RESUMEN
Background: Liver preneoplasia development in rats can be mimicked by an initiation-promotion model that induces the ppearance of altered hepatocyte foci (FAH). Aims: We compare two initiation-promotion models to evaluate the presence of FAH or additional hepatic pathologies in which other organs were affected up to five month post treatment. Material and methods: FAH were induced in male adult Wistar rats with two doses of dietylnitrosamine (DEN, 150 mg/kg bw) followed by 4 doses per week (3 weeks) of 2-acetylaminofluorene (2-AAF, 20 mg/kg bw) or with one dose of DEN (200 mg/kg bw) followed by 2 doses per week (3 weeks) of 2-AAF. DEN 150, DEN 200 and control rats (received the vehicle of the drugs) groups were compared. Rats were euthanized immediately after the last dose of 2-AAF, at 3, 4 and 5 months (n = 3 for euthanasia times per group). Samples of livers, lungs, idneys, pancreatic tissue and small bowel were processed for histological and immunohistochemical analysis. Results: FAH persisted for 5 months in all livers of the DEN groups. Three months after withdrawal of 2-AAF, one rat from DEN 150 group developed fibrosis and 5 months after 2-AAF removal another rat from the same group presented a microscopic hyperplastic nodule. Only the lungs had damages compatible with lesions induced by gavage-related reflux in DEN groups. Conclusion: We concluded that up to five month post treatments, FAH persisted in all the livers from DEN groups; livers from DEN 200 group showed no other hepatic lesions besides FAH, and only the lungs suffered pathological alterations in both treated groups (AU)
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Asunto(s)
Animales , Masculino , Ratas , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/complicaciones , Neoplasias Hepáticas Experimentales/diagnóstico , Dietilnitrosamina/administración & dosificación , Dietilnitrosamina/análisis , 2-Acetilaminofluoreno/administración & dosificación , 2-Acetilaminofluoreno/análisis , Modelos Animales , Hígado/anatomía & histología , Hígado , Hígado/patología , Hepatocitos , Hepatocitos/patología , Proyectos de Investigación/tendencias , Inmunohistoquímica/métodos , InmunohistoquímicaRESUMEN
Changes in lipid metabolism have been associated with tumor promotion in rat liver. Similarities and differences of lipid parameters were investigated using the mycotoxin fumonisin B1 (FB1) and the 2-acetylaminofluorene/partial hepatectomy (AAF/PH) treatments as cancer promoters in rat liver. A typical lipid phenotype was observed, including increased membranal phosphatidylethanolamine (PE) and cholesterol content, increased levels of C16:0 and monounsaturated fatty acids in PE and phosphatidylcholine (PC), as well as a decrease in C18:0 and long-chained polyunsaturated fatty acids in the PC fraction. The observed lipid changes, which likely resulted in changes in membrane structure and fluidity, may represent a growth stimulus exerted by the cancer promoters that could provide initiated cells with a selective growth advantage. This study provided insight into complex lipid profiles induced by two different cancer promoting treatments and their potential role in the development of hepatocyte nodules, which can be used to identify targets for the development of chemopreventive strategies against cancer promotion in the liver.
Asunto(s)
2-Acetilaminofluoreno/administración & dosificación , Fumonisinas/administración & dosificación , Metabolismo de los Lípidos/efectos de los fármacos , Neoplasias Hepáticas Experimentales/metabolismo , Fenotipo , 2-Acetilaminofluoreno/toxicidad , Animales , Colesterol/metabolismo , Ácidos Grasos , Ácidos Grasos Monoinsaturados/metabolismo , Ácidos Grasos Insaturados/metabolismo , Fumonisinas/toxicidad , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Hígado/metabolismo , Neoplasias Hepáticas Experimentales/inducido químicamente , Masculino , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/metabolismo , Ratas , Ratas Endogámicas F344RESUMEN
Liver regeneration may take place after liver injury through replication of hepatocytes or hepatic progenitor cells called oval cells. Interferons (IFN) are natural cytokines with pleiotrophic effects including antiviral and antiproliferative actions. No data are yet available on the physiology and cellular source of natural IFNs during liver regeneration. To address this issue, we have analyzed the levels and biologic activities of IFN-α/IFN-γ in two models of partial hepatectomy. After 2/3rd partial hepatectomy (PH), hepatic levels of IFN-α and IFN-γ declined transiently in contrast to a transient increase of the IFN-γ serum level. After administration of 2-acetylaminofluorene and partial hepatectomy (AAF/PH model), however, both IFN-α and IFN-γ expression were up-regulated in regenerating livers. Again, the IFN-γ serum level was transiently increased. Whereas hepatic IFN-γ was up-regulated early (day 1-5), but not significantly, in the AAF/PH model, IFN-α was significantly up-regulated at later time points in parallel to the peak of oval cell proliferation (days 7-9). Biological activity of IFN-α was shown by activation of IFN-α-specific signal transduction and induction of IFN-α specific-gene expression. We found a significant infiltration of the liver with inflammatory monocyte-like mononuclear phagocytes (MNP) concomitant to the frequency of oval cells. We localized IFN-α production only in MNPs, but not in oval cells. These events were not observed in normal liver regeneration after standard PH. We conclude that IFN-γ functions as an acute-phase cytokine in both models of liver regeneration and may constitute a systemic component of liver regeneration. IFN-α was increased only in the AAF/PH model, and was associated with proliferation of oval cells. However, oval cells seem not to be the source of IFN-α. Instead, inflammatory MNP infiltrating AAF/PH-treated livers produce IFN-α. These inflammatory MNPs may be involved in the regulation of the oval cell compartment through local expression of cytokines, including IFN-α.
