Akira Murakami

Sunrouge is an anthocyanin-rich, new tea cultivar that contains similar levels of catechins as Yabukita, the most popular tea cultivar consumed in Japan. Interestingly, Sunrouge preparations have previously been shown to have more pronounced acetylcholinesterase inhibitory and anticolitis activities than those of Yabukita. In this study, we examined their effects on expressions of self-defensive molecules, including heat shock proteins (HSPs), which are molecular chaperones involved in homeostasis and longevity. Hot water extract from freeze-dried Sunrouge significantly upregulated messenger RNA (mRNA) expressions of HSP40, HSP70, and HSP32 (heme oxygenase-1), with grades greater than those shown by Yabukita. Oral administration of freeze-dried preparation of Sunrouge to male ICR mice at a dose of 1% in the basal diet for 1 month resulted in marked upregulations of several HSP mRNA expressions in mucosa from the gastrointestinal tract, especially the upper small intestine. Again, its efficacy was remarkably higher than that of Yabukita. Moreover, exposure of Caenorhabditis elegans to Sunrouge conferred thermoresistant phenotype, and also resulted in a significant life-span elongation. Taken together, our results suggest that Sunrouge is a unique and promising tea cultivar for regulating self-defense systems.

Accumulated evidence shows that some phytochemicals provide beneficial effects for human health. Recently, a number of mechanistic studies have revealed that direct interactions between phytochemicals and functional proteins play significant roles in exhibiting their bioactivities. However, their binding selectivities to biological molecules are considered to be lower due to their small and simple structures. In this study, we found that zerumbone, a bioactive sesquiterpene, binds to numerous proteins with little selectivity. Similar to heat-denatured proteins, zerumbone-modified proteins were recognized by heat shock protein 90, a constitutive molecular chaperone, leading to heat shock factor 1-dependent heat shock protein induction in hepa1c1c7 mouse hepatoma cells. Furthermore, oral administration of this phytochemical up-regulated heat shock protein expressions in the livers of Sprague-Dawley rats. Interestingly, pretreatment with zerumbone conferred a thermoresistant phenotype to hepa1c1c7 cells as well as to the nematode Caenorhabditis elegans. It is also important to note that several phytochemicals with higher hydrophobicity or electrophilicity, including phenethyl isothiocyanate and curcumin, markedly induced heat shock proteins, whereas most of the tested nutrients did not. These results suggest that non-specific protein modifications by xenobiotic phytochemicals cause mild proteostress, thereby inducing heat shock response and leading to potentiation of protein quality control systems. We considered these bioactivities to be xenohormesis, an adaptation mechanism against xenobiotic chemical stresses. Heat shock response by phytochemicals may be a fundamental mechanism underlying their various bioactivities.

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Curcumin (CUR), a yellow pigment in turmeric, has marked potential for preventing colon cancer. We recently reported that ar‐turmerone (ATM) suppressed nitric oxide (NO) generation in macrophages. In the present study, we explored the molecular mechanisms by which ATM attenuates NO generation and examined the anti‐carcinogenesis activity of turmerones (TUR, a mixture of 5 sesquiterpenes including ATM). Both CUR and ATM inhibited lipopolysaccharide (LPS)‐induced expression of inducible forms of both nitric oxide synthase and cyclooxygenase (iNOS and COX‐2, respectively). A chase experiment using actinomycin D revealed that ATM accelerated the decay of iNOS and COX‐2 mRNA, suggesting a post‐transcriptional mechanism. ATM prevented LPS‐induced translocation of HuR, an AU‐rich element‐binding protein that determines mRNA stability of certain inflammatory genes. In a colitis model, oral administration of TUR significantly suppressed 2% dextran sulfate sodium (DSS)‐induced shortening of the large bowel by 52–58%. We also evaluated the chemopreventive effects of oral feeding of TUR, CUR, and their combinations using a model of dimethylhydradine‐initiated and DSS‐promoted mouse colon carcinogenesis. At the low dose, TUR markedly suppressed adenoma multiplicity by 73%, while CUR at both doses suppressed adenocarcinoma multiplicity by 63–69%. Interestingly, the combination of CUR and TUR at both low and high doses abolished tumor formation. Collectively, our results led to our hypothesis that TUR is a novel candidate for colon cancer prevention. Furthermore, we consider that its use in combination with CUR may become a powerful method for prevention of inflammation‐associated colon carcinogenesis. © 2012 BioFactors, 2013

