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富含亮氨酸α-2 糖蛋白 1 基因干扰白血病 KASUMI-1 细胞凋亡的调控。

Leucine-Rich Alpha-2-Glycoprotein1 Gene Interferes with Regulation of Apoptosis in Leukemia KASUMI-1 Cells.

机构信息

Department of Hematology, Guizhou Provincial People's Hospital, Guiyang, Guizhou, China (mainland).

出版信息

Med Sci Monit. 2018 Nov 20;24:8348-8356. doi: 10.12659/MSM.911249.

Abstract

BACKGROUND Leukemia cells have strong proliferation and anti-apoptosis capabilities. The purpose of this study was to investigate the effect of silencing the leucine-rich alpha-2-glycoprotein1 (LRG1) gene, which was found to regulate tumor proliferation and apoptosis in acute myeloid leukemia (AML) cell lines. MATERIAL AND METHODS Plasmid interference technique was used to silence the LRG1 gene in the KASUMI-1 cell line. The cell counting kit-8 (CCK-8) assay was used to test the effect of transduction on cell viability. Cell cycle and apoptosis were detected by flow cytometry. Western blot and quantitative real-time polymerase chain reaction (RT-qPCR) were applied to detect the expression levels of proteins and mRNA, respectively. RESULTS KASUMI-1 cells with the CD34⁺CD38⁻ phenotype were sorted by flow cytometry. After transfection of the siLRG1 plasmid, the level of LRG1 expression was downregulated and cell viability was reduced. Silencing of LRG1 gene blocked KASUMI-1 cells in G0/G1 phase and promoted apoptosis. Further experiments found that LRG1 gene silencing significantly downregulated cell cycle-associated proteins and anti-apoptotic proteins, while upregulating pro-apoptotic proteins. Downregulation of LRG1 gene expression also inhibits signal transduction of the JAK-STAT pathway. CONCLUSIONS LRG1 gene silencing regulates the expression of cyclin and apoptosis-related proteins to reduce cell viability and promote apoptosis, probably through inhibition of the JAK-STAT pathway.

摘要

背景

白血病细胞具有较强的增殖和抗凋亡能力。本研究旨在探讨沉默富含亮氨酸的α-2-糖蛋白 1(LRG1)基因对急性髓系白血病(AML)细胞系肿瘤增殖和凋亡的调节作用。

材料和方法

采用质粒干扰技术沉默 KASUMI-1 细胞系中的 LRG1 基因。通过细胞计数试剂盒-8(CCK-8)检测转导对细胞活力的影响。通过流式细胞术检测细胞周期和凋亡。Western blot 和定量实时聚合酶链反应(RT-qPCR)分别用于检测蛋白和 mRNA 的表达水平。

结果

通过流式细胞术分选具有 CD34 ⁺ CD38 ⁻ 表型的 KASUMI-1 细胞。转染 siLRG1 质粒后,LRG1 表达水平下调,细胞活力降低。LRG1 基因沉默阻断 KASUMI-1 细胞进入 G0/G1 期并促进细胞凋亡。进一步的实验发现,LRG1 基因沉默显著下调细胞周期相关蛋白和抗凋亡蛋白,同时上调促凋亡蛋白。LRG1 基因表达下调也抑制了 JAK-STAT 通路的信号转导。

结论

沉默 LRG1 基因通过调节细胞周期和凋亡相关蛋白的表达来降低细胞活力并促进凋亡,可能通过抑制 JAK-STAT 通路。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b7ef/6256841/8cc8787b7c90/medscimonit-24-8348-g001.jpg

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