Asunto(s)
Interferón-alfa/metabolismo , Interferón gamma/metabolismo , Regeneración Hepática/fisiología , Células Madre/metabolismo , 2-Acetilaminofluoreno/administración & dosificación , Animales , Proliferación Celular , Células Cultivadas , Hepatectomía , Hepatocitos/metabolismo , Janus Quinasa 1/metabolismo , Células Asesinas Naturales/metabolismo , Masculino , Monocitos/metabolismo , ARN Mensajero , Ratas , Ratas Wistar , Factores de Transcripción STAT/metabolismoRESUMEN
Liver progenitors, so-called oval cells, proliferate remarkably from periportal areas after severe liver injury when hepatocyte regeneration is compromised. These cells invade far into the liver parenchyma. Molecular mechanisms underlying these behaviors of oval cells remain poorly understood. In this study, we treated rats with 2-acetylaminofluorene/carbon tetrachloride to induce hepatic oval cells. By expression microarray analysis, we investigated global gene expression profiles in liver tissue, with an emphasis on adhesion molecules, extracellular matrix proteins, matrix metalloproteinases (MMPs), growth factors/cytokines, and receptors that might contribute to the distinct behaviors of oval cells. Genes upregulated at least twofold were selected. We then performed immunostaining to verify the microarray results and identified expression of MMP-7 and CD44 in oval cells. Staining of cytokeratin (CK)-19, an oval-cell marker, was similar between oval cells located next to periportal areas and those located far within the parenchyma. In contrast, CD44 staining was more intense in the parenchyma than in periportal areas, suggesting a role of CD44 in oval-cell invasion. Moreover, newly differentiated CK-19+ hepatocytes within foci did not show CD44 staining, suggesting that CD44 is related to the undifferentiated oval-cell phenotype. We then investigated oval-cell reactivity in CD44-deficient mice fed an oval cell-inducing diet of 3,5-diethoxycarbonyl-1,4-dihydrocollidine. Results showed significantly reduced oval-cell reactivity in CD44-deficient mice. Thus, oval cells express MMP-7 and CD44, and CD44 appears to play critical roles in the proliferation, invasion, and differentiation of hepatic oval cells in rodents.
Asunto(s)
2-Acetilaminofluoreno/farmacología , Tetracloruro de Carbono/farmacología , Perfilación de la Expresión Génica , Receptores de Hialuranos/metabolismo , Hígado/citología , Hígado/efectos de los fármacos , 2-Acetilaminofluoreno/administración & dosificación , Administración Oral , Animales , Tetracloruro de Carbono/administración & dosificación , Regulación de la Expresión Génica/efectos de los fármacos , Receptores de Hialuranos/genética , Inmunoquímica , Hígado/metabolismo , Regeneración Hepática/efectos de los fármacos , Masculino , Ratas , Ratas Endogámicas F344 , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The aromatic amine 2-acetylaminofluore (2-AAF) is a powerful complete genotoxic rat liver carcinogen that induces tumors without any additional interventions. While the tumor-initiating genotoxic activity of 2-AAF is well established, its tumor-promotion activity is far less understood. It is believed that the tumor-promoting property of 2-AAF is associated with selective enhancement of cell replication and sustained suppression of apoptosis in initiated cells. In the present study, we investigated the underlying mechanisms of tumor-promoting events induced by 2-AAF-exposure. Male Sprague-Dawley rats were fed NIH-31 diet containing 0.02% of 2-AAF for 12 and 24 weeks, and the expression pattern of genes associated with the p53-signaling pathway and microRNA genes was determined in the livers of control and 2-AAF-fed rats. The results indicate that the tumor-promoting property of 2-AAF during hepatocarcinogenesis is associated predominantly with the up-regulation of anti-apoptotic growth-related genes and down-regulation of expression of pro-apoptotic genes. This disrupts the balance between cell proliferation and apoptosis, which leads to consequential unrestricted cell proliferation, especially of initiated cells. Also, the long-term-administration of 2-AAF resulted in disruption of regulatory miR-34a-p53 feed-back loop that mediates apoptosis. This was evidenced by an increased expression of miR-34a in response to genotoxic effects of 2-AAF in the absence of p53 up-regulation, and loss of regulatory control of mir-34a on SIRT1 function. Additionally, the livers of 2-AAF-exposed rats were characterized by the substantial deregulation of expression of miR-18, miR-21, miR-182, and miR-200 family, microRNAs involved in control of apoptosis/cell proliferation and cell-cell contact pathways, two major pathways disrupted during the promotion stage of hepatocarcinogenesis.
