Anticarcinogenic Effects of Dried Citrus Peel in Colon Carcinogenesis Due to Inhibition of Oxidative Stress.

Colorectal cancer is one of the leading causes of death worldwide. Reactive oxygen species produce oxidative stress and contribute to colorectal carcinogenesis. Because dietary citrus has been shown to reduce oxidative stress, we investigated the effects of citrus peel extract at dilutions of 1/200-1/500 on the activity of oxidative-stress-related transcription factors, including AP-1, NF-κB, NRF2, p53, and STAT3, in human colon cancer cell line HCT116 cells using a luciferase reporter gene assay. NRF2 transcriptional activities were 1.8- to 2.0-fold higher than the untreated control value. In addition, NF-κB, p53, and STAT3 transcriptional activities were 12-26% lower than the untreated control value. Administration of dried citrus peel in the diet of F344 rats at a dose of 1,000 ppm prevented the formation of azoxymethane-induced precancerous aberrant crypt foci (ACF) in the colon. The total number of ACF in rats fed with dried citrus peel was reduced to 75% of the control value. Moreover, the levels of oxidative-stress-related markers, reactive carbonyl species, in the serum of F344 rats were significantly reduced following the administration of dried citrus peel. These data suggest that citrus peel possesses an ability to suppress cellular oxidative stress through induction of NRF2, thereby preventing azoxymethane-induced colon carcinogenesis.

Curcumin combined with turmerones, essential oil components of turmeric, abolishes inflammation-associated mouse colon carcinogenesis.

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.

Clinical significance of amphiregulin and epidermal growth factor in colostrum.

BACKGROUND: Colostrum contains a wide variety of crucial nutritional elements including growth factors for newborn infants to adapt to the extrauterine environment. OBJECTIVE: To investigate the clinical significance of epidermal growth factor receptor ligands in milk during the first month of lactation. METHODS: The concentrations of epidermal growth factor (EGF), amphiregulin (AR) and transforming growth factor-α (TGF-α) in milk sampled from a total of 31 normal mothers at days 1-3, 5, and 30 postpartum were examined using ELISA. RESULTS: At days 1-3, the concentration of EGF was extremely high [131.6 ± 20.4 (mean ± SEM) ng/ml] compared to that of AR (4,197.2 ± 1,055.2 pg/ml) or TGF-α (261.7 ± 33.6 pg/ml), while the concentration of AR was significantly elevated compared to that of TGF-α. At days 5 and 30, the concentration of EGF was significantly elevated compared to that of AR or TGF-α. In 16 mothers among the same 31 subjects, samples were longitudinally obtained on days 1, 2, 5, and 30 postpartum. Concentrations of AR were higher on days 1 and 2 and rapidly declined to below 1 ng/ml on day 5, and were maintained at lower levels on day 30. Concentrations of EGF were high on day 1 (greater than 10 ng/ml) but gradually declined by days 2, 5, and 30. Concentrations of TGF-α remained at lower levels of below 1 ng/ml throughout the lactation period from days 1 to 30. CONCLUSION: These results suggested that EGF and amphiregulin in colostrum might contribute to the early stage of development of neonatal gastrointestinal function.

Metronomic doxifluridine chemotherapy combined with the anti-angiogenic agent TNP-470 inhibits the growth of human uterine carcinosarcoma xenografts.

