RESUMO
INTRODUCTION: Xenografts of androgen-independent human DU145 prostate metastatic carcinomas implanted in nu/nu male mice have revealed a significant survival after a prooxidant anticancer treatment consisting of a combination of menadione bisulfite and sodium ascorbate (VK3:VC). METHODS: Implanted samples of diaphragm carcinomas from longest survived mice from either oral, intraperitoneal (IP), or both oral and IP treatment groups were assessed with light, scanning, and transmission electron microscopy to analyze morphologic damages. RESULTS: Compared with previous fine structure data of in vitro untreated carcinomas, the changes induced by oral, IP, and oral with IP VK3:VC treatment dismantled those xenografts with autoschizis, and necrotic atrophy was accomplished by cell's oxidative stress whose injuries were consequent to reactivated deoxyribonucleases and ribonucleases. Tumor destructions resulted from irreversible damages of nucleus components, endoplasmic reticulum, and mitochondria there. Other alterations included those of the cytoskeleton that resulted in characteristic self-excisions named " autoschizis." All these injuries lead resilient cancer cells to necrotic cell death. CONCLUSION: The fine structure damages caused by VK3:VC prooxidant combination in the human DU145 prostate xenografts confirmed those shown in vitro and of other cell lines with histochemistry and biomolecular investigations. These devastations incurred without damage to normal tissues; thus, our data brought support for the above combination to assist in the treatment of prostate cancers and other cancers.
Assuntos
Ácido Ascórbico , Camundongos Nus , Neoplasias da Próstata , Vitamina K 3 , Masculino , Humanos , Animais , Neoplasias da Próstata/patologia , Neoplasias da Próstata/tratamento farmacológico , Ácido Ascórbico/farmacologia , Camundongos , Vitamina K 3/farmacologia , Linhagem Celular Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , Estresse Oxidativo/efeitos dos fármacos , Microscopia Eletrônica de TransmissãoRESUMO
Human prostate carcinoma DU145 cells, androgen-independent malignant cells, implanted in the athymic nu/nu male mouse, developed numerous tumors on peritoneal and retro-peritoneal organs whose growth aspects and vascular supply have yet to be investigated with fine structure techniques. A series of necropsies from moribund implanted mice diaphragms were examined with light, scanning, and transmission electron microscopy. DU145 xenografts installations, far away from the implanted site, were described as the smallest installation to large diaphragm outgrowths in moribund mice. Carcinomas did not show extracellular matrix and, reaching more than 0.15 mm in thickness, they revealed new structures in these outgrowths. Voids to be gland-like structures with mediocre secretion and, unexpectedly, intercellular spaces connected with fascicles of elongated DU145 cells that merged with a vascular supply originated from either the tumor cells and/or some perimysium vessels. In the largest carcinomas, most important vascular invasions coincidently accompanied the mouse lethality, similarly to human cancers. This androgen-independent model would be useful to study tumor outgrowth's changes related to testing anticancer strategy, including anti-angiogenic therapies involving toxicity, simultaneously with those of other vital organs with combined biomolecular and fine structure techniques.
Assuntos
Carcinoma , Neoplasias da Próstata , Androgênios , Animais , Linhagem Celular Tumoral , Diafragma/patologia , Epitélio/patologia , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Nus , Próstata/patologia , Neoplasias da Próstata/patologiaRESUMO
Androgen-independent, human prostate carcinoma cells (DU145) develop into solid, carcinomatous xenotransplants on the diaphragm of nu/nu mice. Tumors encompass at least two poorly differentiated cell types: a rapidly dividing, eosinophilic cell comprises the main cell population and a few, but large basophilic cells able to invade the peritoneal stroma, the muscular tissue, lymph vessels. Poor cell contacts, intracytoplasmic lumina, and signet cells are noted. Lysosomal activities are reflected by entoses and programmed cell deaths forming cribriform carcinomas. In large tumors, degraded cells may align with others to facilitate formation of blood supply routes. Malignant cells would spread via ascites and through lymphatics.
