Electrolyzed Oxidizing Water Modulates the Immune Response in BALB/c Mice Experimentally Infected with Trypanosoma cruzi.

Chagas disease is a major public health problem in Latin America. The mixed Th1/Th2 immune response is required against Trypanosoma cruzi. Electrolyzed oxidizing water (EOW) has been shown to have germicidal efficacy. The objective of this study was to evaluate the EOW effectiveness in T. cruzi-infected BALB/c mice clinically, immunologically, and histologically. The severity of the infection was assessed by parasitaemia, general health condition, mortality, mega syndromes, and histological lesions. IgG, TNF-alpha, IFN-gamma, and IL-1 beta levels were quantified. The EOW administration showed a beneficial effect on parasitaemia, general physical condition, and mortality. High levels of IgG1 at 50 days postinfection were observed. Prophylactic EOW treatment was able to induce a predominantly TH1 immune response based on an IgG2a levels increase at the late acute phase, and a 10-fold increase of INF-gamma in whole acute phase. EOW was able to control the acute phase infection as effectively as benznidazole. Splenomegaly was caused by EOW treatment and lymphadenopathy was stimulated by T. cruzi infection in all groups. Severe tissue damage was not prevented by EOW treatments. Moderate efficacy may be due to immunomodulatory properties and not to a direct toxic effect on the parasite.

Downregulation of sorting nexin 10 is associated with overexpression of miR-30d during liver cancer progression in rats.

As of 2012, liver cancer was the second leading cause of death worldwide, and hepatocellular carcinoma is the most common primary cancer of the liver. The identification of molecules that might be molecular markers or therapeutic targets is urgently needed to improve clinical management. Based on a microarray analysis performed in our laboratory, we selected six genes-namely, ANXA2, DYNLT1, PFKP, PLA2G7, KRT19, and SNX10-as candidates for validation as tumor markers of liver cancer in a rat model. Their patterns of overexpression in preneoplastic lesions and established tumors at 10 different time points between 24 h and 18 months were analyzed to identify putative tumor markers for further studies. We validated the microarray results by quantitative reverse transcription polymerase chain reaction, which revealed high transcriptional expression for five of the genes, consistent with their high protein expression during cancer progression reported in the literature. However, studies of the association of sorting nexin 10 with different types of cancer are limited, prompting further study. The characterization of sorting nexin 10 in preneoplastic lesions and established tumors revealed messenger RNA overexpression and a simultaneous decrease in sorting nexin 10 protein expression. A group of microRNAs related to sorting nexin 10 messenger RNA were selected based on a data analysis conducted using miRDB and microrna.org . An analysis of the expression of these microRNAs revealed an increase in the transcription of microRNA-30d whenever the sorting nexin 10 protein was downregulated. These results suggest that sorting nexin 10 is a potential liver cancer marker exhibiting characteristics of a putative suppressor protein that is likely regulated by microRNA-30d.

Evidence that the anticarcinogenic effect of caffeic acid phenethyl ester in the resistant hepatocyte model involves modifications of cytochrome P450.

Caffeic acid phenethyl ester (CAPE), a natural component of propolis, shows anticarcinogenic properties in the modified resistant hepatocyte model when administered before initiation or promotion of hepatocarcinogenesis process; however, information about the mechanism underlying this chemoprotection is limited. The aim of this work was to characterize the effect of CAPE on cytochrome P450 (CYP), which is involved in diethylnitrosamine (DEN) metabolism during the initiation stage of chemical hepatocarcinogenesis. Male Fischer-344 rats were treated as in the modified resistant hepatocyte model. Liver samples were obtained at four different times: at 12 h after pretreatment with CAPE and at 12 and 24 h and 25 days after DEN administration. Liver damage was determined by histology with hematoxylin and eosin, measurement of total CYP levels and enzyme activity, and gamma-glutamyl transpeptidase-positive (GGT+) staining of hepatocyte foci. CAPE administration prevented DEN-induced necrosis at 24 h. It also decreased O-dealkylation of 7-ethoxy-resorufin (EROD), O-dealkylation of 7-methoxyresorufin (MROD), and 7-pentoxy-resorufin activities at 12 h after its administration and EROD and MROD activities at 12 h after administration of DEN. CAPE treatment decreased GGT+ foci by 59% on day 25. Our results suggest that CAPE modifies the enzymatic activity of CYP isoforms involved in the activation of DEN, such as CYP1A1/2 and CYP2B1/2. These findings describe an alternative mechanism for understanding the ability of CAPE to protect against chemical hepatocarcinogenesis.

Aqueous crude extract of Rhoeo discolor, a Mexican medicinal plant, decreases the formation of liver preneoplastic foci in rats.

There are many plants in Mexico with medicinal properties, some of them used in alternative medicine to treat cancer, such is the case of Rhoeo discolor L. Hér Hance (Commelinaceae family); however, there are not scientific reports that validate their antitumoral property. The present study shows the protective effects of the Rhoeo discolor aqueous crude extract (ACE) against rat liver cancer using the resistant-hepatocyte model. The carcinogenesis protocol consisted on the initiation with N-diethylnitrosamine, followed by the promotion with 2-acetylaminofluorene and a partial hepatectomy. After 24 days, the gamma-glutamyl transpeptidase positive, corresponding to altered hepatocytes foci (AHF), were quantified. Additionally to discard a possible carcinogenic effect of ACE, it was first tested as promoting agent instead 2-acetylaminofluorene, and second, ACE was administered as initiator and promoter instead of the whole carcinogenic treatment. In summary, firstly, ACE administration reduced the number and area of preneoplastic lesions with dose below 20mg/kg body weight and secondly, ACE administration neither presented a promoting or initiator effects nor induced the development of AHF. Results of this investigation justify continuing with further studies of Rhoeo discolor components to develop chemoprevention strategies as an option in the treatment of cancer.