Asunto(s)
2-Acetilaminofluoreno/toxicidad , Apoptosis/efectos de los fármacos , Carcinógenos/toxicidad , Proliferación Celular/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , 2-Acetilaminofluoreno/administración & dosificación , Animales , Apoptosis/fisiología , Carcinógenos/administración & dosificación , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/metabolismo , Neoplasias Hepáticas Experimentales/patología , Masculino , MicroARNs/genética , Ratas , Ratas Sprague-Dawley , Transducción de Señal/fisiología , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Partial hepatectomy or carbon tetrachloride (CCl4) injury, following treatment of rats with 2-acetylaminofluorene (2-AAF) to inhibit proliferation of hepatocytes, induces proliferation of oval cells and possibly their differentiation into nodular foci of hepatocytes when higher doses of 2-AAF are used. Unfortunately, immunohistochemistry in previous studies failed to show oval cell markers in these foci, and thereby to demonstrate the precursor-product relationship between oval cells and hepatocytes. Immunohistochemistry on livers of rats treated with high dose 2-AAF/CCl4 was used. We found 7.6% of the hepatocyte foci were positive for an oval cell marker cytokeratin 19 (CK-19). These foci were positive for alpha-fetoprotein, less positive for carbamoylphosphate synthetase 1, and more positive for laminin in the basement membrane lining. Rarely present transitional foci had weaker expression of CK-19 and discontinuous laminin. Focal hepatocyte differentiation of oval cells was characterized by cell hypertrophy, membranous CK-19, and positive hepatocyte nuclear factor 4 (HNF-4). HNF-4+ small oval cells surrounding CK-19+ foci were frequently seen, suggesting that a paracrine mechanism(s) may be responsible for the enlargement of CK-19+ foci. In conclusions, oval cells appear to differentiate to CK-19+ foci and then to CK-19- foci in the high dose 2-AAF/CCl4 model.
Asunto(s)
2-Acetilaminofluoreno/administración & dosificación , Tetracloruro de Carbono/administración & dosificación , Hepatocitos/metabolismo , Queratina-19/biosíntesis , Regeneración Hepática/efectos de los fármacos , Hígado/efectos de los fármacos , Administración Oral , Animales , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Hepatocitos/efectos de los fármacos , Hepatocitos/patología , Inmunoquímica , Queratina-19/análisis , Hígado/patología , Masculino , Ratas , Ratas Endogámicas F344 , Coloración y Etiquetado , alfa-Fetoproteínas/análisis , alfa-Fetoproteínas/biosíntesisRESUMEN
We studied the effects of C-Phycocyanin (C-PC), a biliprotein from Spirulina platensis on the 2-acetylaminofluorene (2-AAF)-induced expression of MDR1, encoded by the multidrug resistance (MDR1) gene, in mouse macrophage cell line (RAW 264.7). Our experimental and In silico studies revealed a significant inhibition of 2-AAF-induced expression of MDR1 protein in C-PC treated mouse macrophage cell line. MDR1 induction by 2-AAF was dependent on ROS (reactive oxygen species)-Akt (protein kinase B)-NF-kappaB (Nuclear factor kappa B) signaling pathway. Generation of ROS, phosphorylation of Akt and corresponding nuclear translocation of NF-kappaB, the events that play a major role in the induction of MDR1 expression, were decreased significantly in C-PC treated cells. NADPH oxidase inhibitor, DPI (Diphenyl iodide), and pharmacological inhibitor of Akt, Akt inhibitor IV, also showed a reduction in MDR1 expression, although not to the same extent as C-PC mediated inhibition of MDR1 expression. To further understand the mechanism, we created a computational model of the detailed ROS-Akt-NF-kappaB pathway. C-PC was modeled purely as a ROS scavenger and this representation matched the experimental trends accurately. Also the ROS levels determined through In silico investigation showed that C-PC was more effective in reduction of MDR1 expression than inhibitors of NADPH oxidase and Akt. Our experimental and In silico studies collectively suggest that 2-AAF induces MDR1 by ROS dependent pathway and C-PC is a potential negative regulator of MDR1 expression. This down regulation of MDR1 expression, induced by xenobiotics such as 2-AAF, suggests C-PC's usefulness in overcoming the drug resistance in cellular systems.