Assuntos
Adenocarcinoma/ultraestrutura , Carcinoma/ultraestrutura , Neoplasias da Próstata/ultraestrutura , Adenocarcinoma/irrigação sanguínea , Animais , Apoptose , Basófilos/ultraestrutura , Carcinoma/irrigação sanguínea , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Entose , Humanos , Vasos Linfáticos/ultraestrutura , Lisossomos/ultraestrutura , Masculino , Camundongos , Camundongos Nus , Microscopia Eletrônica de Transmissão , Invasividade Neoplásica , Transplante de Neoplasias , Fenótipo , Neoplasias da Próstata/irrigação sanguínea , Células Estromais/ultraestrutura , Transplante HeterólogoRESUMO
We describe the biological activity of some furylbenzo- and naphthoquinones (furylquinones) on hepatocarcinoma cells and healthy rat liver slices. The effects of furylquinones on cancer cells (Transplantable Liver Tumor, TLT) were assessed by measuring cell death (membrane cell lysis); intracellular contents of ATP and GSH and the activity of caspase-3 were used to determine the type of cell death. Most of the furylquinones tested (at a concentration of 25 µg/ml) induced caspase-independent cell death but compound 4 had no cytotoxic effects. The levels of both ATP and GSH were severely affected by quinones 1, 2 and 5, while no effect was observed with compound 4. These cytotoxic properties of quinones are associated with physico-chemical properties as shown by the LUMO energies and lipophilicity. Interestingly, no cytotoxic effects of furylquinones were detected when the in vitro model of precision-cut liver slices (PCLS) was used. Indeed, although CYP2E1 activity was slightly affected, ATP and GSH levels as well as protein synthesis were not modified by furylquinones. Paracetamol, a well-known hepatotoxicant, reduced these parameters by more than 80% compared to control conditions. Taking into account the considerable incidence of adverse-effects induced by most current anticancer drugs, the selective cytotoxicity shown by compounds 1, 2 and 5, in particular that of 1, represents a safety factor that encourages the further development of these quinones as new drugs in cancer therapy.
Assuntos
Antineoplásicos/farmacologia , Quinonas/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/enzimologia , Carcinoma Hepatocelular/patologia , Caspase 3/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocromo P-450 CYP2E1/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Glutationa/metabolismo , Técnicas In Vitro , Fígado/efeitos dos fármacos , Fígado/enzimologia , Fígado/patologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Transplante de Neoplasias , Biossíntese de Proteínas/efeitos dos fármacos , Quinonas/química , Quinonas/uso terapêutico , Ratos , Ratos Wistar , SoftwareRESUMO
Numerous studies suggest that generation of oxidative stress could be useful in cancer treatment. In this study, we evaluated, in vitro and in vivo, the antitumor potential of oxidative stress induced by ascorbate/menadione (asc/men). This combination of a reducing agent (ascorbate) and a redox active quinone (menadione) generates redox cycling leading to formation of reactive oxygen species (ROS). Asc/men was tested in several cell types including K562 cells (a stable human-derived leukemia cell line), freshly isolated leukocytes from patients with chronic myeloid leukemia, BaF3 cells (a murine pro-B cell line) transfected with Bcr-Abl and peripheral blood leukocytes derived from healthy donors. Although these latter cells were resistant to asc/men, survival of all the other cell lines was markedly reduced, including the BaF3 cells expressing either wild-type or mutated Bcr-Abl. In a standard in vivo model of subcutaneous tumor transplantation, asc/men provoked a significant delay in the proliferation of K562 and BaF3 cells expressing the T315I mutated form of Bcr-Abl. No effect of asc/men was observed when these latter cells were injected into blood of mice most probably because of the high antioxidant potential of red blood cells, as shown by in vitro experiments. We postulate that cancer cells are more sensitive to asc/men than healthy cells because of their lack of antioxidant enzymes, mainly catalase. The mechanism underlying this cytotoxicity involves the oxidative cleavage of Hsp90 with a subsequent loss of its chaperone function thus leading to degradation of wild-type and mutated Bcr-Abl protein.
