Phytoestrogens are known to prevent tumor induction. But their molecular mechanisms of action are still unknown. This study aimed to examine the effect of apigenin on proliferation and apoptosis in HER2-expressing breast cancer cells. In our experiments, apigenin inhibited the proliferation of MCF-7 vec and MCF-7 HER2 cells. This growth inhibition was accompanied with an increase of sub G(0)/G(1) apoptotic fractions. Overexpression of HER2 did not confer resistance to apigenin in MCF-7 cells. Apigenin-induced extrinsic apoptosis pathway up-regulating the levels of cleaved caspase-8, and inducing the cleavage of poly (ADP-ribose) polymerase, whereas apigenin did not induce apoptosis via intrinsic mitochondrial apoptosis pathway since this compound did not decrease mitochondrial membrane potential maintaining red fluorescence and did not affect the levels of B-cell lymphoma 2 (BCL2) and Bcl-2-associated X protein. Moreover, apigenin reduced the tyrosine phosphorylation of HER2 (phospho-HER2 level) in MCF-7 HER2 cells, and up-regulated the levels of p53, phospho-p53 and p21 in MCF-7 vec and MCF-7 HER2 cells. This suggests that apigenin induces apoptosis through p53-dependent pathway. Apigenin also reduced the expression of phospho-JAK1 and phospho-STAT3 and decreased STAT3-dependent luciferase reporter gene activity in MCF-7 vec and MCF-7 HER2 cells. Apigenin decreased the phosphorylation level of IκBα in the cytosol, and abrogated the nuclear translocation of p65 within the nucleus suggesting that it blocks the activation of NFκB signaling pathway in MCF-7 vec and MCF-7 HER2 cells. Our study indicates that apigenin could be a potential useful compound to prevent or treat HER2-overexpressing breast cancer.

The potential benefits or the tissue-damaging effects of inflammatory response after central nervous system injuries have long been disputed. Recent studies have noted that substance P (SP), a neuropeptide, plays an important role in the wound-healing process by recruiting bone marrow stem cells to the injured tissue. In this study, we examined whether SP can enhance recovery from spinal cord injury (SCI) in Sprague-Dawley rats through its known function of stem cell mobilization and/or through the modulation of inflammation. We examined proinflammatory and anti-inflammatory cytokines and markers for macrophage subtypes. SP treatment modulated the SCI microenvironment toward a more anti-inflammatory and reparative one by inducing interleukin-10 and M2 macrophages and suppressing inducible nitric oxide synthase and tumor necrosis factor-α. This modulation was achieved at 1 day much earlier than SP-stimulated bone marrow stem cells' mobilization. Early intervention of the devastating inflammatory response by SP treatment caused the lesion cavity to become filled with robust axonal outgrowth that overlaid the M2 macrophages at 2 weeks--all of which culminated in tissue sparing and improvement in functional recovery from the SCI. SP is therefore a potential anti-inflammatory modulator for the treatment of injury-induced inflammatory central nervous system disorders.

Phytoestrogens have been demonstrated to inhibit tumor induction; however, their molecular mechanisms of action have remained elusive. The present study aimed to investigate the effects of a phytoestrogen, apigenin, on proliferation and apoptosis of the human epidermal growth factor receptor 2 (HER2)-expressing breast cancer cell line SKBR3. Proliferation assay, MTT assay, fluorescence-activated cell sorting analysis, western blot analysis, immunocytochemistry, reverse transcription-polymerase chain reaction and ELISA assay were used in the present study. The results of the present study indicated that apigenin inhibited the proliferation of SKBR3 cells in a dose-and time-dependent manner. This inhibition of growth was accompanied by an increase in the sub-G0/G1 apoptotic population. Furthermore, apigenin enhanced the expression levels of cleaved caspase-8 and -3, and induced the cleavage of poly(adenosine diphosphate ribose) polymerase in SKBR3 cells, confirming that apigenin promotes apoptosis via a caspase-dependent pathway. Apigenin additionally reduced the expression of phosphorylated (p)-janus kinase 2 and p-signal transducer and activator of transcription 3 (STAT3), inhibited CoCl2-induced vascular endothelial growth factor (VEGF) secretion and decreased the nuclear localization of STAT3. The STAT3 inhibitor S31-201 decreased the cellular proliferation rate and reduced the expression of p-STAT3 and VEGF. Therefore, these results suggested that apigenin induced apoptosis via the inhibition of STAT3 signaling in SKBR3 cells. In conclusion, the results of the present study indicated that apigenin may be a potentially useful compound for the prevention or treatment of HER2-overexpressing breast cancer.

Breast cancer is the most common cancer for women and is a major cause of mortality in women. Doxorubicin is a generally used chemotherapy drug for breast cancer. However, multidrug resistance of breast cancer interferes with the chemotherapy. We examined whether cucurbitacin D affects doxorubicin resistance of MCF7/ADR breast cancer cells. Cell viability was measured by MTT assay. Levels of p-STAT3, p-NF-κB, IκB, and caspases were measured by Western blot analysis. Nuclear staining of Stat3 and NF-κB was measured by immunocytochemistry. STAT3 and NF-κB transcriptional activity was detected by STAT3 and NF-κB luciferase reporter gene assays. Analysis of cell cycle arrest was performed by flow cytometry. Induction of apoptosis by cucurbitacin D was measured by Annexin V-FITC/propidium iodide assay. More than 90 % of MCF7/ADR cells lived upon treatment with doxorubicin for 24 h. However, upon treatment with cucurbitacin D, cell death was more than 60 %. Co-administration of cucurbitacin D and doxorubicin induced apoptosis, and G2/M cell cycle arrest, and inhibited upregulated Stat3 by doxorubicin on MCF7/ADR cells. Additionally, cucurbitacin D led to an increase in the IκBα level in the cytosol and a decrease in the p-NF-κB level in the nucleus. Finally, cucurbitacin D inhibited translocation of Stat3 and NF-κB and decreased transcriptional activity in the nucleus. Cucurbitacin D decreases cell proliferation and induces apoptosis by inhibiting Stat3 and NF-κB signaling in doxorubicin-resistant breast cancer cells. Cucurbitacin D could be used as a useful compound to treat adriamycin-resistant patients.