Caffeic acid 3,4-dihydroxyphenethyl ester prevents colorectal cancer through inhibition of multiple cancer-promoting signal pathways in 1,2-Dimethylhydrazine/dextran sodium sulphate mouse model.

OBJECTIVE: To elucidate the potential feature and mechanism of the caffeic acid 3,4-dihydroxyphenethyl ester (CADPE) molecule, which can prevent colorectal cancer (CRC) in the 1,2-Dimethylhydrazine (DMH)/dextran sodium sulphate (DSS)-induced mouse model. METHODS: Institute of cancer research (ICR) male mice were injected with 20 mg/kg DMH for a week. After that, 2% DSS was administered in the drinking water for another 7 d. The CADPE treatment was given to the DMH/DSS induced male mice at three different periods until their sacrifice. Histopathological examination was used for observing the CRC development at colonic mucosa. Immunohistochemistry (IHC), blood cells smearing and crypt damage scoring methods were used for investigating the anti-inflammation feature of CADPE related to CRC. The reversing targets searching method was applied with artificial intelligence (AI), computer-aided drug designing (CADD) and Ingenuity Pathway Analysis (IPA) techniques for predicting the potential targets and mechanism of CADPE highly related to CRC. RESULTS: The data indicated that CADPE inhibited CRC tumor development in the colitis-associated DMH/DSS induced mouse model after giving the early treatment. CADPE also impeded the acute inflammation by decreasing the infiltration of neutrophils significantly during the initial stage of CRC development. Finally, our data showed that CADPE prevented CRC by blocking active sites of three pivotal protein targets including epidermal growth factor receptor (EGFR), extracellular signal-regulated kinase (ERK) and mammalian target of rapamycin (mTOR) in two major cancer development pathways. CONCLUSIONS: CADPE effectively prevented CRC at early stage of tumor germination in the DMH/DSS mouse model highly likely due to its anti-acute inflammation characteristic and the ability of blocking EGFR, ERK and mTOR activities in two highly related CRC developing pathways.

Therapeutic role of biogenic silver and gold nanoparticles against a DMH-induced colon cancer model.

Colorectal cancer (CRC) ranks third among fatal diseases afflicting mankind globally due to the shortage of primary detection methods and appropriate choice of drugs. Moreover, current treatments such as chemo drugs and radiotherapies create adverse effects and lead to drug resistance. In this context, recent advances in nanomedicine offer novel clinical solutions for colon cancer therapy. The current study denotes the therapeutic roles of biogenic Abutilon indicum silver and gold nanoparticles (AIAgNPs and AIAuNPs) against a 1, 2-dimethyl hydrazine (DMH)-induced CRC in Wistar rats. Following treatment of nanoparticles (NPs), the CRC rats showed great localization of AIAgNPs and AIAuNPs in colon tumors shown by ICP-OES, indicating their bioavailability. The AIAgNPs and AIAuNPs significantly enhanced cellular antioxidant enzyme levels including catalase, SOD, GSH, GPx and reduced lipid peroxidation (LPO) compared to the standard drug paclitaxel. AIAgNPs and AIAuNPs revealed significant protection against metastasis compared to paclitaxel shown in the histopathological study. The important CRC signaling molecules of the Wnt pathway, the ß-catenin and Tcf-4 levels were significantly downregulated in AIAgNPs and AIAuNPs treated CRC rats compared to paclitaxel. Furthermore, the expression levels of cleaved apoptotic caspase-9, -8, and - 3 and lamins were significantly upregulated in AIAgNPs and AIAuNPs treated CRC rats compared to paclitaxel. This preclinical study provides substantial insights into the anti-colon cancer roles of biogenic NPs and gives an idea for targeting different cancers.

Effect of Diacerein on HOTAIR/IL-6/STAT3, Wnt/ß-Catenin and TLR-4/NF-κB/TNF-α axes in colon carcinogenesis.