Assuntos
Ácido Ascórbico/farmacologia , Proteínas de Fusão bcr-abl/metabolismo , Proteínas Mutantes/metabolismo , Neoplasias/patologia , Estresse Oxidativo/efeitos dos fármacos , Vitamina K 3/farmacologia , Animais , Ácido Ascórbico/química , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citoproteção/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio/farmacologia , Células K562 , Camundongos , Camundongos Nus , Neoplasias/metabolismo , Vitamina K 3/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Oxidative stress generated by ascorbate-driven menadione redox cycling kills MCF7 cells by a concerted mechanism including glycolysis inhibition, loss of calcium homeostasis, DNA damage and changes in mitogen activated protein kinases (MAPK) activities. Cell death is mediated by necrosis rather than apoptosis or macroautophagy. Neither 3-methyladenine nor Z-VAD affects cytotoxicity by ascorbate/menadione (Asc/Men). BAPTA-AM, by restoring cellular capacity to reduce MTT, underlines the role of calcium in the necrotic process. Oxidative stress-mediated cell death is shown by the opposite effects of N-acetylcysteine and 3-aminotriazole. Moreover, oxidative stress induces DNA damage (protein poly-ADP-ribosylation and gamma-H2AX phosphorylation) and inhibits glycolysis. Asc/Men deactivates extracellular signal-regulated kinase (ERK) while activating p38, suggesting an additional mechanism to kill MCF7 cells. Since ascorbate is taken up by cancer cells and, due to their antioxidant enzyme deficiency, oxidative stress should affect cancer cells to a greater extent than normal cells. This differential sensitivity may have clinical applications.
Assuntos
Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Neoplasias da Mama/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Vitamina K 3/farmacologia , Trifosfato de Adenosina/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Combinação de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Glicólise/efeitos dos fármacos , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Ácido Láctico/metabolismo , NAD/metabolismo , Oxirredução , Sais de Tetrazólio/metabolismo , Tiazóis/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
The alterations of deoxyribonuclease DNase activity in cancer cells were the basis of the utilization of mixed vitamins C and K3 in a nontoxic, adjuvant cancer therapy. In order to localize exactly the altered activities of DNase in cancer cells, histochemical methods were utilized. The deficiency of alkaline and acid DNase activity appeared to be characteristic for non-necrotic cells of malignant human and animal tumors. This enzymatic deficiency appeared in experimental carcinogenesis before the phenotypic signs of malignancy. Tumor promoters directly reduced the activity of both DNases. The incidence of spontaneous malignant human and animal tumors appeared to be inversely proportional to the intensity of the activity of both DNases in normal cells and tissues from which these tumors were derived. The fact that alkaline and acid DNase activity was reactivated during the spontaneous and therapeutically induced necrosis of cancer cells suggests that this enzymatic deficiency of DNase activity in cancer cells was due to the action of specific inhibitors of DNases. Characteristic variations of serum alkaline DNase activity in positive responders to therapy, examined in more than 800 cancer-bearing patients, may be the basis for the development of a useful test for therapeutic prognosis and for monitoring of cancer bearing patients. Acid DNase was selectively reactivated in malignant tumor cells by vitamin C (sodium ascorbate), whereas alkaline DNase was reactivated by vitamin K3. Joint vitamin C and K3 administration produced in vitro and in vivo tumor growth inhibition, potentiation and sensitization of chemo- and/or radiotherapy and a decrease in the number of metastases in animals with experimental tumors. Joint vitamin C and K3 administration may be considered as a possible new, non-toxic, adjuvant cancer therapy, which can be easily introduced into the classic protocols of clinical cancer therapy without any supplementary risk for patients.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Desoxirribonucleases/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/enzimologia , Animais , Ácido Ascórbico/administração & dosagem , Quimioterapia Adjuvante , Humanos , Vitamina K 3/administração & dosagemRESUMO
