Bao Shijie, He Dongmei, Zeng Jinrong, Zhang Yanrong, Chen Shengting
Shenzhen Hospital, Southern Medical University, Shenzhen 518110, China.
Departmemt of Hematology, The First Affiliated Hospital, Jinan University, Guangzhou 510290, China.
Ann Transl Med. 2019 Jun;7(12):260. doi: 10.21037/atm.2019.05.32.
This study aimed to investigate the effects of microRNA19a (miR-19a) antisense oligonucleotide (ASODN) on the proliferation and apoptosis of acute myeloid leukemia cells (HL60).
In experiment 1, HL60 cells were divided into the blank control group, the blank transfection group, the scrambled oligonucleotide (SODN) group and the ASODN group. MiR-19a ASODN and SODN were independently transferred into HL60 cells. The miR-19a expression was detected by real-time quantitative RT-PCR (qRT-PCR) after 48-h and 72-h transfection; CCK8 assay was used to detect the proliferation inhibition rate at 48 and 72 h; Hoechst 33258 staining was performed to examine apoptotic cells at 48 h; the apoptosis rate was detected by flow cytometry after AnnexinV/PI staining at 48 and 72 h; the protein expression of E2F1 and Bim was detected by Western blotting at 48 h. In experiment 2, cells were divided into the Ara-C group, the SODN + Ara-C group and the ASODN + Ara-C group. The cell proliferation inhibition rate at 48 and 72 h and apoptosis rate at 72 h were assessed as mentioned above.
MiR-19a expression in the miR-19a ASODN group was lower than in the SODN group and the blank control group (P<0.05). MiR-19a ASODN significantly inhibited the growth of HL60 cells (P<0.05) and increased their apoptosis, and the apoptosis rate peaked at 48 h. The protein expression of E2F1 and Bim in the ASODN group was higher than in the blank control group, blank transfection group and SODN group. In addition, Ara-C further inhibited the growth and induced the apoptosis of miR-19a ASODN-transfected cells (P<0.05) in a time dependent manner. The growth inhibition rate and apoptosis rate in the ASODN + Ara-C group were higher than the sum of those in both Ara-C group and ASODN group.
miRNA-19a ASODN can inhibit the proliferation and induce apoptosis of HL60 cells and may exert synergistic effects with Ara-C on HL60 cells.
本研究旨在探讨微小RNA19a（miR-19a）反义寡核苷酸（ASODN）对急性髓系白血病细胞（HL60）增殖和凋亡的影响。
实验1中，HL60细胞分为空白对照组、空白转染组、乱序寡核苷酸（SODN）组和ASODN组。将miR-19a ASODN和SODN分别转染至HL60细胞。转染48小时和72小时后，采用实时定量逆转录聚合酶链反应（qRT-PCR）检测miR-19a表达；采用CCK8法检测48小时和72小时的增殖抑制率；48小时时进行Hoechst 33258染色检测凋亡细胞；48小时和72小时时，采用膜联蛋白V/碘化丙啶（AnnexinV/PI）染色后通过流式细胞术检测凋亡率；48小时时采用蛋白质印迹法检测E2F1和Bim的蛋白表达。实验2中，细胞分为阿糖胞苷（Ara-C）组、SODN+Ara-C组和ASODN+Ara-C组。如上述方法评估48小时和72小时时的细胞增殖抑制率以及72小时时的凋亡率。
miR-19a ASODN组中miR-19a表达低于SODN组和空白对照组（P<0.05）。miR-19a ASODN显著抑制HL60细胞生长（P<0.05）并增加其凋亡，凋亡率在48小时达到峰值。ASODN组中E2F1和Bim的蛋白表达高于空白对照组、空白转染组和SODN组。此外，Ara-C以时间依赖性方式进一步抑制miR-19a ASODN转染细胞的生长并诱导其凋亡（P<0.05）。ASODN+Ara-C组的生长抑制率和凋亡率高于Ara-C组和ASODN组两者之和。
miRNA-19a ASODN可抑制HL60细胞增殖并诱导其凋亡，且可能与Ara-C对HL60细胞发挥协同作用。