Asunto(s)
2-Acetilaminofluoreno/administración & dosificación , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Macrófagos/metabolismo , Modelos Biológicos , Ficocianina/administración & dosificación , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/fisiología , Animales , Línea Celular , Simulación por Computador , Combinación de Medicamentos , Expresión Génica/efectos de los fármacos , Expresión Génica/fisiología , Macrófagos/efectos de los fármacos , Ratones , Transducción de Señal/efectos de los fármacosRESUMEN
In hepatocarcinogenesis-resistant DRH rats, preneoplastic hepatocytic lesions are smaller than those of usual rats during carcinogenesis. When preneoplastic hepatocytes from DRH and Donryu (original strain of DRH) were reciprocally transplanted into the livers of DRH and Donryu treated with 2-acetylaminofluorene (2-AAF) diet/two-thirds hepatectomy (PH), the Donryu cells formed small colonies within the DRH liver, whereas the DRH cells formed large colonies within the Donryu liver. The DRH liver showed less degree of oval cell proliferation after treatment with 2-AAF and PH, and DRH hepatocytes were more resistant to the growth-inhibitory effect of 2-AAF after PH. Furthermore, DRH hepatocytes were generally resistant to cytotoxicity of hepatotoxins. The tissue environment of the DRH liver, therefore, is less effective for selective growth of preneoplastic hepatocytes during the carcinogen treatment, which is probably a major cause of the hepatocarcinogenesis-resistance in DRH rats.
Asunto(s)
Hepatocitos/patología , Neoplasias Hepáticas Experimentales/patología , Lesiones Precancerosas/patología , 2-Acetilaminofluoreno/administración & dosificación , Animales , Carcinógenos/administración & dosificación , Procesos de Crecimiento Celular/efectos de los fármacos , Predisposición Genética a la Enfermedad , Glutatión Transferasa/biosíntesis , Hepatectomía , Hepatocitos/efectos de los fármacos , Hepatocitos/enzimología , Hepatocitos/trasplante , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/enzimología , Neoplasias Hepáticas Experimentales/genética , Masculino , Trasplante de Neoplasias , Lesiones Precancerosas/inducido químicamente , Lesiones Precancerosas/enzimología , Lesiones Precancerosas/genética , Ratas , Ratas EndogámicasRESUMEN
We previously reported that a sub-necrogenic dose (20 mg/kg) of diethylnitrosamine (DENA) can induce the development of liver cancer when rats undergo a fasting-re-feeding regimen. The present study was undertaken to establish whether fasting followed by re-feeding builds up mechanisms able to trigger liver fibrosis, eventually leading to cirrhosis and cancer. Adult male rats, for fasted 4 days, were given 20 mg/kg of DENA after 1 day of re-feeding; in parallel, consistently fed animals receiving 20 mg/kg (sub-necrogenic) or 200 mg/kg (necrogenic dose) of DENA were used as negative and positive controls, respectively. All three groups were then subjected to the 2-acetylaminofluorene/carbon tetrachloride promoting regimen. Fasting induced moderate apoptosis in liver tissue, as evidenced by increased levels of transforming growth factor-beta1 (TGF-beta1) and Bax proteins and by a dramatic drop in the level of Bcl-2. Subsequent re-feeding caused all changes to revert except TGF-beta1 up-regulation. Histological findings of inflammation and fibrosis were consistently associated with increased production of TGF-beta1, the inflammatory cytokine with the most pronounced profibrogenic action. Thus, up-regulation of TGF-beta1 expression appears as a major mechanism by which the fasting-re-feeding regimen predisposes to initiation and promotion of liver carcinogenesis in rats. Avoiding fasting-re-feeding could be considered in the nutritional status of patients with liver fibrosis.
Asunto(s)
Dietilnitrosamina , Ingestión de Alimentos , Ayuno , Neoplasias Hepáticas Experimentales/inducido químicamente , Factor de Crecimiento Transformador beta/biosíntesis , 2-Acetilaminofluoreno/administración & dosificación , Animales , Apoptosis , Tetracloruro de Carbono/administración & dosificación , Carcinógenos/administración & dosificación , Dietilnitrosamina/administración & dosificación , Cirrosis Hepática Experimental/inducido químicamente , Cirrosis Hepática Experimental/patología , Neoplasias Hepáticas Experimentales/patología , Masculino , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas , Ratas Endogámicas F344 , Factor de Crecimiento Transformador beta1 , Regulación hacia Arriba , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Comparisons on a linear and the Rozman logarithmic scale for dosage versus carcinogenicity in rodents are presented for methyl eugenol (ME), nitrosodiethylamine (NDEA), ethyl carbamate (EC) and 2-acetylaminofluorene (AAF). Each of these chemicals has been shown to be carcinogenic in experimental animals and, in addition, humans are regularly exposed to at least three of these compounds (ME, NDEA, EC) in foods. Although the source of adducts from AAF is not known, the aminofluorene (AF) adduct is present in humans. Plotted on the same graphs are either some doses from common foods (ME, NDEA, EC) or adducts (AF) on human haemoglobin, for perspective, with their thresholds for carcinogenesis in animals. Use of a linear scale when comparing doses administered to animals in studies of carcinogenicity with doses of those same chemicals to which humans are exposed does not provide useful, comparative information. On the other hand, the Rozman logarithmic scale for dose allows one to put these relative doses in perspective. It is also evident that forcing a linear extrapolation through the zero, zero origin does not agree with the experimental data. Further analyses for goodness of fit for these dose responses reveal that the dose response for three of these compounds (ME, NDEA, EC) appears to be linear with the logarithm of the dose. However, AAF appears to be linear with the logarithm of the dose for bladder, but not for liver. It is suggested that the high background incidence of tumours in the BALB/c StCrlfC3Hf/Nctr mouse liver may confound the interpretation of dose response from AAF carcinogenesis in mouse liver.
