摘要
目的探讨复方苦参注射液(CKI)影响HL-60细胞迁移的机制。方法选取HL-60细胞株作为研究对象,根据不同加药方案分为粒细胞集落刺激因子(G-CSF)组(40ng/mL)、CKI组(0.5ng/mL)、G-CSF和CKI联合加药组和空白对照组,共培养48h后采用蛋白质印迹法(WB)和逆转录聚合酶链反应(RT-PCR)分别检测微小RNA-146a(miR-146a)、趋化因子CXC受体4(CXCR4)等信使RNA(mRNA)、CXCR4蛋白表达变化情况;同时采用Transwell小室检测CKI、G-CSF作用前后细胞迁移率变化情况。结果 (1)与空白对照组相比,基质细胞衍生因子-1α(SDF-1α)能促进HL-60细胞的迁移[(2.01±0.13)比(1.05±0.10),P<0.05],而CKI组、G-CSF组、CKI联合GCSF组均能阻断HL-60细胞向SDF-1α迁移[(1.50±0.04)、(1.68±0.08)、(1.27±0.02)比(2.01±0.13),P<0.05]。(2)与空白对照组相比,CKI、G-CSF处理HL60细胞后,CXCR4的mRNA、蛋白表达均受抑[(0.75±0.10)、(0.47±0.09)比(1.00±0.08),P<0.05];与CKI组、G-CSF组相比,CIK联合GCSF组CXCR4的mRNA蛋白表达受抑更明显[(0.24±0.04)比(0.47±0.09)、(0.75±0.10),P<0.05]。(3)与空白对照组比较,G-CSF、CKI处理HL-60细胞后,miR-146a表达显著上调[(5.59±0.46)、(3.53±0.39)比(1.03±0.11),P<0.01];而联合加药组上调更明显[(6.76±0.54)比(5.59±0.46)、(3.53±0.39),P<0.05]。结论 CKI能协同G-CSF上调miR-146a表达,通过降低HL-60细胞表面的CXCR4表达而阻断CXCR4/SDF-1α信号轴,最终导致HL-60细胞体外迁移能力明显下降。
Objective To explore the mechanism of compound Kushen(Sophora flavescens) root injection (CKI) affecting the migration of HL-60 cells.Methods HL-60 cell line was used and divided into 4 groups,positive control group(G-CSF,40ng/mL),experimental group(CKI,0.5ng/mL),G-CSF and CKI combination group,and blank control group,according to different dosage regimen.After 48 hours of co-culture,Western blot and RT-PCR were used to detect the mRNA/protein expression of miR-146a,and CXCR4.Transwell chamber assay was performed to check the migrative activity ofHL-60.Results (1)Compared to the blank control group,SDF-1α(stromal cell-derived factor 1) promoted the migration of HL-60 cells[(2.01±0.13) vs (1.05±0.10),P<0.05),whilethose of CKI,G-CSF,CKI combined with G-CSF blocked the cell migration[(1.50±0.04),(1.68±0.08),and (1.27±0.02) vs (2.01±0.13),P<0.05,respectively].(2)Compared to the blank control group,after the treatment of CKI and G-CSF,the expressions of CXCR4 mRNA and protein were inhibited[(0.75±0.10) and (0.47±0.09) vs (1.00±0.08),P<0.05,respectively];And the difference was more significant in the G-CSF and CKI combination groupcompared tothose in the single drug treatment group[(0.24±0.04) vs (0.47±0.09) and (0.75±0.10),P<0.05,respectively].(3) Compared to the blank control group,after the treatment of G-CSF and CKI,the expression of miR-146a was significantly up-regulated[(5.59±0.46) and (3.53±0.39) vs (1.03±0.11),P<0.01,respectively];And the difference was more significant in the G-CSF and CKI combination group[(6.76±0.54) vs (5.59±0.46) and (3.53±0.39),P<0.05,respectively].Conclusion CKI could co-ordinate with G-CSF to up-regulate the expression of miR-146a,and block the CXCR4/SDF-1α signaling axis by reducing the expression of CXCR4 on the surface of HL-60 cells,which ultimately leads to the decrease of the migrative activity of HL-60 in vitro.
作者
盛贤福
王珺
庄海峰
张宇
陈均法
SHENG Xianfu;WANG Jun;ZHUANG Haifeng;ZHANG Yu;CHEN Junfa(Department of Hematology, The First Affiliated Hospital of Zhejiang University of Traditional Chinese Medicine, Hangzhou, Zhejiang province 310006, China)
出处
《浙江中西医结合杂志》
2019年第1期6-9,共4页
Zhejiang Journal of Integrated Traditional Chinese and Western Medicine
基金
浙江省中医药科技计划(No.2017ZB030)