1. 携带MDR1基因的逆转录病毒载体的安全性研究 2. 大剂量阿霉素化疗联合全反式维甲酸对肝癌小鼠的影响实验研究
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摘要
题目1携带MDR1基因的逆转录病毒载体的安全性研究
目的:探讨携带MDR1基因的逆转录病毒载体在转染入小鼠骨髓单个核细胞(BM-MNCs)前后,转染MDR1基因的BM-MNCs回输或种植在小鼠体内后有无自我复制能力的逆转录病毒产生或致瘤性,为进一步临床应用提供安全性实验依据。
方法: (1)收集产病毒包装细胞PA317-HaMDR1/A的细胞上清液,浓缩后用RT-PCR法检测MDR1和env基因的表达情况;(2)浓缩病毒上清液转染小鼠BM-MNCs后,RT-PCR法检测BM-MNCs中MDR1和env基因的表达情况;(3)20只BALB/c小鼠(经60Co-γ放疗预处理)随机分为两组,每组10只:实验组回输转染MDR1基因的BM-MNCs,对照组回输等量生理盐水,监测外周血白细胞计数变化情况;每月检测骨髓和外周血单个核细胞中MDR1和env基因的表达情况;(4)20只裸鼠随机分为两组,每组10只,实验组在裸鼠腋部皮下注射转染MDR1基因的BM-MNCs,对照组注射等量的生理盐水,观察注射部位有无肿块生成;监测外周血白细胞计数;RT-PCR法检测腋部皮下结缔组织、骨髓和外周血单个核细胞中MDR1和env基因的表达情况;HE染色、电镜检测裸鼠腋部皮下结缔组织结构有无改变。
结果: (1)RT-PCR法在PA317-HaMDR1/A细胞浓缩的病毒上清液中检测到MDR1基因表达的特异性条带,未检测到env基因的表达条带;(2)RT-PCR法在转染的小鼠BM-MNCs中检测到MDR1基因的表达,未检测到env基因的表达;(3)实验组BALB/c小鼠外周血白细胞计数下降较对照组缓慢,在第7天白细胞计数开始回升,在12天后两组差异有显著性意义(P<0.05);RT-PCR法在实验组小鼠骨髓和外周血单个核细胞中均检测到MDR1基因的表达,未检测到env基因的表达;(4)两组裸鼠外周血白细胞计数未见明显异常,差异无统计学意义(P<0.05);RT-PCR法在裸鼠腋部皮下结缔组织、骨髓和外周血单个核细胞中均未检测到MDR1和env基因的表达;透射电镜在裸鼠腋部皮下结缔组织中未检测到逆转录病毒颗粒,结缔组织结构未见明显异常;病理HE染色见结缔组织形态结构无异常,未见病理性核分裂像。
结论:病毒上清液及转染MDR1基因的BM-MNCs回输入体内前后均未检测到有复制能力的逆转录病毒产生;转染MDR1基因的BM-MNCs无致瘤性。
题目2大剂量阿霉素化疗联合全反式维甲酸对肝癌小鼠的影响实验研究
目的:维甲酸类化合物对多种恶性肿瘤具有诱导分化、抑制增殖、诱导凋亡等作用,是一类疗效确切的抗肿瘤药物。本研究将MDR1基因转染入骨髓单个核细胞(BM-MNCs)后回输入肝癌小鼠体内,观察阿阿霉素大剂量化疗联合应用全反式维甲酸(ATRA)后对肝癌组织生长、外周血白细胞计数、肝癌组织P-gp及增殖凋亡因子表达的影响,和外源性MDR1基因在体内的分布情况。
方法:收集产病毒包装细胞PA317-HaMDR1/A的病毒上清液并浓缩,采用病毒上清转染法转染BALB/c小鼠BM-MNCs,RT-PCR法检测小鼠BM-MNCs中MDR1基因的表达情况;将40只肝癌荷瘤BALB/c小鼠随机分为5组:A正常对照组(不照射不回输),B空白对照组(照射并回输等量生理盐水),C阴性对照组(照射并回输未转染MDR1基因的BM-MNCs),D1联合用药组(照射并回输转染MDR1基因的BM-MNCs),D2阿霉素组(照射并回输转染MDR1基因的BM-MNCs),每组8只。B、C、D1组进行大剂量阿霉素化疗同时联合应用ATRA,D2组单用阿霉素大剂量化疗,每三天监测各组肝癌组织的大小、外周血白细胞计数变化情况,每周用FISH和RT-PCR法检测各组小鼠外周血和BM-MNCs、肿瘤组织和重要脏器中MDR1基因的表达和分布情况,每周用免疫组织化学法检测各组肝癌组织P-gp和增殖凋亡因子的表达情况。
结果:(1)RT-PCR法证实外源性MDR1基因可有效地整合到小鼠BM-MNCs基因组中并进行转录表达;RT-PCR法在联合用药组D1和阿霉素组D2肝癌小鼠的外周血和BM-MNCs中均检测到MDR1基因的表达;(2)联合用药组D1组和阿霉素组D2组肝癌小鼠肿瘤组织的瘤重较对照组轻,化疗18天后差异有统计学意义(P<0.05);联合用药组D1组肝癌小鼠的肿瘤组织的瘤重总体较阿霉素组D2组的轻,但差异无统计学意义(P>0.05);(3)联合用药组D1组和阿霉素组D2组肝癌小鼠外周血白细胞计数高于两个对照组,化疗第2周后差异有统计学意义(P<0.01);联合用药组D1组和阿霉素组D2组肝癌小鼠外周血白细胞计数差异无统计学意义(P>0.05);(4)FISH和RT-PCR法在转染组小鼠小鼠重要脏器和肿瘤组织中均未检测到BM-MNCs和外源性MDR1基因定植和表达;(5)随着化疗剂量的增加,联合用药组D1组与阿霉素组D2组比较肝癌组织中P53、bax、PCNA和Ki-67表达的阳性率差异无统计学意义(P>0.05);化疗4周后联合用药组D1组肝癌组织中P-gp表达阳性率高于阿霉素组D2组,增高有统计学意义(P<0.05);化疗3周后联合用药组D1组肝癌组织Bcl-2的表达阳性率均高于阿霉素组D2组,差异有统计学意义(P<0.05)。
结论:(1)通过逆转录病毒载体可在体外将外源性MDR1基因转入小鼠BM-MNCs中,外源性MDR1基因转染回输后在小鼠骨髓和外周血单个核细胞有持续表达;(2)ATRA和阿霉素联用后不能增强或减弱阿霉素化疗的疗效,ATRA和阿霉素联用无协同或拮抗作用;(3)ATRA和阿霉素联用后与单用阿霉素比较,两者对骨髓造血功能的影响差异无统计学意义;(4)外源性MDR1基因转染移植后在小鼠重要脏器和肿瘤组织无表达;(5)全反式维甲酸能增加肝癌组织中P-gp表达的阳性率,增强了肝癌组织的耐药性,推测可能与上调抗凋亡因子Bcl-2的表达有关,而与增殖相关因子PCNA、Ki-67,凋亡相关因子P53、Bax的表达无直接关系。
THE SAFETY RESEARCH OF RETROVIRAL VECTOR WHICH CARRIED MDR1 GENE
Objective : To detecte wether there have producted replication -competent retroviral before or after the MDR1 gene which carried by retroviral vector transfect into BM-MNCs,and the BM-MNCs transfected with MDR1 gene import into mice, and observe wether the BM-MNCs transfected with MDR1 gene have oncogenicity,in order to offer a experimental support for retrovirol as a vector of gene therapy .
