Towards High Capacity Li-ion Batteries Based on Silicon-Graphene Composite Anodes and Sub-micron V-doped LiFePO4 Cathodes.

Lithium iron phosphate, LiFePO4 (LFP) has demonstrated promising performance as a cathode material in lithium ion batteries (LIBs), by overcoming the rate performance issues from limited electronic conductivity. Nano-sized vanadium-doped LFP (V-LFP) was synthesized using a continuous hydrothermal process using supercritical water as a reagent. The atomic % of dopant determined the particle shape. 5 at. % gave mixed plate and rod-like morphology, showing optimal electrochemical performance and good rate properties vs. Li. Specific capacities of >160 mAh g-1 were achieved. In order to increase the capacity of a full cell, V-LFP was cycled against an inexpensive micron-sized metallurgical grade Si-containing anode. This electrode was capable of reversible capacities of approximately 2000 mAh g-1 for over 150 cycles vs. Li, with improved performance resulting from the incorporation of few layer graphene (FLG) to enhance conductivity, tensile behaviour and thus, the composite stability. The cathode material synthesis and electrode formulation are scalable, inexpensive and are suitable for the fabrication of larger format cells suited to grid and transport applications.

The role of microRNA in nutritional control.

MicroRNAs (miRNAs) are one of a growing class of noncoding RNAs that are involved in the regulation of a wide range of metabolic processes including cellular differentiation, cell proliferation and apoptosis. The generation of miRNA is regulated in complex ways, for example by small interfering RNAs (small nucleolar and nuclear RNAs) and various other metabolites. This complexity of control is likely to explain how a relatively small part of the DNA that codes for proteins has enabled the evolution of such complex organisms as mammals. Non-protein-coding DNA is therefore thought to carry the memory of early evolutionary steps that led to progressively complex metabolic controls. Clinically, miRNAs are becoming increasingly important following the recognition that some congenital abnormalities can be traced to defects in miRNA processing. The potential for manipulating metabolism and affecting disease processes by the pharmaceutical or biological targeting of specific miRNA pathways is now being tested. miRNAs are also released into the extracellular milieu after packaging by cells into nano-sized extracellular vesicles. Such vesicles can be taken up by adjacent and possibly more distant cells, thereby allowing coordinated intercellular communication in specific tissues. Extracellular miRNAs found in the blood stream may also serve as novel biomarkers for both diagnosing specific forms of cancer and assessing the likelihood of metastasis, and as powerful prognostic indices for various cancers. Here, we discuss the role of intracellular and extracellular miRNAs in nutritional control of various (patho)physiological processes. In this review, we provide an update of the presentations from the 25th Marabou Symposium (Stockholm, 14-16 June 2013) entitled 'Role of miRNA in health and nutrition', attended by 50 international experts

SUV39H1/H3K9me3 attenuates sulforaphane-induced apoptotic signaling in PC3 prostate cancer cells.

The isothiocyanate sulforaphane is a promising molecule for development as a therapeutic agent for patients with metastatic prostate cancer. Sulforaphane induces apoptosis in advanced prostate cancer cells, slows disease progression in vivo and is well tolerated at pharmacological doses. However, the underlying mechanism(s) responsible for cancer suppression remain to be fully elucidated. In this investigation we demonstrate that sulforaphane induces posttranslational modification of histone methyltransferase SUV39H1 in metastatic, androgen receptor-negative PC3 prostate cancer cells. Sulforaphane stimulates ubiquitination and acetylation of SUV39H1 within a C-terminal nuclear localization signal peptide motif and coincides with its dissociation from chromatin and a decrease in global trimethyl-histone H3 lysine 9 (H3K9me3) levels. Exogenous SUV39H1 expression leads to an increase in H3K9me3 and decreases sulforaphane-induced apoptotic signaling. SUV39H1 is thus identified as a novel mediator of sulforaphane cytotoxicity in PC3 cells. Our results also suggest SUV39H1 dynamics as a new therapeutic target in advanced prostate cancers.

