Pseudolaric acid B induces apoptosis, senescence, and mitotic arrest in human breast cancer MCF-7

Abstract

Aim: The aim of the present study was to investigate the inhibitory effect of pseudolaric acid B (PAB) on human breast cancer MCF-7 cells. Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analysis, morphological changes, acridine orange staining, and agarose gel electrophoresis were applied to detect apoptosis. The percentage of apoptotic and necrotic cells was calculated by the lactate dehydrogenase activity-based cytotoxicity assay; senescence associated (SA)-β-galactosidase activity was detected to evaluate senescence; flow cytometric analysis of propidium iodide staining was carried out to investigate the distribution of cell cycle, and the protein expression was examined by Western blot analysis. Results: During apoptosis, the half maximal inhibitory concentration IC 50 was 3.4 and 1.35 µmol/L at 36 and 48 h after PAB treatment, respectively. The MCF-7 cells exposed to PAB showed typical characteristics of apoptosis, including the morphological changes and DNA fragmentation. The MCF-7 cells treated with 4 µmol/L PAB for 36 h underwent apoptosis, but not necrosis. The apoptosis induced by PAB was independent of the death receptor pathway. The senescent cells became larger and flatter, and the SA-β-galactosidase staining was positive. PAB induced obvious mitotic arrest and it preceded apoptosis and senescence. The expressions of p21 and p53 was upregulated with PAB treatment, and cyclin B1 was upregulated and transported from the cytoplasm to nuclei, and sustained stable levels. Conclusion: PAB induced mitotic arrest in the MCF-7 cells and inhibited proliferation through apoptosis and senescence. The apoptosis was independent of the death receptor pathway.

Pseudolaric acid B activates autophagy in MCF-7 human breast cancer cells to prevent cell death

“…The senescence detection kit was used according to the manufacturer's instructions, as previously described (6). Finally, the cells were observed under a phase contrast microscope (Olympus IX51; Olympus Corporation, Tokyo, Japan) for the development of a blue color.…”

“…DMSO-treated cells were used as a control. PAB doses and incubation times were selected based on a previous study (6).…”

“…Surgery, medication and radiation may be used in the treatment of breast cancer; thus it typically has a favorable prognosis (4,5). Our previous study demonstrated that treatment of MCF-7 human breast cancer cells with PAB led to the induction of mitotic arrest and apoptosis in the majority of cells, while the surviving cells became senescent (6); however, the mechanism by which these cells were directed towards either cell death or survival has not yet been determined.…”

Role of pseudolaric acid B in A549 lung cancer cell proliferation and apoptosis

“…PAB is a diterpene acid isolated from the root and trunk bark of Pseudolarix kaempferi and possesses multiple biological and pharmacological activities, including antifungal, antimicrobial, antifertility and antiangiogenic properties (10)(11)(12)(13)(14)(15). It has been previously confirmed that PAB exhibits an antitumor effect by inducing apoptosis in various types of cancer (16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27). Apoptosis is important in homeostasis maintenance through a balance between cell proliferation and cell death (28).…”

“…In MDA-MB-468 human breast cancer cells, PAB inhibits cell growth by downregulating the hypoxia inducible factor-1α protein (15). In MCF-7 human breast cancer cells, PAB induced apoptosis by activating JNK, inactivating ERK and upregulating P21 and P53 protein expression (21,22). In murine fibrosarcoma L929 cells, PAB promotes proliferation, inhibits apoptosis and arrests the cell cycle at the G2/M phase by downregulating the Bcl-2 and CDC2 proteins (23,24).…”