Uterine carcinosarcoma is a highly aggressive gynecological neoplasm that responds poorly to conventional chemotherapy and radiotherapy. Metronomic chemotherapy is accepted as a new approach for cancer treatment, and its underlying mechanism seems to involve the suppression of angiogenesis. However, the efficacy of metronomic and anti-angiogenic therapies against uterine carcinosarcoma is unknown. The anti-angiogenic effect of doxifluridine was assessed in vitro using human umbilical vein endothelial cells (HUVEC) co-cultured with FU-MMT-1 human uterine carcinosarcoma cells. The antitumor and anti-angiogenic effects of metronomic doxifluridine (delivered via oral gavage) in combination with TNP-470 were evaluated in vivo. Tumor vascularity was assessed by contrast-enhanced color Doppler ultrasound, laser Doppler and microvessel density staining. Doxifluridine suppressed tube formation of HUVEC and vascular endothelial growth factor production by FU-MMT-1 cells. Metronomic doxifluridine alone significantly suppressed tumor growth compared with the untreated (control) group, while metronomic doxifluridine in combination with TNP-470 significantly inhibited tumor growth compared with each treatment alone. A significant reduction of intratumoral vascularity was observed in FU-MMT-1 xenografts following treatment with metronomic doxifluridine in combination with TNP-470, as compared with each treatment alone. Intestinal bleeding was only observed when the maximum tolerated dose of doxifluridine was administered in combination with TNP-470. Metronomic doxifluridine chemotherapy in combination with TNP-470 might be effective for uterine carcinosarcoma without marked toxicity, possibly acting via its potent anti-angiogenic effects. Clinical studies are needed to evaluate the safety and efficacy of this treatment in humans.

Dietary flavonoids as cancer-preventive and therapeutic biofactors.

Flavonoids are present in many plants, and hence, in foods and ingredients derived from them. These polyphenolic compounds have attracted renewed attention as potential anticarcinogens, and the molecular mechanisms of their anticarcinogenic effects and their bioavailability have been extensively explored. In this review, we focus on the major dietary flavonoids; flavones, flavonols, and flavan-3-ols (catechins), and evaluate their roles in cancer prevention. After absorption with or without metabolic conjugation, flavonoids are transported to target organs where they exert their anticarcinogenic activity. The molecular mechanisms of the anticarcinogenic effects of flavonoids include their antagonistic effect on the aryl hydrocarbon receptor (AhR), and regulation of phase I and II drug metabolizing enzymes and phase III transporters. Experimental evidence suggests that flavonoids modulate signal transduction pathways at each stage of carcinogenesis. The interactions between flavonoids and biomolecules in vivo must be investigated in detail to identify specific targets. In addition, the potential side effects should be considered when flavonoid supplements are used for cancer prevention. Therefore, the use of flavonoids as chemopreventive agents should be further investigated to establish safe levels of flavonoid intake.

The modifying effects of green tea polyphenols on acute colitis and inflammation-associated colon carcinogenesis in male ICR mice.

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-1beta 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.

Dietary flavonoids suppress azoxymethane-induced colonic preneoplastic lesions in male C57BL/KsJ-db/db mice.

Obesity is known to be a risk factor for colon carcinogenesis. Although there are several reports on the chemopreventive abilities of dietary flavonoids in chemically induced colon carcinogenesis, those have not been addressed in an obesity-associated carcinogenesis model. In the present study, the effects of 3 flavonoids (chrysin, quercetin and nobiletin) on modulation of the occurrence of putative preneoplastic lesions, aberrant crypt foci (ACF), and beta-catenin-accumulated crypts (BCACs) in the development of colon cancer were determined in male db/db mice with obesity and diabetic phenotypes. Male db/db mice were given 3 weekly intraperitoneal injections of azoxymethane (AOM) to induce the ACF and BCAC. Each flavonoid (100ppm), given in the diet throughout the experimental period, significantly reduced the numbers of ACF by 68-91% and BCAC by 64-71%, as well as proliferation activity in the lesions. Clinical chemistry results revealed that the serum levels of leptin and insulin in mice treated with AOM were greater than those in the untreated group. Interestingly, the most pronounced suppression of development of preneoplastic lesions and their proliferation were observed in the quercetin-fed group, in which the serum leptin level was lowered. Furthermore, quercetin-feeding decreased leptin mRNA expression and secretion in differentiated 3T3-L1 mouse adipocytes. These results suggest that the present dietary flavonoids are able to suppress the early phase of colon carcinogenesis in obese mice, partly through inhibition of proliferation activity caused by serum growth factors. Furthermore, they indicate that certain flavonoids may be useful for prevention of colon carcinogenesis in obese humans.