Colorectal cancer (CRC) is a common malignancy with high mortality and poor prognosis. Diacerein (DIA) is an anti-inflammatory used for treatment of osteoarthritis. We delineated some underlying molecular mechanisms of DIA's anti-carcinogenic effect in CRC using in vivo and in vitro models. Human Caco-2 cells were treated with DIA followed by MTT and Annexin V assays and CRC was experimentally induced using 1,2-dimethylhydrazine. DIA (50 mg/kg/day, orally) was administrated for 8 weeks. The MTT assay confirmed cytotoxic effect of DIA in vitro and Annexin V confirmed its apoptotic effect. DIA resulted in regression of tumour lesions with reduced colonic TLR4, NF-κB and TNF-α protein levels and down-regulated VEGF expression, confirming anti-angiogenic impact. DIA triggered caspase-3 expression and regulated Wnt/ß-Catenin pathway, by apparently interrupting the IL-6/STAT3/ lncRNA HOTAIR axis. In conclusion, DIA disrupted IL-6/STAT3/ lncRNA HOTAIR axis which could offer an effective therapeutic strategy for the management of CRC.

Synbiotic modulates intestinal microbiota metabolic pathways and inhibits DMH-induced colon tumorigenesis through c-myc and PCNA suppression.

The use of probiotic and synbiotic is a promising strategy to modulate the intestinal microbiota, and thereby modify the risk of diseases. In this study, the effect of probiotic VSL#3, isolated or associated with a yacon-based product (PBY), on the functional metabolic pathways of the microbiota, in a colorectal carcinogenesis model, was evaluated. For this, mice induced to carcinogenesis were fed with standard diet AIN-93 M (CON), diet AIN-93 M and VSL#3 (PRO) or diet AIN-93 M with yacon and VSL#3 (SYN). The SYN group showed a highly differentiated intestinal community based on the MetaCyc pathways. Of the 351 predicted functional pathways, 222 differed between groups. Most of them were enriched in the SYN group, namely: amino acid biosynthesis pathways, small molecule biosynthesis pathways (cofactors, prosthetic groups, electron carriers and vitamins) carbohydrate degradation pathways and fermentation pathways. In addition, the synbiotic was able to stimulate the anti-inflammatory immune response and reduce the gene expression of PCNA and c-myc. Thus, we conclude that the synbiotic impacted more significantly the metabolic functions of the microbiota compared to the isolated use of probiotic. We believe that the enrichment of these pathways can exert antiproliferative action, reducing colorectal carcinogenesis. The prediction of the functional activity of the microbiota is a promising tool for understanding the influence of the microbiome on tumor development.

Target-specific micronucleus induction by colon carcinogens: 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and 1,2-dimethylhydrazine.

Genotoxicity occurring at the target organs of carcinogenesis is important for understanding the mechanisms of chemical carcinogenicity and also for setting of threshold estimation. In vivo gene mutations have been evaluated by transgenic animal models in which any organ can be targeted; however, the methodologies that have been applied to assess chromosomal aberrations including micronucleus induction, are organ restricted, (often to bone marrow hematopoietic cells, as a common example). For food and food-related chemicals, the digestive tract is the important target organ as it is the organ of first contact. In the present study, we used 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 1,2-dimethylhydrazine (DMH) as model chemicals of carcinogens primarily targeting the colon. We evaluated the applicability of colon cells and hepatocytes, together with bone marrow cells, in the micronucleus assay. Both model chemicals induced micronuclei in the colon, which is the target organ of these carcinogens, after short- and long-term treatment(s). The results demonstrate the target specificity of micronucleus induction and the assay using organs other than bone marrow will play an important role in understanding the mechanism of carcinogenicity and predicting new carcinogenic agents.

Kumiss Supplementation Reduces Oxidative Stress and Activates Sirtuin Deacetylases by Regulating Antioxidant System.

It was aimed to investigate the effects of kumiss a fermented mare horse beverage on the sirtuin deacetylases in the oxidative stress which had been induced by 1,2-dimethyl hydrazine (DMH). Forty BALB/C male mice were divided into four groups as control, kumiss (2 × 108 cfu/mL), DMH (20 mg/kg), and kumiss + DMH (2 × 108 cfu/mL + 20 mg/kg). At the end of 20-week regimen, SIRT2, SIRT3 protein expressions by western blotting, immunolocalizations, and inhibitory anti-oxidant activity analysis in liver, colon, and kidney tissues were performed. SIRT2 and SIRT3 expressions in DMH group were decreased in liver, colon, and kidney tissues and the decrease further stimulated by kumiss reinforcement. SIRT3, a mitochondrial protein, immunostaining increased in cell nuclei of tissues in response to kumiss treatments. The oxidative stress induced by DMH was determined to increase plasma 8-OH-2-deoxyguanosine, tissue oxidative stress index, and total oxidant capacity levels. Kumiss supplement was identified to reduce these levels and increase tissue total antioxidant capacity and reduced glutathione levels. Clarifying the molecular relationship between intracellular changes in the locations of SIRT2 and SIRT3 and oxidative stress might be important with regards to developing new medical treatments in the future. The kumiss may show a protective effect against DMH-induced damage by regulating the expression of sirtuin proteins and by protecting antioxidant system.