Method: (1)Collected and concentrated the supernatant fluid of PA317-MDR1/A cell, detect the expression of MDR1 gene and env gene by RT-PCR;(2)after transfected mouse bone marrow mononuclear cells (BM-MNCs) with concentrated supernatant of retroviral, detect the expression of MDR1 gene and env gene by RT-PCR;(3)BM-MNCs transfected with MDR1 gene input into BALB/c mouse through vena caudalis primed by 60CO-γ,the control group input tales dose physiological saline, each group have 10 mouses, monitor the white blood cells’(WBC) change, detect the expression of MDR1 and env gene in bone marrow and peripheral blood mononuclearcells;(4)BM-MNCs transfect with MDR1 gene growth in axil subcutaneous, the control group inject physiological saline, each group have 10 mouse,observe wether there have oncologesis; survey wether there have abnormality change by electron microscope; monitor the WBC change; detect the expression of MDR1 and eenv gene in subcutaneous connective tissue of axil of athymic mouse、bone marrow and peripheral blood mononuclear cells.
Result: (1)the MDR1 gene in concentrated supernanant fluid can be detected by RT-PCR assay, but no env gene detected;(2)MDR1 gene was detected in BM-MNCs transfected with MDR1gene by RT-PCR assay, but no env gene detected;(3)the white cell count of experimental group decreased slowly than the control group’s, and there are difference between two groups after 12 days;MDR1 gene was detected in bone marrow and peripheral blood mononuclear cells of the experimental group,there was no env gene expression;(4)The white cell count were no difference between the two group of nude mouse(P<0.05);there were no MDR1 or env gene express in axil subcutaneous connective tissue、bone marrow and peripheral blood mononuclear cells of nude mouse detected by RT-PCR assay; rrtroviral particle can not be detected in axil subcutaneous connective tissue. There was no retroviral particle in oxter connective tissue of node mouse detected by transmission electron microscope;the morphous and structure of the connective tissue have no abnormalities detected by HE staining,and there had no patho-caryocinesia in the connective tissue.
Conclusion : the env gene can not be detected before or after BM-MNCs which carried MDR1 gene import into mouse;the BM-MNCs that transfected MDR1 gene have no oncogenicity
EXPERIMENTAL RESEARCH ON THE OVER DOSE ADRIAMYCIN CHEM COMBINED WITH ALL-TRAN- RATINOIC ACID TO MICE WITH LIVER CANCER
Objective:Retinoid compound have many effectiveness to kinds of malignant tumour ,such as induce differentiation、suppress proliferation、derivn apoptosis、et al,it is an antitumor drug. Our research used BM-MNCs transfected with MDR1 gene import into mouse with liver cancer,to observe the growth of liver cancer、the count of WBC in peripheral blood、the express of P-gp、proliferation and apoptosis factor in liver cancer,the distribute of MDR1 gene in vivo.
Method: Collected and condensed the virus supernatant liquid of incasing cell PA317-HaMDR1/A which product retroviral,using virus supernatant liquid to transfect the BM-MNCs of BALB/c mouse,detected the expression of MDR1 gene in BM-MNCs by RT-PCR; 40 BALB/c mousse with liver cancer divided into 5 groups randomly: A control group(nor exposure or import), B blank(exposure and impore physiological saline),C nagtive control(exposure and impore BM-MNCs non-transfected MDR1 gene)D1 transfected group(exposure and impore BM-MNCs transfected MDR1 gene), D2 transfected group(exposure and impore BM-MNCs transfected MDR1 gene),each group has 8 mouses. B、Cand D1 group treated with Over dose ADM chem Combined with ATRA , D2group treated with over dose ADM chem only, monitor the size of liver cancer、the count of peripheral white blood cells every three days, detect the expression and distribute of MDR1 gene in tumor and the important organs by RT-PCR and FISH,detect the expression of P-gp、proliferation and apoptosis factor in liver cancer every week by immunohistochemical method。
Result:(1) It’s confirm by RT-PCR that exogenous MDR1gene can integrate and express in BM-MNCs of mouse;the expression of MDR1 gene can be detected in peripheral blood and BM-MNCs of liver cancer mouse in both the drug combination D1 and the ADM group D2. (2) the weight of the tumor in the drug combination group D1 and the ADM group D2 lighter than the control groups 18 days after chem (P<0.05), the weight of liver cancer in mouse of D1 group lighter than the D2 group,but the difference has no statisticsal significance (P> 0.05).(3) White cell count in peripheral blood of the mouse in MDR1 transferring group D1 and D2 groups are more than the control groups, There was statistically significant between the two groups 2 weeks later after chemotherapy (P<0.01);but the count of perithal blood whith cell in the drug combination group D1 compared with the ADM D2 group’s have no significant difference (P>0.05);(4)there were no MDR1 gene expression in important organ and tumor detected by FISH and RT-PCR assay;(5) following the increase of the chem dose, the expression of P53、bax、PCNA and Ki-67 in liver cancer between D1 and D2 group have no difference(P>0.05); the expression of P-gp in liver cancer of the drug combination group D1 was higher than the ADM group D2 after chem two weeks(P<0.05); the expression of Bcl-2 in liver cancer of the drug combination D1 group was higher than the ADM group D2 on every week (P<0.05).
Conclusion: (1) The exogenous MDR1 gene can be transfected into BM-MNCs of mouse through retroviral vector, and it can express in BM and PB-MNCs of mouse after transfection;(2) ATRA combined with ADM can’t enhance or attenuate the curative effect of ADM chem.,usingADM combined with ATRA have no synergistic or rivalry effect;(3) ATRA have no influence on the haemopoiesis of bone marrow;(4) there were no extrogen MDR1 gene expressed in important organ and tumor;(5) ATRA can increase the expression of P-gp and reinforce the drug resistance in liver cancer, we suppose that maybe related with up-regulation the expression of Bcl-2, but may not related with the expression of PCNA、Ki-67、P53 and bax.
