Baicalein overcomes tumor necrosis factor-related apoptosis-inducing ligand resistance via two different cell-specific pathways in cancer cells but not in normal cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is one of the most promising candidates for new cancer therapeutics. A current problem is that some cancers still remain resistant to TRAIL. We show for the first time that a naturally occurring flavonoid, baicalein, overcomes TRAIL resistance in cancer cells. The combination of baicalein and TRAIL effectively induced apoptosis in TRAIL-resistant colon cancer SW480 cells. Baicalein up-regulated the expression of death receptor 5 (DR5) among TRAIL receptors at the mRNA and protein levels. Suppression of this up-regulation with small interfering RNA (siRNA) efficiently reduced the apoptosis induced by TRAIL and baicalein, suggesting that the sensitization was mediated through DR5 induction. Moreover, baicalein also overcame TRAIL resistance with DR5 up-regulation in prostate cancer PC3 cells. Of note, the combination of TRAIL and baicalein hardly induced apoptosis in normal human cells, such as blood cells and hepatocytes. Baicalein increased DR5 promoter activity, and this enhanced activity was diminished by mutation of a CCAAT/enhancer-binding protein homologous protein (CHOP)-binding site in SW480 cells. In SW480 cells, CHOP siRNA blocked both functions of baicalein. CHOP expression was induced by baicalein in SW480 cells; however, in PC3 cells, baicalein scarcely induced CHOP and mutation of the CHOP-binding site did not abrogate the DR5 promoter activation by baicalein. Interestingly, baicalein induced reactive oxygen species (ROS) and a ROS scavenger prevented DR5 expression and TRAIL sensitization in PC3 but not SW480 cells. These results indicate that, using two different pathways, baicalein exposes cancer surveillance of TRAIL and overcomes TRAIL resistance in cancer cells.

Kaempferol sensitizes colon cancer cells to TRAIL-induced apoptosis.

Kaempferol is a natural compound contained in edible plants, and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anti-cancer agent. Here, we show for the first time that the combined treatment with kaempferol and TRAIL drastically induced apoptosis in human colon cancer SW480 cells, compared to single treatments. Kaempferol markedly up-regulated TRAIL receptors, DR5 and DR4. DR5 but not DR4 siRNA efficiently blocked apoptosis induced by the co-treatment with kaempferol and TRAIL, indicating that DR5 up-regulation by kaempferol helps to enhance TRAIL actions. Moreover, we examined the combined effect on normal human cells. The co-treatment induced no apoptosis in normal human peripheral blood mononuclear cells and little apoptosis in normal human hepatocytes. These results suggest that kaempferol is useful for TRAIL-based treatments for cancer.

Sesamin, a lignan of sesame, down-regulates cyclin D1 protein expression in human tumor cells.

Sesamin is a major lignan constituent of sesame and possesses multiple functions such as antihypertensive, cholesterol-lowering, lipid-lowering and anticancer activities. Several groups have previously reported that sesamin induces growth inhibition in human cancer cells. However, the nature of this growth inhibitory mechanism remains unknown. The authors here report that sesamin induces growth arrest at the G1 phase in cell cycle progression in the human breast cancer cell line MCF-7. Furthermore, sesamin dephosphorylates tumor-suppressor retinoblastoma protein (RB). It is also shown that inhibition of MCF-7 cell proliferation by sesamin is correlated with down-regulated cyclin D1 protein expression, a proto-oncogene that is overexpressed in many human cancer cells. It was found that sesamin-induced down-regulation of cyclin D1 was inhibited by proteasome inhibitors, suggesting that sesamin suppresses cyclin D1 protein expression by promoting proteasome degradation of cyclin D1 protein. Sesamin down-regulates cyclin D1 protein expression in various kinds of human tumor cells, including lung cancer, transformed renal cells, immortalized keratinocyte, melanoma and osteosarcoma. Furthermore, depletion of cyclin D1 protein using small interfering RNA rendered MCF-7 cells insensitive to the growth inhibitory effects of sesamin, implicating that cyclin D1 is at least partially related to the antiproliferative effects of sesamin. Taken together, these results suggest that the ability of sesamin to down-regulate cyclin D1 protein expression through the activation of proteasome degradation could be one of the mechanisms of the antiproliferative activity of this agent.