HDAC8 and STAT3 repress BMF gene activity in colon cancer cells.

Histone deacetylase (HDAC) inhibitors are undergoing clinical trials as anticancer agents, but some exhibit resistance mechanisms linked to anti-apoptotic Bcl-2 functions, such as BH3-only protein silencing. HDAC inhibitors that reactivate BH3-only family members might offer an improved therapeutic approach. We show here that a novel seleno-α-keto acid triggers global histone acetylation in human colon cancer cells and activates apoptosis in a p21-independent manner. Profiling of multiple survival factors identified a critical role for the BH3-only member Bcl-2-modifying factor (Bmf). On the corresponding BMF gene promoter, loss of HDAC8 was associated with signal transducer and activator of transcription 3 (STAT3)/specificity protein 3 (Sp3) transcription factor exchange and recruitment of p300. Treatment with a p300 inhibitor or transient overexpression of exogenous HDAC8 interfered with BMF induction, whereas RNAi-mediated silencing of STAT3 activated the target gene. This is the first report to identify a direct target gene of HDAC8 repression, namely, BMF. Interestingly, the repressive role of HDAC8 could be uncoupled from HDAC1 to trigger Bmf-mediated apoptosis. These findings have implications for the development of HDAC8-selective inhibitors as therapeutic agents, beyond the reported involvement of HDAC8 in childhood malignancy.

Hierarchically structured titanium foams for tissue scaffold applications.

We present a novel route for producing a new class of titanium foams for use in biomedical implant applications. These foams are hierarchically porous, with both the traditional large (>300µm) highly interconnected pores and, uniquely, wall struts also containing micron scale (0.5-5µm) interconnected porosities. The fabrication method consists of first producing a porous oxide precursor via a gel casting method, followed by electrochemical reduction to produce a metallic foam. This method offers the unique ability to tailor the porosity at several scales independently, unlike traditional space-holder techniques. Reducing the pressure during foam setting increased the macro-pore size. The intra-strut pore size (and percentage) can be controlled independently of macro-pore size by altering the ceramic loading and sintering temperature during precursor production. Typical properties for an 80% porous Ti foam were a modulus of ∼1GPa, a yield strength of 8MPa and a permeability of 350 Darcies, all of which are in the range required for biomedical implant applications. We also demonstrate that the micron scale intra-strut porosities can be exploited to allow infiltration of bioactive materials using a novel bioactive silica-polymer composite, resulting in a metal-bioactive silica-polymer composite.

PKG inhibits TCF signaling in colon cancer cells by blocking beta-catenin expression and activating FOXO4.

Activation of cGMP-dependent protein kinase (PKG) has anti-tumor effects in colon cancer cells but the mechanisms are not fully understood. This study has examined the regulation of beta-catenin/TCF signaling, as this pathway has been highlighted as central to the anti-tumor effects of PKG. We show that PKG activation in SW620 cells results in reduced beta-catenin expression and a dramatic inhibition of TCF-dependent transcription. PKG did not affect protein stability, nor did it increase phosphorylation of the amino-terminal Ser33/37/Thr41 residues that are known to target beta-catenin for degradation. However, we found that PKG potently inhibited transcription from a luciferase reporter driven by the human CTNNB1 promoter, and this corresponded to reduced beta-catenin mRNA levels. Although PKG was able to inhibit transcription from both the CTNNB1 and TCF reporters, the effect on protein levels was less consistent. Ectopic PKG had a marginal effect on beta-catenin protein levels in SW480 and HCT116 but was able to inhibit TCF-reporter activity by over 80%. Investigation of alternative mechanisms revealed that cJun-N-terminal kinase (JNK) activation was required for the PKG-dependent regulation of TCF activity. PKG activation caused beta-catenin to bind to FOXO4 in colon cancer cells, and this required JNK. Activation of PKG was also found to increase the nuclear content of FOXO4 and increase the expression of the FOXO target genes MnSOD and catalase. FOXO4 activation was required for the inhibition of TCF activity as FOXO4-specific short-interfering RNA completely blocked the inhibitory effect of PKG. These data illustrate a dual-inhibitory effect of PKG on TCF activity in colon cancer cells that involves reduced expression of beta-catenin at the transcriptional level, and also beta-catenin sequestration by FOXO4 activation.

