中华小儿外科杂志
中華小兒外科雜誌
중화소인외과잡지
CHINESE JOURNAL OF PEDIATRIC SURGERY
2013年
12期
922-926
,共5页
钟海军%柳龚堡%董岿然%肖现民
鐘海軍%柳龔堡%董巋然%肖現民
종해군%류공보%동규연%초현민
神经母细胞瘤%细胞低氧%RNA,信使
神經母細胞瘤%細胞低氧%RNA,信使
신경모세포류%세포저양%RNA,신사
Neuroblastoma%Cell hypoxia%RNA,messenger
目的 探讨SK-N-SH神经母细胞瘤株在氯化钴模拟缺氧条件下侧群(side population,SP)细胞比例的变化及SP细胞基因表达谱的改变.方法 采用二氯化钴模拟缺氧条件,然后利用Hoechst 33342染料法,通过流式细胞仪紫外激光检测分选功能分选出缺氧及常氧SK-N-SH细胞中的SP细胞,利用Agilent 8×60K人(Human) G3表达谱芯片进行检测,并对基因芯片结果进行分析,以荧光定量PCR验证芯片结果.结果 SK-N-SH细胞株的SP细胞比例约为8.65％,缺氧后降至约2.08％;缺氧SP细胞和常氧SP细胞mRNA及lincRNA的表达谱存在明显差异.差异＞2倍的基因有5077 mRNA、1383 lincRNA、其中上调的有2550 mRNA、572 lincRNA,下调的有2527 mRNA、81 1 lincRNA.差异表达的基因主要涉及细胞周期、DNA复制、p53及MAPK信号通路等生物学过程.荧光定量PCR基因结果与芯片差异基因结果表达趋势一致.结论 缺氧使SP细胞比例下降,并使SP细胞mRNA和lincRNA的表达谱均发生显著改变,提示mRNA和lincRNA共同参与调节神经母细胞瘤SP细胞在缺氧环境中的生物学行为.
目的 探討SK-N-SH神經母細胞瘤株在氯化鈷模擬缺氧條件下側群(side population,SP)細胞比例的變化及SP細胞基因錶達譜的改變.方法 採用二氯化鈷模擬缺氧條件,然後利用Hoechst 33342染料法,通過流式細胞儀紫外激光檢測分選功能分選齣缺氧及常氧SK-N-SH細胞中的SP細胞,利用Agilent 8×60K人(Human) G3錶達譜芯片進行檢測,併對基因芯片結果進行分析,以熒光定量PCR驗證芯片結果.結果 SK-N-SH細胞株的SP細胞比例約為8.65％,缺氧後降至約2.08％;缺氧SP細胞和常氧SP細胞mRNA及lincRNA的錶達譜存在明顯差異.差異＞2倍的基因有5077 mRNA、1383 lincRNA、其中上調的有2550 mRNA、572 lincRNA,下調的有2527 mRNA、81 1 lincRNA.差異錶達的基因主要涉及細胞週期、DNA複製、p53及MAPK信號通路等生物學過程.熒光定量PCR基因結果與芯片差異基因結果錶達趨勢一緻.結論 缺氧使SP細胞比例下降,併使SP細胞mRNA和lincRNA的錶達譜均髮生顯著改變,提示mRNA和lincRNA共同參與調節神經母細胞瘤SP細胞在缺氧環境中的生物學行為.
목적 탐토SK-N-SH신경모세포류주재록화고모의결양조건하측군(side population,SP)세포비례적변화급SP세포기인표체보적개변.방법 채용이록화고모의결양조건,연후이용Hoechst 33342염료법,통과류식세포의자외격광검측분선공능분선출결양급상양SK-N-SH세포중적SP세포,이용Agilent 8×60K인(Human) G3표체보심편진행검측,병대기인심편결과진행분석,이형광정량PCR험증심편결과.결과 SK-N-SH세포주적SP세포비례약위8.65％,결양후강지약2.08％;결양SP세포화상양SP세포mRNA급lincRNA적표체보존재명현차이.차이＞2배적기인유5077 mRNA、1383 lincRNA、기중상조적유2550 mRNA、572 lincRNA,하조적유2527 mRNA、81 1 lincRNA.차이표체적기인주요섭급세포주기、DNA복제、p53급MAPK신호통로등생물학과정.형광정량PCR기인결과여심편차이기인결과표체추세일치.결론 결양사SP세포비례하강,병사SP세포mRNA화lincRNA적표체보균발생현저개변,제시mRNA화lincRNA공동삼여조절신경모세포류SP세포재결양배경중적생물학행위.
Objective To investigate the ratio of side population(SP) cells and their gene expression profile in human neuroblastoma SK-N-SH cell line in mimicked hypoxia induced by CoCl2.Methods Model of hypoxia was established by CoCl2.Sorting and analysis of SP cells and non-side population cells were performed,based on the exclusion of the DNA binding dye,Hoechst 33342,with and without verapamil in FACS.The gene expression profiles (including mRNA and lincRNA)were measured by Agilent microarray.The results were validated by real-time polymerase chain reaction (PCR).Results The ratio of SP cell decreased from 8.65％oo to 2.08％ after hypoxia.The microarray revealed that 2550 mRNAs and 572 lincRNAs were significantly up-regulated and 2527mRNAs and 811 lincRNAs were significantly down-regulated in SP compared to non-SP cell.The analysis of GO and pathway showed that the differentially expressed genes were mainly involved in cell cycle,DNA replication,p53 signal pathway,MAPK signal pathway,et.al.Conclusions The hypoxia down-regulates the ratio of SP cell and has an effect on the mRNA and lincRNA expression profile of SP cells in neuroblastoma SK-N-SH cell line,which suggests that both mRNA and lincRNA are involved in the regulation of the biological behavior of neuroblastoma SP cells in hypoxia.
