Evaluation of the Chemopreventive Activity of Grape Skin Extract Using Medium-term Oral Carcinogenesis Assay Induced by 4-Nitroquinoline 1-Oxide.

BACKGROUND/AIM: The aim of this study was to evaluate the chemoprotective potential of grape skin extract following rat tongue carcinogenesis induced by 4-nitroquinoline 1-oxide (4NQO). MATERIALS AND METHODS: Male Wistar rats were distributed into four groups (n=5, per group): Control Group: free access to commercial diet and drinking water for 12 weeks; 4NQO Group: received 4NQO diluted in drinking water daily, for 12 weeks; Grape Skin Extract Group: free access to water and received grape skin extract incorporated with diet for 12 weeks; 4NQO + Grape Skin Extract Group: received 4NQO in drinking water daily and grape extract incorporated with diet for 12 weeks. RESULTS: Animals treated with grape skin extract revealed a significant reduction in epithelial dysplasia. Also, 8-hydroxy-2'-deoxyguanosine (8-OHdG) and ki-67 immunoexpression was reduced in animals treated with grape skin extract. Western blot analysis showed a significant decrease of p-NFκB p50 and MyD88 protein expression in the groups treated with grape skin extract. Copper-zinc superoxide dismutase, manganese superoxide dismutase, and catalase gene expression did not present any statistically significant differences (p>0.05). CONCLUSION: Grape skin extract displayed chemopreventive activity in oral carcinogenesis assays as depicted by its antioxidant, anti-proliferative and anti-inflammatory properties.

Therapeutical effects of vaccine from Trypanosoma cruzi amastigote surface protein 2 by simultaneous inoculation with live parasites.

The aim of this study was to evaluate the efficacy of vaccine using replication-deficient human recombinant Type 5 replication-defective adenoviruses (AdHu5) carrying sequences of the amastigote surface protein 2 (ASP2) (AdASP2) in mice infected with the Trypanosoma cruzi ( T cruzi) Y strain. A total of 16 A/Sn mice female were distributed into four groups, as follows (n = 4 per group): Group 1 - Control Group (CTRL); Group 2 - Infected Group (TC): animals were infected by subcutaneous route with 150 bloodstream trypomastigotes of T cruzi Y strain; Group 3 - Immunized Group (AdASP-2): animals were immunized by intramuscular injection (im) route with 50 µL of AdSP-2 (2 × 10 8 plaque forming units [pfu]/cam) at day 0; Group 4-Immunized and Infected Group (AdASP-2+TC): animals were immunized by im route with 50 µL of ASP-2 (2 × 10 8 pfu/cam) and infected by T cruzi at the same day (day 0). It was observed a significant decrease of nests in the group that was immunized with AdASP-2 and infected on the same day. Tumor necrosis factor alpha (TNF-α) and inducible nitric oxide synthase (iNOS) gene expressions showed a significant increase in the AdASP-2+TC group when compared to TC group, but it was noted that Cyclooxygenase-2 (Cox-2) was increased in TC group when compared to AdASP-2+TC group. Increase of matrix metalloproteinases-2 (MMP-2) and decrease of MMP-9 immunoexpression in the AdASP-2+TC group was noticed as well. Oxidative DNA damage was present in myocardium for AdASP-2+TC group as a result of 8-hydroxydeoxyguanosine immunoexpression. Taken together, our results highlighted an increased oxidative stress, MMP-2 activity and inflammatory host response promoted by AdASP-2 against T cruzi infection.

Therapeutic effects of vaccine derived from amastigote surface protein-2 (ASP-2) against Chagas disease in mouse liver.

This study investigated the efficacy of the vaccine in liver of mice infected with the Trypanosoma cruzi (T. cruzi) and immunized with AdASP-2. For this purpose, histopathological analysis and gene expression of COX-2, TNF-alpha, TNFR, iNOS, cytochrome C, caspase-3, TLR4, IL-6 and IL10 were evaluated. The following groups were used in this study: Group 1 - Control Group (CTRL) animals received AdßGal vehicle; Group 2 - Infected Group (TC) animals were infected with T. cruzi; Group 3 - Immunized Group (AdASP-2): animals were immunized by AdASP-2 vaccine; Group 4 - Immunized and Infected Group (AdASP-2+TC) animals were infected with T. cruzi and immunized by AdSP-2 vaccine. A significant decrease of amastigote nests was noticed in the group of animals that were immunized with AdASP-2 and infected on the same day. COX-2 and TNF-alpha gene expressions increased in TC group, whereas TNF-alpha decreased in the TC+AdASP-2 group. TNFR expression was high in AdASP-2+TC group. iNOS expression was high for all experimental groups whereas cytochrome C decreased for all experimental groups. Caspase 3 increased in TC and TC+AdASP-2 groups. The gene expression of TLR4 and IL-10 showed an increase in AdASP-2+TC group. Finally, hepatic fibrosis was noticed to TC and AdASP-2 + TC groups. Taken together, our results demonstrated that vaccination with AdASP-2 was effective against the acute phase of experimental Chagas disease as a result of a more powerful and rapid immune response closely related to expression of some inflammatory genes, such as iNOS, TNF-alpha, TLR 4, and IL-10.