Asunto(s)
Carcinógenos/administración & dosificación , Relación Dosis-Respuesta a Droga , Neoplasias/inducido químicamente , 2-Acetilaminofluoreno/administración & dosificación , Animales , Dietilnitrosamina/administración & dosificación , Eugenol/administración & dosificación , Eugenol/análogos & derivados , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratas , Ratas Endogámicas F344 , Factores de Riesgo , Factores Sexuales , Especificidad de la Especie , Factores de Tiempo , Uretano/administración & dosificaciónRESUMEN
BACKGROUND & AIMS: Recruitment and proliferation of Thy-1+ oval cells is a hallmark of liver regeneration after 2-acetylaminofluorene (2-AAF)/partial hepatectomy (PHx) in rats. To understand the molecular mechanism underlying this process, we investigated the role of connective tissue growth factor (CTGF), one of the candidate genes differentially expressed in Thy-1+ oval cells, in this liver injury model. METHODS: Northern and Western analyses were performed to examine the induction of CTGF in total liver homogenate. Quantitative real-time polymerase chain reaction (PCR), immunofluorescent staining, and in situ hybridization were performed to confirm the expression and localization of CTGF in Thy-1+ oval cells. Finally, a known inhibitor of CTGF synthesis, Iloprost, was administered to 2-AAF/PHx treated rats to investigate the effect of Iloprost on oval cell response. RESULTS: CTGF was found to be up-regulated at both the RNA and protein levels and occurred concurrently with an up-regulation of transforming growth factor beta1 (TGF-beta1). Sorted Thy-1+ oval cells expressed a high level of CTGF gene in a quantitative PCR assay. Colocalization of Thy-1 antigen and ctgf signals by in situ hybridization further confirmed that Thy-1+ oval cells were a source of CTGF. Iloprost administration blocked CTGF induction in treated animals but did not affect TGF-beta1 expression. The inhibition of CTGF induction by Iloprost was associated with a significant decrease in oval cell proliferation and a lower level of alpha-fetoprotein expression as compared with control animals. CONCLUSIONS: These results show that CTGF induction is important for robust oval cell response after 2-AAF/PHx treatment in rats.
Asunto(s)
Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Proteínas Inmediatas-Precoces/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Regeneración Hepática/fisiología , 2-Acetilaminofluoreno/administración & dosificación , 2-Acetilaminofluoreno/toxicidad , Animales , Northern Blotting , Western Blotting , Carcinógenos/administración & dosificación , Carcinógenos/toxicidad , Factor de Crecimiento del Tejido Conjuntivo , Hepatectomía/veterinaria , Iloprost/administración & dosificación , Iloprost/farmacología , Hibridación in Situ , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina , Hígado/citología , Regeneración Hepática/genética , Masculino , Reacción en Cadena de la Polimerasa , Ratas , Ratas Endogámicas F344 , Vasodilatadores/administración & dosificación , Vasodilatadores/farmacologíaRESUMEN
The nucleotide excision repair (NER) pathway comprises two sub-pathways, transcription coupled repair (TCR) and global genome repair (GGR). To establish the importance of these separate sub-pathways in tumor suppression, we exposed mice deficient for either TCR (Csb), GGR (Xpc) or both (Xpa) to 300 ppm 2-acetylaminofluorene (in feed, ad libitum) in a unique comparative exposure experiment. We found that cancer proneness was directly linked to a defect in the GGR pathway of NER as both Xpa and Xpc mice developed significantly more liver tumors upon 2-AAF exposure than wild type or Csb mice. In contrast, a defect in TCR appeared to act tumor suppressive, leading to a lower hepatocellular tumor response in Xpa mice (tumor incidence of 25%) as compared to Xpc mice (53% tumor-bearing mice). The link between deficient GGR and tumor proneness was most pronounced in the liver, but this phenomenon was also found in the urinary bladder. As tumor induction by 2-AAF appeared almost exclusively dependent on a defect in GGR, we examined whether gene mutation induction in the non-transcribed lacZ locus could reliably predict tumor risk. Interestingly, however, short-term 2-AAF exposure induced lacZ mutant levels in Csb mice almost as high as those found in Xpa or Xpc mice. This indicates that lacZ mutant frequencies are not correlated with a specific DNA repair defect and eventual tumor outcome, at least not in the experimental design presented here.
