Pomegranate (Punica granatum L.) seed oil (PGO) contains more than 70% cis(c)9,trans(t)11,c13-18:3 as conjugated linolenic acids (CLN). Our previous short-term experiment demonstrated that seed oil from bitter melon (Momordica charantia) (BMO), which is rich in c9,t11,t13-CLN, inhibited the occurrence of colonic aberrant crypt foci (ACF) induced by azoxymethane (AOM). In this study, we investigated the effect of dietary PGO on the development of AOM-induced colonic malignancies and compared it with that of conjugated linoleic acid (CLA). To induce colonic tumors, 6-week old male F344 rats were given subcutaneous injections of AOM (20 mg/kg body weight) once a week for 2 weeks. One week before the AOM treatment they were started on diet containing 0.01%, 0.1%, or 1% PGO or 1% CLA for 32 weeks. Upon termination of the bioassay (32 weeks) colon tumors were evaluated histopathologically. AOM exposure produced colonic adenocarcinoma with an incidence of 81% and multiplicity of 1.88 ± ± ± ±1.54 at week 32. Administration of PGO in the diet significantly inhibited the incidence (AOM + + + +0.01% PGO, 44%, P < < < <0.05; AOM + + + +0.1% PGO, 38%, P < < < <0.01; AOM + + + +1% PGO, 56%) and the multiplicity (AOM + + + +0.01% PGO, 0.56 ± ± ± ±0.73, P < < < <0.01; AOM + + + +0.1% PGO, 0.50 ± ± ± ±0.73, P < < < <0.005; AOM + + + +1% PGO, 0.88 ± ± ± ±0.96, P < < < <0.05) of colonic adenocarcinomas, although a clear dose-response relationship was not observed at these dose levels. CLA feeding also slightly, but not significantly, reduced the incidence and multiplicity of colonic adenocarcinomas. The inhibition of colonic tumors by PGO was associated with an increased content of CLA (c9,t11-18:2) in the lipid fraction of colonic mucosa and liver. Also, administration of PGO in the diet elevated expression of peroxisome proliferator-activated receptor ( olon cancer is one of the leading causes of cancer deaths in Western countries. Globally, more than 875,000 men and women were afflicted with this cancer in 1996 and more than 510,000 died in the same year.

DMT1 (divalent metal transporter 1) is the main iron importer found in animals, and ferrous iron is taken up by cells via DMT1. Once ferrous iron reaches the cytosol, it is subjected to subcellular distribution and delivered to various sites where iron is required for a variety of biochemical reactions in the cell. Until now, the mechanism connecting the transporter and cytosolic distribution had not been clarified. In the present study, we have identified PCBP2 [poly(rC)-binding protein 2] as a DMT1-binding protein. The N-terminal cytoplasmic region of DMT1 is the binding domain for PCBP2. An interaction between DMT1 and PCBP1, which is known to be a paralogue of PCBP2, could not be demonstrated in vivo or in vitro. Iron uptake and subsequent ferritin expression were suppressed by either DMT1 or PCBP2 knockdown. Iron-associated DMT1 could interact with PCBP2 in vitro, whereas iron-chelated DMT1 could not. These results indicate that ferrous iron imported by DMT1 is transferred directly to PCBP2. Moreover, we demonstrated that PCBP2 could bind to ferroportin, which exports ferrous iron out of the cell. These findings suggest that PCBP2 can transfer ferrous iron from DMT1 to the appropriate intracellular sites or ferroportin and could function as an iron chaperone.

