Multidrug resistance protein 1 (MRP1) and P-glycoprotein, which are ATP-dependent multidrug efflux pumps and involved in multidrug resistance of tumor cells, are members of the ATP binding cassette proteins and contain two nucleotide-binding folds (NBFs). P-glycoprotein hydrolyzes ATP at both NBFs, and vanadateinduced nucleotide trapping occurs at both NBFs. We examined vanadate-induced nucleotide trapping in MRP1 stably expressed in KB cell membrane by using 8-azido-[␣-32 P]ATP. Vanadate-induced nucleotide trapping in MRP1 was found to be stimulated by reduced glutathione, glutathione disulfide, and etoposide and to be synergistically stimulated by the presence of etoposide and either glutathione. These results suggest that glutathione and etoposide interact with MRP1 at different sites and that those bindings cooperatively stimulate the nucleotide trapping. Mild trypsin digestion of MRP1 revealed that vanadate-induced nucleotide trapping mainly occurs at NBF2. Our results suggest that the two NBFs of MRP1 might be functionally nonequivalent.Multidrug resistance of tumor cells is a major obstacle to cancer chemotherapy. This phenomenon is frequently associated with the expression of P-glycoprotein and multidrug resistance protein 1 (MRP1), 1 both of which are ATP binding cassette (ABC) proteins. P-glycoprotein and MRP1 function as ATP-dependent efflux pumps that extrude cytotoxic drugs from the cells before they reach their intracellular targets, thus conferring resistance to many structurally dissimilar anticancer drugs, such as the Vinca alkaloids, colchicine, actinomycin D, etoposide, taxol, and anthracyclines (1-5). However, the mechanism of transport for MRP1 could be different from that for P-glycoprotein, because the depletion of intracellular glutathione (GSH) by buthionine sulfoximine results in a complete reversal of resistance to anticancer drugs of some cell lines expressing MRP1 (6, 7), but buthionine sulfoximine has no effect on P-glycoprotein-mediated multidrug resistance. It has been reported that MRP1 transports GSH-S-conjugates such as leukotriene C 4 , glutathione disulfide (GSSG), and 2,4-dinitrophenyl-S-glutathione (8 -11) and that MRP1 mediates ATP-dependent transport of vincristine, daunorubicin, and etoposide in the presence of . From these findings, it has been postulated that MRP1 can actively cotransport GSH and unmodified xenobiotics as well as GSH-S-conjugates.We have reported that MRP1 in membrane from a human MRP1 cDNA transformant can be specifically photoaffinity labeled with 8-azido-[␣-32 P]ATP by vanadate-induced nucleotide trapping (15). Vanadate and Mg 2ϩ were required for trapping of nucleotide, and photoaffinity labeling was inhibited by the excess ADP as well as ATP. These results have suggested that a stable inhibitory complex MRP1⅐MgADP⅐Vi, an analog of the MRP1⅐MgADP⅐P i transition state complex, is formed in the presence of vanadate, as suggested for P-glycoprotein (16). Vanadate-induced nucleotide trapping in P-glycoprotein has been reported to be stimulated by the t...

In the intrinsic pathway of apoptosis, mitochondria play a crucial role by releasing cytochrome c from the intermembrane space into the cytoplasm. Cytochrome c release through Bax/ Bak-dependent channels in mitochondria has been well documented. In contrast, cyclophilin D (CypD), an important component of permeability transition pore-dependent protein release, remains largely undefined, and no apoptogenic proteins that act specifically in a CypD-dependent manner have been reported to date. Here, we describe a novel and evolutionarily conserved protein, apoptogenic protein (Apop). Mouse Apop-1 expression induces apoptotic death by releasing cytochrome c from mitochondria into the cytosolic space followed by activation of caspase-9 and -3. Apop-1-induced apoptosis is not blocked by Bcl-2 or Bcl-x L , inhibitors of Bax/Bak-dependent channels, whereas it is completely blocked by cyclosporin A, an inhibitor of permeability transition pore. Cells lacking CypD were resistant to Apop-induced apoptosis. Moreover, inhibition of Apop expression prevented the cell death induced by apoptosis-inducing substances. Our findings, thus, indicate that the expression of Apop-1 induces apoptosis though CypD-dependent pathway and that Apop-1 plays roles in cell death under physiological conditions.

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According to a 1999 epidemiological survey by the Fatigue Study Group of the Ministry of Health, Labor and Welfare in Japan, 60% (47 million) of workers complained of fatigue. However, there have been no established recommendations for fatigue, and few anti-fatigue medicines or foods have been developed. Physical or mental load induces oxidative stress, resulting in fatigue.Accordingly, antioxidants are candidate anti-fatigue substances. Imidazole dipeptides (carnosine and anserine) have strong antioxidative effects, and chicken breast is rich in such dipeptides. The results of two human studies suggested imidazole dipeptides were effective and useful in attenuating fatigue induced by physical loads and daily activity. In short, imidazole dipeptides are novel anti-fatigue ingredients.

In the present study, we investigated the effects of plasma cluster ions on moisturizing skin and increasing comfort sensation in healthy female volunteers. Thirty-two healthy women who often experienced dry skin participated in a randomized crossover trial. We evaluated skin moisture using the Corneometer CM825 and comfort sensation using the visual analogue scale. Plasma cluster ion concentrations of 25000 pcs/cm 3 increased skin moisture, while concentrations of 7000 pcs/cm 3 increased comfort sensation. These results suggest that plasma cluster ions have an effect on skin moisture contents and comfort sensation.