Asunto(s)
2-Acetilaminofluoreno/toxicidad , Reparación del ADN/genética , Genoma , Neoplasias Hepáticas/inducido químicamente , Transcripción Genética/genética , 2-Acetilaminofluoreno/administración & dosificación , Animales , Análisis Mutacional de ADN , Operón Lac/genética , Neoplasias Hepáticas/patología , Ratones , Ratones Endogámicos C57BL , Mutación/genéticaRESUMEN
The anticancer efficacy of tocotrienol-rich fraction (TRF) was evaluated during diethylnitrosamine (DEN)/2-acetylaminofluorene (AAF)-induced hepatocarcinogenesis in male Sprague-Dawley rats. TRF treatment was carried out for 6 months, and was started 2 weeks before initiation phase of hepatocarcinogenesis. Morphological examination of the livers from DEN/AAF rats showed numerous off-white patches and few small nodules, which were significantly reduced by TRF treatment. Cytotoxic damage by DEN/AAF was estimated by alkaline phosphatase (ALP) release into the plasma from the cell membranes. DEN/AAF caused a twofold increase in the activity of ALP in plasma as compared with normal control rats, and this increase was prevented significantly by TRF treatment. We observed an increase of 79% in liver ALP activity in DEN/AAF rats, which was further increased by another 48% after the administration of TRF. Hepatic activity of glutathione S-transferase (GST) was also increased (3.5-fold) during the induction of hepatic carcinogenesis. Lipid peroxidation and low-density lipoprotein (LDL) oxidation increased threefold following initiation by DEN/AAF as compared with normal control rats. However, TRF treatment to DEN/AAF-treated rats substantially decreased (62-66%) the above parameters and thus limited the action of DEN/AAF. We conclude that long-term intake of TRF could reduce cancer risk by preventing hepatic lipid peroxidation and protein oxidation damage due to its antioxidant actions.
Asunto(s)
Anticolesterolemiantes/química , Antioxidantes/farmacología , Quimioprevención , Neoplasias Hepáticas/prevención & control , Aceites de Plantas/química , Tocotrienoles/farmacología , 2-Acetilaminofluoreno/administración & dosificación , 2-Acetilaminofluoreno/farmacología , Alquilantes/administración & dosificación , Alquilantes/farmacología , Animales , Carcinógenos/administración & dosificación , Carcinógenos/farmacología , Dietilnitrosamina/administración & dosificación , Dietilnitrosamina/farmacología , Peroxidación de Lípido , Masculino , Neoplasias Experimentales , Estrés Oxidativo , Proteínas/metabolismo , Ratas , Ratas Sprague-Dawley , Aceite de Salvado de ArrozRESUMEN
We recently reported that several mono- to tetrachlorinated biphenyls have initiating activity in the livers of Fischer 344 rats. In the present study, we investigated the metabolic activation of one of those compounds, 4-chlorobiphenyl (PCB 3). Monohydroxy (400 micromol/kg), dihydroxy (200 micromol/kg), and quinone (100 micromol/kg) metabolites of PCB 3 were evaluated for their initiating activity. Fischer 344 male rats were fasted for 4 days; 24 h after feeding again, they were injected (ip) with metabolites, vehicle, or diethylnitrosamine (DEN, 20 or 40 mg/kg). All animals were treated with selection agents as follows: three daily p.o. doses of 2-acetylaminofluorene (2-AAF, 30 mg/kg), followed by a single p.o. dose of carbon tetrachloride (2 ml/kg) and three additional daily treatments of 2-AAF. Rats were killed 2 weeks after the last 2-AAF intubation. Livers were evaluated for changes in morphology, and the number and volume of gamma-glutamyl transpeptidase-positive foci were measured. Of the metabolites tested, only one monohydroxy and one quinoid metabolite showed initiating activity. The metabolic activation of PCB 3, therefore, proceeds via parahydroxylation and oxidation to the ortho 3,4-quinone, the ultimate carcinogen. This is the first report to demonstrate that specific PCB metabolites possess initiating activity in the rodent liver in vivo. The results support the conclusion that 4-OH PCB 3 and 3,4-BQ PCB 3 act as proximate and ultimate carcinogenic metabolites resulting from the bioactivation of PCB 3 in rat liver.