2-euryfuryl- and 2-euryfuryl-3-nitro-1,4-benzoquinone Q2 and Q3, prepared via oxidative coupling reactions of sesquiterpene euryfuran 1 to 2-nitro-1,4-benzoquinone and 1,4-benzoquinone, were tested for their cytotoxicity towards TLT cells (a murine hepatoma cell line) in the absence and in the presence of vitamin C. Their cytotoxic profile was completely different. In cells incubated with Q2 (from 1 to 50 microg/ml), cell survival was not modified, both GSH and ATP were depleted to about 50% of control values (at 50 microg/ml); and caspase-3 was activated in a dose-dependent manner. These effects were observed whatever cells were incubated or not in the presence of vitamin C. In the case of Q3, the cytotoxicity was rather unrelated to its concentration but the association of vitamin C plus the highest Q3 concentration (50 microg/ml) results in a strong cell death (more than 60%). At such a concentration, a complete lack of caspase-3 activity was observed, probably due to cell lysis. At lower concentrations of Q3 (1 and 10 microg/ml), caspase-3 activity was lower than that observed in the absence of vitamin C or even under control conditions. Both GSH and ATP were kept fairly constant as compared to control values but in the presence of vitamin C and Q3, at 50 microg/ml, a decrease in their amounts was observed.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Ácido Ascórbico/farmacologia , Benzoquinonas/química , Benzoquinonas/farmacologia , Carcinoma Hepatocelular/patologia , Trifosfato de Adenosina/metabolismo , Animais , Caspase 3/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citoplasma/efeitos dos fármacos , Citoplasma/enzimologia , Ativação Enzimática/efeitos dos fármacos , Glutationa/metabolismo , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , L-Lactato Desidrogenase/metabolismo , CamundongosRESUMO
The effect of oxidative stress induced by the ascorbate/menadione-redox association was examined in K562 cells, a human erythromyeloid leukaemia cell line. Our results show that ascorbate enhances menadione redox cycling, leading to the formation of intracellular reactive oxygen species (as shown by dihydrorhodamine 123 oxidation). The incubation of cells in the presence of both ascorbate/menadione and aminotriazole, a catalase inhibitor, resulted in a strong decrease of cell survival, reinforcing the role of H(2)O(2) as the main oxidizing agent killing K562 cells. This cell death was not caspase-3-dependent. Indeed, neither procaspase-3 and PARP were processed and only a weak cytochrome c release was observed. Moreover, we observed only 23% of cells with depolarized mitochondria. In ascorbate/menadione-treated cells, DNA fragmentation was observed without any sign of chromatin condensation (DAPI and TUNEL tests). The cell demise by ascorbate/menadione is consistent with a necrosis-like cell death confirmed by both cytometric profile of annexin-V/propidium iodide labeled cells and by light microscopy examination. Finally, we showed that a single i.p. administration of the association of ascorbate and menadione is able to inhibit the growth of K562 cells by about 60% (in both tumour size and volume) in an immune-deficient mice model. Taken together, these results reinforced our previous claims about a potential application of the ascorbate/menadione association in cancer therapy.
Assuntos
Ácido Ascórbico/farmacologia , Morte Celular/efeitos dos fármacos , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Estresse Oxidativo/efeitos dos fármacos , Vitamina K 3/farmacologia , Animais , Morte Celular/fisiologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Humanos , Células K562 , Camundongos , Camundongos Nus , Transplante de Neoplasias , Estresse Oxidativo/fisiologia , Vitamina K 3/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
C3H/He and B6C3F1 show much higher liver cancer susceptibility than C57BL/6J mice. We studied the hypothesis that this difference might result from failure of apoptosis. Hepatocarcinogenesis was induced by a single dose of N-nitrosodiethylamine (NDEA), followed by phenobarbital (PB) for up to 90 weeks. We observed (1) earlier appearance of putative preneoplastic foci (PPF), hepatocellular adenoma (HCA), and carcinoma (HCC) in C3H/He than in C57Bl/6J mice and (2) an increase of hepatocellular DNA synthesis in C3H/He and C57Bl/6J mice, compared to normal liver, via PPF and HCA to HCC. PB enhanced DNA synthesis and growth of PPF, in the C3H/He strain only, and of HCA and HCC of both strains. Apoptoses were rare in unaltered livers as well as in preneoplastic lesions, but tended to increase in HCA and HCC of both strains. PB lowered apoptotic activity in PPF of C3H/He mice, but enhanced it in HCA and HCC of C57Bl/6J mice at late stages. In conclusion, the strain difference in growth rates of PPF and tumors is largely determined by higher rates of cell proliferation in C3H/He mice, with and without promotion by PB. Moreover, in C57Bl/6J mice the promoting effect of PB was restricted to HCA and HCC and was not seen in PPF. Apoptosis was generally low and was not a major cause of the strain difference in tumor susceptibility. In contrast with rat liver, inhibition of apoptosis appears to be a minor determinant of tumor promotion in mice.
