CAJ | 학술논문

目的体外观察不同浓度三氧化二砷(As2O3)在不同的诱导时间作用下对3种神经母细胞瘤(NB)细胞株分化与凋亡的作用.方法采用不同浓度(0、1、3、5、7 μmol/L)的As2O3诱导3组NB细胞株SK-N-SH、SK-N-BE2、SH-SY5Y,在同等条件下分别培养24、48、72 h.采用荧光原位杂交技术、流式细胞术、细胞增殖毒性检测法,检测As2O3对3种NB细胞株的细胞生物学影响.结果 5μmol/L As2O3诱导72 h后,SK-N-BE2的MYCN基因扩增数量明显减少;相较于低浓度(1μmol/L) As2O3,随着浓度的增加,3组细胞增殖活力逐渐降低,SH-SY5Y:24 h(chisq=9.666 7,P＜O.05),48 h(chisq=9.666 7,P＜0.05),72 h(chisq=9.512 8,P＜0.05);SK-N-SH:24 h(chisq=10.384 6,P＜O.05,),48 h(chisq=8.641 0,P＜0.05),72 h(chisq=9.461 5,P＜0.05),SK-N-BE2:24 h(chisq=8.435 9,P＜0.05),48 h(chisq=8.641 0,P＜O.05),72 h(chisq=9.545 5,P＜0.05);流式细胞仪检测结果显示,随着As2O3浓度的增加,与空白组相比,细胞早晚期凋亡率明显增加,凋亡率分别为1.6％(0 μmoL/L),3.8％(1μmol/L),6.1％(3μmol/L),10.4％(5μmol/L),40.2％(7μmol/L);细胞周期中G2/M期延长,细胞分裂受到抑制.结论 1.As2O3对NB细胞株具有诱导凋亡,抑制细胞增殖的作用,呈剂量依赖性,不同类型的NB细胞株对As2O3敏感程度不一.2.SK-N-BE2细胞在As2O3诱导后,扩增的MYCN基因具有转阴趋势.3.经As2O3诱导后,3组NB细胞的细胞周期在S期及G2/M期受到阻滞,细胞核酸复制及细胞分裂受到抑制.
목적 체외관찰불동농도삼양화이신(As2O3)재불동적유도시간작용하대3충신경모세포류(NB)세포주분화여조망적작용.방법 채용불동농도(0、1、3、5、7 μmol/L)적As2O3유도3조NB세포주SK-N-SH、SK-N-BE2、SH-SY5Y,재동등조건하분별배양24、48、72 h.채용형광원위잡교기술、류식세포술、세포증식독성검측법,검측As2O3대3충NB세포주적세포생물학영향.결과 5μmol/L As2O3유도72 h후,SK-N-BE2적MYCN기인확증수량명현감소;상교우저농도(1μmol/L) As2O3,수착농도적증가,3조세포증식활력축점강저,SH-SY5Y:24 h(chisq=9.666 7,P＜O.05),48 h(chisq=9.666 7,P＜0.05),72 h(chisq=9.512 8,P＜0.05);SK-N-SH:24 h(chisq=10.384 6,P＜O.05,),48 h(chisq=8.641 0,P＜0.05),72 h(chisq=9.461 5,P＜0.05),SK-N-BE2:24 h(chisq=8.435 9,P＜0.05),48 h(chisq=8.641 0,P＜O.05),72 h(chisq=9.545 5,P＜0.05);류식세포의검측결과현시,수착As2O3농도적증가,여공백조상비,세포조만기조망솔명현증가,조망솔분별위1.6％(0 μmoL/L),3.8％(1μmol/L),6.1％(3μmol/L),10.4％(5μmol/L),40.2％(7μmol/L);세포주기중G2/M기연장,세포분렬수도억제.결론 1.As2O3대NB세포주구유유도조망,억제세포증식적작용,정제량의뢰성,불동류형적NB세포주대As2O3민감정도불일.2.SK-N-BE2세포재As2O3유도후,확증적MYCN기인구유전음추세.3.경As2O3유도후,3조NB세포적세포주기재S기급G2/M기수도조체,세포핵산복제급세포분렬수도억제.
Objective To identify the effect of arsenic trioxide (As2O3) on the differentiation and apoptosis of different types of neuroblastoma(NB) cell lines.Methods The cell lines [SK-N-SH,SK-N-BE2,SH-SY5 Y] were induced with different concentrations (0 μmol/L,1 μmol/L,3 μmol/L,5 μ mol/L and 7 μ mol/L) of arsenic trioxide for 24 h,48 h,72 h under the same conditions.The expression of MYCN gene was examined by fluorescence in situ hybridization assay in SK-N-BE2,cell proliferation,cell cycle and cell apoptosis were detected with cell counting kit-8 (CCK-8) assay and flow cytometry.Results 5 μmol/L of As2O3 inhibited the expression of MYCN gene in SK-N-BE2;CCK-8 assay indicated that As2O3 inhibited the proliferation of NB cell in a dose-and time-dependent manner,the cell proliferation was significantly suppressed compared with the low concentration (1 μ mol/L) after treated with As2O3 by 1 μmol/L,3 μmol/L,5 μmol/L and 7 μmol/L in 24 h,48 h and 72 h,SH-SY5Y:24 h(chisq =9.666 7,P ＜ 0.05),48 h (chisq =9.666 7,P ＜ O.05),72 h (chisq =9.512 8,P ＜ 0.05);SK-N-SH,24 h (chisq =10.38,P＜0.054 6),48 h(chisq=8.641 0,P＜0.05),72 h(chisq=9.461 5,P＜0.05);SK-N-BE2:24 h (chisq =8.435 9,P ＜0.05),48 h(chisq =8.641 O,P ＜0.05),72 h(chisq =9.545 5,P ＜0.05);compared with the control group,the As2O3-treated cells showed increased apoptosis percentage,with the percentage of 1.6％ (0 μmol/L),3.8％ (1 μmol/L),6.1％ (3 μmol/L),10.4％ (5 μmol/L),40.2％ (7 μ mol/L);the cell cycle was arrested at G2/M phase,which prevented cell division.Conclusions (1) As2O3 play an important role on the NB cells proliferation,apoptosis which were dose-and time-dependent manner.(2)As2O3 can inhibit the expression of MYCN gene.(3)As2O3 also could block NB cell cycle at S and G2/M,and inhibit the cell nucleus replication and the As2O3 had different induced effect between different types of NB cell.