weight was significantly lower in HER2shRNApU6 group than those in the mock and negative control group \[（2.17±0.58）vs (3.13±0.38),（3.21±0.89）g\]. Conclusion: HER2 RNAi obviously inhibits the expression of HER2 mRNA and HER2 protein in SKBR3 breast cancer cells, thus increases their sensitivities to epirubicin in vitro and in vivo.
Objective: To observe the sensitivities of breast cancer cells and its implanted tumors to chemotherapeutic drug epirubicin after downregulation of HER2 expression by RNA interference (RNAi). Methods: HER2siRNApU6 vector containing HER2 RNAi was constructed and was used to transfect HER2 positive breast cancer cell line SKBR3. The expression of HER2 mRNA and protein were analyzed by RTPCR and Western blotting, respectively. Transfected SKBR3 cells were treated with different concentrations of epirubicin; the growth of SKBR3 cells and IC50 of epirubicin were observed by MTT. SKBR3 cells were injected into nude mouse to establish breast cancer model; the sensitivity of mouse model to epirubicin was observed after HER2shRNApU6 treatment. Results: Expression of HER2 mRNA and HER2 protein were downregulated in SKBR3 cells after transfection with HER2shRNApU6. Furthermore, the proliferation of SKBR3 cells transfected with HER2shRNApU6 was significantly decreased compared with mock tansfected group（P<0.05）. Chemosensitivity of SKBR3 cells to epirubicin was enhanced after treatment with HER2shRNApU6, with the IC50 values of HER2shRNApU6, mock, and negative tansfected group being 0.25, 3.46 and 3.69 μg/μl, respectively. The tumor weight was significantly lower in HER2shRNApU6 group than those in the mock and negative control group \[（2.17±0.58）vs (3.13±0.38),（3.21±0.89）g\]. Conclusion: HER2 RNAi obviously inhibits the expression of HER2 mRNA and HER2 protein in SKBR3 breast cancer cells, thus increases their sensitivities to epirubicin in vitro and in vivo.
2009, 16(2):156-160.
DOI: 10.3872/j.issn.1007-385X.2009.2.011
Abstract:
Objective: To explore the inhibitory effect of polyamidoamine dendrimer (PAMAMD) STAT3 antisense oligodeoxynucleotide complex (PAMAMasODN) against the proliferation of hepatocarcinoma cell line SMMC7721. Methods: The seventh generation of PAMAM was mixed with STAT3 asODN in room temperature to prepare a PAMAMasODN complex. The morphology of the complex was examined by transmission electron microscope (TEM); the mean diameter of the complex was evaluated using laser particle size analyzer; and its inhibitory effect on the proliferation of SMMC7721 cells was measured by MTT. Cell cycle, apoptosis, and STAT3 mRNA expression in SMMC7721 cells treated with PAMAMasODN were examined by FACS/TUNEL and realtime PCR, respectively. Results: The PAMAMasODN complex was successfully prepared, with even distribution, good fluidity, and a mean diameter of 85.45 nm. MTT assay showed that PAMAMasODN inhibited the proliferation of SMMC7721 cells in a time and concentrationdependent manner, with the highest inhibitory rate being (68.9±3.0)%. FCM and TUNEL assay showed that PAMAMasODN induced apoptosis of SMMC7721 cells and blocked cell cycle at G2/M phase. Realtime PCR showed that the expression of STAT3 mRNA in SMMC7721 cells was significantly decreased after treatment with PAMAMasODN.Conclusion:PAMAM dendrimer can efficiently deliver STAT3 asODN into SMMC7721 cells, significantly inhibit SMMC7721 cells proliferation, and induce apoptosis of hepatocarcinoma SMMC7721 cells.