A single dose of caffeic acid phenethyl ester prevents initiation in a medium-term rat hepatocarcinogenesis model.

AIM: To study of the effect of caffeic acid phenethyl ester (CAPE) on the initiation period in a medium-term assay of hepatocarcinogenesis. METHODS: Male Wistar rats were subjected to a carcinogenic treatment (CT) and sacrificed at 25th d; altered hepatic foci (AHF) were generated efficiently. To a second group of rats a single 20 mg/kg doses of CAPE was given 12 h before initiation with CT and were sacrificed at 25th d. We evaluated the expression of preneoplastic markers as gamma-glutamyltranspeptidase (GGT) and glutathione S-transferase type pi protein (GSTp) by histochemistry, RT-PCR and Western blot analyses, respectively. We measured thiobarbituric acid reactive substances (TBARS) in homogenates of liver and used Unscheduled DNA Synthesis (UDS) assay by incorporation of [3H] thymidine (3HdT) in primary hepatocyte cultures (PHC). RESULTS: At 25th d after CT CAPE reduced the observed increase of GGT+AHF by 84% and liver expression of ggt mRNA by 100%. In case of the GSTp protein, the level was reduced by 90%. As indicative of oxidative stress generated by diethylnitrosamine (DEN) 12 h after its administration, we detected a 68% increase of TBARS. When CAPE was administered before DEN, it completely protected from liver TBARS induction. To have an indication of the sole effect of CAPE on initiation, two carcinogens were tested in a UDS assay in PHC, we used methyl-n-nitrosoguanidine as a direct carcinogen and DEN, as indirect carcinogen. In this assay, genotoxic damage caused by carcinogens was abolished at 5 microM CAPE concentration. CONCLUSION: Our results demonstrated that CAPE possesses anti-genotoxic and antineoplastic capabilities, by an anti-oxidative and free-radical scavenging mechanism.

Gene expression profile related to the progression of preneoplastic nodules toward hepatocellular carcinoma in rats.

In this study, we investigated the time course gene expression profile of preneoplastic nodules and hepatocellular carcinomas (HCC) to define the genes implicated in cancer progression in a resistant hepatocyte model. Tissues that included early nodules (1 month, ENT-1), persistent nodules (5 months, ENT-5), dissected HCC (12 months), and normal livers (NL) from adult rats were analyzed by cDNA arrays including 1185 rat genes. Differential genes were derived in each type of sample (n = 3) by statistical analysis. The relationship between samples was described in a Venn diagram for 290 genes. From these, 72 genes were shared between tissues with nodules and HCC. In addition, 35 genes with statistical significance only in HCC and with extreme ratios were identified. Differential expression of 11 genes was confirmed by comparative reverse transcription-polymerase chain reaction, whereas that of 2 genes was confirmed by immunohistochemistry. Members involved in cytochrome P450 and second-phase metabolism were downregulated, whereas genes involved in glutathione metabolism were upregulated, implicating a possible role of glutathione and oxidative regulation. We provide a gene expression profile related to the progression of nodules into HCC, which contributes to the understanding of liver cancer development and offers the prospect for chemoprevention strategies or early treatment of HCC.

Oxidative stress in carcinogenesis. Correlation between lipid peroxidation and induction of preneoplastic lesions in rat hepatocarcinogenesis.

Oxidative stress during carcinogen metabolism seems to participate in liver tumor production in the rat. N-diethylnitrosamine is an important carcinogen used in liver cancer animal models. This indirect alkylating agent produces DNA-ethyl adducts and oxidative stress. In contrast, N-ethyl-N-nitrosourea, a direct mutagen, which generates DNA-ethyl adducts, does not produce liver tumors in rat unless it is given under oxidative stress conditions such as partial hepatectomy or phenobarbital treatment. To gain insight into the relation between oxidative stress and hepatocarcinogenicity, the induction of preneoplastic liver lesions was compared among three different initiation protocols related to the initiation-promotion-resistant hepatocyte model. In addition, liver lipid peroxidation levels, determined as thiobarituric acid reactive substances were studied early during the initiation stage. Rats initiated with N-ethyl-N-nitrosourea, 25 days after treatment developed fewer and smaller gamma-glutamyl transpeptidase positive preneoplastic lesions than rats initiated with N-diethylnitrosamine. A pre-treatment with the antioxidant quercetin 1 h before N-diethylnitrosamine initiation, significantly prevented development of gamma-glutamyl transpeptidase-positive lesions. Increased lipid peroxidation levels were induced with N-diethylnitrosamine from 3 to 24 h after initiation, while N-ethyl-N-nitrosourea did not induce increments, and importantly, pre-treatment with quercetin decreased lipid peroxidation induced by N-diethylnitrosamine. These results show correlation between lipid peroxidation and hepatocarcinogenicity and support the important role of oxidative stress on liver carcinogenesis.