Wei Sixi, Wang Yating, Chai Qixiang, Fang Qin, Zhang Yaming, Wang Jishi
Department of Hematology, The Affiliated Hospital of Guiyang Medical College, Guiyang, Guizhou 550004, P.R. China ; Hematopoietic Stem Cell Transplantation Center of Guizhou Province, The Affiliated Hospital of Guiyang Medical College, Guiyang, Guizhou 550004, P.R. China ; Department of Clinical Biochemistry, The Affiliated Hospital of Guiyang Medical College, Guiyang, Guizhou 550004, P.R. China.
Department of Hematology, The Affiliated Hospital of Guiyang Medical College, Guiyang, Guizhou 550004, P.R. China ; Hematopoietic Stem Cell Transplantation Center of Guizhou Province, The Affiliated Hospital of Guiyang Medical College, Guiyang, Guizhou 550004, P.R. China.
Exp Ther Med. 2015 Mar;9(3):931-940. doi: 10.3892/etm.2015.2209. Epub 2015 Jan 23.
The aim of this study was to investigate the and effects of targeted heme oxygenase-1 (HO-1) silencing on the proliferation and apoptosis of human acute myelocytic leukemia (AML)-M2 cells. Bone marrow mononuclear cells (BMMNCs) were infected by pRNAi-siHO-1-GFP. The viability of the BMMNCs was determined by cell counting kit-8 (CCK-8) assay following daunorubicin (DNR) treatment. The apoptotic rate was detected by flow cytometry. The expression levels of HO-1 and apoptosis-related genes were detected by quantitative polymerase chain reaction (qPCR) and western blot analysis. An AML-M2 xenograft mouse model was established. The tumor formation outcomes and survival were observed. The leukocyte and platelet counts and hemoglobin levels were monitored, and the copy numbers of AML1/ETO fusion gene were detected by qPCR. pRNAi-siHO-1-GFP silenced the expression of HO-1. DNR inhibited cell viability in a time- and dose-dependent manner. The survival rate of the cells was significantly reduced by infection with pRNAi-siHO-1-GFP. HO-1 expression in the BMMNCs infected with pRNAi-siHO-1-GFP was downregulated, whereas caspase-3, -8 and -9 expression was upregulated compared with that in control BMMNCs. Kasumi-1 cells were successfully inoculated into nude mice. The rats inoculated with pRNAi-siHO-1-GFP-transfected Kasumi-1 cells succumbed to tumors more slowly and survived longer than those inoculated with untransfected Kasumi-1 cells. Furthermore, the leukocyte and platelet counts and hemoglobin levels were higher and the copy numbers of AML1/ETO fusion gene were lower in the former group. HO-1 gene silencing may promote the apoptosis of human M2 leukemic cells by inhibiting a caspase-dependent apoptotic pathway. Targeted silencing of HO-1 is able to inhibit the proliferation and infiltration of leukemic cells in nude mice and thus prolong their survival. The findings provide valuable experimental evidence for the molecular targeted therapy of M2 leukemia.
本研究旨在探讨靶向沉默血红素加氧酶-1（HO-1）对人急性髓细胞白血病（AML）-M2细胞增殖和凋亡的影响。用pRNAi-siHO-1-GFP感染骨髓单个核细胞（BMMNCs）。柔红霉素（DNR）处理后，通过细胞计数试剂盒-8（CCK-8）法测定BMMNCs的活力。采用流式细胞术检测凋亡率。通过定量聚合酶链反应（qPCR）和蛋白质印迹分析检测HO-1及凋亡相关基因的表达水平。建立AML-M2异种移植小鼠模型。观察肿瘤形成结果和生存情况。监测白细胞、血小板计数及血红蛋白水平，通过qPCR检测AML1/ETO融合基因的拷贝数。pRNAi-siHO-1-GFP沉默了HO-1的表达。DNR以时间和剂量依赖性方式抑制细胞活力。感染pRNAi-siHO-1-GFP可显著降低细胞存活率。与对照BMMNCs相比，感染pRNAi-siHO-1-GFP的BMMNCs中HO-1表达下调，而半胱天冬酶-3、-8和-9表达上调。Kasumi-1细胞成功接种到裸鼠体内。接种pRNAi-siHO-1-GFP转染的Kasumi-1细胞的大鼠比接种未转染Kasumi-1细胞的大鼠肿瘤形成更慢，存活时间更长。此外,前一组的白细胞、血小板计数及血红蛋白水平更高，AML1/ETO融合基因的拷贝数更低。HO-1基因沉默可能通过抑制半胱天冬酶依赖性凋亡途径促进人M2白血病细胞凋亡。靶向沉默HO-1能够抑制白血病细胞在裸鼠体内的增殖和浸润，从而延长其生存期。这些发现为M2白血病的分子靶向治疗提供了有价值的实验证据。