Effects of second-generation histamine H1 receptor antagonists on the active avoidance response in rats.

1. The aim of the present study was to establish a new schedule of active avoidance response in rats to estimate the central effects of second-generation histamine H1 receptor antagonists. 2. With the new schedule, a rat was placed into a dark room. A sliding door was opened after a delay of 5 s and, unless the animal moved into the lit room, an electric shock was delivered for 3 s. With the conventional schedule, the sliding door was opened immediately after the rat was placed into the dark room. 3. Ketotifen, at a dose of 50 mg/kg, showed no significant effect on the retrieval of active avoidance response with the conventional schedule. However, with the new schedule, the drug caused significant inhibition of retrieval of the response, even at a dose of 10 mg/kg. 4. Epinastine showed no significant effect on retrieval of the active avoidance response, even at a dose of 50 mg/kg with the new schedule. 5. Cetirizine, at a dose of 50 mg/kg, caused a significant effect, indicating that cetirizine, at this dose, markedly inhibits memory retrieval. 6. Both olopatadine and loratadine had potent effects; at doses of 20 and 50 mg/kg, respectively, these agents showed significant inhibitory effects on retrieval of the response. 7. In conclusion, we have developed a new schedule of active avoidance response that can be used to estimate the central effects of second-generation histamine H1 receptor antagonists.

Effects of NMDA antagonist MK-801 on radial maze performance in Zucker rats.

We examined the participation of the NMDA receptor in the histaminergic system using radial maze performance in Zucker rats. Although pyrilamine caused a significant effect on spatial memory deficit in lean rats, no significant spatial memory deficit was observed in obese rats. On the other hand, MK-801 caused significant spatial memory deficits in obese rats in comparison with lean rats. These results indicate that the histaminergic neuron is not closely related with the radial maze performance in obese rats. In addition, the potent effect of MK-801 observed in obese rats compared with lean rats may be responsible for the activation of NMDA receptors in obese rats.

Low density detergent-insoluble membrane of Xenopus eggs: subcellular microdomain for tyrosine kinase signaling in fertilization.

Protein-tyrosine phosphorylation plays an important role in egg activation signaling at fertilization. We show that in Xenopus, fertilization stimulates a rapid and transient tyrosine phosphorylation of egg proteins, as revealed by immunoblotting with anti-phosphotyrosine antibody. Immunofluorescent microscopic analysis demonstrated that the phosphorylation occurs in cortical area of the egg animal hemisphere. To further characterize subcellular compartment for fertilization-dependent tyrosine kinase signaling, we isolated low density detergent-insoluble membrane (LD-DIM) fraction from Xenopus eggs. The egg LD-DIM was enriched in cholesterol and GM1 ganglioside. It also contained signaling molecules such as Xyk (Xenopus egg Src), Gq alpha, Ras, integrin beta 1 and CD9. Fertilization stimulated tyrosine phosphorylation of Xyk and some other LD-DIM proteins. Remarkably, sperm stimulated tyrosine phosphorylation of the LD-DIM proteins in vitro. The sperm-dependent phosphorylation was sensitive to the tyrosine kinase inhibitors PP2 and genistein. We found that pretreatment of eggs with methyl-beta-cyclodextrin, a cholesterol-binding substance, led to a decrease in cholesterol, Xyk and sperm-induced tyrosine phosphorylation in LD-DIM. In methyl-beta-cyclodextrin-treated eggs, sperm-induced Ca(2+) transient and first cell division were also inhibited. These findings suggest that the egg LD-DIM might serve as subcellular microdomain for tyrosine kinase signaling in Xenopus egg fertilization.