目的:探讨携带MDR1基因的逆转录病毒载体在转染入小鼠骨髓单个核细胞(BM-MNCs)前后,转染MDR1基因的BM-MNCs回输或种植在小鼠体内后有无自我复制能力的逆转录病毒产生或致瘤性,为进一步临床应用提供安全性实验依据。
方法: (1)收集产病毒包装细胞PA317-HaMDR1/A的细胞上清液,浓缩后用RT-PCR法检测MDR1和env基因的表达情况;(2)浓缩病毒上清液转染小鼠BM-MNCs后,RT-PCR法检测BM-MNCs中MDR1和env基因的表达情况;(3)20只BALB/c小鼠(经60Co-γ放疗预处理)随机分为两组,每组10只:实验组回输转染MDR1基因的BM-MNCs,对照组回输等量生理盐水,监测外周血白细胞计数变化情况;每月检测骨髓和外周血单个核细胞中MDR1和env基因的表达情况;(4)20只裸鼠随机分为两组,每组10只,实验组在裸鼠腋部皮下注射转染MDR1基因的BM-MNCs,对照组注射等量的生理盐水,观察注射部位有无肿块生成;监测外周血白细胞计数;RT-PCR法检测腋部皮下结缔组织、骨髓和外周血单个核细胞中MDR1和env基因的表达情况;HE染色、电镜检测裸鼠腋部皮下结缔组织结构有无改变。
结果: (1)RT-PCR法在PA317-HaMDR1/A细胞浓缩的病毒上清液中检测到MDR1基因表达的特异性条带,未检测到env基因的表达条带;(2)RT-PCR法在转染的小鼠BM-MNCs中检测到MDR1基因的表达,未检测到env基因的表达;(3)实验组BALB/c小鼠外周血白细胞计数下降较对照组缓慢,在第7天白细胞计数开始回升,在12天后两组差异有显著性意义(P<0.05);RT-PCR法在实验组小鼠骨髓和外周血单个核细胞中均检测到MDR1基因的表达,未检测到env基因的表达;(4)两组裸鼠外周血白细胞计数未见明显异常,差异无统计学意义(P<0.05);RT-PCR法在裸鼠腋部皮下结缔组织、骨髓和外周血单个核细胞中均未检测到MDR1和env基因的表达;透射电镜在裸鼠腋部皮下结缔组织中未检测到逆转录病毒颗粒,结缔组织结构未见明显异常;病理HE染色见结缔组织形态结构无异常,未见病理性核分裂像。
结论:病毒上清液及转染MDR1基因的BM-MNCs回输入体内前后均未检测到有复制能力的逆转录病毒产生;转染MDR1基因的BM-MNCs无致瘤性。
题目2大剂量阿霉素化疗联合全反式维甲酸对肝癌小鼠的影响实验研究
目的:维甲酸类化合物对多种恶性肿瘤具有诱导分化、抑制增殖、诱导凋亡等作用,是一类疗效确切的抗肿瘤药物。本研究将MDR1基因转染入骨髓单个核细胞(BM-MNCs)后回输入肝癌小鼠体内,观察阿阿霉素大剂量化疗联合应用全反式维甲酸(ATRA)后对肝癌组织生长、外周血白细胞计数、肝癌组织P-gp及增殖凋亡因子表达的影响,和外源性MDR1基因在体内的分布情况。
方法:收集产病毒包装细胞PA317-HaMDR1/A的病毒上清液并浓缩,采用病毒上清转染法转染BALB/c小鼠BM-MNCs,RT-PCR法检测小鼠BM-MNCs中MDR1基因的表达情况;将40只肝癌荷瘤BALB/c小鼠随机分为5组:A正常对照组(不照射不回输),B空白对照组(照射并回输等量生理盐水),C阴性对照组(照射并回输未转染MDR1基因的BM-MNCs),D1联合用药组(照射并回输转染MDR1基因的BM-MNCs),D2阿霉素组(照射并回输转染MDR1基因的BM-MNCs),每组8只。B、C、D1组进行大剂量阿霉素化疗同时联合应用ATRA,D2组单用阿霉素大剂量化疗,每三天监测各组肝癌组织的大小、外周血白细胞计数变化情况,每周用FISH和RT-PCR法检测各组小鼠外周血和BM-MNCs、肿瘤组织和重要脏器中MDR1基因的表达和分布情况,每周用免疫组织化学法检测各组肝癌组织P-gp和增殖凋亡因子的表达情况。
结果:(1)RT-PCR法证实外源性MDR1基因可有效地整合到小鼠BM-MNCs基因组中并进行转录表达;RT-PCR法在联合用药组D1和阿霉素组D2肝癌小鼠的外周血和BM-MNCs中均检测到MDR1基因的表达;(2)联合用药组D1组和阿霉素组D2组肝癌小鼠肿瘤组织的瘤重较对照组轻,化疗18天后差异有统计学意义(P<0.05);联合用药组D1组肝癌小鼠的肿瘤组织的瘤重总体较阿霉素组D2组的轻,但差异无统计学意义(P>0.05);(3)联合用药组D1组和阿霉素组D2组肝癌小鼠外周血白细胞计数高于两个对照组,化疗第2周后差异有统计学意义(P<0.01);联合用药组D1组和阿霉素组D2组肝癌小鼠外周血白细胞计数差异无统计学意义(P>0.05);(4)FISH和RT-PCR法在转染组小鼠小鼠重要脏器和肿瘤组织中均未检测到BM-MNCs和外源性MDR1基因定植和表达;(5)随着化疗剂量的增加,联合用药组D1组与阿霉素组D2组比较肝癌组织中P53、bax、PCNA和Ki-67表达的阳性率差异无统计学意义(P>0.05);化疗4周后联合用药组D1组肝癌组织中P-gp表达阳性率高于阿霉素组D2组,增高有统计学意义(P<0.05);化疗3周后联合用药组D1组肝癌组织Bcl-2的表达阳性率均高于阿霉素组D2组,差异有统计学意义(P<0.05)。
结论:(1)通过逆转录病毒载体可在体外将外源性MDR1基因转入小鼠BM-MNCs中,外源性MDR1基因转染回输后在小鼠骨髓和外周血单个核细胞有持续表达;(2)ATRA和阿霉素联用后不能增强或减弱阿霉素化疗的疗效,ATRA和阿霉素联用无协同或拮抗作用;(3)ATRA和阿霉素联用后与单用阿霉素比较,两者对骨髓造血功能的影响差异无统计学意义;(4)外源性MDR1基因转染移植后在小鼠重要脏器和肿瘤组织无表达;(5)全反式维甲酸能增加肝癌组织中P-gp表达的阳性率,增强了肝癌组织的耐药性,推测可能与上调抗凋亡因子Bcl-2的表达有关,而与增殖相关因子PCNA、Ki-67,凋亡相关因子P53、Bax的表达无直接关系。
THE SAFETY RESEARCH OF RETROVIRAL VECTOR WHICH CARRIED MDR1 GENE
Objective : To detecte wether there have producted replication -competent retroviral before or after the MDR1 gene which carried by retroviral vector transfect into BM-MNCs,and the BM-MNCs transfected with MDR1 gene import into mice, and observe wether the BM-MNCs transfected with MDR1 gene have oncogenicity,in order to offer a experimental support for retrovirol as a vector of gene therapy .
Method: (1)Collected and concentrated the supernatant fluid of PA317-MDR1/A cell, detect the expression of MDR1 gene and env gene by RT-PCR;(2)after transfected mouse bone marrow mononuclear cells (BM-MNCs) with concentrated supernatant of retroviral, detect the expression of MDR1 gene and env gene by RT-PCR;(3)BM-MNCs transfected with MDR1 gene input into BALB/c mouse through vena caudalis primed by 60CO-γ,the control group input tales dose physiological saline, each group have 10 mouses, monitor the white blood cells’(WBC) change, detect the expression of MDR1 and env gene in bone marrow and peripheral blood mononuclearcells;(4)BM-MNCs transfect with MDR1 gene growth in axil subcutaneous, the control group inject physiological saline, each group have 10 mouse,observe wether there have oncologesis; survey wether there have abnormality change by electron microscope; monitor the WBC change; detect the expression of MDR1 and eenv gene in subcutaneous connective tissue of axil of athymic mouse、bone marrow and peripheral blood mononuclear cells.
Result: (1)the MDR1 gene in concentrated supernanant fluid can be detected by RT-PCR assay, but no env gene detected;(2)MDR1 gene was detected in BM-MNCs transfected with MDR1gene by RT-PCR assay, but no env gene detected;(3)the white cell count of experimental group decreased slowly than the control group’s, and there are difference between two groups after 12 days;MDR1 gene was detected in bone marrow and peripheral blood mononuclear cells of the experimental group,there was no env gene expression;(4)The white cell count were no difference between the two group of nude mouse(P<0.05);there were no MDR1 or env gene express in axil subcutaneous connective tissue、bone marrow and peripheral blood mononuclear cells of nude mouse detected by RT-PCR assay; rrtroviral particle can not be detected in axil subcutaneous connective tissue. There was no retroviral particle in oxter connective tissue of node mouse detected by transmission electron microscope;the morphous and structure of the connective tissue have no abnormalities detected by HE staining,and there had no patho-caryocinesia in the connective tissue.
Conclusion : the env gene can not be detected before or after BM-MNCs which carried MDR1 gene import into mouse;the BM-MNCs that transfected MDR1 gene have no oncogenicity
EXPERIMENTAL RESEARCH ON THE OVER DOSE ADRIAMYCIN CHEM COMBINED WITH ALL-TRAN- RATINOIC ACID TO MICE WITH LIVER CANCER
Objective:Retinoid compound have many effectiveness to kinds of malignant tumour ,such as induce differentiation、suppress proliferation、derivn apoptosis、et al,it is an antitumor drug. Our research used BM-MNCs transfected with MDR1 gene import into mouse with liver cancer,to observe the growth of liver cancer、the count of WBC in peripheral blood、the express of P-gp、proliferation and apoptosis factor in liver cancer,the distribute of MDR1 gene in vivo.
