A review of animal model studies of tomato carotenoids,lycopene, and cancer chemoprevention

There are relatively few reports on the cancer chemopreventive effects of lycopene or tomato carotenoids in animal models. The majority, but not all, of these studies indicate a protective effect. Inhibitory effects were reported in two studies using aberrant crypt foci, an intermediate lesion leading to colon cancer, as an end point and in two mammary tumor studies, one using the dimethylbenz(a)anthracene model, and the other the spontaneous mouse model. Inhibitory effects were also reported in mouse lung and rat hepatocarcinoma and bladder cancer models. However, a report from the author's laboratory found no effect in the N-nitrosomethylurea-induced mammary tumor model when crystalline lycopene or a lycopene-rich tomato carotenoid oleoresin was administered in the diet. Unfortunately, because of differences in routes of administration (gavage, intraperitoneal injection, intra-rectal instillation, drinking water, and diet supplementation), species and strain differences, form of lycopene (pure crystalline, beadlet, mixed carotenoid suspension), varying diets (grain-based, casein based) and dose ranges (0.5-500 ppm), no two studies are comparable. It is clear that the majority of ingested lycopene is excreted in the feces and that 1000-fold more lycopene is absorbed and stored in the liver than accumulates in other target organs. Nonetheless, physiologically significant (nanogram) levels of lycopene are assimilated by key organs such as breast, prostate, lung, and colon, and there is a rough dose-response relationship between lycopene intake and blood levels. Pure lycopene was absorbed less efficiently than the lycopene-rich tomato carotenoid oleoresin and blood levels of lycopene in rats fed a grain-based diet were consistently lower than those in rats fed lycopene in a casein-based diet. The latter suggests that the matrix in which lycopene is incorporated is an important determinant of lycopene uptake. A number of issues remain to be resolved before any definitive conclusions can be drawn concerning the anticancer effects of lycopene. These include the following: the optimal dose and form of lycopene, interactions among lycopene and other carotenoids and fat soluble vitamins such as vitamin E and D, the role of dietary fat in regulating lycopene uptake and disposition, organ and tissue specificity, and the problem of extrapolation from rodent models to human populations.     

Dietary astaxanthin inhibits colitis and colitis-associated colon carcinogenesis in mice via modulation of the inflammatory cytokines

Astaxanthin (AX) is one of the marine carotenoid pigments, which possess powerful biological antioxidant, anti-inflammatory and anti-cancer properties. The purpose of this study is to investigate possible inhibitory effect of AX against inflammation-related mouse colon carcinogenesis and dextran sulfate sodium (DSS)-induced colitis in male ICR mice. We conducted two different experiments. In the first experiment, we evaluated the effects of AX at three dose levels, 50, 100 and 200 ppm in diet, on colitis-associated colon carcinogenesis induced by azoxymethane (AOM)/DSS in mice. In the second, the effects of the AX (100 and 200 ppm) in diet on DSS-induced colitis were determined. We found that dietary AX significantly inhibited the occurrence of colonic mucosal ulcers, dysplastic crypts, and colonic adenocarcinoma at week 20. AX-feeding suppressed expression of inflammatory cytokines, including nuclear factor (NF)-κB, tumor necrosis factor (TNF)-α and interleukin (IL)-1β, inhibited proliferation, and induced apoptosis in the colonic adenocarcinomas. Feeding with 200 ppm AX, but not 100 ppm, significantly inhibited the development of DSS-induced colitis. AX feeding (200 ppm in diet) also lowered the protein expression of NF-κB, and the mRNA expression of inflammatory cytokines, including IL-1β, IL-6, and cyclooxygenase (COX)-2. Our results suggest that the dietary AX suppresses the colitis and colitis-related colon carcinogenesis in mice, partly through inhibition of the expression of inflammatory cytokine and proliferation. Our findings suggest that AX is one of the candidates for prevention of colitis and inflammation-associated colon carcinogenesis in humans.     

