Apigenin is a low toxicity and non-mutagenic phytopolyphenol and protein kinase inhibitor. It exhibits anti-proliferating effects on human breast cancer cells. Here we examined several human breast cancer cell lines having different levels of HER2/neu expression and found that apigenin exhibited potent growth-inhibitory activity in HER2/neu-overexpressing breast cancer cells but was much less effective for those cells expressing basal levels of HER2/neu. Induction of apoptosis was also observed in HER2/neu-overexpressing breast cancer cells in a dose-and time-dependent manner. However, the one or more molecular mechanisms of apigenin-induced apoptosis in HER2/neu-overexpressing breast cancer cells remained to be elucidated. A cell survival pathway involving phosphatidylinositol 3-kinase (PI3K), and Akt is known to play an important role in inhibiting apoptosis in response to HER2/neu-overexpressing breast cancer cells, which prompted us to investigate whether this pathway plays a role in apigenininduced apoptosis in HER2/neu-overexpressing breast cancer cells. Our results showed that apigenin inhibits Akt function in tumor cells in a complex manner. First, apigenin directly inhibited the PI3K activity while indirectly inhibiting the Akt kinase activity. Second, inhibition of HER2/neu autophosphorylation and transphosphorylation resulting from depleting HER2/neu protein in vivo was also observed. In addition, apigenin inhibited Akt kinase activity by preventing the docking of PI3K to HER2/HER3 heterodimers. Therefore, we proposed that apigenin-induced cellular effects result from loss of HER2/neu and HER3 expression with subsequent inactivation of PI3K and AKT in cells that are dependent on this pathway for cell proliferation and inhibition of apoptosis. This implies that the inhibition of the HER2/ HER3 heterodimer function provided an especially effective strategy for blocking the HER2/neu-mediated transformation of breast cancer cells. Our results also demonstrated that apigenin dissociated the complex of HER2/neu and GRP94 that preceded the depletion of HER2/neu. Apigenin-induced degradation of mature HER2/neu involves polyubiquitination of HER2/neu and subsequent hydrolysis by the proteasome.Several cancers, including breast cancer, have a lower incidence in Asia than in Western countries (1). This may be attributed to the Asian dietary regimen rich in flavonoid-containing plants, which are thought to be anti-tumorigenic. Among the plant flavonoids, apigenin (4Ј,5,7,-trihydroxyflavone) is a chemopreventive compound (2, 3) and an inhibitor of certain signal transduction pathways (4). It has low toxicity, is non-mutagenic, and is widely distributed in many fruits and vegetables, including parsley, onions, oranges, tea, chamomile, wheat sprouts, and in some seasonings (5, 6). Apigenin has potential uses in cancer prevention and therapy (7), and it possesses growth inhibitory properties against many human cancer cell lines, including breast (8), colon (9), skin (10), thyroid (11), leukemia (12), and prostate car...

Leptin, the adipocyte-secreted hormone that centrally regulates weight control, is known to function as an immunomodulatory regulator. We investigated the signaling pathway involved in IL-6 production caused by leptin in microglia. Microglia expressed the long (OBRl) and short (OBRs) isoforms of the leptin receptor. Leptin caused concentration- and time-dependent increases in IL-6 production. Leptin-mediated IL-6 production was attenuated by OBRl receptor antisense oligonucleotide, PI3K inhibitor (Ly294002 and wortmannin), Akt inhibitor (1L-6-hydroxymethyl-chiro-inositol-2-((R)-2-O-methyl-3-O-octadecylcarbonate)), NF-κB inhibitor (pyrrolidine dithiocarbamate), IκB protease inhibitor (l-1-tosylamido-2-phenylenylethyl chloromethyl ketone), IκBα phosphorylation inhibitor (Bay 117082), or NF-κB inhibitor peptide. Transfection with insulin receptor substrate (IRS)-1 small-interference RNA or the dominant-negative mutant of p85 and Akt also inhibited the potentiating action of leptin. Stimulation of microglia with leptin activated IκB kinase α/IκB kinase β, IκBα phosphorylation, IκBα degradation, p65 phosphorylation at Ser276, p65 and p50 translocation from the cytosol to the nucleus, and κB-luciferase activity. Leptin-mediated an increase of IκB kinase α/IκB kinase β activity, κB-luciferase activity, and p65 and p50 binding to the NF-κB element was inhibited by wortmannin, Akt inhibitor, and IRS-1 small-interference RNA. The binding of p65 and p50 to the NF-κB elements, as well as the recruitment of p300 and the enhancement of histone H3 and H4 acetylation on the IL-6 promoter was enhanced by leptin. Our results suggest that leptin increased IL-6 production in microglia via the leptin receptor/IRS-1/PI3K/Akt/NF-κB and p300 signaling pathway.

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Nerve inflammation plays an important role in the development and progression of neuropathic pain after chronic constrictive injury (CCI). Recent studies have indicated that hypoxia-inducible factor 1α (HIF-1α) is crucial in inflammation. Low-level laser therapy has been used in treating musculoskeletal pain, but rare data directly support its use for neuropathic pain. We investigated the effects of low-level laser on the accumulation of HIF-1α, tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β) in controlling neuropathic pain, as well as on the activation of vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) in promoting functional recovery in a rat CCI model. CCI was induced by placing four loose ligatures around the sciatic nerve of rats. Treatments of low-level laser (660 nm, 9 J/cm(2)) or sham irradiation (0 J/cm(2)) were performed at the CCI sites for 7 consecutive days. The effects of laser in animals with CCI were determined by measuring the mechanical paw withdrawal threshold, as well as the sciatic, tibial, and peroneal function indices. Histopathological and immunoassay analyses were also performed. Low-level laser therapy significantly improved paw withdrawal threshold and the sciatic, tibial, and peroneal functional indices after CCI. The therapy also significantly reduced the overexpressions of HIF-1α, TNF-α, and IL-1β, and increased the amounts of VEGF, NGF, and S100 proteins. In conclusion, a low-level laser could modulate HIF-1α activity. Moreover, it may also be used as a novel and clinically applicable therapeutic approach for the improvement of tissue hypoxia/ischemia and inflammation in nerve entrapment neuropathy, as well as for the promotion of nerve regeneration. These findings might lead to a sufficient morphological and functional recovery of the peripheral nerve.