Characterization of the deformation behavior of intermediate porosity interconnected Ti foams using micro-computed tomography and direct finite element modeling.

Under load-bearing conditions metal-based foam scaffolds are currently the preferred choice as bone/cartilage implants. In this study X-ray micro-computed tomography was used to discretize the three-dimensional structure of a commercial titanium foam used in spinal fusion devices. Direct finite element modeling, continuum micromechanics and analytical models of the foam were employed to characterize the elasto-plastic deformation behavior. These results were validated against experimental measurements, including ultrasound and monotonic and interrupted compression testing. Interrupted compression tests demonstrated localized collapse of pores unfavorably oriented with respect to the loading direction at many isolated locations, unlike the Ashby model, in which pores collapse row by row. A principal component analysis technique was developed to quantify the pore anisotropy which was then related to the yield stress anisotropy, indicating which isolated pores will collapse first. The Gibson-Ashby model was extended to incorporate this anisotropy by considering an orthorhombic, rather than a tetragonal, unit cell. It is worth noting that the natural bone is highly anisotropic and there is a need to develop and characterize anisotropic implants that mimic bone characteristics.

Characterization of the structure and permeability of titanium foams for spinal fusion devices.

Titanium foams produced via the space-holder method are used for spinal fusion devices since their combination of an open-cell structure and bone-like mechanical properties promises potentially excellent bone ingrowth. Earlier studies have indicated that the size of the pores and interconnects must be greater than 100microm for effective bone ingrowth and vascularization. Hence, the quantification of the pore and interconnect size is required for efficient scaffold design. In this study, microcomputed tomography (microCT) was used to obtain the three-dimensional (3D) structure of Ti foams with three levels of porosity (51%, 65% and 78%). Novel algorithms were then applied to quantify both the pore and interconnect size of Ti foams as a function of porosity. All foams possessed a modal pore and interconnect size in excess of 300microm, satisfying the requirement of being greater than 100microm. The pore and interconnect size also dominates the flow properties or permeability of open-cell structures. Therefore, the microCT data was also used to generate a mesh for computational fluid dynamics analysis to predict the permeability. The calculated permeability (117-163x10(-12)m(2) depending on direction) for the Ti foams with 65% porosity was first validated against experimental measurements (98-163x10(-12)m(2)) and then compared to prior authors' measurements in healthy cancellous bovine bone (233-465x10(-12)m(2)). The close match among all the permeability values proves the suitability of the material for biomedical skeletal-implant applications.

Global histone H4 acetylation and HDAC2 expression in colon adenoma and carcinoma.

Chromatin remodeling and activation of transcription are important aspects of gene regulation, but these often go awry in disease progression, including during colon cancer development. We investigated the status of global histone acetylation (by measuring H3, H4 acetylation of lysine residues, which also occur over large regions of chromatin including coding regions and non-promoter sequences) and expression of histone deacetylase 2 (HDAC2) in colorectal cancer (CRC) tissue microarrays using immunohistochemical staining. Specifically, HDAC2 and the acetylation of histones H4K12 and H3K18 were evaluated in 134 colonic adenomas, 55 moderate to well differentiated carcinomas, and 4 poorly differentiated carcinomas compared to matched normal tissue. In addition, the correlation between expression of these epigenetic biomarkers and various clinicopathological factors including, age, location, and stage of the disease were analyzed. HDAC2 nuclear expression was detected at high levels in 81.9%, 62.1%, and 53.1% of CRC, adenomas, and normal tissue, respectively (P = 0.002). The corresponding nuclear global expression levels in moderate to well differentiated tumors for H4K12 and H3K18 acetylation were increased while these levels were decreased in poorly differentiated tumors (P = 0.02). HDAC2 expression was correlated significantly with progression of adenoma to carcinoma (P = 0.002), with a discriminative power of 0.74, when comparing cancer and non-cancer cases. These results suggest HDAC2 expression is significantly associated with CRC progression.