The antioxidant and anti-inflammatory properties of lycopene in mice lungs exposed to cigarette smoke

Lycopene is a carotenoid with known antioxidant and anti-inflammatory properties. We aimed to evaluate the in vitro and in vivo effects of lycopene on reducing the redox imbalance and inflammation induced by cigarette smoke (CS). For the in vitro study, J774A.1 (macrophages) cells were incubated in the presence of 0.5, 1.0, 2.0, 4.0, 8.0, 10.0 and 25 μM of lycopene for 3, 6 and 24 h or in the presence of 0.1%, 0.25%, 0.5%, 0.625%, 1.25%, 2.25%, 5% and 10% cigarette smoke extract (CSE) for 3, 6 and 24 h to assess cell viability and measurement of intracellular reactive oxygen species (ROS). For the in vivo study, 40 mice were divided into 5 groups: a control exposed to ambient air (CG), a vehicle-control group that received 200 μl of sunflower oil by orogastric gavage, a group exposed to CS and two groups administered lycopene (diluted in sunflower oil) at doses of either 25 or 50 mg/kg/day prior to exposure to CS (LY25+CS and LY50+CS). The total treatment time lasted 5 days. A cell viability decrease was observed at 10- and 25-μM concentrations of lycopene in 3, 6 and 24 h compared with CG. There was an increase of ROS production in 24 h in CS compared with CG. Lycopene concentrations of 1 μM and 2 μM were able to reduce the production of ROS in 24 h compared with CS. In the bronchoalveolar lavage fluid, the total number of leukocytes increased in the CS group compared with the control groups (CG). Administration with lycopene at the highest dose suppressed this CS-induced increase in leukocytes. Lipid peroxidation and DNA damage increased in the CS group compared with that in the controls, and this increase was suppressed by lycopene at the highest dose. In contrast, superoxide dismutase activity decreased in the CS group compared with that in the controls. Catalase activity also increased in the CS group compared with that in both control groups, and this increase was suppressed in LY25+CS and LY50+CS. There was an increase in the levels of tumor necrosis factor-α, interferon-γ and interleukin-10 after exposure to CS, and these effects were suppressed by both doses of lycopene. These data elucidate the role of lycopene as an antioxidant and anti-inflammatory agent in these two models of short-term exposure to CS.     

Carotenoid biosynthesis and overproduction in Corynebacterium glutamicum

ABSTRACT: BACKGROUND: Corynebacterium glutamicum contains the glycosylated C50 carotenoid decaprenoxanthin as yellow pigment. Starting from isopentenyl pyrophosphate, which is generated in the non-mevalonate pathway, decaprenoxanthin is synthesized via the intermediates farnesyl pyrophosphate, geranylgeranyl pyrophosphate, lycopene and flavuxanthin. RESULTS: Here, we showed that the genes of the carotenoid gene cluster crtE-cg0722-crtBIYeYfEb are co-transcribed and characterized defined gene deletion mutants. Gene deletion analysis revealed that crtI, crtEb, and crtYeYf, respectively, code for the only phytoene desaturase, lycopene elongase, and carotenoid C45/C50 epsilon-cyclase, respectively. However, the genome of C. glutamicum also encodes a second carotenoid gene cluster comprising crtB2I2-1/2 shown to be co-transcribed, as well. Ectopic expression of crtB2 could compensate for the lack of phytoene synthase CrtB in C. glutamicum DeltacrtB, thus, C. glutamicum possesses two functional phytoene synthases, namely CrtB and CrtB2. Genetic evidence for a crtI2-1/2 encoded phytoene desaturase could not be obtained since plasmid-borne expression of crtI2-1/2 did not compensate for the lack of phytoene desaturase CrtI in C. glutamicum DeltacrtI. The potential of C. glutamicum to overproduce carotenoids was estimated with lycopene as example. Deletion of the gene crtEb prevented conversion of lycopene to decaprenoxanthin and entailed accumulation of lycopene to 0.03 +/- 0.01 mg/g cell dry weight (CDW). When the genes crtE, crtB and crtI for conversion of geranylgeranyl pyrophosphate to lycopene were overexpressed in C. glutamicum DeltacrtEb intensely red-pigmented cells and an 80 fold increased lycopene content of 2.4 +/- 0.3 mg/g CDW were obtained. CONCLUSION: C. glutamicum possesses a certain degree of redundancy in the biosynthesis of the C50 carotenoid decaprenoxanthin as it possesses two functional phytoene synthase genes. Already metabolic engineering of only the terminal reactions leading to lycopene resulted in considerable lycopene production indicating that C. glutamicum may serve as a potential host for carotenoid production.     