Burn injury induces histopathological changes and cell proliferation in liver of rats.

AIM: To investigate effects of severe burn injury (BI) in rat liver through the histopathological and inflammatory markers analysis. METHODS: Forty-two male Wistar rats were distributed into two groups, control (C) and subjected to scald BI (SBI). The animals were euthanized one, four and 14 d post sham or 45% of the total body surface BI. Liver fragments were submitted to histopathological, morphoquantitative (hepatocyte area and cell density), ciclooxigenase-2 (COX-2) immunoexpression, and gene expression [real-time polymerase chain reaction for tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS) and caspase-3] methods. RESULTS: Histopathological findings showed inflammatory process in all periods investigated and hepatocyte degeneration added to increased amount of connective tissue 14 d post injury. Hepatocyte area, the density of binucleated hepatocytes and density of sinusoidal cells of SBI groups were increased when compared with control. COX-2 immunoexpression was stronger in SBI groups. No differences were found in TNF-α, iNOS and caspase-3 gene expression. CONCLUSION: BI induces histopathological changes, upregulation of COX-2 immunoexpression, and cell proliferation in liver of rats.

Antioxidant activity of apple extract protects against rat tongue carcinogenesis induced by 4-nitroquinoline 1-oxide.

Several studies have shown that apple (Malus sp.) has many components able to exert chemopreventive activity. The aim of this study was to evaluate the chemopreventive potential of apple extract following medium-term oral carcinogenesis assay induced by 4-nitroquinoline 1-oxide (4NQO) by means of histopathological analysis and gene expression of antioxidant enzymes, such as CuZnSOD, MnSOD and catalase. A total of 30 male Wistar rats were distributed into five groups, as follows (n = 6 per group): Group 1 - negative control group (non-treated group); Group 2 - received 4NQO during 8 weeks in drinking water and treated with apple extract by gavage between the 1st and 4th weeks daily (initiation phase); Group 3 - received 4NQO for 8 weeks in drinking water and treated with apple extract by gavage between the 5th and 8th weeks daily (promotion phase); Group 4 - received apple extract by gavage for eight consecutive weeks only; and Group 5 - received 4NQO for 8 weeks in drinking water daily. Histopathological analysis revealed that apple extract protect oral lesions induced by 4NQO at initiation or promotion phase. Higher gene expression of CuZnSOD and MnSOD enzymes were noticed in groups treated with apple extract as well. Taken together, our results demonstrate that the apple extract is able to modulate medium-term oral carcinogenesis assay as a result of antioxidant activity.

Temporal study following burn injury in young rats is associated with skeletal muscle atrophy, inflammation and altered myogenic regulatory factors.

INTRODUCTION: Burn injury (BI) greater than 40% has been associated with protein catabolism and it is characterized by a hypermetabolic response followed for muscle loss. OBJECTIVE: The purpose of this study was to investigate the temporal effects of extensive experimental BI in the skeletal muscle distant from lesion, through morphological analysis, expression of genes related to muscle atrophy, inflammation and the myogenic regulatory factors. MATERIALS AND METHODS: A total of 60 young male wistar rats were distributed into two groups, control (C) and subjected to scald burn injury (SBI). The animals were euthanized 1, 4 and 14 days post-sham or 45% of the total body surface BI. The medial head of gastrocnemii muscles were submitted to histopathological, morphometric (muscle fibers area and density), MyoD and myogenin immunoexpression, and gene expression for TNF-α, iNOS and E3 ubiquitin ligases (MuRF1 and MAFbx). RESULTS: Histopathological findings were consistent with increased amount of connective tissue and inflammatory process. Muscle fiber area of SBI groups was smaller than C and no differences were found in fiber muscle density. TNF-α was higher in SBI groups, one and 14 days post-injury; iNOS expression was higher on the first and fourth day post-injury. MuRF-1 was higher on the day four and MAFbx on the day 14. CONCLUSION: In conclusion, BI causes inflammation, atrophy and myogenesis stimulation in muscle as a result of systemic host response.