Bcl-2 overexpression in PhIP-induced colon tumors: cloning of the rat Bcl-2 promoter and characterization of a pathway involving beta-catenin, c-Myc and E2F1.

Beta-catenin/T-cell factor (Tcf) signaling is constitutively active in the majority of human colorectal cancers, and there are accompanying changes in Bcl-2 expression. Similarly, 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (PhIP)-induced colon tumors in the rat have increased beta-catenin and elevated Bcl-2. To examine the possible direct transcriptional regulation of rat Bcl-2 by beta-catenin/Tcf, we cloned and characterized the corresponding promoter region and found 70.1% similarity with its human counterpart, BCL2. Bcl-2 promoter activity was increased in response to LiCl and exogenous beta-catenin, including oncogenic mutants of beta-catenin found in PhIP-induced colon tumors. Protein/DNA arrays identified E2F1, but not beta-catenin/Tcf, as interacting most strongly with the rat Bcl-2 promoter. Exogenous E2F1 increased the promoter activity of rat Bcl-2, except in mutants lacking the E2F1 sites. As expected, beta-catenin induced its downstream target c-Myc, as well as E2F1 and Bcl-2, and this was blocked by siRNA to c-Myc or E2F1. These findings suggest an indirect pathway for Bcl-2 over-expression in PhIP-induced colon tumors involving beta-catenin, c-Myc and E2F1.

Ultraviolet radiation-induced non-melanoma skin cancer in the Crl:SKH1:hr-BR hairless mouse: augmentation of tumor multiplicity by chlorophyllin and protection by indole-3-carbinol.

Over 1 million new cases of ultraviolet radiation-induced non-melanoma skin cancers (NMSC) per year now occur in the USA and the incidence of these diseases continues to increase. New preventative strategies are required. The hypothesis tested was that dietary administration of the putative cancer chemopreventatives sodium-copper-chlorophyllin (Chlor) or indole-3-carbinol (I3C) would inhibit UV-induced skin carcinogenesis in the Crl:SKH1:hr-BR hairless mouse. Groups of 20 mice were pre-fed isocaloric/isonutritive 20% corn-oil AIN-76a based diets that contained either Chlor (1.52 g%), I3C (5.08 g%) or no chemopreventative (control) for 2 weeks followed by exposure of their dorsal skin to a 10 week incremental, sub-erythemal, carcinogenic simulated solar UV exposure regime. Feeding was continued for the duration of the experiment. Matched non-UV exposed dietary groups were also included in the experimental design. The diets had no significant (p > 0.05) effect on body weight, feed consumption, cutaneous methanol-extractable UV photoprotective substances or on cutaneous UV-reflective characteristics. By day 180, UV-irradiated mice fed the Chlor had a significantly (p < 0.05) higher tumor multiplicity (33.6 +/- 4.72; mean +/- SEM) than UV-irradiated control animals (22.8 +/- 4.25). UV-irradiated mice fed I3C had a significantly (p < 0.001) lower tumor multiplicity (13.0 +/- 2.42) than that of both the UV-irradiated control and UV-irradiated Chlor-fed mice. The Chlor or I3C diets did not significantly (p > 0.05) affect UV-induced systemic suppression of contact hypersensitivity responses. These results demonstrate augmentation of the UV-induced cutaneous carcinogenic process by dietary chlorophyllin and protection from this carcinogenic process by indole-3-carbinol via mechanisms that do not involve changes in skin optical properties, modulation of photoimmunosuppression or caloric/nutrient effects.