Evaluation of immunologic and intestinal effects in rats administered an E 171-containing diet, a food grade titanium dioxide (TiO2).

The toxicity of dietary E 171, a food grade titanium dioxide was evaluated. A recent study reported rats receiving E 171 in water developed inflammation and aberrant crypt foci (ACF) in the gastrointestinal tract. Here, rats received food containing E 171 (7 or 100 days). The 100-day study included feeding E 171 after dimethylhydrazine (DMH) or vehicle only pretreatment. Food consumption was similar between treatment groups with maximum total cumulative E 171 exposure being 2617 mg/kg in 7 days and 29,400 mg/kg in 100 days. No differences were observed due to E 171 in the percentage of dendritic, CD4+ T or Treg cells within Peyer's patches or the periphery, or in cytokine production in plasma, sections of jejunum, and colon in 7- or 100-day E 171 alone fed rats. Differences were observed for IL-17A in colon (400 ppm E 171 + DMH) and IL-12p70 in plasma (40 ppm E 171 + DMH). E 171 had no effect on histopathologic evaluations of small and large intestines, liver, spleen, lungs, or testes, and no effects on ACF, goblet cell numbers, or colonic gland length. Dietary E 171 administration (7- or 100-day), even at high doses, produced no effect on the immune parameters or tissue morphology.

Markedly enhanced micronucleus induction by 1,2-dimethylhydrazine dihydrochloride in colonic cells of rats with bacterial colonization in the intestine.

To investigate how intestinal bacteria affect host cytogenetic alterations in the early initiation step of colon carcinogenesis, we conducted a comet assay and micronucleus (MN) test using germ-free (GF) and conventionalized (Cvd) rats after a single subcutaneous injection of the carcinogen, 1,2-dimethylhydrazine dihydrochloride (DMH). DNA damage was also determined in the liver in comet assays, as DMH is metabolized and activated in this organ. The time-response patterns of DNA damage in the liver and colon were similar in both rats, and maximum values were observed at 3 h after the treatment. In contrast, the maximum frequency of micronucleated (MNed) colonic cells was markedly higher in the Cvd rats than in the GF rats and was observed after 72 h and 120 h, respectively. The frequency of MNed cells in non-treated animals was similar in the GF and Cvd rats. In addition, we determined time-responses in the incidence of apoptosis and cell proliferation indices, i.e., the numbers of BrdU-labeled cells, mitotic cells in the crypts, and crypt column heights, using histological sections of the colons in these rats. Maximal incidence of apoptosis was observed at 6 and 24 h in the Cvd and GF rats, respectively. The value in the Cvd rats tended to be higher than that in the GF rats. Cell proliferation was suppressed until 24 and 48 h in the Cvd and GF rats, respectively, and increased subsequently. The rebound response of cell proliferation was more pronounced and occurred earlier in the Cvd rats than that in the GF rats. We demonstrated that cytogenetic alterations other than DNA damage, particularly the MNed colonic cell induction by DMH, were markedly enhanced in rats with bacterial colonization in the intestine compared to those in GF rats.

Precancerous ACF induction affects their regional distribution forsaking oxidative stress implication in 1,2-dimethylhydrazine-induced colon carcinogenesis model.