Mimosa (Mimosa caesalpiniifolia) prevents oxidative DNA damage induced by cadmium exposure in Wistar rats.

The Mimosa (Mimosa caesalpiniifolia) is a plant native from South America; it is used in the traditional medicine systems for treating bacterial, fungal, parasitic and inflammatory conditions. The aim of this study was to evaluate the antigenotoxic and antioxidant activities induced by mimosa (M. caesalpiniifolia) in multiple rodent organs subjected to intoxication with cadmium chloride. A total of 40 Wistar rats (8 weeks old, 250 g) were distributed into eight groups (n = 5), as follows: Control group (non-treated group, CTRL); Cadmium exposed group (Cd); cadmium exposure and treated with extract at 62.5 mg/kg/day; cadmium exposure and treated with extract at 125 mg/kg/day; cadmium exposure and treated with extract at 250 mg/kg/day; cadmium exposure and treated with ethyl acetate fraction at 62.5 mg/kg/day. For evaluating the toxicogenetic potential of mimosa, two groups were included in the study being treated with extract at 250 mg/kg/day and acetate fraction of mimosa at 62 mg/kg/day, only. Extract of mimosa at concentrations of 62.5 and 125 mg decreased DNA damage in animals intoxicated with cadmium when compared to cadmium group. In a similar manner, treatment with ethyl acetate fraction of mimosa at 62.5 mg concentration in animals previously exposed to cadmium reduced genetic damage in peripheral blood cells. In a similar manner, the treatment with ethyl acetate fraction reduced DNA damage in liver cells. Oxidative DNA damage was reduced to animals exposed to cadmium and treated with 125 mg of extract as well as those intoxicated to cadmium and treated with 62.5 of acetate fraction of mimosa. Taken together, our results indicate that mimosa prevents genotoxicity induced by cadmium exposure in liver and peripheral blood cells of rats as a result of antioxidant activity.

Antimutagenic and antigenotoxic potential of grape juice concentrate in blood and liver of rats exposed to cadmium.

The aim of this study was to evaluate the antimutagenic and antigenotoxic potential of grape juice concentrate in rodent organs exposed to cadmium chloride intoxication. A total of 15 Wistar rats were distributed into three groups (n = 5), as follows: control group (CTRL; nontreated group), cadmium group (Cd), and cadmium-grape juice group (Cd + GJ). Exposed animals received intraperitoneal injection of cadmium chloride (1.2 mg/kg body weight) diluted in water and, after 15 days, Cd + GJ group received grape juice concentrate for 15 days, by gavage (0.8 mL, 1.18 mg of polyphenols kg(-1) day(-1)). Grape juice concentrate was able to decrease genotoxic effects induced by cadmium in peripheral blood and liver cells as depicted by single cell gel (comet) and micronucleus assays. A decrease for anti-8-hydroxy-20-deoxyguanosine (8OHdG) expression in hepatocytes of animals exposed to cadmium and treated with grape juice concentrate was also detected. Higher CuZn-SOD activity was observed in liver cells of the Cd + GJ group. No remarkable differences were seen regarding Mn-SOD activity among groups. Taken together, our results demonstrate that grape juice concentrate was able to exert antimutagenic and antigenotoxic activities in blood and liver cells of rats exposed to cadmium.

Genotoxicity and cytotoxicity induced by municipal effluent in multiple organs of Wistar rats.

The aim of this study was to evaluate cytotoxicity and genotoxicity in multiple organs of rats induced by municipal effluent released by submarine outfall in city of Santos. A total of 20 male Wistar rats were exposed to effluents by drinking water ad libitum at concentrations of 0, 10, 50, and 100 % for 30 days. Microscopic analysis revealed severe lesions such as necrosis and hemorrhagic areas in liver and kidney from animals exposed to effluent at 50 and 100 % concentration. DNA damage in peripheral blood, liver, and kidney cells were detected by comet assay at higher concentrations of effluent. Moreover, a decrease DNA repair capacity was detected in liver cells. Significant statistical differences (p<0.05) for micronucleated cells from liver were noticed at 50 % concentration of effluent. Taken together, our results demonstrate that municipal effluent is able to induce cytotoxicity and genotoxicity in multiple organs of Wistar rats.

