In this study to screen for stable, high Taxolproducing cell lines（CL5, CL12, and CL21） of Taxus cuspidata, stem tissues were used to induce calli, which were then subcultured nine times to establish suspension cell...In this study to screen for stable, high Taxolproducing cell lines（CL5, CL12, and CL21） of Taxus cuspidata, stem tissues were used to induce calli, which were then subcultured nine times to establish suspension cell cultures. From 97 cell lines obtained from conditioned cultures, 10 cell lines with high Taxol content were selected. Stability analyses on solid and liquid B5 media were then used to obtain lines that stably produced high levels of Taxol. Fresh biomass and Taxol production of the ninth generation became stable. Taxol content of selected CL5, CL12, and CL21 samples was 0.0448, 0.0477, and0.0428% of dry mass（DW）, respectively. Proliferation of CL5, CL12 and CL21 was 346.3, 382.5, and 409.2%,respectively. From work over about 2 years, the three cell lines appear suitable for mass production of Taxol,promoting the industrialisation and commercial-scale production of Taxol using cell culture.展开更多
目的:了解两种卵巢癌紫杉醇耐药株(OC3/PIX3及OC3/PIX5)和其亲代细胞紫杉醇敏感株(OC3)染色体DNA拷贝数的差异变化,探讨耐药细胞的分子遗传学改变。方法:运用比较基因组杂交(comparative genom ic hybrid ization,CGH)技术,将OC3/PIX3及...目的:了解两种卵巢癌紫杉醇耐药株(OC3/PIX3及OC3/PIX5)和其亲代细胞紫杉醇敏感株(OC3)染色体DNA拷贝数的差异变化,探讨耐药细胞的分子遗传学改变。方法:运用比较基因组杂交(comparative genom ic hybrid ization,CGH)技术,将OC3/PIX3及OC3/PIX5与其亲代细胞株OC3的染色体基因组进行比较分析。结果:3种细胞株的染色体2pter-p21、3q、5p、9均有遗传物质表达增加;10号染色体表达增加仅见于OC3。染色体1,4,6的遗传物质在3种细胞株的表达均有减少。3p的减少发生在OC3和OC3/PIX5。仅在OC3细胞中见到7q、8p减少。OC3和OC3/PIX5出现染色体10q22过度扩增。最有意义的变化是OC3/PIX3细胞染色体2p22的过度扩增。结论:2p22拷贝数过度扩增可能与卵巢癌紫杉醇耐药相关。展开更多
目的建立紫杉醇耐药细胞株LoVo/Taxol,探讨结肠癌细胞紫杉醇获得性耐药的可能机制。方法反复用紫杉醇处理结肠癌LoVo细胞,诱导建立结肠癌紫杉醇耐药细胞株LoVo/Taxol;通过MTT法检测细胞药物敏感性,光学显微镜和透射电镜观察细胞形态学变...目的建立紫杉醇耐药细胞株LoVo/Taxol,探讨结肠癌细胞紫杉醇获得性耐药的可能机制。方法反复用紫杉醇处理结肠癌LoVo细胞,诱导建立结肠癌紫杉醇耐药细胞株LoVo/Taxol;通过MTT法检测细胞药物敏感性,光学显微镜和透射电镜观察细胞形态学变化,流式细胞术检测细胞周期分布及细胞凋亡水平,实时荧光定量PCR法测定细胞中多药耐药基因MDR1、MRP1、LRP、GST-π和TopoⅡ的表达。结果建立的紫杉醇耐药细胞株LoVo/Taxol对紫杉醇的耐药指数为42,为高度耐药;对阿霉素及5-氟脲嘧啶也产生强烈的交叉耐药,耐药指数分别为796和187。相较于亲本细胞LoVo,LoVo/Taxol细胞形态发生明显的变化,G2/M期阻滞和细胞凋亡率明显降低(P<0.05)。在五种经典的耐药相关蛋白中MRP1、MDR1基因m RNA的转录水平分别提高了37倍和2倍,LRP、GST-π基因m RNA的转录水平分别下降了11倍和3倍(P均<0.05),To Po II基因转录水平无明显变化。结论成功建立的人结肠癌紫杉醇耐药细胞株LoVo/Taxol为典型的多药耐药细胞模型,其耐药机制可能与MRP1基因的过表达有关。展开更多
