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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

Folate deficiency provides protection against colon carcinogenesis in DNA polymerase beta haploinsufficient mice.

Aging and DNA polymerase beta deficiency (beta-pol(+/-)) interact to accelerate the development of malignant lymphomas and adenocarcinoma and increase tumor bearing load in mice. Folate deficiency (FD) has been shown to induce DNA damage repaired via the base excision repair (BER) pathway. We anticipated that FD and BER deficiency would interact to accelerate aberrant crypt foci (ACF) formation and tumor development in beta-pol haploinsufficient animals. FD resulted in a significant increase in ACF formation in wild type (WT) animals exposed to 1,2-dimethylhydrazine, a known colon and liver carcinogen; however, FD reduced development of ACF in beta-pol haploinsufficient mice. Prolonged feeding of the FD diet resulted in advanced ACF formation and liver tumors in wild type mice. However, FD attenuated onset and progression of ACF and prevented liver tumorigenesis in beta-pol haploinsufficient mice, i.e. FD provided protection against tumorigenesis in a BER-deficient environment in all tissues where 1,2-dimethylhydrazine exerts its damage. Here we show a distinct down-regulation in DNA repair pathways, e.g. BER, nucleotide excision repair, and mismatch repair, and decline in cell proliferation, as well as an up-regulation in poly(ADP-ribose) polymerase, proapoptotic genes, and apoptosis in colons of FD beta-pol haploinsufficient mice.

Non steroidal anti-inflammatory drugs modulate the physicochemical properties of plasma membrane in experimental colorectal cancer: a fluorescence spectroscopic study.

According to "fluid-mosaic model," plasma membrane is a bilayer constituted by phospholipids which regulates the various cellular activities governed by many proteins and enzymes. Any chemical, biochemical, or physical factor has to interact with the bilayer in order to regulate the cellular metabolism where various physicochemical properties of membrane, i.e., polarization, fluidity, electrostatic potential, and phase state may get affected. In this study, we have observed the in vivo effects of a pro-carcinogen 1,2-dimethylhydrazine dihydrochloride (DMH) and the two non steroidal anti-inflammatory drugs (NSAIDs); sulindac and celecoxib on various properties of the plasma membrane of colonocytes, i.e., electric potential, fluidity, anisotropy, microviscosity, lateral diffusion, and phase state in the experimentally induced colorectal cancer. A number of fluorescence probes were utilized like membrane fluidity and anisotropy by 1,6-diphenyl-1,3,5-hexatriene, membrane microviscosity by Pyrene, membrane electric potential by merocyanine 540, lateral diffusion by N-NBD-PE, and phase state by Laurdan. It is observed that membrane phospholipids are less densely packed and therefore, the membrane is more fluid in case of carcinogenesis produced by DMH than control. But NSAIDs are effective in reverting back the membrane toward normal state when co-administered with DMH. The membrane becomes less fluid, composed of low electric potential phospholipids whose lateral diffusion is being prohibited and the membrane stays mostly in relative gel phase. It may be stated that sulindac and celecoxib, the two NSAIDs may exert their anti-neoplastic role in colorectal cancer via modifying the physicochemical properties of the membranes.

PI3-kinase/Wnt association mediates COX-2/PGE(2) pathway to inhibit apoptosis in early stages of colon carcinogenesis: chemoprevention by diclofenac.

In addition to having anti-inflammatory properties, non-steroidal anti-inflammatory drugs (NSAIDs) inhibit neoplastic cell proliferation by inducing apoptosis. Inhibition of cyclooxygenase-2 (COX-2) seemed to be the principal target of NSAIDs, as it is overexpressed in several cancers and catalyzes the synthesis of prostaglandin E2 (PGE2), the critical pro-inflammatory molecule. A major role for phosphatidylinositol-3 kinase (PI3-kinase) pathway activation in human tumors has been more recently established. The present study explored the role of PI3-kinase and Wnt molecular pathways in COX-2 and PGE2 production as well as NSAIDs' chemopreventive effect in colon cancer. 1,2-dimethylhydrazine (DMH) was used for experimental colon cancer model in rat and diclofenac as the preferential COX-2 selective chemopreventive agent. Expression of caspase-3 and caspase-9 was checked in the colonic tissue by immunofluorescence. A decrease was seen in their expressions, indicative of inhibition of apoptosis in the present model. COX-2 mRNA expression as well as PGE2 levels was elevated after DMH treatment; however, COX-1 mRNA expression was unaltered as seen by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. DMH also activated PI3-kinase, Akt, Wnt, and ß-catenin expressions but reduced the glycogen synthase kinase-3ß (GSK-3ß) levels. Co-administration of diclofenac with DMH increased the mRNA expression of GSK-3ß while inactivating PI3-kinase, Akt, Wnt, and ß-catenin. The study suggests that activation of PI3-kinase and Wnt signaling is associated with COX-2/PGE2 production and in turn inhibition of apoptosis in colon cancer, while diclofenac targeted these pathways to restore apoptosis in the present system.

