目的:探究尿石素A(urolithinA，UA)对三阴性乳腺癌细胞恶性生物学行为的影响以及对蛋白激酶B(protein kinase B，AKT)/雷帕霉素靶蛋白(mammalian target of rapamycin，mTOR)信号通路的调控机制。方法:采用CCK-8 法对人正常乳腺上皮细胞株MCF-10A 和三阴性乳腺癌细胞 MDA-MB-231 探索UA的安全性和毒性;将人三阴性乳腺癌细胞MDA-MB-231 随机分为MDA-MB-231 组、MDA-MB-231 细胞+尿石素A(UA)组、MDA-MB-231 细胞+AKT激动剂(SC79)组、MDA-MB-231 细胞+尿石素A+AKT激动剂(UA+SC79)组，采用平板克隆实验检测各组细胞的增殖活性;采用划痕实验和 Tr-answell实验检测各组细胞的迁移和侵袭能力;采用流式细胞术检测各组细胞的凋亡情况;采用 Western blot检测各组细胞中AKT/mTOR信号通路相关蛋白的活化水平。结果:确定7。5μmoL/L的UA既对正常乳腺细胞安全又对乳腺癌细胞有毒性，为本实验所用浓度。相比于 MDA-MB-231 组，UA 组克隆形成数量(105。67±9。18)、细胞迁移率(12。92±2。13)、侵袭细胞数(56。33±3。68)以及细胞内p-AKT/AKT(0。12±0。02)和p-mTOR/mTOR值(0。16±0。03)均降低(P＜0。05)，而细胞凋亡率(22。1±1。75)升高(P＜0。05);相比于MDA-MB-231 组，SC79 组克隆形成数量(222±17。57)、细胞迁移率(62。7±6。14)、侵袭细胞数(233。33±15。43)以及细胞内p-AKT/AKT(0。58±0。07)和p-mTOR/mTOR值(0。74±0。08)均升高(P＜0。05)，而细胞凋亡率(4。57±0。57)降低(P＜0。05);SC79 的加入逆转了 UA 对 MDA-MB-231 细胞恶性生物学行为的抑制作用(P＜0。05)。结论:UA能够通过抑制AKT/mTOR信号通路抑制三阴性乳腺癌细胞的增殖、迁移、侵袭和抗凋亡等恶性生物学行为。
Effect of Urolithin A on Malignant Biological Behavior of Triple-Negative Breast Cancer Cells via AKT/mTOR Signaling Pathway
Objective:To investigate the effect of urolithin A(UA)on the malignant biological behavior of triple-negative breast cancer(TNBC)cells and its regulatory mechanism on the protein kinase B(AKT)/mammalian target of rapamycin(mTOR)signaling pathway.Methods:The safety and toxicity of UA were as-sessed on human normal breast epithelial cell line MCF-10A and TNBC cell line MDA-MB-231 using the CCK-8 assay.MDA-MB-231 cells were randomly divided into the following groups:MDA-MB-231,MDA-MB-231+UA,MDA-MB-231+AKT activator(SC79),and MDA-MB-231+UA+AKT activator(UA+SC79).The proliferation activity of each group was detected by colony formation assay.Cell migration and in-vasion abilities were examined by wound healing and Transwell assays.Apoptosis was assessed by flow cytome-try.The activation levels of AKT/mTOR signaling pathway-related proteins were detected by Western blot.Results:UA at a concentration of 7.5μmoL/L was determined to be safe for normal breast cells and toxic to breast cancer cells.Compared to the MDA-MB-231 group,the UA group showed decreased colony formation(105.67±9.18),cell migration rate(12.92±2.13),invasion cell count(56.33±3.68),and p-AKT/AKT(0.12±0.02)and p-mTOR/mTOR(0.16±0.03)levels(P＜0.05),while the apoptosis rate(22.1±1.75)increased(P＜0.05).In contrast,the SC79 group showed increased colony formation(222±17.57),cell mi-gration rate(62.7±6.14),invasion cell count(233.33±15.43),and p-AKT/AKT(0.58±0.07)and p-mTOR/mTOR(0.74±0.08)levels(P＜0.05),and decreased apoptosis rate(4.57±0.57)(P＜0.05).The addition of SC79 reversed the inhibitory effects of UA on the malignant biological behavior of MDA-MB-231 cells(P＜0.05).Conclusion:UA can inhibit the proliferation,migration,invasion,and anti-apoptotic activ-ities of TNBC cells by suppressing the AKT/mTOR signaling pathway.
Urolithin AProtein kinase BMammalian target of rapamycinTriple-negative breast cancer
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