1)成都中医药大学附属医院药剂科,成都 610072;2)北京康仁堂药业有限公司中药配方颗粒关键技术国家地方联合工程研究中心,北京 101301
四川省科技厅基础项目(2019YJ0614)资助。
1)Pharmacy Department, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu 610072, China;2)National and Local Joint Engineering Research Center for Key Technology of Chinese Medicinal Composition Granules, Beijing Tcmages Pharmaceutical Co., Ltd., Beijing 101301, China
This work was supported by a grant from Basic Project of Sichuan Provincial Department of Science and Technology(2019YJ0614).
目的 白扁豆总皂苷是中药白扁豆经过提取分离纯化步骤制备得到,关于白扁豆的总皂苷成分如何影响前列腺癌细胞系PC-3细胞的生长情况缺少研究。因此,有必要探讨白扁豆总皂苷对前列腺癌细胞系PC-3细胞生长的机制研究。方法 本研究采用CCK8方法检测不同浓度的白扁豆总皂苷对前列腺癌细胞系PC-3细胞生长的影响。利用转录组学分析白扁豆总皂苷抑制前列腺癌细胞系PC-3细胞生长的分子机制,并且进一步通过实时定量PCR(qRT-PCR)和蛋白质免疫印迹(Western blot)实验对相关差异基因的表达进行验证。利用Western blot和CCK8检测白扁豆总皂苷处理过表达醛脱氢酶7家族成员A1(ALDH7A1)的PC-3细胞存活率。结果 随着白扁豆总皂苷浓度升高,前列腺癌细胞PC-3的存活率显著下降,白扁豆总皂苷的IC50值为1 086 mg/L。转录组学测序结果显示,与对照组相比,白扁豆总皂苷处理的细胞中有2 360个差异表达基因,其中1 982个基因上调,378个基因下调。基因功能注释(GO)结果显示,差异表达基因显著富集到与有丝分裂纺锤体检查点(mitotic spindle checkpoint)、纺锤体组装检查点(spindle assembly checkpoint)等一系列跟癌症的发生发展密切相关的生物学过程。此外,基因组京都百科全书(KEGG)分析结果也显示,差异表达基因富集在肿瘤代谢等信号通路。进一步对其中的差异基因进行验证,结果显示,与对照组相比,白扁豆总皂苷处理的前列腺癌细胞中ALDH7A1、甘氨酸C-乙酰转移酶(GCAT)和磷酸甘油酸变位酶家族成员4(PGAM4)的蛋白质表达水平明显降低(P<0.05),而二甲基甘氨酸脱氢酶(DMGDH)和胱硫醚β合成酶样(CBSL)的蛋白质表达水平显著升高(P<0.001)。体外细胞实验结果表明,白扁豆总皂苷通过下调前列腺癌细胞中ALDH7A1的表达抑制PC-3细胞生长。结论 白扁豆总皂苷可能通过下调ALDH7A1表达从而在体外抑制前列腺癌细胞的生长。
Objective The total saponins of Baibiandou (Lablab Semen Album) are prepared from the traditional Chinese medicine Lablab Semen Album through extraction, separation and purification steps. Although there are studies on the targeting and treatment of cancer with Xiaobiandou (Lens culinaris) lectin in nude mice with human liver cancer transplants, there is a lack of research on how the total saponins of Lablab Semen Album affect the growth of the prostate cancer cell line PC-3. Therefore, it is necessary to explore the mechanism through which total saponins of Lablab Semen Album impact the growth of the prostate cancer cell line PC-3.Methods In this paper, the CCK8 method was adopted to test the effect of different concentrations of Lablab Semen Album total saponins on the growth of PC-3 cells. Transcriptomics was used to analyze the molecular mechanism of the inhibition of the growth of PC-3 cells by Lablab Semen Album total sponins, and the related differential gene expression was verified by real-time quantitative PCR and Western blot experiments. Western blot and CCK8 assays were used to detect the survival rate of total saponins in Lablab Semen Album-treated PC-3 cells overexpressing ALDH7A1.Results With increasing concentration of Lablab Semen Album total saponins, the survival rate of PC-3 cells was reduced significantly, and the IC50 value of Lablab Semen Album total saponins was 1 086 mg/L. The transcriptomics sequencing results showed that compared to the control group, there were 2 360 differentially expressed genes in the cells treated with Lablab Semen Album total saponins, of which 1 982 genes were upregulated and 378 genes were downregulated. The GO results showed that the differentially expressed genes were significantly enriched in biological processes closely related to cancer onset and development. In addition, the KEGG analysis results also showed that the differentially expressed genes were enriched in the signaling pathway of tumor metabolism. When the differential genes were further verified, the results showed that compared to the control group, the protein expression levels of ALDH7A1, GCAT and PGAM4 in the prostate cancer cells treated with Lablab Semen Album total saponins were obviously reduced (P<0.05), whereas the protein expression levels of DMGDH and CBSL were significantly increased (P<0.001). According to the results of in vitro cell experiments, the downregulated expression of ALDH7A1 in prostate cancer cells by total saponins of Lablab Semen Album inhibited their growth.Conclusion Lablab Semen Album total saponins may inhibit the growth of prostate cancer cells in vitro through the expression of ALDH7A1.
袁强华,韩君.基于转录组学的白扁豆总皂苷抑制前列腺癌细胞系PC-3细胞生长的机制研究[J].生物化学与生物物理进展,2022,49(8):1597-1606
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