Method: Collected and condensed the virus supernatant liquid of incasing cell PA317-HaMDR1/A which product retroviral,using virus supernatant liquid to transfect the BM-MNCs of BALB/c mouse,detected the expression of MDR1 gene in BM-MNCs by RT-PCR; 40 BALB/c mousse with liver cancer divided into 5 groups randomly: A control group(nor exposure or import), B blank(exposure and impore physiological saline),C nagtive control(exposure and impore BM-MNCs non-transfected MDR1 gene)D1 transfected group(exposure and impore BM-MNCs transfected MDR1 gene), D2 transfected group(exposure and impore BM-MNCs transfected MDR1 gene),each group has 8 mouses. B、Cand D1 group treated with Over dose ADM chem Combined with ATRA , D2group treated with over dose ADM chem only, monitor the size of liver cancer、the count of peripheral white blood cells every three days, detect the expression and distribute of MDR1 gene in tumor and the important organs by RT-PCR and FISH,detect the expression of P-gp、proliferation and apoptosis factor in liver cancer every week by immunohistochemical method。
Result:(1) It’s confirm by RT-PCR that exogenous MDR1gene can integrate and express in BM-MNCs of mouse;the expression of MDR1 gene can be detected in peripheral blood and BM-MNCs of liver cancer mouse in both the drug combination D1 and the ADM group D2. (2) the weight of the tumor in the drug combination group D1 and the ADM group D2 lighter than the control groups 18 days after chem (P<0.05), the weight of liver cancer in mouse of D1 group lighter than the D2 group,but the difference has no statisticsal significance (P> 0.05).(3) White cell count in peripheral blood of the mouse in MDR1 transferring group D1 and D2 groups are more than the control groups, There was statistically significant between the two groups 2 weeks later after chemotherapy (P<0.01);but the count of perithal blood whith cell in the drug combination group D1 compared with the ADM D2 group’s have no significant difference (P>0.05);(4)there were no MDR1 gene expression in important organ and tumor detected by FISH and RT-PCR assay;(5) following the increase of the chem dose, the expression of P53、bax、PCNA and Ki-67 in liver cancer between D1 and D2 group have no difference(P>0.05); the expression of P-gp in liver cancer of the drug combination group D1 was higher than the ADM group D2 after chem two weeks(P<0.05); the expression of Bcl-2 in liver cancer of the drug combination D1 group was higher than the ADM group D2 on every week (P<0.05).
Conclusion: (1) The exogenous MDR1 gene can be transfected into BM-MNCs of mouse through retroviral vector, and it can express in BM and PB-MNCs of mouse after transfection;(2) ATRA combined with ADM can’t enhance or attenuate the curative effect of ADM chem.,usingADM combined with ATRA have no synergistic or rivalry effect;(3) ATRA have no influence on the haemopoiesis of bone marrow;(4) there were no extrogen MDR1 gene expressed in important organ and tumor;(5) ATRA can increase the expression of P-gp and reinforce the drug resistance in liver cancer, we suppose that maybe related with up-regulation the expression of Bcl-2, but may not related with the expression of PCNA、Ki-67、P53 and bax.
引文
[1] Barquinero J, Eixarch H, Perez-Melgosa M.Retroviral vectors: new applications for an old tool[J].Gene Ther.2004,11(Suppl1):3~9
[2] Bastone P, Lochelt M. Kinetics and characteristics of replication-competent revertants derived from self-inactivating foamy virus vectors[J].Gene Ther. 2004,11(5):465~473
[3] Laufs S, Gentner B, Nagy KZ,et al.Retroviral vector integration occurs in preferred genomic targets of human bone marrow-repopulating cells[J].Blood. 2003, 101(6):2191~2198
[4] Wu XL, Li Y, Crise B,et al. Weak Palindromic Consensus Sequences Are a Common Feature Found at the Integtes of ration Target SiMany Retroviruses[J].J Virol. 2005,79(8): 5211~5214
[5]王珊,金先庆,杨纯正,等.MDR1基因转染K562细胞株及人骨髓干祖细胞的研究[J].中华小儿外科杂志.2000,21(4):238~41
[6] Choi K, Frommei TO, Stem RK, et al. Multidrug resistance after Retroviral transfer of the human MDR1 gene correlates with P-glycoprotein density in the plasma membrane and is not affected by cytotoxic selection[J]. Proc Natl Acad Sci USA.1991, 88:7386~90
[7] De Angioletti M,Rovira A,Sadelain M,et al.Frequency of missense mutation in coding region of a eukaryotic gene transferred by retroviral victors[J].J vjrol. 2002,76(4):1991~1994
[8] Yang S,Delgado R,King SR,et al.Generation of retroviral vector for clinical studies using transient transfection[J].Hum Gene Ther. 1999,10(1):l23~132
[9] Zhao J,Pincaewaki J,Gomez-Roman VR,et al.Improved protection of rhesus macaques against intrarectal simian immunodeficiency virus SIV(mac251) challenge by a replication-competent Ad5hr-SIVenv/rev and Ad5hr-SIVgag recombinant priming/gp120 boosting regimen[J].J Virol.2003,77(15):8354~8365
[10] RICHARD A. MORGAN, KENNETH CORNETTA,and W. FRENCH ANDERSON. Applications of the Polymerase Chain Reaction in Retroviral- -Mediated Gene Transfer and the Analysis of Gene-Marked Human TIL Cells[J]. HUMAN GENE THERAPY.1990,1:135~149
[11]孙敏,涂新明,丛姑,等.猴D型反转录病毒巢式PCR检测方法的建立[J].中国实验动物学报.2006,14(4):287~289
[12] Wu X,Li Y,Crise B,et al.Weak palindromic consensus sequences are a common feature found at the integration target sites of many retroviruses [J].J Virol. 2005,79(8):5211~5214
[13] Abonour R, Williams DA, Einhorn L. Efficient retrovirus-mediated transfer of the multidrug resistance 1 gene into autologous human long-term repopulating hematopoietic stem cells [J]. Nat Med. 2000,6(6):652~658
[14] Carpinteiro A, Peinert S, Ostertag W, et al. Genetic protection of repopulating hematopoietic cells with an improved MDR1-retrovirus allows administration of intensified chemotherapy following stem cell transplantation in mice [J].Int J Cancer.2002,98(5):785~792
[15] Modlich U, Kustikova OS, Schmidt M, et al. Leukemias following retroviral transfer of multidrug resistance 1 (MDR1) are driven by combinatorial insertional mutagenesis [J]. Blood. 2005,105(11):4235~4246
[16] Bunting K, Zhou Sh, Lu TH, et al. Enforced P-glycoprotein pump function in murine bone marrow cells results in expansion of side population stem cells in vitro and repopulating cells in vivo [J].Blood. 2000,96(3):902~909
