Department of Pediatric, The Second People's Hospital of Jingzhou, Jingzhou 434000, Hubei Province, China.
Department of Nursing, The Third People's Hospital of Jingzhou, Jingzhou 434000, Hubei Province, China.
Cell Mol Biol (Noisy-le-grand). 2023 Feb 28;69(2):157-161. doi: 10.14715/cmb/2023.69.2.26.
This study was performed to analyze the biological behavior of childhood leukemia cells regulated by miR-708 by binding to the 3' UTR end of the target gene and reducing the level of the target gene. In this regard, human leukemia Jurkat cell lines were selected and divided into a control group, miR-708 overexpression group and miR-708 inhibition group. MTT assay was used to detect the cell proliferation inhibition rate, flow cytometry was used to detect the apoptosis rate and cell cycle change, the scratch test was used to detect the cell migration capacity, and Western Blot assay was used to detect the expression of CNTFR, apoptosis and JAK/STAT pathway related proteins. To verify the binding site of miR-708 and target gene CNTFR. The results showed that the cell proliferation inhibition rate, apoptosis rate, G1 phase ratio, Bax protein, and CNTFR protein in the miR-708 overexpression group were significantly lower than those in the control group at each time point, while the S phase ratio, Bcl-2 protein, cell migration ability, JAK3 and STAT3 protein were significantly higher than those in the control group (P<0.05). The results of the miR-708 inhibition group were contrary to those of the miR-708 overexpression group. The binding sites of miR-708 and CNTFR were predicted by TargetScan bioinformatics software. It was found that there were two binding sites of miR-708 and CNTFR, 394-400 bp and 497-503 bp respectively. In conclusion, miR-708 can reduce the expression of CNTFR by binding to the target gene CNTFR3' UTR, activate the JAK/STAT pathway to regulate apoptosis-related proteins, reduce apoptosis, and enhance the migration ability of leukemia cells.
本研究通过分析 miR-708 与靶基因 3'UTR 末端结合,降低靶基因水平,从而调控儿童白血病细胞的生物学行为。为此,选择人白血病 Jurkat 细胞系,分为对照组、miR-708 过表达组和 miR-708 抑制组。采用 MTT 法检测细胞增殖抑制率,流式细胞术检测细胞凋亡率和细胞周期变化,划痕实验检测细胞迁移能力,Western blot 法检测 CNTFR、凋亡及 JAK/STAT 通路相关蛋白表达。验证 miR-708 与靶基因 CNTFR 的结合位点。结果显示,miR-708 过表达组各时间点细胞增殖抑制率、凋亡率、G1 期比例、Bax 蛋白、CNTFR 蛋白均明显低于对照组,而 S 期比例、Bcl-2 蛋白、细胞迁移能力、JAK3、STAT3 蛋白明显高于对照组(P<0.05)。miR-708 抑制组结果与 miR-708 过表达组相反。TargetScan 生物信息学软件预测 miR-708 与 CNTFR 的结合位点。发现 miR-708 与 CNTFR 有两个结合位点,分别为 394-400bp 和 497-503bp。结论:miR-708 可通过与靶基因 CNTFR3'UTR 结合,降低 CNTFR 表达,激活 JAK/STAT 通路,调节凋亡相关蛋白,减少细胞凋亡,增强白血病细胞迁移能力。