Aqueous and ethanolic extract fractions from the Brazilian propolis suppress azoxymethane-induced aberrant crypt foci in rats.

We investigated the effects of the two fractions, aqueous (AEP) and ethanolic extracts of propolis (EEP) of the Brazilian propolis on azoxymethane (AOM)-induced aberrant crypt foci (ACF). Male Wistar Hannover (GALAS) rats were administered two weekly subcutaneous injections of AOM (20 mg/kg bw) and fed with diets mixed with AEP (100, 500 and 1,000 ppm) or EEP (500 and 1,000 ppm) for 4 weeks, starting one week before the first dosing of AOM. The modifying effects of the test extracts on ACF formation were assessed by counting the incidence and multiplicity of ACF at week 4. Proliferation cell nuclear antigen (PCNA)-labeling nuclei and apoptotic index were also immunohistochemically determined. Dietary supplementation with AEP and EEP significantly reduced the multiplicity of ACF with the effect of EEP being more potent than AEP. In the ACF and their surrounding non-lesional crypts, significantly lowered cell proliferation was observed in the rats, administered with AOM, and the extracts, while neither fraction affected the apoptotic index. Our findings suggest that AEP and EEP possess a chemopreventive ability in the early phase of colon carcinogenesis through the modulation of cell proliferation.

Suppressive effects of nobiletin on hyperleptinemia and colitis-related colon carcinogenesis in male ICR mice.

Adipocytokines are a group of adipocyte-secreted proteins that have significant effects on the metabolism of lipids and carbohydrates, as well as numerous other processes. A number of recent studies have indicated that some adipocytokines may significantly influence the proliferation of malignant cells in vitro, whereas it remains unclear whether they have similar roles in vivo. In this study, we determined serum levels of adipocytokines in mice with azoxymethane (AOM)- and dextran sulfate sodium (DSS)-induced colon carcinogenesis. Five-week-old ICR mice were given a single intraperitoneal injection of AOM followed by 1% DSS in drinking water for 7 days. Nobiletin (NOB), a citrus flavonoid, was given in the diet (100 p.p.m) for 17 weeks. Thereafter, the incidence and number of colon tumors and serum concentration of adipocytokines were determined at the end of week 20. The serum leptin level in AOM/DSS-treated mice was six times higher than that in untreated mice, whereas there were no significant differences in the levels of triglycerides, adiponectin and interleukin-6. Feeding with NOB abolished colonic malignancy and notably decreased the serum leptin level by 75%. Further, NOB suppressed the leptin-dependent, but not independent, proliferation of HT-29 colon cancer cells and decreased leptin secretion through inactivation of mitogen-activated protein kinase/extracellular signaling-regulated protein kinase, but not that of adiponectin in differentiated 3T3-L1 mouse adipocytes in a dose-dependent manner. Taken together, our results suggest that higher levels of leptin in serum promote colon carcinogenesis in mice, whereas NOB has chemopreventive effects against colon carcinogenesis, partly through regulation of leptin levels.

9trans,11trans conjugated linoleic acid inhibits the development of azoxymethane-induced colonic aberrant crypt foci in rats.

We investigated the effects of 9trans,11trans (9t,11t)-conjugated linoleic acid (CLA) isomer on azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) in rats. Male F344 rats were given 2 weekly subcutaneous injections of AOM (20 mg/kg bw) to induce colonic ACF. They also were fed a diet containing either 0.01%, 0.1%, or 1% 9t,11t-CLA for 4 wk starting 1 wk before the first dosing of AOM. The group that received a diet supplemented with 9t,11t-CLA had a significantly lower number of ACF/colon in comparison to the AOM alone group in a dose-dependent manner up to 0.1%. Furthermore, treatment with 9t,11t-CLA induced apoptosis and suppressed cell proliferation activity in the non-lesional crypts. The downregulation of cyclooxygenase-2 and cyclin D1 and the activation of peroxisome proliferators activated receptor gamma were observed in the colonic mucosa of rats fed a diet supplemented with 9t,11t-CLA. Our findings thus provide some novel insight into the chemopreventive effect of 9t,11t-CLA against preinitiation as well as postinitiation stages of colorectal carcinogenesis.

