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黄芪-莪术组合通过调节上皮-间质转化抑制结肠癌HT-29细胞的增殖、迁移和侵袭

[Astragali Radix-Curcumae Rhizoma combination inhibits proliferation, migration, and invasion of colon cancer HT-29 cells by regulating EMT].

作者信息

Yang Qi, Sun Zheng, Zhu Yi-Miao, Xiang Dong-Yang, Zhang Qun-Yao, Wang Fang, Yang Gang, Yang Hao, Tang De-Cai, Wu Xiao-Yu

机构信息

the First School of Clinical Medicine, Nanjing University of Chinese Medicine Nanjing 210023, China.

Affiliated Hospital of Nanjing University of Chinese Medicine Nanjing 210004, China.

出版信息

Zhongguo Zhong Yao Za Zhi. 2023 Feb;48(3):736-743. doi: 10.19540/j.cnki.cjcmm.20221102.701.

Abstract

This study aims to investigate the effect of Astragali Radix-Curcumae Rhizoma(AC) combination on the proliferation, migration, and invasion of colon cancer HT-29 cells based on epithelial-mesenchymal transition(EMT). HT-29 cells were respectively treated with 0, 3, 6 and 12 g·kg(-1) AC-containing serum for 48 h. The survival and growth of cells were measured by thiazole blue(MTT) colorimetry, and the proliferation, migration, and invasion of cells were detected by 5-ethynyl-2'-deoxyuridine(EdU) test and Transwell assay. Cell apoptosis was examined by flow cytometry. The BALB/c nude mouse model of subcutaneous colon cancer xenograft was established, and then model mice were classified into blank control group, 6 g·kg(-1) AC group, and 12 g·kg~(-1) AC group. The tumor weight and volume of mice were recorded, and the histopathological morphology of the tumor was observed based on hematoxylin-eosin(HE) staining. The expression of apoptosis-associated proteins B-cell lymphoma-2-associated X protein(Bax), cysteine-aspartic acid protease-3(caspase-3), and cleaved caspase-3, and EMT-associated proteins E-cadherin, MMP9, MMP2 and vimentin in HT-29 cells and mouse tumor tissues after the treatment of AC was determined by Western blot. The results showed that cell survival rate and the number of cells at proliferation stage decreased compared with those in the blank control group. The number of migrating and invading cells reduced and the number of apoptotic cells increased in the administration groups compared with those in the blank control group. As for the in vivo experiment, compared with the blank control group, the administration groups had small tumors with low mass and shrinkage of cells and karyopycnosis in the tumor tissue, indicating that the AC combination may improve EMT. In addition, the expression of Bcl2 and E-cadherin increased and the expression of Bax, caspase-3, cleaved caspase-3, MMP9, MMP2, and vimentin decreased in HT-29 cells and tumor tissues in each administration group. In summary, the AC combination can significantly inhibit the proliferation, invasion, migration, and EMT of HT-29 cells in vivo and in vitro and promote the apoptosis of colon cancer cells.

摘要

本研究旨在基于上皮-间质转化(EMT)探讨黄芪-莪术(AC)药对结肠癌HT-29细胞增殖、迁移及侵袭的影响。将HT-29细胞分别用含0、3、6和12 g·kg⁻¹ AC的血清处理48 h。采用噻唑蓝(MTT)比色法检测细胞存活及生长情况,通过5-乙炔基-2'-脱氧尿苷(EdU)试验和Transwell试验检测细胞增殖、迁移及侵袭能力。采用流式细胞术检测细胞凋亡情况。建立皮下结肠癌异种移植BALB/c裸鼠模型,然后将模型小鼠分为空白对照组、6 g·kg⁻¹ AC组和12 g·kg⁻¹ AC组。记录小鼠肿瘤重量和体积,并基于苏木精-伊红(HE)染色观察肿瘤组织病理形态学变化。采用蛋白质免疫印迹法检测AC处理后HT-29细胞及小鼠肿瘤组织中凋亡相关蛋白B细胞淋巴瘤-2相关X蛋白(Bax)、半胱天冬酶-3(caspase-3)及裂解的caspase-3,以及EMT相关蛋白E-钙黏蛋白、基质金属蛋白酶9(MMP9)、基质金属蛋白酶2(MMP2)和波形蛋白的表达。结果显示,与空白对照组相比,各给药组细胞存活率及增殖期细胞数量降低。各给药组迁移及侵袭细胞数量减少,凋亡细胞数量增加。在体内实验中,与空白对照组相比,各给药组肿瘤体积小、质量轻,肿瘤组织细胞萎缩、核固缩,提示AC药对可能改善EMT。此外,各给药组HT-29细胞及肿瘤组织中Bcl2和E-钙黏蛋白表达增加,Bax、caspase-3、裂解的caspase-3、MMP9、MMP2和波形蛋白表达降低。综上所述,AC药对可显著抑制HT-29细胞体内外增殖、侵袭、迁移及EMT,并促进结肠癌细胞凋亡。

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