[17] Buske C, Humphries RK. Homeobox genes in leukemogenesis [J]. Int J Hematol. 2000,71(4):301~308
[18] Schiedlmeier B, Klump H, Will E, et al. High level ectopic HOXB4 expression confers a profound in vivo competitive growth advantage to human cord blood CD34+ cell,but impairs lymphomyeloid differentiation [J]. Blood. 2003, 101(5): 1759~1768
[19]张秀亚,王珊,李圆,等.多药耐药基因转染对荷瘤小鼠骨髓造血细胞的保护作用[J].第三军医大学学报.2008,30(23):2164~2167
[20]李圆,王珊,王江波,等.多药耐药基因转染骨髓造血细胞移植小鼠的安全性研究[J].中华实验外科杂志.2008.25(5):658~660
[21] Hesdorffer C, Ayello J, Ward M, et al. Phase I trial of retroviral-mediated transfer of the human MDR1 gene as marrow chemoprotection in patients undergoing high-dose chemotherapy and autologous stem-cell transplantation[J].Clin Oncol. 1998,16(1):165~172
[22] O'Shaughnessy JA, Cowan KH, Nienhuis AW, et al. Retroviral mediated transfer of the human multidrug resistance gene (MDR-1) into hematopoietic stem cells during autologous transplantation after intensive chemotherapy for metastatic breast cancer[J]. Hum Gene Ther. 1994,5(7):891~911
[23] Cowan KH, Moscow JA, Huang H, et al. Paclitaxel chemotherapy after autologous stem-cell transplantation and engraftment of hematopoietic cells transduced with a retrovirus containing the multidrug resistance complementary DNA (MDR1) in metastatic breast cancer patients[J]. Clin Cancer Res. 1999, 5(7): 1619~1628
[24] Rahman Z, Kavanagh J, Champlin R. et al. Chemotherapy immediately following autologous stem-cell transplantation in patients with advanced breast cancer[J]. Clin Cancer Res. 1998,4(11):2717~2721
[25] Sugimoto Y, Tsukahara S, Sato S, et al. Drug-selected co-expression of P- glycoprotein and gp91 in vivo from an MDR1-bicistronic retrovirus vector Ha-MDR-IRES-gp91 [J]. Gene Med. 2003,5(5):366~376
[1]王珊,金先庆,杨纯正,等.MDR1基因转染K562细胞株及人骨髓干祖细胞的研究[J].中华小儿外科杂志.2000,21(4):238~41
[2] Wang Y, Jin XQ, Wang S, et a1.Therapeutic efficacy and bone marrow protection of the mdr1 gene and over-dose chemotherapy with doxorubicin for rabbits with hepatocarcinoma[J]. Hepatobiliary Pancreat Dis Int.2006,5(4): 545~51
[3] Chun-Bao Guo, Ying-Cun Li, Xian-Qing Jin. Chemoprotection effect of retroviral vector encoding multidrug resistance 1 gene to allow intensified chemotherapy in vivo[J]. Cancer Chemother Pharmacol (2006) 58: 40~49
[4]张秀亚,王珊,李圆,等.多药耐药基因转染对荷瘤小鼠骨髓造血细胞的保护作用[J].第三军医大学学报.2008,30(23):2164~2167
[5]安淑华,金先庆,解启莲,等。多药耐药基因转染的脐血细胞对白血病小鼠化疗的骨髓保护作用[J].中华血液学杂志.2005:26(2):82~85
[6] DIMBERG A ,BAHRAM F , KARLBERG I , et al . Retinoicacid-induced cell cycle arrest of human myeloid cell lines is associated wit h sequential down-regulation of c-Myc and cyclin Eand post t ranscription up-regulation of p27 (kipⅠ)[J].Blood.2002,99(6):2199~2206
[7]夏璐,鲍英,袁耀宗,等.全反式维甲酸诱导胰腺癌细胞凋亡机制研究[J].中华消化杂志.2004,24(7):391~394
[8] ABU J ,BATUWANGALA M , HERBERT K, et al . Retinoicacid and retinoid receptors :potential chemopreventive and therapeutic role in cervical cancer[J]. Lancet Oncol,2005,6(9):712~720
[9]方建林,郑传胜,冯敢生,等.全反式维甲酸对Walker-256肝癌细胞分化及凋亡的诱导作用[J].世界华人消化杂志.2008,16(26):2929~2934
[10]王珊,金先庆,杨纯正,等.探讨MDR1基因转染K562细胞的最适条件[J].中华小儿外科杂志.2002,23(6):564~565.
[11] Rentala S, Sagar Balla MM, Khurana S, et al.MDR1 gene expression enhances long-term engraftibility of cultured bone marrow cells[J]. BBRC.2005,335(3), 957~964
[12]曹晓伟,唱浩,季峻松,等.阿霉素对肝癌细胞核因子κB活化及细胞周期基因D1表达的影响[J].临床荟萃.2008,23(16):1153~1156
[13] Unter D, Sirioorn K, Jisnuson S, et al. A prospective study of the intake of vitamins C,E and A and the risk of breast cancer[J].N Engl J Med.1993, 329: 234~240
[14] Zhao J, Pinczewski J, Gomez-Roman VR, et al.Improved protection of rhesus macaques against intrarectal simian immunodeficiency virus SIV(mac251) chall- enge by a replication-competent Ad5hr-SIVenv/rev and Ad5hr-SIVgag recom- binant priming/gp120 boosting regimen[J].J Virol. 2003, 77(15):8354 ~8365
[15] Carpinteiro A, Peinert S, Ostertag W,et al. Genetic protection of repopulating hematopoirtic cells with an improved mdr1-retrovirus allows administration of intensified chemotherapy following stem cell transplantation in mice[J].Int J Cancer.2002,98(5):785~792
[16] Carllen DF, Baker E, Smmers RN, et al. Localization of the human multidrug resistance gene,mdr1,to 7q21.1[J]. Hum Genet. 1987,77:142~7
[17] Alexander J. Smith, Ardy van Helvoort, Gerrit van Meer,et al.MDR3 P-glycoprotein, a Phosphatidylcholine Translocase,Transports Several Cytotoxic Drugs and Directly Interacts withDrugs as Judged by Interference with Nucleotide Trapping[J].THE JOURNAL OF BIOLOGICAL CHEMISTRY. 2000,275(31):23530~23539
[18] Harunobu Tahara, Hiroyuki Kusuhara, Eiichi Fuse,et al. P-GLYCOPROTEIN PLAYS A MAJOR ROLE IN THE EFFLUX OF FEXOFENADINE IN THE SMALL INTESTINE AND BLOOD-BRAIN BARRIER, BUT ONLY A LIMITED ROLE IN ITS BILIARY EXCRETION [J].DRUG METABOLISM AND DISPOSITION.2005,33,(7):963~968
[19] Diana Kuhnke1,Gabriele Jedlitschky1,Markus Grube1,et al. MDR1- P-Glycoprotein (ABCB1) Mediates Transport of Alzheimer’s Amyloid-βPeptides-Implications for the Mechanisms of AβClearance at the Blood–Brain Barrier[J]. Brain Pathol.2007,17:347~353
[20] Geromin D,Bourge JF,SouolieA,et al.Glycoprotein 170 induces platelet -activating factor receptor membrane expression and confers tumor cell hypersensitivity to NK-dependent cell lysis[J].J Immunol.2004,172(6): 3604 ~ 3611
[21] Ling V. Multidrug resistance: molecular mechanisms and clinical relevance [J].Cancer chemother pharmacol,1997,40:S3~8
[22] Bennett S,Matthew L,Gayle E,et a1.Selective drug restsrant human osteosarcoma cell lines[J].J Clinical Orthopedics and Related Reseach. 2001, 383,259~267
[23] Kars MD, Iseri OD, Gunduz U, et al. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds [J]. Anticancer Res.2006,26(6):4559~4568
[24] Stromskaya TP, Rybalkina EY, Shtil AA,et al. Influence of exogenous RAR alpha gene on MDR1 expression and P-glycoprotein function in human and rodent cell lines[J]. Br J Cancer.1998,77(11):1718~25
[25]方燕,黄必军,梁启万.原发性肝癌p53基因高频缺失的双色荧光原位杂交证据[J].中国病理生理杂志.2003,19(3):306~309
[26] Zolota V,Batistatou A,Tsamandas A,et al.Immunohistochemical expression of TGF-betal,p2l WAF1,p53,Ki67,and angiogenesis in gastric carcinomas:a clinicopathologic study[J].Int J Gastrointest Cancer.2002,32(2-3):83-9
[27] Jason ON,Michae1 M,Pam S et a1.Promises and challenges of targeting Bcl-2 anti-apoptotic proteins for cancertherapy[J].Biochimica and Biophysica Acta. 2004,1705(1):43-51