Glycine- and proline-rich glycoprotein regulates the balance between cell proliferation and apoptosis for ACF formation in 1,2-dimethylhydrazine-treated A/J mice.

The objective of this study was to investigate the chemopreventive potentials of glycine- and proline-rich glycoprotein (SNL glycoprotein, 150-kDa) isolated from Solanum nigrum Linne on formation of colonic aberrant crypt foci (ACF) induced by 1,2-dimethylhydrazine (DMH, 20 mg/kg) in A/J mice. Administration of SNL glycoprotein inhibited phosphorylation of extracellular signal-regulated kinase (ERK), expression of colonic proliferating cell nuclear antigen (PCNA), and frequency of colonic ACF in DMH-stimulated mice colon carcinogenesis. In addition, SNL glycoprotein increased expression of cyclin-dependent kinase inhibitors (p21(WAF/Cip1) and p27(Kip1)), whereas reduced expression of precursor form of apoptosis-related proteins [pro-caspase-3 and pro-poly(ADP-ribose)polymerase (PARP)] in the mice. Interestingly, the results in this study revealed that SNL glycoprotein has suppressive effects on activity of nuclear factor-kappa B (NF-kappaB), whereas it has stimulatory effect on the expression of p53, accompanying inhibitory effects on expression of NF-kappaBp50, inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin (IL)-6, and tumor necrosis factor (TNF)-alpha in DMH-stimulated ACF formation. Also, SNL glycoprotein has inhibitory effects on the formation of thiobarbituric acid reactive substances (TBARS), on the production of inducible nitric oxide (NO), and on the release of lactate dehydrogenase (LDH) in the mice plasma. Collectively, our findings in this study suggest that SNL glycoprotein has chemopreventive activity via modulation of cell proliferation and apoptosis in DMH-treated A/J mice.

Membrane fluidity and surface changes during initiation of 1,2 dimethylhydrazine-induced colon carcinogenesis: protection by zinc.

The present study evaluated the modulatory effects of zinc on colonic membrane fluidity and surface abnormalities following 1,2 dimethylhydrazine (DMH)-induced colon carcinogenesis. Rats were segregated into four groups: normal control, DMH treated, zinc treated, DMH + zinc treated. Colon carcinogenesis was initiated through weekly subcutaneous injections of DMH (30 mg/kg body weight) for 8 weeks. Zinc (in the form of zinc sulphate) was supplemented to rats at a dose level of 227 mg/L in drinking water, ad libitum, for the entire duration of the study. Brush border membranes (BBM) were isolated from the colon of rats and the fluidity parameters were assessed by steady-state fluorescence polarization technique using the membrane extrinsic fluorophore 1,6-diphenyl-1,3,5-hexatriene (DPH). The translational diffusion was measured by using the excimer formation of pyrene incorporated in the membrane. The results demonstrated a significant increase in the polarization and anisotropy, accompanied by an increase in order parameter in the membrane preparations from the colon of DMH-injected rats. Further, studies with pyrene fluorophore indicated a marked decrease in membrane microviscosity following DMH treatment. However, the alterations in membrane fluorescence polarization and the fluidity parameters were completely restored following zinc treatment. Drastic alterations in colon surface were noticed after 8 weeks of DMH treatment. However, zinc treatment to DMH-treated rats greatly restored normalcy in the colonic surface. The study concludes that zinc has a strong membrane stabilizing effect and thus has a positive beneficial effect against chemically induced colonic preneoplastic progression in rats.

Effect of Neem (Azadirchta indica) on serum glycoprotein contents of rats administered 1,2 dimethylhydrazine.

The present study was designed to investigate the effects of aqueous Azadirchta Indica leaf extract (AAILE) on serum glycoprotein contents and tumor incidence rate in colon of rats subjected to Dimethylhydrazine (DMH) treatment. Forty rats were divided equally and randomly into four groups viz., Group I (normal control), Group II (DMH-treated), Group III (AAILE) and Group IV (DMH + AAILE treated). Group II and IV animals were injected subcutaneously every week with DMH (30 mg/kg b.wt.) for two durations of 10 and 20 weeks. AAILE was given orally three times a week on alternate days (100 mg/kg b.wt.) to animals belonging to groups III and IV. Blood samples were drawn from all the animals by ocular vein puncture every month for the estimation of Total Sialic Acid (TSA) and Lipid Bound Sialic Acid (LSA), which served as markers for the cancer. No incidence of tumor was recorded in the animals given DMH treatment for 10 weeks. However, DMH treatment for 20 weeks showed 100% tumor incidence. Animals treated with DMH for both the time durations showed a significant increase in the levels of TSA in comparison to normal control, which however were decreased significantly following AAILE supplementation. There was no significant difference between LSA levels of DMH-treated animals and normal controls. The present study suggested that supplementation of AAILE in cancer-bearing animals attenuates considerably the molecular events that initiate the development of tumors.

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.

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.