Chemopreventive activity of apple extract following medium-term oral carcinogenesis assay induced by 4-nitroquinoline-1-oxide.

OBJECTIVE: The aim of this study was to evaluate the chemopreventive activity of an apple extract following medium-term oral carcinogenesis assay induced by 4-nitroquinoline-1-oxide (4NQO). METHODS: A total of 30 male Wistar rats were distributed into five groups as follows (n=6 per group): Group 1, negative control group (non-treated group); Group 2, received 4NQO during 8 weeks in drinking water and treated with apple extract at 1% by gavage between the first and fourth weeks daily (initiation phase); Group 3, received 4NQO for 8 weeks in drinking water and treated with apple extract by gavage at 1% between the fifth and eighth weeks daily (promotion phase); Group 4, received apple extract at 1% by gavage for 8 consecutive weeks only; and Group 5, received 4NQO for 8 weeks in drinking water daily. RESULTS: Histopathological analysis revealed decreased hyperplasic lesions in Group 2 when compared with Group 5. Likewise, decreased dysplastic lesions in Group 3 were observed when compared with Group 5. In Groups 2 and 3, decreased COX-2 and TNF-alpha gene expressions were observed when compared with Group 5. Cytochrome c and caspase 3 levels increased in Groups 2 and 3 when compared with Group 5. CONCLUSION: In conclusion, our results demonstrate that apple extract suppresses rat tongue carcinogenesis as a result of anti-inflammatory activity and apoptosis through the intrinsic mitochondrial pathway.

Effective targeting of the epidermal growth factor receptor (EGFR) for treating oral cancer: a promising approach.

Oral cancer is a serious problem growing in incidence in many parts of the world; it is considered the sixth most common cancer and despite sophisticated surgical and radiotherapeutic modalities, oral squamous cell carcinoma, which represents 90% of oral cancers, is characterized by poor prognosis and a low survival rate. The Epidermal growth factor receptor family of receptor tyrosine kinases (RTK) comprises of four distinct receptors: the EGFR (also known as ErbB-1/HER1), ErbB-2 (neu, HER2), ErbB-3 (HER3) and ErbB-4 (HER4). Several studies have been published on the role of EGFR in the pathogenesis of oral carcinoma. The aim of the present review is to describe the role of EGFR pathway in oral cancer with special focus on its role during the carcinogenesis process as a result of therapeutic approaches of EGFR in oral cancer. The EGFR is a 170-kDa cell-surface protein involved in many biological processes, such as proliferation, migration, DNA synthesis and adhesion. Overexpression of EGFR results in a poor prognosis in oral cancer and its activation is associated with the malignant phenotype, inhibition of apoptosis and increased metastatic potential. EGFR variations and mutations have been correlated with tumor formation, and possibly alter the therapeutic efficacy of EGFR inhibitors.

Anti-tumor activity of grape juice concentrate in the rat tongue two-stage initiation-promotion protocol induced by 4-nitroquinoline 1-oxide.

The aim of this study was to evaluate the anti-tumor activity of grape juice concentrate following medium-term oral carcinogenesis assay induced by 4-nitroquinoline 1-oxide (4NQO). A total of 30 male Wistar rats were distributed into five groups, as follows (n = 6 per group): Group 1 - negative control group (non-treated group); Group 2 - received grape juice concentrate at 1% dose by gavage for eight consecutive weeks; Group 3 - received 4NQO for 8 weeks at 20 ppm dose in drinking water daily; Group 4 - received 4NQO at 20 ppm dose during 8 weeks in drinking water and treated with grape juice concentrate at 1% dose orally by gavage for first 4 weeks after 4-NQO administration; Group 5 - received 4NQO at 20 ppm dose for 8 weeks in drinking water and treated with grape juice concentrate at 1% dose orally by gavage between the 5th and 8th weeks daily. Histopathological analysis revealed a decrease in hyperplasic and dysplastic lesions in Group 4. Groups 4 and 5 showed decreased COX-2 and TNF-alpha and eNOS gene expression. Grape juice concentrate also increased SOD Cu/Zn and catalase expression. However, Ki-67 immunoexpression was reduced at the promotion step of oral carcinogenesis (G5). Taken together, our results demonstrate that grape juice concentrate modulates rat tongue carcinogenesis as a result of anti-inflammatory activity, antioxidant activity and down-regulation of oral cells proliferation.