Wang Miao, Meng Jing-yan, He Su-fei
College of Traditional Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, 300193, China.
Chin J Integr Med. 2014 Oct;20(10):751-7. doi: 10.1007/s11655-014-1812-8. Epub 2014 May 6.
To investigate the antiproliferative and anti-metastasis effect of Xihuang Pill (, XP) on human colorectal cancer cell and to explore the molecular mechanism by which it produces the effects.
Highly metastatic human colorectal cancer cell line LoVo was treated with low-, medium-, and highdose XP-containing serum (XP-L, XP-M, XP-H) groups for 48 h, cells intervened with no drug rat serum and PD98059 [extracellular signal-regulated kinase (ERK) inhibitor] as negative and positive controls (NC and PC) groups. Cell proliferation assay was made using cell counting kit-8 (CCK8). The 8 μm pore-size transwell chamber and 4', 6-diamidino-2-phenylindole (DAPI) staining were applied to examine the ability of invasion and migration of the cells. The protein expression of ERK1/2, zinc fifi nger E-box-binding homeobox 1 (ZEB1), Scrib and lethal giant larvae homolog 2 (Lgl2) was detected by Western blotting while the relative mRNA quantity of E-cadherin, N-cadherin, Occludin and junctional adhesion molecule-1 (JAM1) was measured by realtime fluorescent quantitative polymerase chain reaction (RT-qPCR).
XP induced a dose-dependent suppression on the proliferation of LoVo cells (P <0.05 or P<0.01), with the inhibition rates varied from 27.30% to 31.08%. Transwell assay showed that when preprocessed with PD98059 and XP-containing serum, the number of cells that passed the filter decreased significantly compared with that of NC group (P <0.05 or P<0.01). Moreover, XP inhibited the protein expression of ERK1/2 and ZEB1 (P <0.05); and up-regulated the protein expression of Scrib and Lgl2 (P <0.05). The mRNA levels of E-cadherin, Occludin and JAM1 of the XP intervened groups and PC group markedly ascended (P <0.05) while that of N-cadherin showed a descending tendency (P>0.05).
XP intervention suppressed the ability of proliferation, invasion and migration of the LoVo cells. Regulating ZEB1-SCRIB Loop so as to recover epithelial phenotype and apical junctional complex might be one of the mechanisms by which XP produces the anti-metastasis effect.
探讨西黄丸（XP）对人结肠癌细胞的抗增殖和抗转移作用，并探究其作用的分子机制。
用含低、中、高剂量XP的血清（XP-L、XP-M、XP-H）处理高转移性人结肠癌细胞系LoVo 48小时，用无药大鼠血清和PD98059[细胞外信号调节激酶（ERK）抑制剂]干预细胞作为阴性和阳性对照（NC和PC）组。使用细胞计数试剂盒-8（CCK8）进行细胞增殖测定。应用8μm孔径的Transwell小室和4',6-二脒基-2-苯基吲哚（DAPI）染色检测细胞的侵袭和迁移能力。通过蛋白质印迹法检测ERK1/2、锌指E盒结合同源框1（ZEB1）、Scrib和致死巨幼虫同源物2（Lgl2）的蛋白表达，同时通过实时荧光定量聚合酶链反应（RT-qPCR）测定E-钙黏蛋白、N-钙黏蛋白、闭合蛋白和连接黏附分子-1（JAM1）的相对mRNA量。
XP对LoVo细胞的增殖具有剂量依赖性抑制作用（P<0.05或P<0.01），抑制率在27.30%至31.08%之间。Transwell实验表明，用PD98059和含XP血清预处理后，穿过滤膜的细胞数量与NC组相比显著减少（P<0.05或P<0.01）。此外，XP抑制ERK1/2和ZEB1的蛋白表达（P<0.05）；并上调Scrib和Lgl2的蛋白表达（P<0.05）。XP干预组和PC组的E-钙黏蛋白、闭合蛋白和JAM1的mRNA水平明显升高（P<0.05），而N-钙黏蛋白的mRNA水平呈下降趋势（P>0.05）。
XP干预抑制了LoVo细胞的增殖、侵袭和迁移能力。调节ZEB1-SCRIB环路以恢复上皮表型和顶端连接复合体可能是XP产生抗转移作用的机制之一。