Sunrouge, an anthocyanin‐rich tea, has similar levels of catechins as “Yabukita,” the most popular green tea cultivar consumed in Japan. Green tea polyphenols (GTPs) have attracted interest due to their potent antioxidative activities combined with a lack of side effects in humans at normal consumption levels. However, we previously reported that high doses (0.5 and 1%) of dietary GTPs can result in deterioration of colitis and failed to prevent colon carcinogenesis in inflamed colons. In the present study, we determined the inhibitory effects of Sunrouge on colitis in dextran sodium sulfate (DSS)‐treated and untreated control mice. Five‐week‐old female ICR mice were administered a single dose of Yabukita or Sunrouge (extracts in 1 mL distilled water) via a stomach tube for 3 weeks. After 1 week of treatment, the mice were divided into four groups (two Yabukita and two Sunrouge groups) and given drinking water with or without 3% DSS for 2 weeks, then they were euthanized. Those treated with DSS developed watery diarrhea and bloody stools, and showed body weight loss, spleen hypertrophy, and shortening of the colon, as well as deteriorations in survival rate, liver function, colon mucosal interleukin‐1β level and expression of phase II detoxification enzyme mRNA. Sunrouge improved these DSS‐induced symptoms, at least in part, whereas Yabukita showed either no effect or adverse effects in regard to some those parameters. It is suggested that the differences between Yabukita and Sunrouge on DSS‐induced colitis might be due to the high levels of anthocyanins found in Sunrouge tea. © 2012 International Union of Biochemistry and Molecular Biology, Inc.

Phytochemicals are generally defined as secondary metabolites in plants that play crucial roles in their adaptation to a variety of environmental stressors. There is a great body of compelling evidence showing that these metabolites have pronounced potentials for regulating and modulating human health and disease onset, as shown by both experimental and epidemiological approaches. Concurrently, enormous efforts have been made to elucidate the mechanism of actions underlying their biological and physiological functions. For example, the pioneering work of Tachibana et al. uncovered the receptor for (−)-epigallocatechin-3-gallate (EGCg) as the 67 kDa laminin receptor, which was shown to partially mediate the functions of EGCg, such as anti-inflammatory, anti-allergic, and anti-proliferative activities. Thereafter, several protein kinases were identified as binding proteins of flavonoids, including myricetin, quercetin, and kaempferol. Isothiocyanates, sulfur-containing phytochemicals present in cruciferous plants, are well known to target Keap1 for activating the transcription factor Nrf2 for inducing self-defensive and anti-oxidative gene expression. In addition, we recently identified CD36 as a cell surface receptor for ursolic acid, a triterpenoid ubiquitously occurring in plants. Importantly, the above mentioned target proteins are indispensable for phytochemicals to exhibit, at least in part, their bioactivities. Nevertheless, it is reasonable to assume that some of the activities and potential toxicities of metabolites are exerted via their interactions with unidentified, off-target proteins. This notion may be supported by the fact that even rationally designed drugs occasionally display off-target effects and induce unexpected outcomes, including toxicity. Here we update the current status and future directions of research related to target molecules of food phytochemicals.

Phenethyl isothiocyanate (PEITC), a constituent of many cruciferous vegetables, is well known to have versatile physiological activities, including chemopreventive effects. On the other hand, its anti‐inflammatory effects are poorly reported. Nitric oxide (NO) is associated with a wide variety of inflammatory diseases. In this study, we investigated the effects of PEITC on NO production in LPS‐activated peritoneal macrophages from ICR mice. The signaling pathway of LPS‐induced NO production was examined using neutralizing antibodies [anti‐interferon (IFN)‐γ and anti‐interleukin (IL‐12)] and specific protein kinase inhibitors, as well as others. The activity of PEITC toward NOx production was assessed in mice that received LPS via intraperitoneal administration. The neutralizing antibody of anti‐IFN‐γ, but not anti‐IL‐12, suppressed LPS‐induced NO production by 90%. LY294002, a specific inhibitor of phosphoinositide‐3‐kinase, suppressed Akt and IFN‐γ mRNA expression up‐regulated by LPS, whereas PEITC exhibited a similar inhibition profile. Furthermore, oral administration of PEITC significantly suppressed the serum concentration of NOx in ICR mice. Our results suggest that PEITC suppresses LPS‐induced NO production via inhibition of Akt activation and the resultant decrease in expression of IFN‐γ. This is one of the first reports to demonstrate a marked anti‐inflammatory effect of PEITC following its oral administration.

Reactive oxygen species (ROS) have been implicated as mediators of intestinal inflammation and carcinogenesis. Although green tea polyphenols (GTP) have anticancer property as antioxidants they also generate ROS in vitro. In this study, we investigated the modifying effects of GTP on dextran sulfate sodium (DSS)‐induced acute colitis and on 1,2‐dimethylhydrazine (DMH) and DSS‐induced colon carcinogenesis in male ICR mice. At sacrifice after 6 days, the colon shortening induced by 2% DSS was unchanged by 0.1% and 0.25% GTP, but increased by 0.5% and 1% GTP‐containing diet. The expression of interleukin‐1β and macrophage‐migration inhibitory factor in the DSS + 0.1% GTP group were lower than the DSS alone group, whereas the expression levels were increased in the DSS + 0.5% GTP and DSS + 1% GTP groups when compared with the DSS alone group. In a subsequent experiment to determine the effects of 0.01–1% GTP on inflammation‐associated colon carcinogenesis induced by DMH/DSS, 0.5 and 1% doses of GTP failed to prevent the development of multiple colon tumors, rather, they tended to increase it. Our results thus indicate that the modifying effects of GTP on DSS‐induced acute colitis and DMH/DSS‐induced colon carcinogenesis depends upon its dosage and the expression of proinflammatory cytokines.