Objective: To explore the inhibitory effect of polyamidoamine dendrimer (PAMAMD) STAT3 antisense oligodeoxynucleotide complex (PAMAMasODN) against the proliferation of hepatocarcinoma cell line SMMC7721. Methods: The seventh generation of PAMAM was mixed with STAT3 asODN in room temperature to prepare a PAMAMasODN complex. The morphology of the complex was examined by transmission electron microscope (TEM); the mean diameter of the complex was evaluated using laser particle size analyzer; and its inhibitory effect on the proliferation of SMMC7721 cells was measured by MTT. Cell cycle, apoptosis, and STAT3 mRNA expression in SMMC7721 cells treated with PAMAMasODN were examined by FACS/TUNEL and realtime PCR, respectively. Results: The PAMAMasODN complex was successfully prepared, with even distribution, good fluidity, and a mean diameter of 85.45 nm. MTT assay showed that PAMAMasODN inhibited the proliferation of SMMC7721 cells in a time and concentrationdependent manner, with the highest inhibitory rate being (68.9±3.0)%. FCM and TUNEL assay showed that PAMAMasODN induced apoptosis of SMMC7721 cells and blocked cell cycle at G2/M phase. Realtime PCR showed that the expression of STAT3 mRNA in SMMC7721 cells was significantly decreased after treatment with PAMAMasODN.Conclusion:PAMAM dendrimer can efficiently deliver STAT3 asODN into SMMC7721 cells, significantly inhibit SMMC7721 cells proliferation, and induce apoptosis of hepatocarcinoma SMMC7721 cells.
LIU Liming
,
JIN Ningyi
,
LI Xiao
,
TIAN Mingyao
,
YANG Encheng
,
LIU Yan
,
ZHAO Cuiqing
,
ZHANG Jinshuang
,
QIAN Aidong
2009, 16(2):161-164.
DOI: 10.3872/j.issn.1007-385X.2009.2.012
Abstract:
Objective:To investigate the antitumor effects of Apoptin against human hepatocellular carcinoma HepG2 cells and implanted H22 tumors in C57BL/6 mice. Methods: C57BL/6 mice were used to establish H22bearing models. pVAX1Apoptin was intratumorally injected and the inhibition of tumor growth was observed. Recombinant plasmid pVAX1Apoptin was transfected into HepG2 cells by Lipofectamine. The expression of Apoptin protein in HepG2 cells was examined by Western blotting. Antitumor effect of pVAX1Apoptin on HepG2 cells was measured by MTT assay, and the apoptosis of HepG2 cells after transfected with pVAX1Apoptin was observed by AO/EB staining.Results:Apoptin gene was effectively expressed in HepG2 cells after transfection with pVAX1Apoptin. pVAX1Apoptin induced apoptosis of HepG2 cells and inhibited the growth of HepG2 cells, with the suppression rate being 69.28% at 48 h after transfection. Intratumoral injection of pVAX1Apoptin significantly inhibited tumor growth, with tumor inhibition rate being 46.71% and mice survival rate being 40% at 39 d after injection. Conclusion: pVAX1Apoptin can inhibit the proliferation of HepG2 cells, and intratumoral injection of pVAX1Apoptin can greatly inhibit tumor growth in vivo.
Objective:To investigate the antitumor effects of Apoptin against human hepatocellular carcinoma HepG2 cells and implanted H22 tumors in C57BL/6 mice. Methods: C57BL/6 mice were used to establish H22bearing models. pVAX1Apoptin was intratumorally injected and the inhibition of tumor growth was observed. Recombinant plasmid pVAX1Apoptin was transfected into HepG2 cells by Lipofectamine. The expression of Apoptin protein in HepG2 cells was examined by Western blotting. Antitumor effect of pVAX1Apoptin on HepG2 cells was measured by MTT assay, and the apoptosis of HepG2 cells after transfected with pVAX1Apoptin was observed by AO/EB staining.Results:Apoptin gene was effectively expressed in HepG2 cells after transfection with pVAX1Apoptin. pVAX1Apoptin induced apoptosis of HepG2 cells and inhibited the growth of HepG2 cells, with the suppression rate being 69.28% at 48 h after transfection. Intratumoral injection of pVAX1Apoptin significantly inhibited tumor growth, with tumor inhibition rate being 46.71% and mice survival rate being 40% at 39 d after injection. Conclusion: pVAX1Apoptin can inhibit the proliferation of HepG2 cells, and intratumoral injection of pVAX1Apoptin can greatly inhibit tumor growth in vivo.
2009, 16(2):165-169.