Colon cancer is thought to develop in a stepwise fashion. In this study, the relationship between aberrant crypt foci (ACF) regional distribution and oxidative stress evolution was studied in a murine model of initial colon carcinogenesis induced by dimethylhydrazine (DMH). Mice were given 2 weekly subcutaneous injections of DMH (20 mg/kg) and killed at the 10th, 12th or 14th week. ACF was scored for number, distribution and crypt multiplicity after methylene-blue coloration and histologically analyzed afterwards. Oxidative stress evaluation was assessed through myeloperoxidase activity (MPO), nitric oxide (NO), L-ornithine and malondialdehyde (MDA) levels as well as antioxidant CAT, SOD and GSH. DMH treatment showed a shift from small to large ACF but also from distal to proximal colon between week 10 and 14 (p < 0.05). This was further illustrated histologically with crypt disruption and mucin depletion. Oxidative stress imbalance was observed in all DMH-treated groups. All markers (MPO, MDA and NO) peaked at week 12 (p < 0.01) and decreased at week 14 (p < 0.05) while L-ornithine decreased through all protocol (p < 0.01). Antioxidants decreased in all points (p < 0.05) but only GSH increased at week 14 (p < 0.05). This work provided insight to response-patterns of oxidative stress between distal and proximal colon, showing for the first time a decreasing implication during the development process and suggesting other inflammatory, immunologic or microbiota implication as factors to be considered during chemotherapy approaches.

Orientin, a C-glycosyl dietary flavone, suppresses colonic cell proliferation and mitigates NF-κB mediated inflammatory response in 1,2-dimethylhydrazine induced colorectal carcinogenesis.

Orientin, a C-glycosyl dietary flavone profusely found in rooibos tea and passion fruit have gained much attention owing to their multiple pharmacological potentials. The present study intends to investigate the anti-proliferative and anti-inflammatory efficacy of Orientin in 1,2-dimethyl hydrazine (DMH) induced colorectal cancer (CRC) in rats. Animals were arbitrarily segmented into six groups and fed with high-fat diet. Group 1 served as control. Group 2 received weekly subcutaneous injections of DMH (20 mg/kg b.w.), for first 15 weeks. Group 3 administered with Orientin (10 mg/kg b.w., i.p.) whereas Groups 4-6 treated with Orientin in three phases, namely initiation (along with DMH), post-initiation (post-DMH injection) and entire period. Orientin ameliorates tumor marker levels significantly (p < 0.05) and reinstates the histological changes induced by DMH. The proliferative markers (PCNA and Ki67) were observed to be suppressed significantly (p < 0.05) in Orientin treated rats. Orientin abrogates (p < 0.05) the inflammatory mast cells and diminishes the expression of pro-inflammatory NF-κB and cytokines (TNF-α and IL-6). It also down-regulates over expression of inflammatory inducible enzymes (iNOS and COX-2) significantly (p < 0.05) and further substantiated by GLIDE XP and QPLD studies. Overall results promptly elucidate the anti-proliferative and anti-inflammatory efficacy of Orientin against CRC. Orientin can be developed as a promising chemotherapeutic agent, on further validation of other molecular mechanisms.

Inositol hexaphosphate suppresses colorectal cancer cell proliferation via the Akt/GSK-3ß/ß-catenin signaling cascade in a 1,2-dimethylhydrazine-induced rat model.

Colorectal cancer (CRC) is common worldwide, and most treatments for CRC have undesirable side effects. Many researchers have demonstrated that inositol hexaphosphate (IP6) has potent anticarcinogenic activity against CRC and no apparent toxicity to normal cells. However, the underlying mechanism is still unclear. In this study, we investigated the anticancer and anti-proliferative properties of IP6 in CRC and its possible mechanisms during this chemopreventive process. We examined the expression of genes related to the PI3K/Akt and Wnt pathways at the transcriptional and translational levels in a DMH-induced rat CRC model following IP6 administration. In addition, we also conducted cell proliferation analysis. The results demonstrated that IP6 could inhibit tumors, in terms of tumor incidence, number, weight and volume in DMH-induced rats. Additionally, Akt and c-Myc mRNA levels were significantly decreased. IP6 was also shown to downregulate Akt, pAkt, pGSK-3ß, and c-Myc protein expression and upregulate pß-catenin protein expression. Furthermore, tumor tissues from IP6-treated rats showed decreased proliferation. In conclusion, the anti-proliferative effect of IP6 may be related to crosstalk between the PI3K/Akt and Wnt pathways, revealing a potential mechanism of CRC inhibition by IP6 in our model.

Frequency and Duration Modulate Anticarcinogenic Effects of a Physical Training in the Colon.