Asunto(s)
Compuestos de Bifenilo/efectos adversos , Compuestos de Bifenilo/metabolismo , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/prevención & control , 2-Acetilaminofluoreno/administración & dosificación , Adenoma de Células Hepáticas/inducido químicamente , Administración Oral , Animales , Compuestos de Bifenilo/química , Peso Corporal/efectos de los fármacos , Tetracloruro de Carbono/administración & dosificación , Carcinógenos/efectos adversos , Carcinógenos/química , Carcinógenos/metabolismo , Dietilnitrosamina/administración & dosificación , Dietilnitrosamina/efectos adversos , Esquema de Medicación , Evaluación Preclínica de Medicamentos/métodos , Hidroxilación , Inyecciones Intraperitoneales , Intubación Intratraqueal , Hígado/química , Hígado/efectos de los fármacos , Hígado/patología , Masculino , Modelos Biológicos , Tamaño de los Órganos/efectos de los fármacos , Quinonas/efectos adversos , Quinonas/química , Quinonas/metabolismo , Ratas , Ratas Endogámicas F344 , Factores de Tiempo , gamma-Glutamiltransferasa/biosíntesis , gamma-Glutamiltransferasa/químicaRESUMEN
Combined treatment with different agents represents a promising approach in cancer chemoprevention. Therefore, it is useful to assess in preclinical models the efficacy of combinations that are selected by taking into account mechanistic considerations. We designed 2 studies evaluating the interaction between N-acetylcysteine (NAC) and sodium selenite (Se), both given with the drinking water to Balb/c mice, in modulating clastogenic effects in bone marrow polychromatic erythrocytes. In a first study, a single i.p. injection of urethane considerably enhanced the frequency of micronucleated cells. While NAC produced a significant inhibition, Se further enhanced urethane clastogenicity. When given in combination at the same doses, NAC prevented the adverse effect of Se. In a second study, a single i.p. injection of 2-acetylaminofluorene enhanced the frequency of micronucleated cells. Se did not reduce this effect to a significant extent, while NAC produced a dose-dependent inhibition. When tested at the lower dose in combination with Se, the protective effect of NAC was unchanged. Especially in association with Se, NAC also prevented the toxicity of 2-acetylaminofluorene by normalizing the ratio of polychromatic to normochromatic erythrocytes. In conclusion, NAC attenuated the clastogenicity of both urethane and 2-acetylaminofluorene and the toxicity of this aromatic amine. In addition, NAC prevented the clastogenic and toxic effects resulting from the interaction of Se with urethane. Together with the findings of previous studies, it appears that, besides its intrinsic protective properties in carcinogenesis, NAC is capable of attenuating the adverse effects of several cytotoxic drugs and chemopreventive agents.
Asunto(s)
2-Acetilaminofluoreno/farmacología , Acetilcisteína/farmacología , Antimutagênicos/farmacología , Selenito de Sodio/farmacología , Uretano/farmacología , 2-Acetilaminofluoreno/administración & dosificación , Acetilcisteína/administración & dosificación , Animales , Antimutagênicos/administración & dosificación , Interacciones Farmacológicas , Células Madre Hematopoyéticas , Masculino , Ratones , Ratones Endogámicos BALB C , Pruebas de Micronúcleos , Selenito de Sodio/administración & dosificación , Uretano/administración & dosificaciónAsunto(s)
Pruebas de Carcinogenicidad/métodos , Neoplasias Experimentales/inducido químicamente , 2-Acetilaminofluoreno/administración & dosificación , 2-Acetilaminofluoreno/toxicidad , Carcinógenos/administración & dosificación , Carcinógenos/toxicidad , Interpretación Estadística de Datos , Relación Dosis-Respuesta a Droga , Modelos BiológicosAsunto(s)
Pruebas de Carcinogenicidad/métodos , Neoplasias Experimentales/inducido químicamente , 2-Acetilaminofluoreno/administración & dosificación , 2-Acetilaminofluoreno/toxicidad , Carcinógenos/administración & dosificación , Carcinógenos/toxicidad , Interpretación Estadística de Datos , Relación Dosis-Respuesta a Droga , Modelos BiológicosRESUMEN
Vegetables, natural products of plant origin and numerous non-nutritive dietary constituents have been shown to play a salutary role in cancer chemoprevention. The present study aims to evaluate the chemopreventive efficacy of the methanol fraction of Asteracantha longifolia seed extract against development of 2-acetylaminofluorene (2-AAF)-selected gamma-glutamyl transpeptidase (gamma-GT)-positive foci following diethylnitrosamine (DEN) initiation. Treatment of rats with doses 200 and 400 mg/kg body weight of methanol extract of A. longifolia seeds on alternate days, subsequent to carcinogen treatment, for 6 weeks significantly reduced the incidence and size distribution of gamma-GT-positive foci and tumor formation. Administration of A. longifolia seeds significantly (P<0.001) ameliorated the activities of antioxidant enzymes, glutathione peroxidase (GPx) and catalase (CAT), in a dose-dependent manner. Prophylactic administration of seed extract simultaneous to 2-AAF in the diet, at same doses, significantly suppressed 2-AAF and partial hepatectomy (PH)-induced ornithine decarboxylase (ODC) activity and [(3)H]thymidine incorporation into hepatic DNA, in a dose-dependent manner. Assimilation of the quantitative foci data together with the findings of the modulation of tumor promoting markers give ample evidence to the anti-tumor promoting potential of A. longifolia seeds against chemically-induced hepatocarcinogenesis in Wistar rats.