Assuntos
Apoptose/fisiologia , Proliferação de Células/efeitos dos fármacos , Cocarcinogênese , Neoplasias Hepáticas Experimentais/patologia , Fígado/patologia , Lesões Pré-Cancerosas/patologia , Animais , Apoptose/efeitos dos fármacos , Carcinógenos/toxicidade , Diferenciação Celular/efeitos dos fármacos , DNA/biossíntese , Dietilnitrosamina/toxicidade , Fígado/efeitos dos fármacos , Neoplasias Hepáticas Experimentais/induzido quimicamente , Masculino , Camundongos , Camundongos Endogâmicos , Fenobarbital/toxicidade , Lesões Pré-Cancerosas/induzido quimicamente , Especificidade da EspécieRESUMO
Since the higher redox potential of quinone molecules has been correlated with enhanced cellular deleterious effects, we studied the ability of the association of ascorbate with several quinones derivatives (having different redox potentials) to cause cell death in K562 human leukaemia cell line. The rationale is that the reduction of quinone by ascorbate should be dependent of the quinone half-redox potential thus determining if reactive oxygen species (ROS) are formed or not, leading ultimately to cell death or cell survival. Among different ROS that may be formed during redox cycling between ascorbate and the quinone, the use of different antioxidant compounds (mannitol, desferal, N-acetylcysteine, catalase and superoxide dismutase) led to support H2O2 as the main oxidizing agent. We observed that standard redox potentials, oxygen uptake, free ascorbyl radical formation and cell survival were linked. The oxidative stress induced by the mixture of ascorbate and the different quinones decreases cellular contents of ATP and GSH while caspase-3-like activity remains unchanged. Again, we observed that quinones having higher values of half-redox potential provoke a severe depletion of ATP and GSH when they were associated with ascorbate. Such a drop in ATP content may explain the lack of activation of caspase-3. In conclusion, our results indicate that the cytotoxicity of the association quinone/ascorbate on K562 cancer cells may be predicted on the basis of half-redox potentials of quinones.
Assuntos
Apoptose/efeitos dos fármacos , Ácido Ascórbico/farmacologia , Caspases/metabolismo , Leucemia/metabolismo , Leucemia/patologia , Quinonas/metabolismo , Trifosfato de Adenosina/metabolismo , Ácido Ascórbico/química , Ácido Ascórbico/metabolismo , Ácido Ascórbico/toxicidade , Caspase 3 , Linhagem Celular Tumoral , Radicais Livres/metabolismo , Glutationa/metabolismo , Humanos , Oxirredução/efeitos dos fármacos , Oxigênio/metabolismo , Vitamina K 3/química , Vitamina K 3/metabolismoRESUMO
Dietary treatment with inulin or oligofructose incorporated in the basal diet for experimental animals: (I) reduced the incidence of mammary tumors induced in Sprague-Dawley rats by methylnitrosourea; (II) inhibited the growth of transplantable malignant tumors in mice; (III) decreased the incidence of lung metastases of a malignant tumor implanted intramuscularily in mice. (IV) Moroever, besides such cancer risk reduction effects, dietary treatment with inulin or oligofructose significantly potentiated the effects of subtherapeutic doses of six cytotoxic drugs commonly utilized in human cancer treatment. (V) The same prebiotics potentiated the effects of radiotherapy on solid form of TLT tumors to a statistically very high level. Such dietary treatment, with the inulin or oligofructose potentiating the effects of cancer therapy, might be introduced into classic protocols of human cancer treatment as a new, non-toxic and easily applicable adjuvant cancer therapy without any additional risk to patients.
Assuntos
Adjuvantes Imunológicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Inulina/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Oligossacarídeos/farmacologia , Animais , Neoplasias da Mama/induzido quimicamente , Dieta , Feminino , Injeções Intramusculares , Neoplasias Hepáticas/patologia , Masculino , Metilnitrosoureia , Camundongos , Transplante de Neoplasias , Ratos , Ratos Sprague-Dawley , Transplante HeterólogoRESUMO
Vitamin C (VC) and vitamin K(3) (VK(3)) administered in a VC:VK(3) ratio of 100:1 exhibit synergistic antitumor activity and preferentially kill tumor cells by autoschizis, a novel type of necrosis characterized by exaggerated membrane damage and progressive loss of organelle-free cytoplasm through a series of self-excisions. During this process, the nucleus becomes smaller, cell size decreases one-half to one-third of its original size, and most organelles surround an intact nucleus in a narrow rim of cytoplasm. While the mitochondria are condensed, tumor cell death does not result from ATP depletion. However, vitamin treatment induces a G(1)/S block, diminishes DNA synthesis, increases H(2)O(2) production, and decreases cellular thiol levels. These effects can be prevented by the addition of catalase to scavenge the H(2)O(2). There is a concurrent 8- to 10-fold increase in intracellular Ca(2+) levels. Electrophoretic analysis of DNA reveals degradation due to the caspase-3-independent reactivation of deoxyribonuclease I and II (DNase I, DNase II). Redox cycling of the vitamins is believed to increase oxidative stress until it surpasses the reducing ability of cellular thiols and induces Ca(2+) release, which triggers activation of Ca(2+)-dependent DNase and leads to degradation of DNA. Recent experiments indicate that oral VC:VK(3) increases the life-span of tumor-bearing nude mice and significantly reduces the growth rate of solid tumors without any significant toxicity by reactivating DNase I and II and inducing autoschizis. This report discusses the mechanisms of action employed by these vitamins to induce tumor-specific death by autoschizis.
