Zheng Li, Wu Yu, Wang Fu, Shi Hui, Xu Bo, Yang Aihua
Pharmaceutical Department, Affiliated Maternity and Child Health Care Hospital of Nantong University, Nantong, Jiangsu 226018, China.
Pharmaceutical Department, Nantong Hospital of traditional Chinese Medicine, Nantong, Jiangsu 226001, China.
J Oncol. 2022 May 14;2022:1138851. doi: 10.1155/2022/1138851. eCollection 2022.
To study the pharmacological activity and the mechanism of action of natural compounds derived from 6,7-dimethoxycoumarin on the differentiation of human chronic myeloid leukemia K562 cells.
We use MTT assay (Sigma-Aldrich, USA) to detect cell viability; use flow cytometry to analyze DNA content for cell cycle analysis; use benzidine staining to synthesize hemoglobin to determine K562 cell differentiation; use western blot analysis and qPCR to detect the expression levels of FOX03, P27, CDK4, and their phosphorylation; and use the AOBS laser scanning confocal system (Leica, Wetzlar, Germany) to analyze and quantify the number of positive green spots. The statistical methods used are one-way analysis of variance (ANOVA) and Dunnett's test to analyze within and between groups.
In order to explore the effect of 6,7-dimethoxycoumarin on the differentiation of K562 cell erythrocytes, it was concluded that 6,7-dimethoxycoumarin promotes the differentiation of K562 cell erythrocytes; the proliferation of K562 cells was detected by MTT method, and the results showed that 6,7-dimethoxycoumarin can inhibit the proliferation of K562 cells; to evaluate the effect of 6,7-dimethoxycoumarin on the proliferation of K562 cells, the results showed that 6,7-dimethoxycoumarin increased the expression of FOXO3, P27, CDK4, and CDK65, and decreased the phosphorylation of CDK4 and CDK6 proteins. To further explore the effect of knocking out FOXO3 on cell differentiation, the results show that 6,7-dimethoxycoumarin can reduce the differentiation and proliferation of K562 cells by increasing the expression of FOXO3.
This study extended the understanding of the pharmacological activity of 6,7-dimethoxycoumarin and may provide a potential new target for the treatment of chronic myelogenous leukemia. However, we still need to further study the specific molecular capabilities of 6.7 dimethylcoumarin to understand their possible capture mechanism.
研究6,7 - 二甲氧基香豆素衍生的天然化合物对人慢性髓性白血病K562细胞分化的药理活性及作用机制。
我们使用MTT法（美国西格玛奥德里奇公司）检测细胞活力；使用流式细胞术分析DNA含量以进行细胞周期分析；使用联苯胺染色法合成血红蛋白以确定K562细胞分化；使用蛋白质免疫印迹分析和定量聚合酶链反应检测FOX03、P27、CDK4及其磷酸化的表达水平；并使用AOBS激光扫描共聚焦系统（德国韦茨拉尔徕卡公司）分析和量化绿色阳性斑点的数量。所采用的统计方法是单因素方差分析（ANOVA）和Dunnett检验以分析组内和组间情况。
为探究6,7 - 二甲氧基香豆素对K562细胞向红细胞分化的影响得出，6,7 - 二甲氧基香豆素促进K562细胞向红细胞分化；采用MTT法检测K562细胞的增殖情况结果显示，6,7 - 二甲氧基香豆素可抑制K562细胞增殖；评估6,7 - 二甲氧基香豆素对K562细胞增殖的影响结果显示，6,7 - 二甲氧基香豆素增加了FOXO3、P27、CDK4和CDK65蛋白表达，降低了CDK4和CDK6蛋白的磷酸化水平。为进一步探究敲除FOXO3对细胞分化的影响，结果表明6,7 - 二甲氧基香豆素可通过增加FOXO3表达降低K562细胞的分化和增殖能力。
本研究拓展了对6,7 - 二甲氧基香豆素药理活性的认识，可能为慢性粒细胞白血病的治疗提供一个潜在的新靶点。然而，我们仍需进一步研究6,7 - 二甲基香豆素的具体分子功能，以了解其可能的捕获机制。
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