Asunto(s)
Anticarcinógenos/farmacología , Neoplasias Hepáticas Experimentales/prevención & control , Hígado/efectos de los fármacos , Extractos Vegetales/farmacología , Semillas/química , 2-Acetilaminofluoreno/administración & dosificación , Alquilantes , Animales , Anticarcinógenos/uso terapéutico , Carcinógenos/administración & dosificación , Catalasa/metabolismo , Dietilnitrosamina , Relación Dosis-Respuesta a Droga , Glutatión Peroxidasa/metabolismo , Hígado/enzimología , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/tratamiento farmacológico , Masculino , Ornitina Descarboxilasa/metabolismo , Extractos Vegetales/uso terapéutico , Ratas , Ratas Wistar , Timidina , Factores de Tiempo , gamma-Glutamiltransferasa/efectos de los fármacos , gamma-Glutamiltransferasa/metabolismoRESUMEN
The genotoxic effects of 2-acetylaminofluorene (AAF) alone cannot explain the formation of rat liver tumors. It has been proposed that mitochondrial effects are associated with its tumor-promoting properties. These mitochondrial effects are thought to trigger apoptosis and regenerative proliferation, which alters the liver lobule in a cirrhosis-like manner. A situation is generated which favors the growth of initiated cells. To test this sequence of events, the dose dependence of early effects with time was studied. Male Wistar rats received 50, 100, 200, 400, or 800 ppm AAF in the diet and the following endpoints were analyzed at 2, 4, 8, and 16 weeks of feeding: apoptotic cell death, cell proliferation, GST-P-positive foci (placental form of glutathione S-transferase), and morphological alterations. Hydrolyzable hemoglobin adducts were used as a dosimeter for metabolic activation after 2 weeks of feeding. The hemoglobin adduct levels increased linearly with dose. With the conventional carcinogenic concentration of 200-ppm AAF in the diet, the number of apoptoses increased first, predominantly in the periportal area (2 weeks). Cell proliferation followed with some delay (4 weeks). This reflects regenerative tissue response and not the growth of initiated cells, because the number of enzyme-altered foci was still extremely low at that time. Foci developed only later when the morphology had changed. With 50 ppm AAF in the diet, a no-effect level had not been reached for any of the endpoints, but foci developed much more gradually than with higher doses. Unexpectedly, the proliferative response stabilized at 8 weeks with a labeling index of 12-17, with all AAF concentrations. The observed sequence of events supports the hypothesis. It is concluded that (1) The proliferation of initiated cells-defined as promotable cells-is largely determined by the cellular environment, such as morphologically altered liver. (2) The morphological alterations in rat liver result from imperfect regeneration from toxic effects. (3) Imperfect regeneration results from limitations in the possibilities to adapt to chemical stress. (4) If these limits are overwhelmed and morphology has changed to a certain extent, preneoplastic foci develop; this occurs much faster, at least, than without these changes.
Asunto(s)
2-Acetilaminofluoreno/toxicidad , Carcinógenos/toxicidad , 2-Acetilaminofluoreno/administración & dosificación , Animales , Apoptosis/efectos de los fármacos , Carcinógenos/administración & dosificación , División Celular/efectos de los fármacos , Dieta , Relación Dosis-Respuesta a Droga , Glutatión Transferasa/metabolismo , Hemoglobinas/metabolismo , Masculino , Neoplasias Experimentales/inducido químicamente , Neoplasias Experimentales/patología , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
Oval cells which appear in the liver after hepatic injuries are suspected to be progenitor cells for both hepatocytes and bile duct cells. Oval cell isolated from the livers of the hamsters treated with diethylnitrosamine and 2-acetylaminofluorene and infected with Clonorchis sinensis (CS). cultured for 2 weeks and evaluated for differentiation and plasticity by electron microscopy and immunohistochemistry. In the CS-uninfected group, glycogen granules and peroxisomes were noted in the cells that were cultured for 2 weeks. Starting at 1 week postculture, immunoreactivity of the cells to cytokeratin 19 markedly decreased but that to albumin and alpha-fetoprotein gradually increased. This means that oval cells isolated from hamsters that were not infected with CS differentiated toward hepatocyte lineage. However, in the CS-infected group, cultured cells contained numerous rough endoplasmic reticulum and showed immunoreactivity that was generally in reverse to that of CS-uninfected group, meaning that cells isolated following CS infection were primed by CS and differentiated toward bile duct cell lineage. The results of this study suggested that oval cells are indeed bipolar progenitor cells for hepatocytes and bile duct cells and can differentiate toward either lineage depending upon the priming factor.