Assuntos
Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Morte Celular/fisiologia , Neoplasias/patologia , Estresse Oxidativo/efeitos dos fármacos , Vitamina K/farmacologia , Animais , Antioxidantes/química , Antioxidantes/uso terapêutico , Ácido Ascórbico/química , Ácido Ascórbico/uso terapêutico , Humanos , Necrose , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Vitamina K/química , Vitamina K/uso terapêuticoRESUMO
PURPOSE: In the liver, transforming growth factor beta-1 (TGF-beta1) constitutes a major negative growth regulating factor involved in the control of cell numbers; failure of this control mechanism has been associated with the development of liver cancer. Since no reports on the in vivo effects of exogenously administered TGF-beta1 on apoptosis in liver tumors have been published yet, we studied hepatocyte sensitivity to the proapoptotic action of TGF-beta1 in stages of chemically induced mouse liver carcinogenesis. METHODS: Mouse liver carcinogenesis was initiated by a single dose of N-nitrosodiethylamine (NDEA, 90 mg/kg b.w., i.p.) to 5-week-old B6C3F1 mice. After 2 weeks, mice received either standard diet or a diet containing phenobarbital (PB, 90 mg/kg b.w) for 85 weeks. Four hours before being killed mice received a single dose of TGF-beta1 (56 microg or 200 microg TGF-beta1/kg of b.w., injected into the tail vein). Quantitative histological analysis of mitosis and apoptosis in normal liver tissue (NL), putative preneoplastic foci (PPF), hepatocellular adenoma (HCA), and hepatocellular carcinoma (HCC) was performed on H&E-stained liver sections. RESULTS: In NDEA and NDEA + PB-treated mice, NL exhibited a very low incidence of apoptosis and mitosis, which increased in HCA and HCC. In the lesions apoptoses ranged between 0.03 and 0.6%. Two hundred micrograms of TGF-beta1/kg stimulated apoptoses in NL as well as in neoplastic lesions (significant increase in NL, HCA, and HCC); the most pronounced proapoptotic action of TGF-ss1 was observed in lesions of NDEA+PB pretreated mice (about 1.7%). Fifty-six microg TGF-beta1/kg had no detectable effect on apoptosis. CONCLUSION: These observations indicate that during chemically induced liver carcinogenesis in B6C3F1 mice basal rates of apoptoses in adenoma and carcinoma are higher than in normal liver and can be further increased by a proapoptotic cytokine.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Fator de Crescimento Transformador beta/farmacologia , Adenoma/tratamento farmacológico , Animais , Carcinoma/tratamento farmacológico , Camundongos , Fator de Crescimento Transformador beta1RESUMO
The tumor growth-inhibiting and chemo-potentiating effects of vitamin C and K(3)combinations have been demonstrated both in vitro and in vivo. The purpose of this study was to investigate the influence of orally administered vitamin C and K(3) on the metastasis of mouse liver tumor (T.L.T.) cells implanted in C3H mice. Adult male C3H mice were given water containing vitamin C and K3 (15 g/0.15 g dissolved in 1000 ml) beginning 2 weeks before tumor transplantation until the end of the experiment. T.L.T. cells (106) were implanted intramuscularly in the right thigh of mice. All mice were sacrificed 42 days after tumor transplantation. Primary tumor, lungs, lymph nodes and other organs or tissues suspected of harboring metastases were macroscopically examined. Samples of primary tumors, their local lymph nodes, lungs and main organs such as liver, kidneys, spleen were taken for histological examination. Forty-two percent of control mice exhibited lung metastases and 27% possessed metastases in local lymph nodes whereas 24% of vitamin-treated mice exhibited lung metastases and 10% possessed local lymph nodes metastases. The total number of lung metastases was 19 in control group and 10 in vitamin C and K(3)-treated mice. Histopathological examination of the metastatic tumors from the vitamin-treated mice revealed the presence of many tumor cells undergoing autoschizic cell death. These results demonstrate that oral vitamin C and K(3) significantly inhibited the metastases of T.L.T. tumors in C3H mice. At least a portion of this inhibition was due to tumor cell death by autoschizis.