[28] Konopleva M, Tsao T, Ruvolo P, et al. Novel triterpenoid CDDO-Me is a potent inducer of apoptosis and differentiation in acute myelogenous leukemia[J]. Blood. 2002 ,99(1):326~335
[29]郑建勇,李开宗,王为忠,等.阿霉素诱导人肝癌细胞凋亡机制的研究[J].中国普通外科杂志.2005,14(7):509~511
[30]李成荣,付劲荣.维甲酸导致多药耐药的机制初探[J].中华肿瘤杂志.2001,23(6):441~443
[31] MAGA G,HUBSCHER U.Proliferating cell nuclear antigen(PCNA):a dancer with many partners[J].J Cel Sci.003,1 16(15):3051~3060
[32] Potrunol R,Zizzo N,Zito AF,et al.Microvascular deneity and endothelial area correlate with Ki-67 proliferative rate in the canine non-Hodgkin’s lymphoma spontaneous mode[J]. Leuk Lymphom.2006,Jun 47(6):1138~1143
[1]Kars MD, Iseri OD, Gunduz U, et al. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds[J]. Anticancer Res.2006,26(6):4559~4568
[2]Benlloch M,Ortega A, Ferrer P.Acceleration of glutathione efflux and inhibition of gamma -glutamyltranspeptidase sensitize metastatic B16 melanoma cells to endothelium-induced cytotoxicity[J].Journal of Biological Chemistry. 2005, 280(8):6950~6959
[3]Brooks T,Minderman H, Kieran L, et a1.Taxane-based reversal agents modulate drug resistance mediated by P-glycoprotein,multidrng resistance protein,and breast cancer resistance protein[J].Cancer Ther.2003,2:1195
[4]Okada M,Nishio W ,Sakamoto T,et a1.Effect of tumor size on prognosis in patients with non-small cell lung cancer:the role of segmentectomy as a type of lesser resection [J].J Thorac Cardiovasc Surg.2005,129(1):87~93
[5]Greijer AE, de Jong MC, Scheffer GL, et al. Hypoxia-induced acidification causes mitoxantrone resistance not mediated by drug transporters in human breast cancer cells[J]. Cell Oncol.2005,27(1):43~49
[6]Ribosomal proteins S13 and L23 promote multidrug resistance in gastric cancer cells by suppressing drug-induced apoptosis[J]. Exp Cell Res. 2004,296(2):337~346
[7]Sampath J, Sun D, KiddⅥ,et a1.Mutant p53 cooperates with ET3 and selectively up - regulates human MDR1 not MRP1[J].J Biol Chem.2001,276(42):39359
[8] Thomas H,Coley HM.Overcoming multidrug resistance in cancer;an update on the clinical strategy of inhibiting P-glycopmtein[J].Cancer Control.2003,10(2):159~165
[9] Munoz Martinez F,Reyes CP,Perez Lomas AL,et a1.Insights into the molecular mechanism of action of celastraceae sesquiterpenes as specific,non-transported inhibitors of human P-glycoprotein[J].Biochim Biophys Acta.2006,1758(1):98~110
[10]Globisch C,Pajeva IK,Wiese M.Structure-activity relationships of a series of tariqu- idar analogs as multidrug resistance modulators[J].Bioorg Med Chem.2006,14(5): 1588~1598
[11]J WANG S,FOLKESA,CHUCKOWREEI,et a1.Studies On pyrrolopy-rimidines as selective inhibitors Of multidrug-resistance-associated protein in multidrug resistance[J].J Med Chem.2004,47:1329~1338
[12] BIENYAHIA B,HUGUAT S,DECIEYES X,et a1.Multidrug resistance- associaied protein MRP1 expression in human gliomas:Chemosensitization tovincristine and etoposide by indomethacin in human glioma cell lines overexpressing MRP1[J].J Neurooncol.2004,66:65~70
[13]Kitazono M,Okumura H,Ikeda R.Reversal of LRP-associated drug resistance in colon carcinoma SW-620 cells[J].Int J Cancer.2001,91(1):126~131
[14]Burg D,Filippov DV ,Hermanns R,et al.Peptidomimetic glutathi- olle analogues as novel gammaGT stable GST inhibitors[J].Bioorg Med Chem, 2002, 10(1):195~205.
[15]Mistry P,Stewart AJ,Dangerfield W,et al.In vitro and in vivo characterization of XR11576,a novel,orally active,dual inhibitor of to poisomeraseⅠandⅡ[J]. Anticancer Drugs,2002,13(1):15~28
[16]Chan JY,Chu AC,Fung KP.Inhibition of P-glycoprotein expression and reversal of drug resistance of human hepatoma HepG2 cells by multidrug resistance gene mdrl ontiscnse RNA[J].Life Sci.2000,67:2l17~2l24
[17]Irie A, Matsumoto K, Anderegg B, et al.Growth inhibition efficacy anadenovirus expressing dual therapeutic genes,wild type p53,and anti-erbB2 ribozyme,against human bladder cancer cells[J].Cancer genes Ther.2006,13(3):298~305
[18]Kobayashi H,Takemura Y,Miyachi H.Novel approaches to reversing anti-cancer drug resistance using gene-specific therapeutics[J].Hum Cell.2001,14:172~184
[19]Kostenko EV, Laktionov PP,VIassov VV ,et a1.Downregulation of PGY1/MDR1 mRNA level in human KB cells by antisense oligonucleotide conjugates : RNA accessibility in vitro and intracellular antisensc activity[J].Bioehim Biophys Acta. 2002,l576:l43~147
[20]Eferth T,Futseher BW, Osieka R.5-Aza dine modu1ates the response of senstive and mdtidrug-resistant K562 leukemic cells to cytostatic drugs[J].Blood Cells Mol Dis.2001,27(3):637
[21] Barzon L,Bonaguro R,Castagliuolo I,et a1.Gene therapy of thyreid cancer via retrovirally-driven combined expression of human interleukin-2 and herpes simplex virus thymidine kinase[J].Eur J Endocrinol.2003,148:73~80
[2] Bastone P, Lochelt M. Kinetics and characteristics of replication-competent revertants derived from self-inactivating foamy virus vectors[J].Gene Ther. 2004,11(5):465~473
[3] Laufs S, Gentner B, Nagy KZ,et al.Retroviral vector integration occurs in preferred genomic targets of human bone marrow-repopulating cells[J].Blood. 2003, 101(6):2191~2198
[4] Wu XL, Li Y, Crise B,et al. Weak Palindromic Consensus Sequences Are a Common Feature Found at the Integtes of ration Target SiMany Retroviruses[J].J Virol. 2005,79(8): 5211~5214
[5]王珊,金先庆,杨纯正,等.MDR1基因转染K562细胞株及人骨髓干祖细胞的研究[J].中华小儿外科杂志.2000,21(4):238~41
[6] Choi K, Frommei TO, Stem RK, et al. Multidrug resistance after Retroviral transfer of the human MDR1 gene correlates with P-glycoprotein density in the plasma membrane and is not affected by cytotoxic selection[J]. Proc Natl Acad Sci USA.1991, 88:7386~90
[7] De Angioletti M,Rovira A,Sadelain M,et al.Frequency of missense mutation in coding region of a eukaryotic gene transferred by retroviral victors[J].J vjrol. 2002,76(4):1991~1994
[8] Yang S,Delgado R,King SR,et al.Generation of retroviral vector for clinical studies using transient transfection[J].Hum Gene Ther. 1999,10(1):l23~132
[9] Zhao J,Pincaewaki J,Gomez-Roman VR,et al.Improved protection of rhesus macaques against intrarectal simian immunodeficiency virus SIV(mac251) challenge by a replication-competent Ad5hr-SIVenv/rev and Ad5hr-SIVgag recombinant priming/gp120 boosting regimen[J].J Virol.2003,77(15):8354~8365
