CAJ | 학술논문

目的：研究二烯丙基二硫( diallyl disulfide, DADS)诱导人白血病HL-60细胞周期阻滞及其分子机制。方法采用细胞计数、软琼脂克隆形成实验及流式细胞术观察DADS对HL-60细胞生长抑制与周期阻滞效应。 Western blot检测DADS对HL-60细胞Chk1/2以及下游分子的影响。结果细胞计数显示,60、120μmol · L-1 DADS处理后,其群体倍增时间从19．14 h增加到35．03、71．82 h ( P<0．05)。软琼脂克隆形成实验表明,30、60、90、120μmol · L-1 DADS 对HL-60细胞克隆形成率的抑制率分别为35．06%、62．10%、93．79%、99．35%(P<0．05)。流式细胞术检测显示,60和120μmol·L-1 DADS分别作用HL-60细胞24 h后,DADS可呈时间与浓度依赖性诱导 HL-60细胞 G2/M 期阻滞( P <0．05)。60μmol·L-1 DADS处理HL-60细胞后,p-Chk1可呈时间依赖性上调(P<0．05),而Chk1与Chk2总蛋白和p-Chk2无改变(P>0．05)。并且,Cdc25C、CyclinB1和CDK1分别呈时间依赖性下调( P<0．05),但14-3-3蛋白表达没有改变( P>0．05)。结论 DADS能够抑制HL-60细胞增殖,并通过Chk1/Cdc25C/CyclinB1/CDK1通路阻滞HL-60细胞于G2/M期。
목적：연구이희병기이류( diallyl disulfide, DADS)유도인백혈병HL-60세포주기조체급기분자궤제。방법채용세포계수、연경지극륭형성실험급류식세포술관찰DADS대HL-60세포생장억제여주기조체효응。 Western blot검측DADS대HL-60세포Chk1/2이급하유분자적영향。결과세포계수현시,60、120μmol · L-1 DADS처리후,기군체배증시간종19．14 h증가도35．03、71．82 h ( P<0．05)。연경지극륭형성실험표명,30、60、90、120μmol · L-1 DADS 대HL-60세포극륭형성솔적억제솔분별위35．06%、62．10%、93．79%、99．35%(P<0．05)。류식세포술검측현시,60화120μmol·L-1 DADS분별작용HL-60세포24 h후,DADS가정시간여농도의뢰성유도 HL-60세포 G2/M 기조체( P <0．05)。60μmol·L-1 DADS처리HL-60세포후,p-Chk1가정시간의뢰성상조(P<0．05),이Chk1여Chk2총단백화p-Chk2무개변(P>0．05)。병차,Cdc25C、CyclinB1화CDK1분별정시간의뢰성하조( P<0．05),단14-3-3단백표체몰유개변( P>0．05)。결론 DADS능구억제HL-60세포증식,병통과Chk1/Cdc25C/CyclinB1/CDK1통로조체HL-60세포우G2/M기。
Aim To study the effects of cycle arrest and molecular mechanism in human leukemia HL-60 cells induced by diallyl disulfide ( DADS ) . Methods Cell count, colony formation in soft agar experiments and flow cytometry analysis were employed to observe the DADS-induced cell growth inhibition and the effect of cycle arrest in HL-60 cells. The expressions of Chk1/2 and its downstream element in HL-60 cells were detected by Western blot. Results Cell count revealed that population doubling time increased to 35. 03 h and 71. 82 h, respectively, from 19. 14 h in HL-60 cells treated with 60 and 120 μmol·L-1 DADS ( P<0. 05 ) . Colony formation in soft agar experiments showed that colony formation inhibition rate of HL-60 cells exposed to 30, 60, 90 and 120μmol·L-1 DADS increased to 35. 06%, 62. 10%, 93. 79% and 99. 35%, respectively ( P<0. 05 ) . Flow cytometry a-nalysis exhibited that HL-60 cells treated with 60 and 120 μmol · L-1 DADS for 24 h and 48 h arrested in G2/M phase in a concentration-and time-dependent manner ( P <0. 05 ) . Western blot disclosed that the expression of p-Chk1 increased in a time-dependent manner ( P <0. 05 ); however, Chk1, Chk2 and p-Chk2 were not changed in HL-60 cells treated with 60μmol·L-1 DADS (P >0. 05). The expression of Cdc25C, CyclinB1 and CDK1 decreased after treated with 60 μmol·L-1 DADS in a time-dependent manner ( P<0. 05 ) , but the expression of 14-3-3 protein did not change ( P>0. 05 ) . Conclusion DADS can in-hibit the proliferation of HL-60 cells, and induce G2/M arrest through Chk1/Cdc25 C/CyclinB1/CDK1 path-way.