DOI: 10.3872/j.issn.1007-385X.2009.2.013
Abstract:
Objective:To investigate the inhibitory effects of antistathmin monoclonal antibody combined paclitaxel on the proliferation of human hepatocellular carcinoma cell lines HepG2.Methods: HepG2 cells were treated with antistathmin monoclonal antibody, paclitaxel or their combinations; untreated cells served as control. 24, 48, 72, and 96 h after exposure, the numbers and morphology of cells in different groups were observed under inverted microscope. Proliferation and apoptosis of HepG2 cells in different groups were studied by MTT and Annexin V/PI staining, respectively. Results: The numbers of HepG2 cells were decreased in all treated groups; and the cells in these groups showed morphological changes: some with round shape, some with nuclear chromatin condensation; but HepG2 cells in the control group did not show abnormal morphology. Antistathmin monoclonal antibody, paclitaxe alone or in combinations dosedependently inhibited the proliferation of HepG2 cells, and the inhibitory rate in the combination group was significantly higher than those in the two single drug groups (P<0.05), suggesting a synergistic effect between the two drugs (P<0.05). Antistathmin monoclonal antibody, paclitaxe alone or in combinations induced apoptosis of HepG2 cells, and the apoptosis in the combination group was higher than those in the two single drug groups (P<0.05). Conclusion: Antistathmin monoclonal antibody, paclitaxe alone or in combination can inhibit proliferation and induce apoptosis of HepG2 cells, and a synergistic effect is observed between antistathmin monoclonal antibody and paclitaxe.
Objective:To investigate the inhibitory effects of antistathmin monoclonal antibody combined paclitaxel on the proliferation of human hepatocellular carcinoma cell lines HepG2.Methods: HepG2 cells were treated with antistathmin monoclonal antibody, paclitaxel or their combinations; untreated cells served as control. 24, 48, 72, and 96 h after exposure, the numbers and morphology of cells in different groups were observed under inverted microscope. Proliferation and apoptosis of HepG2 cells in different groups were studied by MTT and Annexin V/PI staining, respectively. Results: The numbers of HepG2 cells were decreased in all treated groups; and the cells in these groups showed morphological changes: some with round shape, some with nuclear chromatin condensation; but HepG2 cells in the control group did not show abnormal morphology. Antistathmin monoclonal antibody, paclitaxe alone or in combinations dosedependently inhibited the proliferation of HepG2 cells, and the inhibitory rate in the combination group was significantly higher than those in the two single drug groups (P<0.05), suggesting a synergistic effect between the two drugs (P<0.05). Antistathmin monoclonal antibody, paclitaxe alone or in combinations induced apoptosis of HepG2 cells, and the apoptosis in the combination group was higher than those in the two single drug groups (P<0.05). Conclusion: Antistathmin monoclonal antibody, paclitaxe alone or in combination can inhibit proliferation and induce apoptosis of HepG2 cells, and a synergistic effect is observed between antistathmin monoclonal antibody and paclitaxe.
2009, 16(2):170-174.
DOI: 10.3872/j.issn.1007-385X.2009.2.014
Abstract:
Objective:To investigate the expression of indoleamine 2,3dioxygenase (IDO) in dendritic cells derived from chronic myeloid leukemia（CMLDCs）and to study the influence of IDO inhibition on the function of CMLDCs. Methods: The expression of IDO mRNA in dendritic cells derived from 17 patients with chronic myeloid leukemia was detected by RTPCR. The phenotypes of CMLDCs were analyzed by flow cytometry. The immature CMLDCs (imDCs) and the mature CMLDCs (mDCs) were used as stimulating cells and autologous Tlymphocytes were used as reactive cells for a mixed lymphocyte reaction system. IL12 concentration was detected by ELISA kit; mix lymphocyte reaction was analyzed by MTT assay. Results: It was demonstrated that DCs derived from bone marrow mononuclear cells of CML displayed a typical morphology of DCs. Expressions of costimulatory molecules on DCs, such as CD80, CD86, CD83 and HLADR, except for CD1a, were obviously higher after maturation (P<0.05) and were not influenced by 1Methyltroptophan(1MT, an inhibitor of IDO). Inhibition IDO activity in mature and immature DCs by 1MT significantly enhanced their abilities to activate T cells proliferation and to produce IL12 (P<0.05,P<0.01).Conclusion: Inhibition of IDO activity in CMLDCs can increase their abilities to produce IL12 and activate autologous T cells. Negative regulation of DCs by IDO paves a way for DCbased leukemia immunotherapy.
