西北药学杂志
西北藥學雜誌
서북약학잡지
2014年
5期
472-477
,共6页
易岚%伍尤华%谭晖%何洁%李林蔚%单健%苏琦
易嵐%伍尤華%譚暉%何潔%李林蔚%單健%囌琦
역람%오우화%담휘%하길%리림위%단건%소기
DADS%Rac2%活性氧%凋亡%K562细胞
DADS%Rac2%活性氧%凋亡%K562細胞
DADS%Rac2%활성양%조망%K562세포
DADS%Rac2%reactive oxygen species%apoptosis%K562 cells
目的：研究二烯丙基二硫（diallyl disulfide ，DADS ）诱导人白血病K562细胞凋亡的分子机制。方法 Real-time PCR检测Rac2基因mRNA水平；Western blot检测Rac2、JNK、p38等蛋白的表达；基因沉默Rac2蛋白的表达；流式细胞术检测凋亡细胞百分率以及细胞内的活性氧（reactive oxygen species ，ROS）水平；琼脂糖凝胶电泳检测晚期凋亡。结果随着DADS质量浓度的增加，Rac2基因mRNA水平明显上调（P＜0．05）。与未干扰组相比，siRac2 RNA组凋亡率明显降低（P＜0．05）。Rac2 siR-NA干扰的DADS诱导的 K562细胞并未出现梯状条带。与未干扰组相比，siRac2 RNA组在5．0，10．0和15．0 mg · L-1 DADS作用于K562细胞8 h后，荧光强度显著降低（P＜0．05）。Western blot分析结果显示：与对照组相比，10．0和15．0 mg · L-1 DADS作用于K562细胞24 h后，蛋白 Rac2的表达水平明显上调（P＜0．05）；用 siRNA抑制Rac2蛋白的表达后，JNK1的表达显著降低，p38和磷酸化的p38的表达在Rac2干扰前后没有明显变化。结论 Rac2与DADS诱导K562细胞凋亡、活性氧的产生密切相关。活化的Rac2选择性地激活JN K信号通路而不是p38信号通路导致细胞凋亡。
目的：研究二烯丙基二硫（diallyl disulfide ，DADS ）誘導人白血病K562細胞凋亡的分子機製。方法 Real-time PCR檢測Rac2基因mRNA水平；Western blot檢測Rac2、JNK、p38等蛋白的錶達；基因沉默Rac2蛋白的錶達；流式細胞術檢測凋亡細胞百分率以及細胞內的活性氧（reactive oxygen species ，ROS）水平；瓊脂糖凝膠電泳檢測晚期凋亡。結果隨著DADS質量濃度的增加，Rac2基因mRNA水平明顯上調（P＜0．05）。與未榦擾組相比，siRac2 RNA組凋亡率明顯降低（P＜0．05）。Rac2 siR-NA榦擾的DADS誘導的 K562細胞併未齣現梯狀條帶。與未榦擾組相比，siRac2 RNA組在5．0，10．0和15．0 mg · L-1 DADS作用于K562細胞8 h後，熒光彊度顯著降低（P＜0．05）。Western blot分析結果顯示：與對照組相比，10．0和15．0 mg · L-1 DADS作用于K562細胞24 h後，蛋白 Rac2的錶達水平明顯上調（P＜0．05）；用 siRNA抑製Rac2蛋白的錶達後，JNK1的錶達顯著降低，p38和燐痠化的p38的錶達在Rac2榦擾前後沒有明顯變化。結論 Rac2與DADS誘導K562細胞凋亡、活性氧的產生密切相關。活化的Rac2選擇性地激活JN K信號通路而不是p38信號通路導緻細胞凋亡。
목적：연구이희병기이류（diallyl disulfide ，DADS ）유도인백혈병K562세포조망적분자궤제。방법 Real-time PCR검측Rac2기인mRNA수평；Western blot검측Rac2、JNK、p38등단백적표체；기인침묵Rac2단백적표체；류식세포술검측조망세포백분솔이급세포내적활성양（reactive oxygen species ，ROS）수평；경지당응효전영검측만기조망。결과수착DADS질량농도적증가，Rac2기인mRNA수평명현상조（P＜0．05）。여미간우조상비，siRac2 RNA조조망솔명현강저（P＜0．05）。Rac2 siR-NA간우적DADS유도적 K562세포병미출현제상조대。여미간우조상비，siRac2 RNA조재5．0，10．0화15．0 mg · L-1 DADS작용우K562세포8 h후，형광강도현저강저（P＜0．05）。Western blot분석결과현시：여대조조상비，10．0화15．0 mg · L-1 DADS작용우K562세포24 h후，단백 Rac2적표체수평명현상조（P＜0．05）；용 siRNA억제Rac2단백적표체후，JNK1적표체현저강저，p38화린산화적p38적표체재Rac2간우전후몰유명현변화。결론 Rac2여DADS유도K562세포조망、활성양적산생밀절상관。활화적Rac2선택성지격활JN K신호통로이불시p38신호통로도치세포조망。
Objective To research the molecular mechanism of diallyl disulfide (DADS )-induced apoptosis in human leukemia K562 cells .Methods mRNA levels of Rac2 were detected by real-time PCR ;Rac2 ,JNK and p38 levels were measured by Western blot .Treatment of K562 cells with siRNAs resulted in inhibition of Rac2 expression .Flow cytometry method was used to deter-mine the percentage of apoptosis cells and DNA agarose electrophoresis was used to determine the late stage of apoptosis .Levels of ROS were measured by 2′,7′-dichlorofluorescein diacetate (DCFH-DA) fluorescence .Results There was a significant up regu-lation of the mRNA level of Rac2 in K562 cells after the treatment with different concentration of DADS (P<0 .05) .Compared with the control group ,there was a significant up regulation of Rac2 protein in K562 cells treated with 5 .0 and 10 .0 mg · L -1 DADS for 24 h (P<0 .05) .Treatment of cells with DADS for 24 h markedly reduced the percentage of apoptotic cells after inhibi-tion of Rac2 with siRNA .SiRNA-treated groups showed normal bands on agarose gel electrophoresis .After suppression of Rac2 , ROS production was markedly reduced after DADS treatment (P<0 .05) .The expression of JNK1 in K562 cells decreased when Rac2 expression was inhibited with siRNA ,while there was no change in p38 and p-p38 .Conclusion These results indicate Rac2 have a critical role in DADS-induced apoptosis and ROS production in DADS-induced K562 cells .Rac2 selectively activated the JNK pathway ,not the p38 pathway in DADS-induced apoptosis in K562 cells .