K-ras mutations in mouse lung tumors of extreme age: independent of paternal preconceptional exposure to chromium(III) but significantly more frequent in carcinomas than adenomas

Preconceptional exposure of male NIH Swiss mice to chromium(III) chloride resulted in increased incidence of neoplastic and non-neoplastic changes in their progeny, including lung tumors in females [Toxicol. Appl. Pharmacol. 158 (1999) 161-176]. Since mutations in the K-ras protooncogene are frequent, early changes in mouse lung tumors, we investigated possible mutational activation of this gene as a mechanism for preconceptional carcinogenesis by chromium(III). These offspring had lived until natural death at advanced ages (average 816+/-175 days for controls, 904+/-164 for progeny of chromium-treated fathers). Mutations of K-ras, analyzed by single-strand conformation polymorphism and sequencing, were, in codon 12, wild type GGT (glycine), to GAT (aspartic acid); to GTT (valine); and to CGT (arginine); and in codon 61, wild-type CAA (glutamine), to CGA (arginine). K-ras mutation frequencies in lung tumors were very similar in control progeny (4/14) and in progeny of chromium-treated fathers (5/15). Thus, germline mutation or tendency to spontaneous mutation in K-ras does not seem to be part of the mechanism of preconceptional carcinogenesis here. However, an additional interesting observation was that K-ras mutations were much more frequent in lung carcinomas (8/16) than in adenomas (1/13) (P=0.02), for all progeny combined. This was not related to age of the tumor-bearing mice or the size of the tumors. K-ras mutations may contribute to malignant tumor progression during aging, of possible relevance to the putative association of such mutations with poor prognosis of human lung adenocarcinomas.     

Modulatory effect of Piperine on mitochondrial antioxidant system in Benzo(a)pyrene-induced experimental lung carcinogenesis.

Chemoprevention has emerged as a very effective preventive measure against carcinogenesis. Many bioactive compounds present in edible as well in herbal plants have revealed their cancer chemopreventive potential. In the present study, our goal was to investigate the impact of piperine, a principle ingredient of pepper, on alterations of mitochondrial antioxidant system and lipid peroxidation in Benzo(a)pyrene (B(a)P) induced experimental lung carcinogenesis. Oral supplementation of piperine (50 mg/kg body weight) effectively suppressed lung carcinogenesis in B(a)p induced mice as revealed by the decrease in the extent of mitochondrial lipid peroxidation and concomitant increase in the activities of enzymatic antioxidants (superoxide dismutase, catalase and glutathione peroxidase) and non enzymatic antioxidant (reduced glutathione, vitamin E and vitamin C) levels when compared to lung carcinogenesis bearing animals. Our data suggests that piperine may extent its chemopreventive effect by modulating lipid peroxidation and augmenting antioxidant defense system.     

Inhibition of gut- and lung-derived serotonin attenuates pulmonary hypertension in mice