Objective:To investigate the expression of indoleamine 2,3dioxygenase (IDO) in dendritic cells derived from chronic myeloid leukemia（CMLDCs）and to study the influence of IDO inhibition on the function of CMLDCs. Methods: The expression of IDO mRNA in dendritic cells derived from 17 patients with chronic myeloid leukemia was detected by RTPCR. The phenotypes of CMLDCs were analyzed by flow cytometry. The immature CMLDCs (imDCs) and the mature CMLDCs (mDCs) were used as stimulating cells and autologous Tlymphocytes were used as reactive cells for a mixed lymphocyte reaction system. IL12 concentration was detected by ELISA kit; mix lymphocyte reaction was analyzed by MTT assay. Results: It was demonstrated that DCs derived from bone marrow mononuclear cells of CML displayed a typical morphology of DCs. Expressions of costimulatory molecules on DCs, such as CD80, CD86, CD83 and HLADR, except for CD1a, were obviously higher after maturation (P<0.05) and were not influenced by 1Methyltroptophan(1MT, an inhibitor of IDO). Inhibition IDO activity in mature and immature DCs by 1MT significantly enhanced their abilities to activate T cells proliferation and to produce IL12 (P<0.05,P<0.01).Conclusion: Inhibition of IDO activity in CMLDCs can increase their abilities to produce IL12 and activate autologous T cells. Negative regulation of DCs by IDO paves a way for DCbased leukemia immunotherapy.
2009, 16(2):175-180.
DOI: 10.3872/j.issn.1007-385X.2009.2.015
Abstract:
Objective: To evaluate the efficacy and safety of capecitabine combined with irinotecan or rhendostatin（endostar）for the treatment of oxaliplatinresistant patients with advanced colorectal cancer. Methods: Fortyfive patients with oxaliplatinresistant advanced colorectal cancer were included in this study. Twentyfive of them were treated with capecitabine combined with irinotecan and 20 of them were treated with capecitabine combined with endostar. Results:The followup period was 321 months. The response rate(RR) was 32.0% in irinotecan treated patients; the clinical benefit rate (CBR) was 72.0%; time of tumor progression (TTP) was 6.2 (95％CI: 3.125~8.905) months. The RR was 55.0% in endostar treated patients; the CBR was 90.0% and TTP was 10.6 (95％CI: 7.876~12.962) months. There was significant difference between the two groups concerning the above three parameters (P<0.05). The overall survival periods (OS) of the two groups were 15.2 (95％CI: 12.576~17.842) and 16.1 (95％CI:13.988~18.234) months, respectively, with no significant difference (P>0.05). The quality of life (QOL) was improved in 2 cases (80%), kept stable in 6 cases (24.0%), and decreased in 17 cases (68.0%) in irinotecan treated group; the numbers in the endostar treated group were 12 (60.0%), 6 (30.0%), and 2 (10.0%), respectively, with significant difference between the two groups(P<0.01). The incidences of drugrelated adverse events, neutropenia and diarrhea, were higher in the irinotecan group than in the endostar group (P<0.01). Conclusion: Capecitabine combined with irinotecan or endostar are alternative therapies for oxaliplatinresistant patients with advanced colorectal cancer, and capecitabine combined with endostar is more effective and safe.
Objective: To evaluate the efficacy and safety of capecitabine combined with irinotecan or rhendostatin（endostar）for the treatment of oxaliplatinresistant patients with advanced colorectal cancer. Methods: Fortyfive patients with oxaliplatinresistant advanced colorectal cancer were included in this study. Twentyfive of them were treated with capecitabine combined with irinotecan and 20 of them were treated with capecitabine combined with endostar. Results:The followup period was 321 months. The response rate(RR) was 32.0% in irinotecan treated patients; the clinical benefit rate (CBR) was 72.0%; time of tumor progression (TTP) was 6.2 (95％CI: 3.125~8.905) months. The RR was 55.0% in endostar treated patients; the CBR was 90.0% and TTP was 10.6 (95％CI: 7.876~12.962) months. There was significant difference between the two groups concerning the above three parameters (P<0.05). The overall survival periods (OS) of the two groups were 15.2 (95％CI: 12.576~17.842) and 16.1 (95％CI:13.988~18.234) months, respectively, with no significant difference (P>0.05). The quality of life (QOL) was improved in 2 cases (80%), kept stable in 6 cases (24.0%), and decreased in 17 cases (68.0%) in irinotecan treated group; the numbers in the endostar treated group were 12 (60.0%), 6 (30.0%), and 2 (10.0%), respectively, with significant difference between the two groups(P<0.01). The incidences of drugrelated adverse events, neutropenia and diarrhea, were higher in the irinotecan group than in the endostar group (P<0.01). Conclusion: Capecitabine combined with irinotecan or endostar are alternative therapies for oxaliplatinresistant patients with advanced colorectal cancer, and capecitabine combined with endostar is more effective and safe.