The disposition and metabolism of 2-amino-3-methylimidazo-[4,5-f]quinoline in the F344 rat at high versus low doses of indole-3-carbinol.

Indole-3-carbinol (I3C), a compound found in cruciferous vegetables, inhibits the formation of DNA adducts, colonic aberrant crypts, and tumors in rats given heterocyclic amines, such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). Previous mechanism studies indicated that I3C induces cytochromes P4501A1 (CYP1A1) and CYP1A2, as well as phase 2 pathways, leading to enhanced metabolism and excretion of IQ. However, the chemopreventive activity is dependent on the dose of I3C, and at low doses which do not induce CYP1A activity, there is evidence for increased IQ-DNA adduct formation in vivo. The present study examined the fate of IQ in the rat and the profile of urinary metabolites across a broad range of I3C doses. Male F344 rats were given a single injection of I3C by oral gavage, at a dose equivalent to that received from a single daily exposure to 0, 5, 10, 25, 50, 100, 200, 500 or 1000 ppm I3C in the diet, or they were given the 1000-ppm-equivalent dose of I3C for 14 consecutive days. Subsequently, each rat was given 14C-labeled IQ (5 mg/kg; 0.1 mCi/kg) and the animal was sacrificed 8 h later. With increasing I3C, there was a dose-dependent decrease in IQ-associated radiolabel in several systemic tissues, and an increase in the radiolabel eliminated via the feces. In the urine, there was a dose-dependent increase in IQ-5-O-glucuronide and IQ-5-O-sulfate metabolites, and a concomitant decrease in the IQ-sulfamate at intermediate and high doses of I3C. However, 5- and 10 ppm-equivalent doses of I3C enhanced the levels of IQ-sulfamate compared with controls, possibly due to the high ratio of hepatic CYP1A2 versus CYP1A1 activities at these I3C doses. The possible significance of the low versus high dose effects are discussed in the context of ongoing clinical trials with I3C and the reported chemopreventive mechanisms in vivo.

Potent antimutagenic activity of white tea in comparison with green tea in the Salmonella assay.

There is growing interest in the potential health benefits of tea, including the antimutagenic properties. Four varieties of white tea, which represent the least processed form of tea, were shown to have marked antimutagenic activity in the Salmonella assay, particularly in the presence of S9. The most active of these teas, Exotica China white tea, was significantly more effective than Premium green tea (Dragonwell special grade) against 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and four other heterocyclic amine mutagens, namely 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (4,8-DiMeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2). Mechanism studies were performed using rat liver S9 in assays for methoxyresorufin O-demethylase (MROD), a marker for the enzyme cytochrome P4501A2 that activates heterocyclic amines, as well as Salmonella assays with the direct-acting mutagen 2-hydroxyamino-3-methylimidazo[4,5-f]quinoline (N-hydroxy-IQ). White tea at low concentrations in the assay inhibited MROD activity, and attenuated the mutagenic activity of N-hydroxy-IQ in the absence of S9. Nine of the major constituents found in green tea also were detected in white tea, including high levels of epigallocatechin-3-gallate (EGCG) and several other polyphenols. When these major constituents were mixed to produce "artificial" teas, according to their relative levels in white and green teas, the complete tea exhibited higher antimutagenic potency compared with the corresponding artificial tea. The results suggest that the greater inhibitory potency of white versus green tea in the Salmonella assay might be related to the relative levels of the nine major constituents, perhaps acting synergistically with other (minor) constituents, to inhibit mutagen activation as well as "scavenging" the reactive intermediate(s).

A comparison of whole wheat, refined wheat and wheat bran as inhibitors of heterocyclic amines in the Salmonella mutagenicity assay and in the rat colonic aberrant crypt focus assay.