Decreasing the bioavailability of serotonin (5-HT) by inhibiting its biosynthesis may represent a useful adjunctive treatment of pulmonary hypertension (PH). We assessed this hypothesis using LP533401, which inhibits the rate-limiting enzyme tryptophan hydroxylase 1 (Tph1) expressed in the gut and lung, without inhibiting Tph2 expressed in neurons. Mice treated repeatedly with LP533401 (30-250 mg/kg per day) exhibited marked 5-HT content reductions in the gut, lungs, and blood, but not in the brain. After a single LP533401 dose (250 mg/kg), lung and gut 5-HT contents decreased by 50%, whereas blood 5-HT levels remained unchanged, suggesting gut and lung 5-HT synthesis. Treatment with the 5-HT transporter (5-HTT) inhibitor citalopram decreased 5-HT contents in the blood and lungs but not in the gut. In transgenic SM22-5-HTT+ mice, which overexpress 5-HTT in smooth muscle cells and spontaneously develop PH, 250 mg/kg per day LP533401 or 10 mg/kg per day citalopram for 21 days markedly reduced lung and blood 5-HT levels, right ventricular (RV) systolic pressure, RV hypertrophy, distal pulmonary artery muscularization, and vascular Ki67-positive cells (P < 0.001). Combined treatment with both drugs was more effective in improving PH-related hemodynamic parameters than either drug alone. LP533401 or citalopram treatment partially prevented PH development in wild-type mice exposed to chronic hypoxia. Lung and blood 5-HT levels were lower in hypoxic than in normoxic mice and decreased further after LP533401 or citalopram treatment. These results provide proof of concept that inhibiting Tph1 may represent a new therapeutic strategy for human PH.     

A COX-2 inhibitor,nimesulide, inhibits chemically-induced rat tongue carcinogenesis through suppression of cell proliferation activity and COX-2 and iNOS expression

The modifying effects of a cyclooxygenase (cox)-2 selective inhibitor nimesulide on tongue carcinogenesis were investigated in male F344 rats initiated with 4-nitroquinoline-1-oxide (4-NQO). The cell proliferation activity measured by proliferating cell nuclear antigen (PCNA)-positive index and apoptotic index, and the immunohistochemical expression of COX-2, and inducible nitric oxide synthase (iNOS) in the tongue mucosa or neoplasms were also examined for mechanistic analysis of modifying effects of nimesulide on tongue carcinogenesis. All animals except those treated with nimesulide alone and untreated rats were given 20 ppm 4-NQO in drinking water for 8 weeks to induce tongue neoplasms. Starting 1 week after the cessation of 4-NQO exposure, rats given 4-NQO were fed the experimental diets containing nimesulide (100 and 400 ppm) for 22 weeks. At week 32, the incidence of tongue squamous cell carcinoma was significantly reduced by feeding of the diet containing 400 ppm nimesulide. Feeding of nimesulide significantly decreased polyamine content and PCNA-labeling index in tongue carcinoma. Apoptotic index in tongue carcinoma was increased by feeding of nimesulide. In addition, nimesulide feeding reduced COX-2 and iNOS expression in the tongue dysplasia and neoplasms. These results suggest that 400 ppm nimesulide in diet, when given during the promotion phase, exerts chemopreventive ability against 4-NQO-induced tongue tumorigenesis through inhibition of cell proliferation activity in conjunction with modification of COX-2 and iNOS expression of the target lesions.     

The citrus flavonone hesperetin inhibits growth of aromatase-expressing MCF-7 tumor in ovariectomized athymic mice

Aromatase is responsible for the rate-determining reaction in estrogen synthesis and is a prime target for treating estrogen-receptor-positive breast cancer. Previous in vitro study has demonstrated that apigenin (APG), naringenin (NGN) and hesperetin (HSP) are three of the most potent natural aromatase inhibitors. Because the enzyme inhibition could potentially block breast cancer development, we employed an established postmenopausal breast cancer model to examine the chemopreventive effect of these flavonoids in vivo. Athymic mice were ovariectomized and transplanted with aromatase-overexpressing MCF-7 cells. Dietary administration of HSP at 1000 ppm and 5000 ppm significantly deterred the xenograft growth, while a null effect was observed in mice treated with APG or NGN. Further study illustrated that plasma estrogen in HSP-treated mice was reduced. Messenger RNA expression of the estrogen-responsive gene pS2 was also decreased in the tumors of mice treated with 1000 and 5000 ppm HSP. On the other hand, western analysis indicated that cyclin D1, CDK4 and Bcl-x(L) were reduced in the tumors. This study suggested that HSP could be a potential chemopreventive agent against breast carcinogenesis through aromatase inhibition.     