[10] RICHARD A. MORGAN, KENNETH CORNETTA,and W. FRENCH ANDERSON. Applications of the Polymerase Chain Reaction in Retroviral- -Mediated Gene Transfer and the Analysis of Gene-Marked Human TIL Cells[J]. HUMAN GENE THERAPY.1990,1:135~149
[11]孙敏,涂新明,丛姑,等.猴D型反转录病毒巢式PCR检测方法的建立[J].中国实验动物学报.2006,14(4):287~289
[12] Wu X,Li Y,Crise B,et al.Weak palindromic consensus sequences are a common feature found at the integration target sites of many retroviruses [J].J Virol. 2005,79(8):5211~5214
[13] Abonour R, Williams DA, Einhorn L. Efficient retrovirus-mediated transfer of the multidrug resistance 1 gene into autologous human long-term repopulating hematopoietic stem cells [J]. Nat Med. 2000,6(6):652~658
[14] Carpinteiro A, Peinert S, Ostertag W, et al. Genetic protection of repopulating hematopoietic cells with an improved MDR1-retrovirus allows administration of intensified chemotherapy following stem cell transplantation in mice [J].Int J Cancer.2002,98(5):785~792
[15] Modlich U, Kustikova OS, Schmidt M, et al. Leukemias following retroviral transfer of multidrug resistance 1 (MDR1) are driven by combinatorial insertional mutagenesis [J]. Blood. 2005,105(11):4235~4246
[16] Bunting K, Zhou Sh, Lu TH, et al. Enforced P-glycoprotein pump function in murine bone marrow cells results in expansion of side population stem cells in vitro and repopulating cells in vivo [J].Blood. 2000,96(3):902~909
[17] Buske C, Humphries RK. Homeobox genes in leukemogenesis [J]. Int J Hematol. 2000,71(4):301~308
[18] Schiedlmeier B, Klump H, Will E, et al. High level ectopic HOXB4 expression confers a profound in vivo competitive growth advantage to human cord blood CD34+ cell,but impairs lymphomyeloid differentiation [J]. Blood. 2003, 101(5): 1759~1768
[19]张秀亚,王珊,李圆,等.多药耐药基因转染对荷瘤小鼠骨髓造血细胞的保护作用[J].第三军医大学学报.2008,30(23):2164~2167
[20]李圆,王珊,王江波,等.多药耐药基因转染骨髓造血细胞移植小鼠的安全性研究[J].中华实验外科杂志.2008.25(5):658~660
[21] Hesdorffer C, Ayello J, Ward M, et al. Phase I trial of retroviral-mediated transfer of the human MDR1 gene as marrow chemoprotection in patients undergoing high-dose chemotherapy and autologous stem-cell transplantation[J].Clin Oncol. 1998,16(1):165~172
[22] O'Shaughnessy JA, Cowan KH, Nienhuis AW, et al. Retroviral mediated transfer of the human multidrug resistance gene (MDR-1) into hematopoietic stem cells during autologous transplantation after intensive chemotherapy for metastatic breast cancer[J]. Hum Gene Ther. 1994,5(7):891~911
[23] Cowan KH, Moscow JA, Huang H, et al. Paclitaxel chemotherapy after autologous stem-cell transplantation and engraftment of hematopoietic cells transduced with a retrovirus containing the multidrug resistance complementary DNA (MDR1) in metastatic breast cancer patients[J]. Clin Cancer Res. 1999, 5(7): 1619~1628
[24] Rahman Z, Kavanagh J, Champlin R. et al. Chemotherapy immediately following autologous stem-cell transplantation in patients with advanced breast cancer[J]. Clin Cancer Res. 1998,4(11):2717~2721
[25] Sugimoto Y, Tsukahara S, Sato S, et al. Drug-selected co-expression of P- glycoprotein and gp91 in vivo from an MDR1-bicistronic retrovirus vector Ha-MDR-IRES-gp91 [J]. Gene Med. 2003,5(5):366~376
[1]王珊,金先庆,杨纯正,等.MDR1基因转染K562细胞株及人骨髓干祖细胞的研究[J].中华小儿外科杂志.2000,21(4):238~41
[2] Wang Y, Jin XQ, Wang S, et a1.Therapeutic efficacy and bone marrow protection of the mdr1 gene and over-dose chemotherapy with doxorubicin for rabbits with hepatocarcinoma[J]. Hepatobiliary Pancreat Dis Int.2006,5(4): 545~51
[3] Chun-Bao Guo, Ying-Cun Li, Xian-Qing Jin. Chemoprotection effect of retroviral vector encoding multidrug resistance 1 gene to allow intensified chemotherapy in vivo[J]. Cancer Chemother Pharmacol (2006) 58: 40~49
[4]张秀亚,王珊,李圆,等.多药耐药基因转染对荷瘤小鼠骨髓造血细胞的保护作用[J].第三军医大学学报.2008,30(23):2164~2167
[5]安淑华,金先庆,解启莲,等。多药耐药基因转染的脐血细胞对白血病小鼠化疗的骨髓保护作用[J].中华血液学杂志.2005:26(2):82~85
[6] DIMBERG A ,BAHRAM F , KARLBERG I , et al . Retinoicacid-induced cell cycle arrest of human myeloid cell lines is associated wit h sequential down-regulation of c-Myc and cyclin Eand post t ranscription up-regulation of p27 (kipⅠ)[J].Blood.2002,99(6):2199~2206
[7]夏璐,鲍英,袁耀宗,等.全反式维甲酸诱导胰腺癌细胞凋亡机制研究[J].中华消化杂志.2004,24(7):391~394
[8] ABU J ,BATUWANGALA M , HERBERT K, et al . Retinoicacid and retinoid receptors :potential chemopreventive and therapeutic role in cervical cancer[J]. Lancet Oncol,2005,6(9):712~720
[9]方建林,郑传胜,冯敢生,等.全反式维甲酸对Walker-256肝癌细胞分化及凋亡的诱导作用[J].世界华人消化杂志.2008,16(26):2929~2934
[10]王珊,金先庆,杨纯正,等.探讨MDR1基因转染K562细胞的最适条件[J].中华小儿外科杂志.2002,23(6):564~565.
[11] Rentala S, Sagar Balla MM, Khurana S, et al.MDR1 gene expression enhances long-term engraftibility of cultured bone marrow cells[J]. BBRC.2005,335(3), 957~964
[12]曹晓伟,唱浩,季峻松,等.阿霉素对肝癌细胞核因子κB活化及细胞周期基因D1表达的影响[J].临床荟萃.2008,23(16):1153~1156
[13] Unter D, Sirioorn K, Jisnuson S, et al. A prospective study of the intake of vitamins C,E and A and the risk of breast cancer[J].N Engl J Med.1993, 329: 234~240
[14] Zhao J, Pinczewski J, Gomez-Roman VR, et al.Improved protection of rhesus macaques against intrarectal simian immunodeficiency virus SIV(mac251) chall- enge by a replication-competent Ad5hr-SIVenv/rev and Ad5hr-SIVgag recom- binant priming/gp120 boosting regimen[J].J Virol. 2003, 77(15):8354 ~8365
[15] Carpinteiro A, Peinert S, Ostertag W,et al. Genetic protection of repopulating hematopoirtic cells with an improved mdr1-retrovirus allows administration of intensified chemotherapy following stem cell transplantation in mice[J].Int J Cancer.2002,98(5):785~792
[16] Carllen DF, Baker E, Smmers RN, et al. Localization of the human multidrug resistance gene,mdr1,to 7q21.1[J]. Hum Genet. 1987,77:142~7
[17] Alexander J. Smith, Ardy van Helvoort, Gerrit van Meer,et al.MDR3 P-glycoprotein, a Phosphatidylcholine Translocase,Transports Several Cytotoxic Drugs and Directly Interacts withDrugs as Judged by Interference with Nucleotide Trapping[J].THE JOURNAL OF BIOLOGICAL CHEMISTRY. 2000,275(31):23530~23539
[18] Harunobu Tahara, Hiroyuki Kusuhara, Eiichi Fuse,et al. P-GLYCOPROTEIN PLAYS A MAJOR ROLE IN THE EFFLUX OF FEXOFENADINE IN THE SMALL INTESTINE AND BLOOD-BRAIN BARRIER, BUT ONLY A LIMITED ROLE IN ITS BILIARY EXCRETION [J].DRUG METABOLISM AND DISPOSITION.2005,33,(7):963~968