Assuntos
Ácido Ascórbico/administração & dosagem , Neoplasias Hepáticas Experimentais/patologia , Metástase Neoplásica/prevenção & controle , Vitamina K 3/administração & dosagem , Adjuvantes Imunológicos/administração & dosagem , Animais , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Masculino , Camundongos , Camundongos Endogâmicos C3H , Transplante de NeoplasiasRESUMO
Non-toxic, dietary treatment with oligofructose or inulin clearly inhibited the growth of a transplantable mouse liver tumor (TLT) and potentiated its chemotherapy. Thus, it appeared interesting to investigate the possible radiotherapy-potentiating effects of the same dietary treatment. Dietary treatment with 15% oligofructose or inulin incorporated in the basal diet was started four weeks before intramuscular transplantation of TLT tumor cells into young adult male mice of the NMRI strain and was continued until the end of the experiment. When the tumors reached approximately 1000 mm3 they were irradiated with a single X-ray dose of 5 to 20 Gy. Tumor dimensions were measured twice weekly and their mean volume per group of mice was compared to the control groups fed the basal diet. This non-toxic dietary treatment with oligofructose or inulin potentiated the effects of radiotherapy at an optimal dose of 10 Gy to a statistically very highly significant (p < 0.0001) level. They were similar for oligofructose and inulin. The introduction of such non-toxic adjuvant treatment potentiating the effect of cancer radiotherapy in classical protocols of human cancer treatment appears to be possible and without any additional risk for the patients.
Assuntos
Inulina/farmacologia , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Neoplasias Hepáticas Experimentais/radioterapia , Oligossacarídeos/farmacologia , Radiossensibilizantes/farmacologia , Animais , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Terapia Combinada , Dieta , Relação Dose-Resposta à Radiação , Inulina/toxicidade , Neoplasias Hepáticas Experimentais/patologia , Masculino , Camundongos , Oligossacarídeos/toxicidade , Radiossensibilizantes/toxicidadeRESUMO
Deficiency of alkaline and acid DNase is a hallmark in all non-necrotic cancer cells in animals and humans. These enzymes are reactivated at early stages of cancer cell death by vitamin C (acid DNase) and vitamin K(3) (alkaline DNase). Moreover, the coadministration of these vitamins (in a ratio of 100:1, for C and K(3), respectively) produced selective cancer cell death. Detailed morphological studies indicated that cell death is produced mainly by autoschizis, a new type of cancer cell death. Several mechanisms are involved in such a cell death induced by CK(3), they included: formation of H(2)O(2) during vitamins redox cycling, oxidative stress, DNA fragmentation, no caspase-3 activation, and cell membrane injury with progressive loss of organelle-free cytoplasm. Changes in the phosphorylation level of some critical proteins leading to inactivation of NF-kappaB appear as main intracellular signal transduction pathways. The increase knowledge in the mechanisms underlying cancer cells death by CK(3) may ameliorate the techniques of their in vivo administration. The aim is to prepare the introduction of the association of vitamins C and K(3) into human clinics as a new, non-toxic adjuvant cancer therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Morte Celular/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Ácido Ascórbico/administração & dosagem , Morte Celular/fisiologia , Quimioterapia Adjuvante , DNA/metabolismo , Sinergismo Farmacológico , Inibidores Enzimáticos/farmacologia , Humanos , NF-kappa B/metabolismo , Neoplasias/patologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Fosforilação , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Células Tumorais Cultivadas , Vanadatos/farmacologia , Vitamina K 3/administração & dosagemRESUMO
Microscopic aspects, densitometric evaluation of Feulgen-stained DNA, and gel electrophoresis of total DNA have been used to elucidate the effects of 1, 2, and 3 h VC (ascorbic acid), VK3 (menadione), and combined VC:VK3 treatments on the cellular and nuclear morphology and DNA content of a human ovarian carcinoma cell line (MDAH 2774). Optical densitometry showed a significant decrease in cancer cell DNA content directly related to VC and VC:VK3 treatments while VK3 and VC:VK3 treated cells exhibited cytoskeletal changes that included self-excision of cytoplasmic pieces with no membranous organelles. Nuclei decreased in size and exhibited poor contrast consistent with progressive decondensation of their chromatin. Degraded chromatin was also detected in cytoplasmic autophagosomes. Nucleoli segregated their components and fragmented into small pieces. Gel electrophoretic analysis of total DNA revealed evidence of generalized DNA degradation specific to treated tumor cells. These results are consistent with previous observations [Scanning 20 (1998a) 564; Ultrastruct. Pathol. 25 (2001b) 183; J. Histochem. Cytochem. 49 (2001) 109] which demonstrated that the VC:VK3 combination induced autoschizic cell death by a series of cytoplasmic excisions without organelles along with specific nuclear ultrastructural damage.