A multi-mineral natural product inhibits liver tumor formation in C57BL/6 mice

C57BL/6 mice were maintained for up to 18 months on high-fat and low-fat diets with or without a multi-mineral supplement derived from the skeletal remains of the red marine algae Lithothamnion calcareum. Numerous grossly observable liver masses were visible in animals on the "western-style" high-fat diet sacrificed at 12 and 18 months. The majority of the masses were in male mice (20 out of 100 males versus 3 out of 100 females; p = 0.0002). There were more liver masses in animals on the high-fat diet than on the low-fat diet (15 out of 50 on high-fat versus 5 out of 50 on low-fat; p = 0.0254). The multi-mineral supplement reduced the number of liver masses in mice on both diets (3 out of 25 male mice in the low-fat diet group without the supplement versus 1 out of 25 mice with supplement; 12 of 25 male mice in the high-fat diet group without the supplement versus 3 of 25 mice with supplement [p = 0.0129]). Histological evaluation revealed a total of 17 neoplastic lesions (9 adenomas and 8 hepatocellular carcinomas), and 18 pre-neoplastic lesions. Out of eight hepatocellular carcinomas, seven were found in unsupplemented diet groups. Steatosis was widely observed in livers with and without grossly observable masses, but the multi-mineral supplement had no effect on the incidence of steatosis or its severity. Taken together, these findings suggest that a multi-mineral-rich natural product can protect mice against neoplastic and pre-neoplastic proliferative liver lesions that may develop in the face of steatosis.     

Lycopene and the lung

The human lung, due to the oxidative and ozone stress to which it is exposed, is particularly vulnerable to oxidative damage. Concentrations of dietary antioxidants in the lung epithelial lining and lining fluids may provide protection against oxidative damage. A randomized clinical trial was conducted to study the effects of supplemental, carotenoid-rich vegetable juice (V-8) on lung function macrophage levels of carotenoids and in moderating ozone-induced lung damage. Healthy young adults (n = 23) were exposed to 0.4 ppm ozone in a chamber for 2 hr after either 2 weeks of antioxidant supplementation (including one can of V-8 juice daily) or placebo. Mean lung concentrations of lycopene increased by 12%, and lung epithelial cell DNA damage as measured by the Comet Assay decreased 20% in supplemented subjects. No change in peripheral blood lymphocyte DNA damage was observed as evidenced by no change in mean comet area or length in supplemented or placebo subjects. We were not able to separate the effects of lycopene from other carotenoids or antioxidants administered in this study; however, lycopene is the predominant carotenoid in V-8 (it represents 88% of total carotenoids). A review of the epidemiologic literature providing evidence for the effect of lycopene (diet or serum) or tomatoes on the risk of lung cancer reveals 27 observational epidemiologic studies (18 case-control and nine cohort studies) reporting relative risk (RR) estimates. RR estimates for cohort studies ranged from 0.63 to 1.24 (mean RR = 0.93, SD = 0.16). Odds ratios (OR) for case-control studies ranged from 0.27 to 0.93 (mean OR = 0.61, SD = 0.16). Both plasma levels (RR = 1.01, OR = 0.37) and estimated intakes of lycopene from dietary sources (mean RR = 0.93, RR range = 0.80-1.05; mean OR = 0.67, OR range = 0.27-0.93) were examined. Seventeen studies, three of which were cohorts, reported their results at the level of tomato consumption rather than, or in addition to, lycopene consumption (mean RR = 0.89, RR range = 0.63-1.24; mean OR = 0.61, OR range = 0.37-0.80). The published epidemiologic literature shows an interaction between study design and the relationship between lycopene and/ or tomatoes and risk of lung cancer. Overall, cohort studies did not show an association, whereas case-control studies showed a decreased risk with greater consumption of lycopene and tomatoes. Although lycopene can be found in the human lung, and there is evidence, albeit weak, for a protective association with lung cancer, its biologic role remains to be elucidated.     