2009, 16(2):181-186.
DOI: 10.3872/j.issn.1007-385X.2009.2.016
Abstract:
Objective: To observe the expression of ATPase F1α in colorectal cancer (CRC) tissues and in LoVo cells, and to discuss its clinical significance. Methods: Expression of ATPase F1α protein in 44 CRC specimens and their adjacent normal tissues (August 2007 to December 2007, Changhai Hospital) and ATPase F1α mRNA in 8 colorectal cancer tissues and their adjacent normal tissues were examined by immunohistochemistry EnVision assay and RTPCR, respectively. Expression of ATPase F1α on the cell surface of LoVo cells was observed by immunofluorescence. The inhibitory effect of antiATPase F1α antibody on the proliferation of LoVo cells was evaluated by CCK8 assay. Results: Expression of ATPase F1α in the 44 CRCs were significantly higher than those in the adjacent normal tissues as detected by immunohistochemistry (P<0.01). Thirtyfive samples (79.5%) in 44 CRCs and 15 samples (34.1%) in their adjacent normal tissues showed moderate to high expression of ATPase F1α. Expression of ATPase F1α mRNA in 8 CRCs was significantly higher than those in their adjacent normal tissues (P<0.01). Positive rate of ATPase F1α mRNA in CRCs was significantly higher than that of CEA in the same patients before surgery (P<0.01). There was no correlation of ATPase F1α expression with the age, gender, tumor stage, lymphoid metastasis, remote metastasis, tumor location and tumor differentiation of colorectal cancer (P>0.05). Expression of ATPase F1α was observed on the cell surface of LoVo cells, and antiATPase F1α antibody significantly inhibited the proliferation of LoVo cells (P<0.01). Conclusion: ATPase F1α is highly expressed in colorectal cancer tissues and cell line. As a tumor association antigen, ATPase F1α may serve as a new target in colorectal cancer immunotherapy.
Objective: To observe the expression of ATPase F1α in colorectal cancer (CRC) tissues and in LoVo cells, and to discuss its clinical significance. Methods: Expression of ATPase F1α protein in 44 CRC specimens and their adjacent normal tissues (August 2007 to December 2007, Changhai Hospital) and ATPase F1α mRNA in 8 colorectal cancer tissues and their adjacent normal tissues were examined by immunohistochemistry EnVision assay and RTPCR, respectively. Expression of ATPase F1α on the cell surface of LoVo cells was observed by immunofluorescence. The inhibitory effect of antiATPase F1α antibody on the proliferation of LoVo cells was evaluated by CCK8 assay. Results: Expression of ATPase F1α in the 44 CRCs were significantly higher than those in the adjacent normal tissues as detected by immunohistochemistry (P<0.01). Thirtyfive samples (79.5%) in 44 CRCs and 15 samples (34.1%) in their adjacent normal tissues showed moderate to high expression of ATPase F1α. Expression of ATPase F1α mRNA in 8 CRCs was significantly higher than those in their adjacent normal tissues (P<0.01). Positive rate of ATPase F1α mRNA in CRCs was significantly higher than that of CEA in the same patients before surgery (P<0.01). There was no correlation of ATPase F1α expression with the age, gender, tumor stage, lymphoid metastasis, remote metastasis, tumor location and tumor differentiation of colorectal cancer (P>0.05). Expression of ATPase F1α was observed on the cell surface of LoVo cells, and antiATPase F1α antibody significantly inhibited the proliferation of LoVo cells (P<0.01). Conclusion: ATPase F1α is highly expressed in colorectal cancer tissues and cell line. As a tumor association antigen, ATPase F1α may serve as a new target in colorectal cancer immunotherapy.
17 Expression of Survivin gene in gastric carcinoma and its relationship with Bcl-2 and Bax expression
2009, 16(2):187-190.
