Department of Orthopedics, Affiliated Hospital of Inner Mongolia University for The Nationalities, Tongliao, Inner Mongolia 028007, P.R. China.
Mol Med Rep. 2020 Nov;22(5):3962-3968. doi: 10.3892/mmr.2020.11471. Epub 2020 Sep 1.
Previous studies have demonstrated that calycosin is a natural phytoestrogen with a similar structure to estrogen, which can inhibit cell proliferation and induce apoptosis in a variety of tumors. Calycosin exerts potential pharmacological effects on osteosarcoma cells by inducing apoptosis. The aim of the present study was to elucidate the specific molecular mechanism of calycosin‑induced apoptosis in osteosarcoma cells. Cell proliferation was determined by an MTT assay. Annexin V/PI and JC‑1 staining were used to detect apoptosis and mitochondrial dysfunction, respectively, by flow cytometry. Western blot analysis was used to detect the expression of caspases or mitochondrial proteins. The results revealed that calycosin reduced the cell viability of human osteosarcoma 143B cells, induced apoptosis and increased the loss of mitochondrial membrane potential (MMP). In addition, calycosin increased the expression of the proapoptotic antiapoptotic proteins cleaved caspase‑3, cleaved caspase‑9, cleaved poly(ADP‑ribose) polymerase and Bcl‑2‑associated X protein (Bax), and decreased the expression of the antiapoptotic proapoptotic protein B‑cell lymphoma‑2 (Bcl‑2), thus altering the Bax/Bcl‑2 ratio. In addition, the expression levels of cytochrome c were markedly decreased in the mitochondria and increased in the cytoplasm following calycosin treatment. Furthermore, calycosin treatment induced p38‑mitogen‑activated protein kinase (MAPK) phosphorylation, whereas the p38‑MAPK inhibitor BIRB 796 markedly reversed cell viability, apoptosis and loss of MMP in 143B cells. These results suggested that calycosin inhibited osteosarcoma 143B cell growth via p38‑MAPK regulation of mitochondrial‑dependent intrinsic apoptotic pathways.
先前的研究表明，毛蕊异黄酮是一种与雌激素结构相似的天然植物雌激素，它可以抑制多种肿瘤的细胞增殖并诱导细胞凋亡。毛蕊异黄酮通过诱导细胞凋亡对骨肉瘤细胞发挥潜在的药理作用。本研究旨在阐明毛蕊异黄酮诱导骨肉瘤细胞凋亡的具体分子机制。通过 MTT 测定法测定细胞增殖。通过流式细胞术分别用 Annexin V/PI 和 JC-1 染色检测细胞凋亡和线粒体功能障碍。通过 Western blot 分析检测半胱天冬酶或线粒体蛋白的表达。结果表明，毛蕊异黄酮降低人骨肉瘤 143B 细胞的细胞活力，诱导细胞凋亡并增加线粒体膜电位（MMP）的丧失。此外，毛蕊异黄酮增加了促凋亡蛋白 cleaved caspase-3、cleaved caspase-9、cleaved poly(ADP-ribose) polymerase 和 Bcl-2 相关 X 蛋白（Bax）的表达，降低了抗凋亡蛋白 B 细胞淋巴瘤-2（Bcl-2）的表达，从而改变了 Bax/Bcl-2 比值。此外，毛蕊异黄酮处理后，线粒体中细胞色素 c 的表达水平明显降低，细胞质中细胞色素 c 的表达水平增加。此外，毛蕊异黄酮处理诱导 p38-丝裂原激活的蛋白激酶（MAPK）磷酸化，而 p38-MAPK 抑制剂 BIRB 796 显著逆转了 143B 细胞的细胞活力、细胞凋亡和 MMP 丧失。这些结果表明，毛蕊异黄酮通过 p38-MAPK 调节线粒体依赖性内在凋亡途径抑制骨肉瘤 143B 细胞生长。