[19] Diana Kuhnke1,Gabriele Jedlitschky1,Markus Grube1,et al. MDR1- P-Glycoprotein (ABCB1) Mediates Transport of Alzheimer’s Amyloid-βPeptides-Implications for the Mechanisms of AβClearance at the Blood–Brain Barrier[J]. Brain Pathol.2007,17:347~353
[20] Geromin D,Bourge JF,SouolieA,et al.Glycoprotein 170 induces platelet -activating factor receptor membrane expression and confers tumor cell hypersensitivity to NK-dependent cell lysis[J].J Immunol.2004,172(6): 3604 ~ 3611
[21] Ling V. Multidrug resistance: molecular mechanisms and clinical relevance [J].Cancer chemother pharmacol,1997,40:S3~8
[22] Bennett S,Matthew L,Gayle E,et a1.Selective drug restsrant human osteosarcoma cell lines[J].J Clinical Orthopedics and Related Reseach. 2001, 383,259~267
[23] Kars MD, Iseri OD, Gunduz U, et al. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds [J]. Anticancer Res.2006,26(6):4559~4568
[24] Stromskaya TP, Rybalkina EY, Shtil AA,et al. Influence of exogenous RAR alpha gene on MDR1 expression and P-glycoprotein function in human and rodent cell lines[J]. Br J Cancer.1998,77(11):1718~25
[25]方燕,黄必军,梁启万.原发性肝癌p53基因高频缺失的双色荧光原位杂交证据[J].中国病理生理杂志.2003,19(3):306~309
[26] Zolota V,Batistatou A,Tsamandas A,et al.Immunohistochemical expression of TGF-betal,p2l WAF1,p53,Ki67,and angiogenesis in gastric carcinomas:a clinicopathologic study[J].Int J Gastrointest Cancer.2002,32(2-3):83-9
[27] Jason ON,Michae1 M,Pam S et a1.Promises and challenges of targeting Bcl-2 anti-apoptotic proteins for cancertherapy[J].Biochimica and Biophysica Acta. 2004,1705(1):43-51
[28] Konopleva M, Tsao T, Ruvolo P, et al. Novel triterpenoid CDDO-Me is a potent inducer of apoptosis and differentiation in acute myelogenous leukemia[J]. Blood. 2002 ,99(1):326~335
[29]郑建勇,李开宗,王为忠,等.阿霉素诱导人肝癌细胞凋亡机制的研究[J].中国普通外科杂志.2005,14(7):509~511
[30]李成荣,付劲荣.维甲酸导致多药耐药的机制初探[J].中华肿瘤杂志.2001,23(6):441~443
[31] MAGA G,HUBSCHER U.Proliferating cell nuclear antigen(PCNA):a dancer with many partners[J].J Cel Sci.003,1 16(15):3051~3060
[32] Potrunol R,Zizzo N,Zito AF,et al.Microvascular deneity and endothelial area correlate with Ki-67 proliferative rate in the canine non-Hodgkin’s lymphoma spontaneous mode[J]. Leuk Lymphom.2006,Jun 47(6):1138~1143
[1]Kars MD, Iseri OD, Gunduz U, et al. Development of rational in vitro models for drug resistance in breast cancer and modulation of MDR by selected compounds[J]. Anticancer Res.2006,26(6):4559~4568
[2]Benlloch M,Ortega A, Ferrer P.Acceleration of glutathione efflux and inhibition of gamma -glutamyltranspeptidase sensitize metastatic B16 melanoma cells to endothelium-induced cytotoxicity[J].Journal of Biological Chemistry. 2005, 280(8):6950~6959
[3]Brooks T,Minderman H, Kieran L, et a1.Taxane-based reversal agents modulate drug resistance mediated by P-glycoprotein,multidrng resistance protein,and breast cancer resistance protein[J].Cancer Ther.2003,2:1195
[4]Okada M,Nishio W ,Sakamoto T,et a1.Effect of tumor size on prognosis in patients with non-small cell lung cancer:the role of segmentectomy as a type of lesser resection [J].J Thorac Cardiovasc Surg.2005,129(1):87~93
[5]Greijer AE, de Jong MC, Scheffer GL, et al. Hypoxia-induced acidification causes mitoxantrone resistance not mediated by drug transporters in human breast cancer cells[J]. Cell Oncol.2005,27(1):43~49
[6]Ribosomal proteins S13 and L23 promote multidrug resistance in gastric cancer cells by suppressing drug-induced apoptosis[J]. Exp Cell Res. 2004,296(2):337~346
[7]Sampath J, Sun D, KiddⅥ,et a1.Mutant p53 cooperates with ET3 and selectively up - regulates human MDR1 not MRP1[J].J Biol Chem.2001,276(42):39359
[8] Thomas H,Coley HM.Overcoming multidrug resistance in cancer;an update on the clinical strategy of inhibiting P-glycopmtein[J].Cancer Control.2003,10(2):159~165
[9] Munoz Martinez F,Reyes CP,Perez Lomas AL,et a1.Insights into the molecular mechanism of action of celastraceae sesquiterpenes as specific,non-transported inhibitors of human P-glycoprotein[J].Biochim Biophys Acta.2006,1758(1):98~110
[10]Globisch C,Pajeva IK,Wiese M.Structure-activity relationships of a series of tariqu- idar analogs as multidrug resistance modulators[J].Bioorg Med Chem.2006,14(5): 1588~1598
[11]J WANG S,FOLKESA,CHUCKOWREEI,et a1.Studies On pyrrolopy-rimidines as selective inhibitors Of multidrug-resistance-associated protein in multidrug resistance[J].J Med Chem.2004,47:1329~1338
[12] BIENYAHIA B,HUGUAT S,DECIEYES X,et a1.Multidrug resistance- associaied protein MRP1 expression in human gliomas:Chemosensitization tovincristine and etoposide by indomethacin in human glioma cell lines overexpressing MRP1[J].J Neurooncol.2004,66:65~70
[13]Kitazono M,Okumura H,Ikeda R.Reversal of LRP-associated drug resistance in colon carcinoma SW-620 cells[J].Int J Cancer.2001,91(1):126~131
[14]Burg D,Filippov DV ,Hermanns R,et al.Peptidomimetic glutathi- olle analogues as novel gammaGT stable GST inhibitors[J].Bioorg Med Chem, 2002, 10(1):195~205.
[15]Mistry P,Stewart AJ,Dangerfield W,et al.In vitro and in vivo characterization of XR11576,a novel,orally active,dual inhibitor of to poisomeraseⅠandⅡ[J]. Anticancer Drugs,2002,13(1):15~28
[16]Chan JY,Chu AC,Fung KP.Inhibition of P-glycoprotein expression and reversal of drug resistance of human hepatoma HepG2 cells by multidrug resistance gene mdrl ontiscnse RNA[J].Life Sci.2000,67:2l17~2l24
[17]Irie A, Matsumoto K, Anderegg B, et al.Growth inhibition efficacy anadenovirus expressing dual therapeutic genes,wild type p53,and anti-erbB2 ribozyme,against human bladder cancer cells[J].Cancer genes Ther.2006,13(3):298~305
[18]Kobayashi H,Takemura Y,Miyachi H.Novel approaches to reversing anti-cancer drug resistance using gene-specific therapeutics[J].Hum Cell.2001,14:172~184
[19]Kostenko EV, Laktionov PP,VIassov VV ,et a1.Downregulation of PGY1/MDR1 mRNA level in human KB cells by antisense oligonucleotide conjugates : RNA accessibility in vitro and intracellular antisensc activity[J].Bioehim Biophys Acta. 2002,l576:l43~147
[20]Eferth T,Futseher BW, Osieka R.5-Aza dine modu1ates the response of senstive and mdtidrug-resistant K562 leukemic cells to cytostatic drugs[J].Blood Cells Mol Dis.2001,27(3):637
[21] Barzon L,Bonaguro R,Castagliuolo I,et a1.Gene therapy of thyreid cancer via retrovirally-driven combined expression of human interleukin-2 and herpes simplex virus thymidine kinase[J].Eur J Endocrinol.2003,148:73~80