DOI: 10.3872/j.issn.1007-385X.2009.2.017
Abstract:
Objective:To investigate the expression of Survivin gene in gastric carcinoma and its relationship with Bcl2 and Bax expression, as well as their roles in the development and progression of gastric carcinoma. Methods: Fiftyfour paraffin blocks were obtained from patients with gastric carcinoma receiving surgery. The diagnosis of these patients was confirmed by pathology in our Hospital from 20052007; no patients received chemoradiation therapy before surgery. Fifteen benign gastricmucosa tissues were used as control. SP immunohistochemistry was used to detect Survivin, Bcl2 and Bax expression in gastric carcinoma tissues, and Survivin in 15 benign gastric mucosa tissues. Results: Survivin was positive in 39 of the 54 tissues, with a positive rate of 72.2%. Survivin expression was not detected in the 15 benign gastricmucosa tissues. Expression of Survivin in gastric carcinoma tissues was correlated with the invasion, lymph nodes metastasis and TNM stage of gastric carcinoma, but not with patients′ sex and age, tumor size, remote metastasis or cell histological stage. Positive rate of Survivin expression in Bcl2 positive gastric carcinoma tissues was 81.8% (27/33) and the rate was 63.6% (14/22) in Bcl2 negative gastric carcinoma tissues. Survivin expression in gastric carcinoma tissues was correlated with the expression of Bcl2 but not with Bax (P<0.05). Conclusion: Expression of Survivin in gastric carcinoma tissues is correlated with the invasion, lymph node metastasis and TNM stage of gastric carcinoma. Survivin expression in gastric carcinoma tissues is positively correlated with the expression of Bcl2 but not with Bax (P<005).
Objective:To investigate the expression of Survivin gene in gastric carcinoma and its relationship with Bcl2 and Bax expression, as well as their roles in the development and progression of gastric carcinoma. Methods: Fiftyfour paraffin blocks were obtained from patients with gastric carcinoma receiving surgery. The diagnosis of these patients was confirmed by pathology in our Hospital from 20052007; no patients received chemoradiation therapy before surgery. Fifteen benign gastricmucosa tissues were used as control. SP immunohistochemistry was used to detect Survivin, Bcl2 and Bax expression in gastric carcinoma tissues, and Survivin in 15 benign gastric mucosa tissues. Results: Survivin was positive in 39 of the 54 tissues, with a positive rate of 72.2%. Survivin expression was not detected in the 15 benign gastricmucosa tissues. Expression of Survivin in gastric carcinoma tissues was correlated with the invasion, lymph nodes metastasis and TNM stage of gastric carcinoma, but not with patients′ sex and age, tumor size, remote metastasis or cell histological stage. Positive rate of Survivin expression in Bcl2 positive gastric carcinoma tissues was 81.8% (27/33) and the rate was 63.6% (14/22) in Bcl2 negative gastric carcinoma tissues. Survivin expression in gastric carcinoma tissues was correlated with the expression of Bcl2 but not with Bax (P<0.05). Conclusion: Expression of Survivin in gastric carcinoma tissues is correlated with the invasion, lymph node metastasis and TNM stage of gastric carcinoma. Survivin expression in gastric carcinoma tissues is positively correlated with the expression of Bcl2 but not with Bax (P<005).
2009, 16(2):191-194.