Diet supplemented with citrus unshiu segment membrane suppresses chemically induced colonic preneoplastic lesions and fatty liver in male db/db mice.

The modulatory effects of dietary citrus unshiu segment membrane (CUSM) on the occurrence of aberrant crypt foci (ACF) and beta-catenin accumulated crypts (BCACs) were determined in male C57BL/KsJ-db/db (db/db) mice initiated with azoxymethane (AOM). Male db/db, db/+ and +/+ mice were given 5 weekly subcutaneous injections of AOM (15 mg/kg body weight), and then they were fed the diet containing 0.02%, 0.1% or 0.5% CUSM for 7 weeks. At Week 12, a significant increase in the numbers of ACF and BCAC was noted in the db/db mice in comparison with the db/+ and +/+ mice. Feeding with CUSM caused reduction in the frequency of ACF in all genotypes of mice and the potency was high in order of the db/db mice, db/+ mice and +/+ mice. The number of BCACs was also reduced by feeding with CUSM, thus resulting in a 28-61% reduction in the db/db mice, possibly due to suppression of cell proliferation activity in the lesions by feeding with CUSM-containing diet. Clinical chemistry revealed a low serum level of triglyceride in mice fed CUSM. In addition, CUSM feeding inhibited fatty metamorphosis and fibrosis in the liver of db/db mice. Our findings show that CUSM in the diet has a chemopreventive ability against the early phase of AOM-induced colon carcinogenesis in the db/db as well as db/+ and +/+ mice, indicating potential use of CUSM in cancer chemoprevention in obese people.

Catalpa seed oil rich in 9t,11t,13c-conjugated linolenic acid suppresses the development of colonic aberrant crypt foci induced by azoxymethane in rats.

Catalpa (Catalpa ovata) seed oil (CPO) is a unique oil that contains a high amount of 9trans,11trans,13cis-conjugated linolenic acid. In the present study, we investigated whether dietary administration with CPO affects the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) in male F344 rats to elucidate its possible cancer chemopreventive efficiency. Also, the effect of CPO on the fatty acid composition of liver tissue and colonic mucosa, the serum levels of total cholesterol and triglyceride, and the mRNA expression of cyclooxygenase (COX)-2 in the colonic mucosa were measured. In addition, the cell proliferation activity and apoptotic index in the colonic mucosa were estimated immunohistochemically. Animals were given two weekly subcutaneous injections of AOM (20 mg/kg body weight). They also received the experimental diet containing 0.01%, 0.1% or 1% CPO for 4 weeks, starting one week before the first dosing of AOM. AOM exposure produced a substantial number of ACF (99+/-28) at the end of the study (week 4). Dietary administration of CPO reduced the number of ACF (AOM + 0.01% CPO, 32+/-11, P<0.001; AOM + 0.1% CPO, 35+/-18, P<0.001; AOM + 1% CPO, 18+/-10, P<0.001). 9t,11t-conjugated linoleic acid was detected in the liver tissue and colonic mucosa of rats fed the CPO-containing diet. Additionally, dietary administration with CPO decreased the serum triglyceride level and the expression of COX-2 mRNA in the colonic mucosa. The indices of cell proliferation and apoptosis in the colonic mucosa of rats treated with AOM and 1% CPO have significant differences when compared with the AOM alone group. These findings suggest the possible chemopreventive activity of CPO in the early phase of colon carcinogenesis.