Embryonic stem (ES) cells can differentiate into many cell types and are expected to be useful for tissue engineering. Recent reports have shown that ES cells can differentiate into insulin-producing cells in response to the transient expression of the pdx-1 gene, after the removal of feeder cells. To investigate the lineage of insulin-producing cells and their in vitro differentiation, we introduced the ␤geo gene, encoding a ␤-galactosidase-neomycin phosphotransferase fusion protein under the control of the mouse insulin 2 promoter, into ES cells that had been adapted to feederfree culture, and analyzed insulin gene expression during their in vitro differentiation. We also examined the expression of transcription factors that are related to the differentiation of the pancreas. X-gal staining analysis revealed ␤-galactosidase-positive cells on the surface and in the center of the embryoid body that proliferated during differentiation. Glucose-responsive insulin-producing cells, derived from our feeder-free ES cells, expressed insulin 2, pdx-1, Pax4, and Isl1 and also the glucagon, somatostatin, and PP genes. Moreover, the genes encoding p48, amylase, and carboxypeptidase A were also expressed. These results suggest that ES cells can differentiate not only into endocrine cells but also into exocrine cells of the pancreas, without the initiation of pdx-1 expression. Diabetes 52:1163-1168, 2003 M ouse embryonic stem (ES) cells are continuous cell lines derived directly from the fetal founder tissue of the preimplantation embryo. They can be expanded in culture while retaining the functional attributes of pluripotent early embryo cells. Their capacity for multilineage differentiation is reproduced in culture, as ES cells can differentiate into a wide range of well-defined cell types. Cell transplantation to restore tissue function after disease or injury is in theory applicable to a huge variety of human diseases. Thus, the use of lineage-restricted differentiation techniques developed for ES cells will promote future cell therapy. Moreover, an in vitro ES cell differentiation system will be valuable for research into early embryonic cell lineage. We and other groups have shown that ES cells can differentiate into the endoderm cell lineage (1-5). Recently, several groups have reported that ES cells can be induced to differentiate into insulin-producing cells These reports indicate that ES cells can differentiate into insulin-producing cells, but the cell lineage of these insulin-producing cells is still unknown. Indeed, insulin gene expression has been observed not only in the pancreas but also in the yolk sac and embryonic brain in vivo (6,7). To assess the cell lineage of insulin-producing cells during their differentiation from ES cells, we have developed ES cell lines carrying a lacZ-neomycin phosphotransferase fusion gene (␤geo) under the mouse insulin 2 gene promoter and analyzed the lacZ expression pattern of clusters of differentiating ES cells. Moreover, we have examined the expression of a series of transc...

Zerumbone (ZER), present in subtropical ginger Zingiber zerumbet Smith, possesses anti-growth and anti-inflammatory properties in several human cancer cell lines. ZER also down-regulates the cyclooxygenase-2 and inducible nitric oxide synthase expression via modulation of nuclear factor (NF)-jB activation in cell culture systems. These findings led us to investigate whether ZER is able to inhibit carcinogenesis in the colon and lung, using 2 different preclinical mouse models. In Exp. 1, a total of 85 male ICR mice were initiated using a single intraperitoneal (i.p.) injection with azoxymethane (AOM, 10 mg/kg bw) and promoted by 1.5% dextran sulfate sodium (DSS) in drinking water for 7 days for rapid induction of colonic neoplasms. Animals were then fed the diet containing 100, 250 or 500 ppm ZER for 17 weeks. In Exp. 2, a total of 50 female A/J mice were given a single i.p. injection of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (10 lmol/mouse) to induce lung proliferative lesions. They were then fed the diet mixed with 100, 250 or 500 ppm ZER for 21 weeks. At the termination of the experiments (wk 20 of Exp. 1 and wk 22 of Exp. 2), all animals were subjected to complete necropsy examination to determine the pathological lesions in both tissues. Oral administration of ZER at 100, 250 and 500 ppm significantly inhibited the multiplicity of colonic adenocarcinomas. The treatment also suppressed colonic inflammation. In the lung carcinogenesis, ZER feeding at 250 and 500 ppm significantly inhibited the multiplicity of lung adenomas in a dose-dependent manner. Feeding with ZER resulted in inhibition of proliferation, induction of apoptosis, and suppression of NFjB and heme oxygenase (HO)-1 expression in tumors developed in both tissues. Our findings suggest that dietary administration of ZER effectively suppresses mouse colon and lung carcinogenesis through multiple modulatory mechanisms of growth, apoptosis, inflammation and expression of NFjB and HO-1 that are involved in carcinogenesis in the colon and lung. ' 2008 Wiley-Liss, Inc.Key words: zerumbone; colon; lung; carcinogenesis; chemoprevention A sesquiterpenoid, zerumbone (ZER), is a major constituent of the subtropical ginger plant Zingiber zerumbet Smith. The essential oil of the rhizomes contains large amount of ZER and is used as an anti-inflammatory medicine. 1 Recent studies revealed several biological properties of ZER that may be responsible for inhibition of carcinognesis. They include suppression of skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced Epstein-Barr virus activation in Raji cells, 1 inhibition of free radical generation, inhibition of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2 expression, inhibition of tumor necrosis factor (TNF)-a-release in activated leukocytes and induction of apoptosis in human colonic adenocarcinoma (ADC) cell lines. 2 In vivo studies demonstrated that dietary feeding with ZER markedly suppressed dextran sulfate sodium (DSS)-induced acute colitis in mice 3 and a putat...

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