Department of Hepatobiliary Surgery, the First Affiliated Hospital of Gannan Medical University, Ganzhou, China.
Gannan Medical University, Ganzhou, China.
Turk J Gastroenterol. 2020 Mar;31(3):264-271. doi: 10.5152/tjg.2020.18800.
BACKGROUND/AIMS: To investigate the effect and the possible mechanism of lanthanum citrate on the proliferation and apoptosis of human hepatocellular carcinoma (HCC) cell line SMMC-7721 through the Hedgehog (Hh) signaling pathway.
Different concentrations of lanthanum citrate and KAAD-cyclopamine (the Hh signaling pathway representative inhibitor) were used to treat SMMC-7721 cells. Cell proliferation was detected using Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assays. Cell apoptosis was detected using flow cytometry analysis of Annexin V-FITC/ propidium iodide (PI). The protein expressions of regulatory genes, such as cell cycle protein D1 (CyclinD1), cyclin-dependent kinase inhibitor 1 (p21), cysteinyl aspartate specific proteinase 3 (Caspase-3), B-cell lymphoma-2 (Bcl-2), glioma-associated oncogene homolog 1 (Gli1), and sonic hedgehog (Shh) were quantified using Western blot assays. The mRNA expressions of Gli1 and Shh were tested using quantitative real-time polymerase chain reaction (qRT-PCR) assays and the protein expressions of Gli1 and Shh were determined using immunofluorescence assays.
The Annexin V-FITC and PI double staining results revealed that the 0.1 mM lanthanum citrate group and the 15 µM KAAD-cyclopamine group had both increased the apoptosis rate of SMMC-7721 cells. Both lanthanum citrate and KAAD-cyclopamine downregulated the protein expressions of CyclinD1, Bcl-2, Gli1, and Shh and upregulated the protein expressions of p21 and Caspase-3. Additionally, the immunofluorescence results revealed that the protein expressions of Gli1 and Shh were significantly decreased in both the lanthanum citrate group and the KAAD-cyclopamine group compared to the control group.
Lanthanum citrate inhibits proliferation and promotes apoptosis in HCC SMMC-7721 cells by suppressing the Hh signaling pathway.
背景/目的：通过 Hedgehog（Hh）信号通路研究柠檬酸镧对人肝癌细胞系 SMMC-7721 的增殖和凋亡的影响及其可能的机制。
用不同浓度的柠檬酸镧和 KAAD-环巴胺（Hh 信号通路代表性抑制剂）处理 SMMC-7721 细胞。用 MTT 法检测细胞增殖。用 Annexin V-FITC/PI 流式细胞术分析检测细胞凋亡。用 Western blot 法检测周期蛋白蛋白 D1（CyclinD1）、细胞周期蛋白依赖性激酶抑制剂 1（p21）、半胱天冬氨酸特异性蛋白酶 3（Caspase-3）、B 细胞淋巴瘤-2（Bcl-2）、神经胶质瘤相关癌基因同源物 1（Gli1）和 sonic hedgehog（Shh）等调节基因的蛋白表达。用 qRT-PCR 法检测 Gli1 和 Shh 的 mRNA 表达，用免疫荧光法检测 Gli1 和 Shh 的蛋白表达。
Annexin V-FITC/PI 双染结果显示，0.1mM 柠檬酸镧组和 15µM KAAD-环巴胺组均增加了 SMMC-7721 细胞的凋亡率。柠檬酸镧和 KAAD-环巴胺均下调了 CyclinD1、Bcl-2、Gli1 和 Shh 的蛋白表达，上调了 p21 和 Caspase-3 的蛋白表达。此外，免疫荧光结果显示，与对照组相比，柠檬酸镧组和 KAAD-环巴胺组 Gli1 和 Shh 的蛋白表达均显著降低。
柠檬酸镧通过抑制 Hh 信号通路抑制 HCC SMMC-7721 细胞的增殖并促进其凋亡。
