许小婷,汪月,王琳,曾宪卓.旋转式生物反应器培养细胞因子诱导的杀伤细胞研究[J].转化医学杂志,2020,9(4):233-237
旋转式生物反应器培养细胞因子诱导的杀伤细胞研究
The study of culture of cytokine-induced killer cells by rotating wall-vessel bioreactor
  
DOI:
中文关键词:  免疫细胞  细胞因子诱导的杀伤细胞  细胞活率  肿瘤杀伤力  旋转式生物反应器
英文关键词:Immune cell  Cytokine-induced killer (CIK) cells  Cell viability  Tumor lethality  Rotating wall-vessel bioreactor (RWV)
基金项目:深圳市海外高层次人才创新创业专项资金项目(KQCY2015072914455645);深圳市财政专项资金股权投资项目(GQYCZZ20160427145010)
作者单位
许小婷 深圳爱生再生医学科技有限公司 
汪月 深圳爱生再生医学科技有限公司 
王琳 深圳爱生再生医学科技有限公司 
曾宪卓 深圳爱生再生医学科技有限公司 
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中文摘要:
      目的采用旋转式生物反应器(rotating wall-vessel bioreactor,RWV)与传统的T-Flask培养瓶培养人外周血细胞因子诱导的杀伤(cytokine-induced killer,CIK)细胞,比较CIK细胞的增殖能力、细胞因子分泌情况、细胞杀伤力。方法提取健康志愿者的外周血,分离出外周单个核细胞(peripheral blood mononuclear cell,PBMC),体外诱导成CIK细胞,分别接种到T-Flask培养瓶及RWV中培养。采用台酚蓝染色法检测细胞活率并绘制其生长曲线;ELISA法检测各组CIK细胞的IL-2,IFN-γ,TNF-α分泌情况;乳酸脱氢酶测定法检测CIK细胞对靶细胞的杀伤力。结果与传统的T-Flask培养瓶培养CIK细胞比较,RWV培养CIK细胞增殖数量明显多且生长力旺盛;使用RWV培养CIK细胞能使其分泌更多的IL-2,IFN-γ,TNF-α细胞因子;RWV组培养的CIK细胞对肿瘤细胞的杀伤力更强。结论RWV培养CIK细胞更能提高其增殖能力、细胞因子分泌能力及肿瘤杀伤活力,同时可解决传统细胞培养数量少、稳定性差、易污染、工作量大及周期长等问题,有效地降低临床治疗成本。
英文摘要:
      ObjectiveThe purpose of this study was to culture human peripheral blood cytokine-induced killer (CIK) cells using rotating wall-vessel bioreactor (RWV) and conventional culture T-Flask to compare the proliferation capacity, factor secretion and cell killing power of CIK cells between the two groups. MethodsPeripheral blood of healthy volunteers was extracted, peripheral blood mononuclear cells (PBMC) were isolated and induced into CIK cells in vitro, and inoculated into T-Flask and RWV at a density of 1×107/mL to observe the morphological characteristics. The cell viability was measured by phenol blue staining and its growth curve was drawn. The secretion of IL-2, IFN-γ and TNF-α factors in each group of CIK cells was detected by ELISA. Using the lactate dehydrogenase assay to detect the lethality of CIK cells on target cells. ResultsCompared with traditional culture T-Flask, although the morphology of the CIK cells that was cultured in RWV was not significantly different, the amount of CIK cells cultured in RWV was significantly higher than that of the conventional culture T-Flask (P<0.05). And the cells were more stronger and vigorous than the traditional culture T-Flask. The growth factor secretion of each experimental group showed that the use of RWV to culture CIK cells could secrete more IL-2, IFN-γ, and TNF-α cytokines. The CIK cells cultured in the RWV group had stronger lethality to tumor cells. ConclusionThis study reveals that the RWV culture can improve the proliferation ability, cell growth factor secretion and tumor killing activity of CIK cells, and that can solve the problem of small number of traditional cell culture, poor stability, easy pollution, and large workload and long periods and so on. This will effectively reduce the cost of clinical treatment.
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