Refined wheat, unrefined whole wheat, and wheat bran were studied for their ability to protect against heterocyclic amines (HCAs) in vitro and in vivo. Wheat bran, which binds HCAs in vitro, as well as refined wheat and unrefined whole wheat, inhibited the mutagenic activities of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) when they were co-incubated and the supernatant (minus grain) was added to the Salmonella assay. The water-soluble fraction alone from refined and unrefined wheat, but not bran, also inhibited against these mutagens in vitro. In vivo, AIN-93G diets containing refined wheat or unrefined wheat were examined for their ability to inhibit IQ-induced colonic aberrant crypt foci (ACF) in the Fischer 344 rat. A slight increase in the number of AC/ACF (aberrant crypts/ACF) was seen after 16 weeks in rats treated post-initiation with refined wheat (P < 0.05), and fewer foci with two or three aberrant crypts (ACF-2) were found in rats given unrefined whole wheat post-initiation compared with animals treated with the same diet during the initiation phase (P < 0.05). There was no significant difference in the profile of IQ urinary metabolites or excretion of promutagens 0-48 h after carcinogen dosing, and grains had no effect on hepatic cytochrome P4501A1 (CYP1A1), CYP1A2, aryl sulfotransferase or N-acetyltransferase activities; however, a slightly higher UDP-glucuronosyl transferase activity was observed in rats fed unrefined wheat compared with refined wheat diets (P < 0.05). Thus, despite their antimutagenic activities in vitro, only marginal effects were seen with refined and unrefined wheat in vivo with respect to hepatic enzyme activities, carcinogen metabolism and IQ-induced ACF in the rat colon.

Colonic cell proliferation, apoptosis and aberrant crypt foci development in rats given 2-amino-3-methylimidaz.

Sodium-copper chlorophyllin (CHL) inhibits the formation of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ)- and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced colonic aberrant crypt foci (ACF) and tumours in the F344 rat when it is given simultaneously with either carcinogen. However, CHL reportedly increased the incidence of dimethylhydrazine (DMH)-induced colon tumours in the same species when administered post-initiation. In the present study, rats were given IQ (130 mg/kg body weight, by oral gavages on alternating days) for 2 weeks, starting in experiment week 3, and one week after the final IQ dose rats received CHL treatment until the study was terminated at 16 weeks. Compared with animals given carcinogen alone, the mean number of IQ-induced ACF per colon was reduced significantly by 1% (w/v) CHL in the drinking water (P < 0.05), whereas 0.1% and 0.01% CHL had no effect. These CHL concentrations increased in a dose-related manner both the terminal deoxynucleotidyl transferase-mediated nick end labelling (TUNEL) and bromodeoxyuridine (BrdU) labelling indices in the distal colon. However, the lowest concentration tested, 0.001% CHL, increased the mean number of IQ-induced ACF per colon (P < 0.05), and increased the BrdU labelling index without a concomitant change in TUNEL. These studies indicated that 0.001% CHL promoted IQ-ACF due to deregulation of the homeostatic balance between cell birth and apoptosis in the colonic mucosa, whereas higher concentrations of CHL had either no effect or protected against IQ-induced ACF by causing dose-related increases in the overall rate of cell turnover in the colon.

beta-Catenin mutation in rat colon tumors initiated by 1,2-dimethylhydrazine and 2-amino-3-methylimidazo[4,5-f]quinoline, and the effect of post-initiation treatment with chlorophyllin and indole-3-carbinol.