Genes from a Dietzia sp. for synthesis of C40 and C50 beta-cyclic carotenoids

Dietzia sp. CQ4 accumulated the C(40) beta-cyclic carotenoids (canthaxanthin and echinenone) and the C(50) beta-cyclic carotenoid (C.p.450 monoglucoside). A plant-type lycopene beta-cyclase gene crtL was identified for beta-cyclization of the C(40) carotenoids. A carotenoid synthesis gene cluster was identified away from the crtL gene, which contained the crtEBI genes for the synthesis of lycopene followed by the lbtABC genes for lycopene elongation and beta-cyclization of the C(50) carotenoids. This C(50) beta-cyclic carotenoid synthesis gene cluster from Dietzia sp. CQ4 showed high homology with the gene clusters for synthesizing the C(50) epsilon-cyclic carotenoids (decaprenoxanthin and glucosides) from Corynebacterium glutamicum and Agromyces mediolanus. One unique feature of the C(50) beta-cyclic carotenoid synthesis genes in Dietzia sp. CQ4 was that the gene encoding a C(50) carotenoid beta-cyclase subunit and the gene encoding the lycopene elongase appeared to be fused as a single gene (lbtBC). Expression of the gene (lbtA) encoding another subunit of the C(50) carotenoid beta-cyclase and the lbtBC gene in lycopene-accumulating Escherichia coli produced almost exclusively the C(50) beta-cyclic carotenoid C.p.450. One gene (crtX) with high homology to glycosyl transferases was transcribed in the opposite orientation downstream of the lbtBC gene. The crtX gene was likely involved in C.p.450 glucosylation in Dietzia sp. CQ4. The pathway analogous to the synthesis of the C(50) epsilon-cyclic carotenoids was proposed for the synthesis of the C(50) beta-cyclic carotenoids.     

N-acetil-l-cysteine and 2-amino-2-thiiazoline N-acetyl-l-cysteinate as a possible cancer chemopreventive agents in murine models

The aim of this study was to investigate N-acetyl-cysteine (NAC) and its 2-amino-2-thiazoline salt (NACAT) as potential chemopreventive agents on experimentally induced lung tumours by urethane (U) in mice. Female BALB/c mice were used. U was given by intraperitoneal injections during 2 weeks (single dose - 10 mg/mouse, total - 50 mg/mouse). Mice were treated daily per os with NAC 1/10 LD50, NACAT 1/10 or 1/100 LD50 starting 2 weeks prior U administration, then during U treatment and thereafter for 2 months. The duration of experiment was 4 months. The results showed that NAC (1000 mg/kg) reduced the lung tumour incidence to 30% that of controls, P < or = 0.05. Most effective of NACAT was 100 mg/kg dose; it reduced an average of lung adenomas per mouse by 26%, P < or = 0.05, but lower dose (10 mg/kg) was less effective. In order to achieve similar chemopreventive effect (approximately 30%) on mice, it is necessary to use 0.38 mM/kg of NACAT or 6.13 mM/kg of NAC. It means that 16 times less of NACAT is required, if calculated by molar concentration. In general, NAC and NACAT have a moderate chemopreventive effect on lung tumorigenesis induced by urethane in mice.     

Characteristics of selenazolidine prodrugs of selenocysteine: toxicity, selenium levels, and glutathione peroxidase induction in A/J mice

We have previously reported the synthesis and characterization of two new classes of selenazolidine-4(R)-carboxylic acids (2-oxo and 2-methyl-SCAs) (OSCA and MSCA, respectively), as well as the "parent" compound, selenazolidine-4(R)-carboxylic acid (SCA, selenaproline). These compounds were designed as prodrugs of L-selenocysteine with potential application in cancer chemoprevention or other clinical uses. We will be exploring the chemopreventive activity of the new compounds in the well-established A/J mouse model of tobacco-induced lung carcinogenesis. The objectives of the present study were to investigate several fundamental biochemical endpoints after selenazolidine administration compared with other selenium-containing agents. Groups of mice were fed either AIN-76A diet alone or the diet supplemented with the following selenium compounds (ppm Se): sodium selenite (5), L-selenomethionine (3.75), L-selenocystine (15), Se-methyl-L-selenocysteine (3), MSCA (5, 10, or 15), OSCA (5, 10, or 15), or SCA (5, 10, or 15). After 28 days of supplementation, toxicity of the selenazolidines was not evident, as measured by outward appearance and behavior, body and organ weight changes, and histological evaluation of liver and lung tissue. Select treatment groups showed significant increases in selenium levels in blood and tissues. Increased activity of selenium-dependent glutathione peroxidase (GPx) in blood and liver illustrated that the selenazolidines provided a source of biologically-available selenium.     