Physical exercise has proven protective against colon carcinogenesis. We sought to clarify whether the frequency and duration of physical training were key factors for its anticarcinogenic effects on the colon. Either sedentary or physically trained male Wistar rats (n=82) were either exposed or not to the carcinogen dimethylhidrazine (DMH). The first protocol investigated whether swimming for 60 min in different frequencies modulates antipreneoplastic effects of physical training. Another protocol then explored whether the duration for training 5 times a week impacts on the development of colon preneoplastic lesions. After 8 weeks, serum and colon samples were collected and analyzed afterwards. Swimming once a week for 60 min did not promote those anticarcinogenic effects found in rats trained 5 times weekly. Such weekly sustained physical training not only decreased the development of colon preneoplastic, but also epithelial proliferation, and subepithelial cyclooxygenase 2 (COX-2) expression. Interestingly, a 5 time per week training for less than 60 min was not as protective against colon carcinogenesis as swimming for 90 min. This 90 min training indeed reduced serum cholesterol and triglycerides levels, as well as colonic lipid peroxidation in carcinogen-exposed rats. Our collective data suggest anticarcinogenic effects of physical exercises are potentially promoted when training 5 times a week for at least 60 min.

Pioglitazone hydrochloride: chemopreventive potential and development of site-specific drug delivery systems.

The aim of this study was to investigate the potential of pioglitazone hydrochloride as a promising anticancer agent and then to design and evaluate the colon-targeted delivery system. The role of pioglitazone hydrochloride as a promising anticancer agent was evaluated by in vitro cell line studies and in vivo 1,2-dimethylhydrazine-induced colon carcinogenesis in rats. In order to deliver the drug at site of action, i.e. colon, drug embedded in matrices containing a release retarding polymer (HPMC K4M) and a polysaccharide (locust bean gum) were prepared. These matrix systems were further enteric coated with Eudragit®S100 to minimize the premature drug release in the upper segments of the GIT. In vitro dissolution studies were performed in absence and presence of rat caecal contents on selected batches and samples were analyzed using a validated RP-HPLC method. Hence, the studies led to the conclusion that successful site-specific delivery systems of pioglitazone hydrochloride were developed to improve its therapeutic efficacy in the management of colorectal cancer.

Chemopreventive effects of (-)-hinokinin against 1,2-dimethylhydrazine-induced genotoxicity and preneoplastic lesions in rat colon.

(-)-Hinokinin (1) is a dibenzylbutyrolactone lignan obtained by the partial synthesis of (-)-cubebin. This study reports the antigenotoxic and anticarcinogenic potential of 1 by the comet and aberrant crypt focus assays in the peripheral blood and colon of 4-5-week-old Wistar rats, respectively. The rats were exposed to 1,2-dimethylhydrazine (40 mg/kg) and were treated by gavage with doses of 10, 20, and 40 mg/kg of 1. The results showed that the dose of 40 mg/kg was neither genotoxic nor carcinogenic. In the comet assay, all 1 doses displayed antigenotoxic effects. In addition, this compound (20 and 40 mg/kg) exhibited an anticarcinogenic effect in the aberrant crypt focus assay.

[Hematological effects of the protein kinase inhibitor maleimide derivative in dimethylhydrazin E-induced colorectal carcinogenesis of rats].

The effect of the protein kinase inhibitor maleimide derivative (MI-, 1, 1-(4-Cl-benzyl)-3-CI-4-(CF3-phenylamino)- 1H-pyrrole-2,5-dione) on blood cells of rats with 1,2-dimethylhydrazine (DMH)-induced colon carcinogenesis has been studied. Administration of MI-I1 at 2.7 mg/kg for 20 weeks on DMH-induced carcinogenesis prevents anemia, which is a consequence of cancer and complicates it. This is confirmed by a reduction in the numberofreticulocytes (0,19(0,15;0,21)x 10(12)/1)and restoration of mean corpuscular hemoglobin (18,02 (17,44;19,03) pg) and mean corpuscular hemoglobin concentration (309,42 (292,38;318,27) g/L) to a control value (0,17 (0,15;0,19), 18,31 (17,95;18,45), 310,78 (306,25;316,18), respectively) in contrast to the group DMH (0,28 (0,24;0,39); 17,50 (17,00;17,96); 288,10 (284,71;303,73), respectively). MI-I normalizes the number of monocytes (1,40 (0,95;2,50)x 10(9)/L) and platelets (646,32 (575,23;700,50)xl10(9)/L) in the blood after 26 weeks ofexperiment; in the DMH group, the values are significantly higher (1,97 (1,52;2,58), 783,90 (687,64;922,27), respectively) as compared to control group (1,23 (0,94; 1,68), 629,34 (590,19;711,48), respectively). MI- 1 reduces the involvement of these cells in the tumors progression and metastasis. Reduction of the monocytosis and thrombocytosis may be mediated by: 1) a decrease in the number and size of tumors and, consequently, the influence of their cytokines on hematopoietic tissue; 2) suppression of proliferation and differentiation of hematopoietic progenitor cells through inhibiting of receptor protein kinases of vascular endothelial and epidermal growth factors and non-receptor PDKI-, Src- and Syk- kinases, that are involved in hematopoiesis and carcinogenesis.