基金supported by the ‘‘12th Five Year Plan’’ National Science and Technology in Rural Area(Nos.2013AA103005-04 and 2012AA10A506-04)Changchun City Science and Technology Development Program(No.2014174)+1 种基金Changchun City Science and Technology Support Program(No.2014NK002)Graduate Innovation Fund of Jilin University(No.2016172)
文摘In this study to screen for stable, high Taxolproducing cell lines（CL5, CL12, and CL21） of Taxus cuspidata, stem tissues were used to induce calli, which were then subcultured nine times to establish suspension cell cultures. From 97 cell lines obtained from conditioned cultures, 10 cell lines with high Taxol content were selected. Stability analyses on solid and liquid B5 media were then used to obtain lines that stably produced high levels of Taxol. Fresh biomass and Taxol production of the ninth generation became stable. Taxol content of selected CL5, CL12, and CL21 samples was 0.0448, 0.0477, and0.0428% of dry mass（DW）, respectively. Proliferation of CL5, CL12 and CL21 was 346.3, 382.5, and 409.2%,respectively. From work over about 2 years, the three cell lines appear suitable for mass production of Taxol,promoting the industrialisation and commercial-scale production of Taxol using cell culture.
文摘目的:了解两种卵巢癌紫杉醇耐药株(OC3/PIX3及OC3/PIX5)和其亲代细胞紫杉醇敏感株(OC3)染色体DNA拷贝数的差异变化,探讨耐药细胞的分子遗传学改变。方法:运用比较基因组杂交(comparative genom ic hybrid ization,CGH)技术,将OC3/PIX3及OC3/PIX5与其亲代细胞株OC3的染色体基因组进行比较分析。结果:3种细胞株的染色体2pter-p21、3q、5p、9均有遗传物质表达增加;10号染色体表达增加仅见于OC3。染色体1,4,6的遗传物质在3种细胞株的表达均有减少。3p的减少发生在OC3和OC3/PIX5。仅在OC3细胞中见到7q、8p减少。OC3和OC3/PIX5出现染色体10q22过度扩增。最有意义的变化是OC3/PIX3细胞染色体2p22的过度扩增。结论:2p22拷贝数过度扩增可能与卵巢癌紫杉醇耐药相关。
文摘目的建立紫杉醇耐药细胞株LoVo/Taxol,探讨结肠癌细胞紫杉醇获得性耐药的可能机制。方法反复用紫杉醇处理结肠癌LoVo细胞,诱导建立结肠癌紫杉醇耐药细胞株LoVo/Taxol;通过MTT法检测细胞药物敏感性,光学显微镜和透射电镜观察细胞形态学变化,流式细胞术检测细胞周期分布及细胞凋亡水平,实时荧光定量PCR法测定细胞中多药耐药基因MDR1、MRP1、LRP、GST-π和TopoⅡ的表达。结果建立的紫杉醇耐药细胞株LoVo/Taxol对紫杉醇的耐药指数为42,为高度耐药;对阿霉素及5-氟脲嘧啶也产生强烈的交叉耐药,耐药指数分别为796和187。相较于亲本细胞LoVo,LoVo/Taxol细胞形态发生明显的变化,G2/M期阻滞和细胞凋亡率明显降低(P<0.05)。在五种经典的耐药相关蛋白中MRP1、MDR1基因m RNA的转录水平分别提高了37倍和2倍,LRP、GST-π基因m RNA的转录水平分别下降了11倍和3倍(P均<0.05),To Po II基因转录水平无明显变化。结论成功建立的人结肠癌紫杉醇耐药细胞株LoVo/Taxol为典型的多药耐药细胞模型,其耐药机制可能与MRP1基因的过表达有关。