Carcinogens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 1,2-dimethylhydrazine (DMH) induce colon tumors in the rat that contain mutations in beta-catenin, but the pattern of mutation differs from that found in human colon cancers. In both species, mutations affect the glycogen synthase kinase-3beta consensus region of beta-catenin, but whereas they directly substitute critical Ser/Thr phosphorylation sites in human colon cancers, the majority of mutations cluster around Ser33 in the rat tumors. Two dietary phytochemicals, chlorophyllin and indole-3-carbinol, given post-initiation, shifted the pattern of beta-catenin mutations in rat colon tumors induced by IQ and DMH. Specifically, 17/39 (44%) of the beta-catenin mutations in groups given carcinogen plus modulator were in codons 37, 41 and 45, and substituted critical Ser/Thr residues directly, as seen in human colon cancers. None of the tumors from groups given carcinogen alone had mutations in these codons. Interestingly, many of the mutations that substituted critical Ser/Thr residues in beta-catenin were from a single group given DMH and 0.001% chlorophyllin, in which a statistically significant increase in colon tumor multiplicity was observed compared with the group given DMH only. These tumors had marked over-expression of cyclin D1, c-myc and c-jun mRNA and c-Myc and c-Jun proteins were strongly elevated compared with tumors containing wild-type beta-catenin. The results indicate that the pattern of beta-catenin mutations in rat colon tumors can be influenced by exposure to dietary phytochemicals administered post-initiation, and that the mechanism might involve the altered expression of beta-catenin/Tcf/Lef target genes.

Post-initiation effects of chlorophyllin and indole-3-carbinol in rats given 1,2-dimethylhydrazine or 2-amino-3-methyl- imidazo.

Chlorophyllin (CHL) is a water-soluble derivative of chlorophyll, the ubiquitous pigment in green and leafy vegetables, whereas indole-3-carbinol (I3C) is present in cruciferous vegetables such as cabbage, broccoli and cauliflower. In rats initiated with 1,2-dimethylhydrazine (DMH), CHL and I3C reportedly promoted or enhanced the incidence of colon tumors when they were administered after, or during and after the carcinogen exposure, respectively. The same compounds given post-initiation inhibited the formation of colonic aberrant crypts induced by heterocyclic amines, such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), but tumor suppression was not examined in the latter studies. In the present investigation, male F344 rats were treated with IQ or DMH during the first 5 weeks of a 1 year study; IQ was given in the diet (0.03%), whereas DMH was administered once a week by s.c. injection (20 mg/kg body wt). Beginning 1 week after the last dose of IQ or DMH until sacrifice, rats received 0.001, 0.01 or 0.1% (w/v) CHL in the drinking water or 0.001, 0.01 or 0.1% I3C in the diet. Compared with controls given carcinogen alone, 0.1% I3C treatment suppressed the multiplicity of IQ-induced colon tumors, and CHL inhibited in a dose-related manner the incidence of IQ-induced liver tumors. However, 0.001% CHL increased significantly the multiplicity of DMH-induced colon tumors while having no effect on the colon tumors induced by IQ. These results indicate that both the choice of carcinogen as well as the dose of the tumor modulator can be important determinants of the events that occur during post-initiation exposure to CHL or I3C. Based on the present findings and data in the literature, it is possible for CHL and I3C to act as tumor promoters or anticarcinogens, depending upon the test species, initiating agent and exposure protocol.

Inhibition by white tea of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine-induced colonic aberrant crypts in the F344 rat.

There is growing interest in the potential health benefits of tea, including the anticarcinogenic properties. We report here that white tea, the least processed form of tea, is a potent inhibitor of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced colonic aberrant crypts in the rat. Male Fischer 344 rats were treated for 8 wk with white tea (2% wt/vol) or drinking water alone, and on alternating days in experimental Weeks 3 and 4 the animals were given PhIP (150 mg/kg body wt p.o.) or vehicle alone. At the end of the study there were 5.65 +/- 0.81 and 1.31 +/- 0.27 (SD) aberrant crypt foci per colon in groups given PhIP and PhIP + white tea, respectively (n = 12, P < 0.05). No changes were detected in N-acetyltransferase or arylsulfotransferase activities compared with controls, but there was marked induction of ethoxyresorufin O-deethylase, methoxyresorufin O-demethylase, and UDP-glucuronosyltransferase after treatment with white tea. Western blot revealed corresponding increases in cytochrome P-450 1A1 and 1A2 proteins. Enzyme assays and Western blot also revealed induction of glutathione S-transferase by white tea. There was less parent compound and 4'-hydroxy-PhIP but more PhIP-4'-O-glucuronide and PhIP-4'-O-sulfate in the urine from rats given PhIP + white tea than in urine from animals given carcinogen + drinking water. The results indicate that white tea inhibits PhIP-induced aberrant crypt foci by altering the expression of carcinogen-metabolizing enzymes, such that there is increased ring hydroxylation at the 4' position coupled with enhanced phase 2 conjugation.