Probiotic Escherichia coli CFR 16 producing pyrroloquinoline quinone (PQQ) ameliorates 1,2-dimethylhydrazine-induced oxidative damage in colon and liver of rats.

Inflammation of the gastrointestinal tract is associated with reactive oxygen species (ROS) genesis. Alleviation of oxidative stress is achieved by using antioxidants and probiotics. Present study investigates a synergistic effect of the probiotic Escherichia coli CFR 16 containing Vitreoscilla haemoglobin gene (vgb), green fluorescent protein (gfp) gene and pyrroloquinoline quinone (pqq) gene cluster on oxidative stress induced by 1,2-dimethylhydrazine (DMH). Adult virgin Charles foster male rats (3-4 months) weighing 200-250 g were administered with DMH (25 mg/kg body weight, s.c.) twice a week for eight consecutive weeks. Rats receiving only DMH dose showed increased lipid peroxidation in liver and intestinal tissues with reduced activity of antioxidant enzymes, i.e. superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). Oral dose of E. coli CFR 16::vgb-gfp harbouring pqq gene cluster increased rat faecal PQQ concentration by twofold, reduced lipid peroxidation and retained SOD, CAT and GPx activities close to normal levels in liver and colonic tissues following DMH treatment. In addition, significant protection was found in colonic histological sections of these rat groups. This study demonstrates a protective efficacy in the following order: E. coli CFR 16 < E. coli CFR 16::vgb-gfp < vitamin C = PQQ < E. coli CFR 16::vgb-gfp (pqq).

Calcium inhibits promotion by hot dog of 1,2-dimethylhydrazine-induced mucin-depleted foci in rat colon.

Epidemiology suggests that processed meat is associated with colorectal cancer risk, but few experimental studies support this association. We have shown that a model of cured meat made in a pilot workshop promotes preneoplastic lesions, mucin-depleted foci (MDF) in the colon of rats. This study had two aims: to check if real store-bought processed meats also promote MDF, and to test if calcium carbonate, which suppresses heme-induced promotion, can suppress promotion by processed meat. A 14-day study was done to test the effect of nine purchased cured meats on fecal and urinary biomarkers associated with heme-induced carcinogenesis promotion. Fecal water from rats given hot dog or fermented raw dry sausage was particularly cytotoxic. These two cured meats were thus given to rats pretreated with 1,2-dimethylhydrazine, to evaluate their effect on colorectal carcinogenesis. After a 100-days feeding period, fecal apparent total N-nitroso compounds (ATNC) were assayed and colons were scored for MDF. Hot dog diet increased fecal ATNC and the number of MDF per colon compared with the no-meat control diet (3.0 ± 1.7 vs. 1.2 ± 1.4, p < 0.05). In a third study, addition of calcium carbonate (150 µmol/g) to the hot dog diet decreased the number of MDF/colon and fecal ATNC compared with the hot dog diet without calcium carbonate (1.2 ± 1.1 vs. 2.3 ± 1.4, respectively, p < 0.05). This is the first experimental evidence that a widely consumed processed meat promotes colon carcinogenesis in rats. It also shows that dietary prevention of this detrimental effect is possible.

Alterations in mitochondrial membrane in chemopreventive action of fish oil.