DOI: 10.3872/j.issn.1007-385X.2009.2.018
Abstract:
新型辅助性T细胞Th17在分化和功能特征上较传统的辅助性T细胞Th1和Th2存在显著的不同。分泌效应分子IL17A/F和IL22的Th17细胞与机体抗胞外菌感染和自身免疫疾病免疫病理的关系密切。Th17细胞的分化发育很大程度上依赖于局部微环境的影响，其中活化DC或其他细胞提供的特定细胞因子诱导微环境尤为重要。小鼠系统中，TGFβ和IL6的共同刺激是Th17细胞分化的始动因素，自分泌细胞因子IL21参与其分化的正反馈调节，而IL23在维持其后续的细胞扩增和存活中有重要作用，转录因子ROR γt、ROR α、STAT3和IRF4参与其中转录水平的调节。此外，Th17细胞的分化发育还受到机体严密的调控。人Th17细胞分化的相关研究证实其中的机制和小鼠存在相似性。深入研究Th17细胞的分化发育将有助于认识Th17细胞在抗感染免疫和自身免疫疾病中的免疫病理机制，也有利于相应治疗靶点的选择。
新型辅助性T细胞Th17在分化和功能特征上较传统的辅助性T细胞Th1和Th2存在显著的不同。分泌效应分子IL17A/F和IL22的Th17细胞与机体抗胞外菌感染和自身免疫疾病免疫病理的关系密切。Th17细胞的分化发育很大程度上依赖于局部微环境的影响，其中活化DC或其他细胞提供的特定细胞因子诱导微环境尤为重要。小鼠系统中，TGFβ和IL6的共同刺激是Th17细胞分化的始动因素，自分泌细胞因子IL21参与其分化的正反馈调节，而IL23在维持其后续的细胞扩增和存活中有重要作用，转录因子ROR γt、ROR α、STAT3和IRF4参与其中转录水平的调节。此外，Th17细胞的分化发育还受到机体严密的调控。人Th17细胞分化的相关研究证实其中的机制和小鼠存在相似性。深入研究Th17细胞的分化发育将有助于认识Th17细胞在抗感染免疫和自身免疫疾病中的免疫病理机制，也有利于相应治疗靶点的选择。
2009, 16(2):195-198.
DOI: 10.3872/j.issn.1007-385X.2009.2.019
Abstract:
异基因造血干细胞移植(allogeneic hematopoietic stem cell transplantation, alloHSCT)是目前治愈血液系统恶性疾病的有效手段。由于次要组织相容性抗原（minor histocompatibility antigens, mHags）本身的特性及其免疫学效应，在alloHSCT后的移植物抗宿主病（graft versus host disease, GVHD）和移植物抗白血病效应（graft versus leukemia effect, GVL）中都发挥了重要的作用。研究发现，一些mHags分布广泛，在各种细胞中均有表达,如HA3、HA4等；另外一些则限制性表达在造血细胞起源的细胞表面，如HA1、HA2等。目前正在研究利用mHags分布的差异应用于alloHSCT中，以达到加强GVL和减少GVHD的目的。
异基因造血干细胞移植(allogeneic hematopoietic stem cell transplantation, alloHSCT)是目前治愈血液系统恶性疾病的有效手段。由于次要组织相容性抗原（minor histocompatibility antigens, mHags）本身的特性及其免疫学效应，在alloHSCT后的移植物抗宿主病（graft versus host disease, GVHD）和移植物抗白血病效应（graft versus leukemia effect, GVL）中都发挥了重要的作用。研究发现，一些mHags分布广泛，在各种细胞中均有表达,如HA3、HA4等；另外一些则限制性表达在造血细胞起源的细胞表面，如HA1、HA2等。目前正在研究利用mHags分布的差异应用于alloHSCT中，以达到加强GVL和减少GVHD的目的。
2009, 16(2):199-202.
DOI: 10.3872/j.issn.1007-385X.2009.2.020
Abstract:
肝素酶（heparanase）是一种能降低细胞表面硫酸乙酰肝素蛋白聚糖（heparan sulfate proteoglycans，HSPG）的内切糖苷酶。它参与细胞表面细胞外基质（extracellular matrix，ECM）的降解和重塑，促进多配体蛋白聚糖1（syndecan1）的合成和释放；它在内涵体和溶酶体等细胞器中具有管家基因的功能，广泛表达于人类的肿瘤细胞中，在侵袭型的动物肿瘤细胞中也过度表达，因此它的过度表达被认为能促进肿瘤的浸润与转移。最近的研究显示，肝素酶能释放硫酸乙酰肝素依赖的生长因子并能产生大量有活性的硫酸乙酰肝素片段，破坏、降解细胞外基底膜屏障，促进肿瘤细胞扩散和转移，并加快血管生成，对肿瘤患者的预后造成了不利的影响。现在，越来越多的肝素酶抑制剂（PI88、肝素、肽类、硫酸昆布多糖、RNA干扰和苏拉明等）正在被开发出来，并显示出良好的前景。深入研究肝素酶在肿瘤转移和调节中的作用可能会带来肿瘤治疗的新手段。
肝素酶（heparanase）是一种能降低细胞表面硫酸乙酰肝素蛋白聚糖（heparan sulfate proteoglycans，HSPG）的内切糖苷酶。它参与细胞表面细胞外基质（extracellular matrix，ECM）的降解和重塑，促进多配体蛋白聚糖1（syndecan1）的合成和释放；它在内涵体和溶酶体等细胞器中具有管家基因的功能，广泛表达于人类的肿瘤细胞中，在侵袭型的动物肿瘤细胞中也过度表达，因此它的过度表达被认为能促进肿瘤的浸润与转移。最近的研究显示，肝素酶能释放硫酸乙酰肝素依赖的生长因子并能产生大量有活性的硫酸乙酰肝素片段，破坏、降解细胞外基底膜屏障，促进肿瘤细胞扩散和转移，并加快血管生成，对肿瘤患者的预后造成了不利的影响。现在，越来越多的肝素酶抑制剂（PI88、肝素、肽类、硫酸昆布多糖、RNA干扰和苏拉明等）正在被开发出来，并显示出良好的前景。深入研究肝素酶在肿瘤转移和调节中的作用可能会带来肿瘤治疗的新手段。
2009, 16(2):203-206.