No mutations found in exon 2 of gene p16CDKN2A during rat tongue carcinogenesis induced by 4-nitroquinoline-1-oxide

The medium-term tongue carcinogenesis assay is a useful model for studying oral squamous cell carcinomas phase by phase. The present study aimed to investigate mutations in exon 2 of gene p16CDKN2A during rat tongue carcinogenesis induced by 4-nitroquinoline 1-oxide (4NQO) using direct DNA-sequencing method. A total of 30 male Wistar rats were treated with 4-nitroquinoline 1-oxide (4NQO) in drinking water for 4, 12, and 20 weeks at 50 ppm dose. Ten animals were used as negative control. No histopathological changes in tongue epithelia were observed among controls or in the group treated for 4 weeks with 4NQO. Following 12-week treatment, hyperplasia and epithelial dysplasia were found in mild and moderate forms. At 20 weeks, the tongue presented moderate and/or severe oral dysplasia and squamous cell carcinoma, with squamous cell carcinoma in the majority of animals. No mutations were found in any experimental period evaluated that corresponded to normal oral mucosa, hyperplasia, dysplasia and squamous cell carcinomas. Taken together, our results suggest that p16CDKN2A mutations in exon 2 are not involved in the multistep tongue carcinogenesis of Wistar rats induced by 4NQO.     

Vascular pressure effects on lung edema formation after glass bead embolism

The canine lung lobe was embolized with 100-micron glass beads before lobectomy and blood anticoagulation. The lobe was isolated, ventilated, and pump-perfused with blood at an arterial pressure (Pa) of about 50 (high pressure, HP, n = 9) or 25 Torr (low pressure, LP, n = 9). Rus/PVR, the ratio of upstream (Rus) to total lobar vascular resistance (PVR), was determined by venous occlusion and the isogravimetric capillary pressure technique. The capillary filtration coefficient (Kf), an index of vascular permeability, was obtained from rate of lobe weight gain during stepwise capillary pressure (Pc) elevation. The embolized lobes became more edematous than nonembolized controls, (C, n = 11), (P less than 0.05), with Kf values of 0.20 +/- 0.04, 0.25 +/- 0.06, and 0.07 +/- 0.01 ml X min-1 X Torr-1 X 100 X g-1 in LP, HP, and C, respectively (P less than 0.05). The greater Rus/PVR in embolized lobes (P less than 0.05) protected the microvessels and, although Pc was greater in HP than in controls (P less than 0.05), Pc did not differ between HP and LP (P greater than 0.05). Although indexes of permeability did not differ between embolized groups (P greater than 0.05), HP became more edematous than LP (P less than 0.05). The greater edema in HP did not appear due to a greater imbalance of Starling forces across the microvessel wall or to vascular recruitment. At constant Pc and venous pressure, elevating Pa from 25 to 50 Torr in embolized lobes resulted in greater edema to suggest fluid filtration from precapillary vessels.     

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 β-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 (100 ppm), 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.     

Apigenin--strong cytostatic and anti-angiogenic action in vitro contrasted by lack of efficacy in vivo.

The cancer chemopreventive agent apigenin also has strong cytostatic and anti-angiogenic effects in vitro. We now investigated its efficacy against experimental Lewis lung carcinomas (LLC), C-6 gliomas and DHDK 12 colonic cancers in vivo. Tumour bearing mice received 50 mg/kg/day apigenin in three different galenical formulations during 12 days in 8-hourly intervals. Only weak effects of apigenin on the size and the number of new tumour blood vessels of both established and newly transplanted tumours were recorded although the intratumoural necrosis was elevated (45 +/- 15% vs. 20 +/- 7% (control), p < 0.05%). These results contrast sharply with the high in vitro sensitivity of LLC, C-6, DHDK 12 and endothelial cells to apigenin where complete growth suppression occurs at concentrations beyond 30 g/ml. Possible causes are discussed.