Dilber Yağmur, Çeker Hanife Tuğçe, Öztüzün Aleyna, Çırçırlı Bürke, Kırımlıoğlu Esma, Barut Zerrin, Aslan Mutay
Department of Medical Biochemistry, Faculty of Medicine, Akdeniz University, Antalya 07070, Turkey.
Department of Medical Biotechnology, Institute of Health Sciences, Akdeniz University, Antalya 07070, Turkey.
Pharmaceuticals (Basel). 2024 Nov 21;17(12):1564. doi: 10.3390/ph17121564.
The aim of this study was to determine the effect of Sparstolonin B (SsnB) on cell proliferation and apoptosis in human breast cancer (MCF-7) and human ovarian epithelial cancer (OVCAR-3) cell lines in the presence and absence of estradiol hemihydrate (ES). Phosphoinositol-3 kinase (PI3K), phosphorylated protein kinase B alpha (p-AKT), phosphorylated mTOR (mechanistic target of rapamycin) signaling proteins, and sphingomyelin/ceramide metabolites were also measured within the scope of the study. The anti-proliferative effects of SsnB therapy were evaluated over a range of times and concentrations. Cell proliferation was determined by measuring the Proliferating Cell Nuclear Antigen (PCNA). PCNA was quantified by ELISA and cell distribution was assessed by immunofluorescence microscopy. MTT analysis was used to test the vitality of the cells, while LC-MS/MS was used to analyze the amounts of ceramides (CERs), sphingosine-1-phosphate (S1P), and sphingomyelins (SMs). TUNEL labeling was used to assess apoptosis, while immunofluorescence staining and enzyme-linked immunosorbent assay (ELISA) were used to measure the levels of PI3K, p-AKT, and p-mTOR proteins. Sparstolonin B administration significantly decreased cell viability in MCF-7 and OVCAR-3 cells both in the presence and absence of ES, while it did not cause toxicity in healthy human fibroblasts. In comparison to controls, cancer cells treated with SsnB showed a significant drop in the levels of S1P, PI3K, p-AKT, and p-mTOR. In cancer cells cultured with SsnB, a significant increase in intracellular concentrations of C16-C24 CERs and apoptosis was observed. SsnB downregulated the levels of S1P, PI3K, p-AKT, and p-mTOR while reducing cell proliferation and promoting ceramide buildup and apoptosis.
本研究的目的是确定在存在和不存在半水合雌二醇（ES）的情况下，蛇床子素B（SsnB）对人乳腺癌（MCF-7）和人卵巢上皮癌（OVCAR-3）细胞系中细胞增殖和凋亡的影响。在本研究范围内，还测定了磷酸肌醇-3激酶（PI3K）、磷酸化蛋白激酶Bα（p-AKT）、磷酸化mTOR（雷帕霉素作用靶点）信号蛋白以及鞘磷脂/神经酰胺代谢物。在一系列时间和浓度范围内评估了SsnB治疗的抗增殖作用。通过测量增殖细胞核抗原（PCNA）来确定细胞增殖。通过酶联免疫吸附测定（ELISA）对PCNA进行定量，并通过免疫荧光显微镜评估细胞分布。MTT分析用于测试细胞活力，而液相色谱-串联质谱（LC-MS/MS）用于分析神经酰胺（CERs）、鞘氨醇-1-磷酸（S1P）和鞘磷脂（SMs）的含量。TUNEL标记用于评估细胞凋亡，而免疫荧光染色和酶联免疫吸附测定（ELISA）用于测量PI3K、p-AKT和p-mTOR蛋白的水平。无论有无ES，给予蛇床子素B均显著降低了MCF-7和OVCAR-3细胞的活力，而对健康人成纤维细胞未产生毒性。与对照组相比，用SsnB处理的癌细胞中S1P、PI3K、p-AKT和p-mTOR的水平显著下降。在用SsnB培养的癌细胞中，观察到C16 - C24 CERs的细胞内浓度显著增加以及细胞凋亡。蛇床子素B下调了S1P、PI3K、p-AKT和p-mTOR的水平，同时减少细胞增殖并促进神经酰胺积累和细胞凋亡。