Mitochondria are major regulators of pathways related to tumorigenesis; therefore, mitochondrial membrane characteristics and associated cell signaling events were evaluated with different ratios of fish oil (FO) and corn oil (CO) in experimental colon carcinogenesis. Treatment with carcinogen 1,2-dimethylhydrazine (DMH) altered reactive oxygen species (ROS), Ca(2+), and membrane characteristics, which resulted in an elevation in apoptosis in initiation phase and reduction in post-initiation phase. FO+CO(2.5:1)+DMH treatment, however, altered mitochondrial membrane parameters, ROS, and Ca(2+) to increase apoptosis in both phases, whereas FO+CO(1:1)+DMH treatment enhanced apoptosis only in post-initiation phase suggesting that FO supplementation in higher ratio has better chemopreventive efficacy.

Sustained proliferation and resistance to apoptosis after a cytotoxic insult are early alterations in rat colon carcinogenesis.

To study the early alterations in carcinogenesis, we determined apoptosis and proliferation in rat mucin depleted foci (MDF), precancerous lesions in the colon under basal conditions and 24 h after treatment with 1,2-dimethylhydrazine (DMH), which induces apoptosis in the colon. Spontaneous apoptosis in MDF was higher than in normal mucosa (Apoptotic Index was 1.61 ± 0.30 and 0.21 ± 0.02 in MDF and normal mucosa, respectively, mean ± SE, p < 0.05). DMH (30 and 75 mg/kg) increased apoptosis in both normal mucosa and MDF (up to 20 times higher compared to basal levels in normal mucosa, but only two times in MDF). MDF had a higher and deregulated pattern of proliferation along the crypt compared to normal mucosa. After DMH, proliferation in normal mucosa was significantly depressed, but it did not vary in MDF. Survivin-Birc5 regulating apoptosis and proliferation was significantly over-expressed (RT-qPCR and immunohistochemistry experiments) in MDF vs. normal mucosa, but did not vary in response to DMH. The expression of the pro-apoptotic protein Bak did not vary in normal mucosa and MDF. Since inflammation is present in MDF, which may hamper apoptosis, we studied the effect of pre-treatment with aspirin (600 ppm in the diet for 10 days). No significant effects of aspirin were observed. In conclusion, MDF had a higher spontaneous apoptosis and proliferation coupled with a reduced response to apoptotic stimuli from cytotoxic compounds. Survivin over-expression in MDF indicates that this is an early event in colon carcinogenesis and suggests that down-regulation of Survivin may represent a strategy for cancer prevention.

Bimodal visualization of colorectal uptake of nanoparticles in dimethylhydrazine-treated mice.

AIM: To investigate colorectal uptake of solid lipid nanoparticles (SLNs) in mice receiving different doses of 1,2-dimethylhydrazine (DMH) using magnetic resonance (MR) and laser-scanning confocal fluorescence microscope (LSCFM) imaging. METHODS: Eight mice were sacrificed in a pilot study to establish the experimental protocol and to visualize colorectal uptake of SLNs in normal mice. Gadopentetate dimeglumine and fluorescein isothiocyanate (FITC)-loaded SLN (Gd-FITC-SLN) enemas were performed on mice receiving DMH for 10 wk (group 1, n = 9) or 16 wk (group 2, n = 7) and FITC-SLN enema was performed on 4 DMH-treated mice (group 3). Pre- and post-enema MR examinations were made to visualize the air-inflated distal colorectum. Histological and LSCFM examinations were performed to verify colorectal malignancy and to track the distribution of SLNs. RESULTS: Homogeneous enhancement and dense fluorescence (FITC) deposition in colorectal wall were observed in normal mice and 1 DMH-treated mouse (group 1) on fluid attenuated inversion recovery (FLAIR) and LSCFM images, respectively. Heterogeneous mural enhancement was found in 6 mice (4 in group 1; 2 in group 2). No visible mural enhancement was observed in the other mice. LSCFM imaging revealed linear fluorescence deposition along the colorectal mucosa in all groups. Nine intraluminal masses and one prolapsed mass were detected by MR imaging with different enhancement modes and pathologies. Interstitial FITC deposition was identified where obvious enhancement was observed in FLAIR images. Bladder imaging agent accumulations were observed in 11 of 16 DMH-treated mice of groups 1 and 2. CONCLUSION: There are significant differences in colorectal uptake and distribution of SLNs between normal and DMH-treated mice, which may provide a new mechanism of contrast for MR colonography.