DOI: 10.3872/j.issn.1007-385X.2009.2.021
Abstract:
α干扰素（interferonα，IFNα）是一种重要的治疗性细胞因子，通过调节细胞周期、抑制原癌基因、调控细胞黏附和血管生成等机制发挥抗肿瘤效应。然而患者对IFNα反应性的差异限制了其在临床的广泛使用。信号转导通路中一些关键因子如STAT1、STAT3、P48、SOCS1等的异常表达，与肿瘤的IFNα抗性机制相关；某些原癌基因Bcl2、cmyc、EVI1等的过度表达也参与IFNα抗性形成；此外DNA甲基转移酶、组蛋白修饰与肿瘤IFNα抗性的发生密切相关。因此，对这些预测IFNα临床疗效有潜在价值的分子标志物进行多因素分析，选择适合IFNα治疗的敏感病例，设计个体化的治疗方案，将成为今后肿瘤治疗的一个发展方向。
α干扰素（interferonα，IFNα）是一种重要的治疗性细胞因子，通过调节细胞周期、抑制原癌基因、调控细胞黏附和血管生成等机制发挥抗肿瘤效应。然而患者对IFNα反应性的差异限制了其在临床的广泛使用。信号转导通路中一些关键因子如STAT1、STAT3、P48、SOCS1等的异常表达，与肿瘤的IFNα抗性机制相关；某些原癌基因Bcl2、cmyc、EVI1等的过度表达也参与IFNα抗性形成；此外DNA甲基转移酶、组蛋白修饰与肿瘤IFNα抗性的发生密切相关。因此，对这些预测IFNα临床疗效有潜在价值的分子标志物进行多因素分析，选择适合IFNα治疗的敏感病例，设计个体化的治疗方案，将成为今后肿瘤治疗的一个发展方向。
2009, 16(2):207.
DOI: 10.3872/j.issn.1007-385X.2009.2.022
Abstract:
黑素瘤是起源于黑素细胞或其母细胞的恶性肿瘤，对放化疗不敏感，一旦远处转移，难以控制，预后极差。黑素瘤因存在很强的免疫源性，一直是生物治疗的研究热点。以往研究发现,LAK、TIL等过继性细胞以及黑素瘤抗原相关免疫瘤苗等都有一定的治疗效果。近年来随着免疫编辑学说的提出，新一代肿瘤免疫治疗方案，如抗CTLA4抗体、针对肿瘤抗原的IgG抗体、Toll样受体9激动剂、白喉毒素/IL2融合蛋白等开展了一系列临床试验，并取得了可喜的进步。
黑素瘤是起源于黑素细胞或其母细胞的恶性肿瘤，对放化疗不敏感，一旦远处转移，难以控制，预后极差。黑素瘤因存在很强的免疫源性，一直是生物治疗的研究热点。以往研究发现,LAK、TIL等过继性细胞以及黑素瘤抗原相关免疫瘤苗等都有一定的治疗效果。近年来随着免疫编辑学说的提出，新一代肿瘤免疫治疗方案，如抗CTLA4抗体、针对肿瘤抗原的IgG抗体、Toll样受体9激动剂、白喉毒素/IL2融合蛋白等开展了一系列临床试验，并取得了可喜的进步。
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Supported by:Beijing E-Tiller Technology Development Co., Ltd.