Assuntos
Ácido Ascórbico/farmacologia , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/patologia , DNA de Neoplasias/metabolismo , Neoplasias Ovarianas/patologia , Vitamina K 3/farmacologia , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Linhagem Celular , Linhagem Celular Tumoral , Núcleo Celular/ultraestrutura , Proliferação de Células/efeitos dos fármacos , Cromatina/efeitos dos fármacos , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Fragmentação do DNA/efeitos dos fármacos , DNA de Neoplasias/ultraestrutura , Feminino , Humanos , Membranas Intracelulares/efeitos dos fármacos , Membranas Intracelulares/metabolismo , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/ultraestrutura , Espécies Reativas de Oxigênio/metabolismoRESUMO
Implanted human, androgen-independent prostatic carcinoma cells (DU145) into athymic (NCr nu/nu) mice produce diverse tumors on the peritoneal surfaces of many organs. Light and ultrastructural observations show that the mesothelial covering these surfaces are typically microvilli-coated, squamous cells or secretory cuboidal cells. The peritoneal regions colonized by tumors lack mesothelial cells and are covered by actively replicating carcinoma cells that grow as poorly differentiated cell clusters made of cell aggregates to somewhat compact spheroids covered with pleiomorphic microvilli and containing an undifferentiated vascular supply. These xenografts clusters invade the diaphragm and develop into tumors with both a basal solid aspect and an upper region of cribriform morphology. Furthermore, each tumor contains two cell types: (1) a poorly differentiated clear cell type, which grows into intraperitoneal tumors and (2) a large, basophilic cell type, which invades the peritoneal stroma of organs, including of the diaphragm.
Assuntos
Proliferação de Células , Peritônio/patologia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/fisiopatologia , Animais , Linhagem Celular Tumoral , Epitélio/patologia , Epitélio/fisiopatologia , Epitélio/ultraestrutura , Humanos , Masculino , Camundongos , Camundongos Nus , Microvilosidades/patologia , Microvilosidades/ultraestrutura , Invasividade Neoplásica/patologia , Invasividade Neoplásica/fisiopatologia , Invasividade Neoplásica/ultraestrutura , Neoplasias Peritoneais/fisiopatologia , Neoplasias Peritoneais/secundário , Neoplasias Peritoneais/ultraestrutura , Peritônio/fisiopatologia , Peritônio/ultraestrutura , Neoplasias da Próstata/ultraestrutura , Transplante HeterólogoRESUMO
Increase in cytosolic calcium concentration ([Ca2+](c)), release of endoplasmic reticulum (ER) calcium ([Ca2+](er)) and ER stress have been proposed to be involved in oxidative toxicity. Nevertheless, their relative involvements in the processes leading to cell death are not well defined. In this study, we investigated whether oxidative stress generated during ascorbate-driven menadione redox cycling (Asc/Men) could trigger these three events, and, if so, whether they contributed to Asc/Men cytoxicity in MCF-7 cells. Using microspectrofluorimetry, we demonstrated that Asc/Men-generated oxidative stress was associated with a slow and moderate increase in [Ca2+](c), largely preceding permeation of propidium iodide, and thus cell death. Asc/Men treatment was shown to partially deplete ER calcium stores after 90 min (decrease by 45% compared to control). This event was associated with ER stress activation, as shown by analysis of eIF2 phosphorylation and expression of the molecular chaperone GRP94. Thapsigargin (TG) was then used to study the effect of complete [Ca2+](er) emptying during the oxidative stress generated by Asc/Men. Surprisingly, the combination of TG and Asc/Men increased ER stress to a level considerably higher than that observed for either treatment alone, suggesting that [Ca2+](er) release alone is not sufficient to explain ER stress activation during oxidative stress. Finally, TG-mediated [Ca2+](er) release largely potentiated ER stress, DNA fragmentation and cell death caused by Asc/Men, supporting a role of ER stress in the process of Asc/Men cytotoxicity. Taken together, our results highlight the involvement of ER stress and [Ca2+](er) decrease in the process of oxidative stress-induced cell death in MCF-7 cells.