Increased susceptibility of adult rats to azoxymethane-induced aberrant crypt foci.

The purpose of this study was to compare azoxymethane-induced aberrant crypt foci development in the colons of young and adult rats. Young (4 weeks of age) and adult (50 weeks of age) Sprague-Dawley rats were treated with two weekly injections of azoxymethane or saline. Rats were killed either 6 or 14 weeks following the first injection, and the number, size and location of aberrant crypt foci were determined. At both the 6- and 14-week time points, the number of aberrant crypt foci in older rats was significantly greater than in young rats (P<0.01). A higher percentage of aberrant crypt foci were found in the region from the mid-colon to the cecum in older rats as compared to young rats. Colonic cell proliferation was evaluated using bromodeoxyuridine immunohistochemistry. Colonic cell proliferation indices in the rectal, mid-colon and cecal regions of young and older rats were similar in young compared to adult rats. Ten large ACF from each group were screened for mutations in the beta-catenin gene (Ctnnb1) by PCR single strand conformation polymorphism. No mutations were detected. These results demonstrate that older female rats are more susceptible to the induction of aberrant crypt foci by azoxymethane than young female rats. Differences in colonic cell proliferation or beta-catenin mutations in these two age groups do not appear to be responsible for differences in aberrant crypt foci development.

Effect of carcinogen dose fractionation, diet and source of F344 rat on the induction of colonic aberrant crypts by 2-amino-3-methylimidazo[4,5-f]quinoline.

Carcinogen dose fractionation, diet and source of laboratory animal were examined as variables in the induction of colonic aberrant crypt foci (ACF) by the heterocyclic amine 2-amino-3-methylimidazo [4, 5-f]quinoline (IQ). In the first experiment, male F344 rats from the National Cancer Institute (NCI rats) were fed AIN-93G diet and, starting in the third week, IQ was given by gavage on alternating days, the total carcinogen dose of 105 mg being fractionated proportionally over 2, 4, 8 or 14 weeks. Only the high dose (2 week) treatment with IQ was effective for the induction of ACF at 16 weeks, producing on average 3.8 ACF/colon versus 0.5 ACF/colon in all other groups (P < 0.05). The 2 week IQ dosing protocol was used in a second experiment in which male F344 rats from Simonsen Laboratories (SN) or NCI were fed AIN-93G, AIN-76A or chow diet. On average, SN rats on chow diet had twice the number of aberrant crypts compared with NCI rats given the same diet and three to four times as many aberrant crypts as NCI rats fed AIN diets. Hepatic cytochrome P4501A1 (CYP1A1) levels were essentially unaffected by diet, but methoxyresorufin O-demethylase activities and CYP1A2 protein levels were increased 2- to 3-fold in animals fed chow versus AIN diets. During the 2 week period of carcinogen administration, IQ markedly induced CYP1A proteins and negated the differences among groups related to diet. No consistent diet-related changes were detected in the activities of aryl sulfotransferase or N-acetyltransferase, but UDP-glucuronosyltransferase activities were elevated 2- to 3-fold in rats given chow versus AIN diets. In summary, high dose treatment with IQ was required for the induction of ACF, rats on the chow diet had more aberrant crypts than those given AIN diets and male F344 rats purchased from different vendors and fed chow diet differed with respect to their sensitivity to induction of ACF.