RESUMO
INTRODUCTION: This study aimed to examine the relationship between gender, apolipoprotein E (APOE) genotype, and mesial temporal atrophy in mild cognitive impairment (MCI) with and without progression to Alzheimer's disease (AD). METHODS: We evaluated 236 MCI patients with (n = 121) and without (n = 115) AD progression. Longitudinal MRI-based hippocampal volumes (HV) and entorhinal cortex (ERC) thickness were obtained. The Clinical Dementia Rating Sum of Boxes (CDR-SB) score was used to assess disease severity. RESULTS: We found a significant effect of APOE, gender, and clinical course (stable MCI versus MCI-AD progression) on HV. There was a significant effect of clinical course and APOE, but not gender, on ERC. Baseline HV and APOE4 status predicted MCI-AD progression in women. Baseline ERC and APOE4 status predicted MCI-AD progression in men. There were significant differences in CDR-SB scores between patients with and without MCI-AD progression, but not between males and females, or APOE4 carriers and non-carriers. CONCLUSIONS: HV, but not ERC, is strongly influenced by gender in MCI. The effects of gender and APOE4 on neuroimaging biomarkers have potentially important implications in the prediction of MCI-AD progression and should be taken into account in clinical trials.
Assuntos
Doença de Alzheimer/epidemiologia , Doença de Alzheimer/genética , Apolipoproteínas E/genética , Disfunção Cognitiva/epidemiologia , Disfunção Cognitiva/genética , Córtex Entorrinal/patologia , Idoso , Doença de Alzheimer/patologia , Atrofia/epidemiologia , Atrofia/patologia , Causalidade , Disfunção Cognitiva/patologia , Comorbidade , Progressão da Doença , Feminino , Seguimentos , Predisposição Genética para Doença/epidemiologia , Predisposição Genética para Doença/genética , Humanos , Incidência , Estudos Longitudinais , Masculino , Polimorfismo de Nucleotídeo Único/genética , Fatores de Risco , Distribuição por Sexo , Estados Unidos/epidemiologiaRESUMO
There is substantial evidence that the tumor promoter 4 beta-12-O-tetradecanoylphorbol-13-acetate (TPA) elicits enhanced arachidonic acid release and its metabolism to prostaglandins and lipoxygenase products in many cell types. The goal of this study was to determine whether 4 alpha-12-O-tetradecanoylphorbol-13-acetate (4 alpha TPA), a stereoisomer of TPA, can induce arachidonic acid release and whether it is by the same mechanism as release induced by TPA. The finding that 10 micrograms/ml 4 alpha TPA produces a response comparable with 1 microgram/ml TPA and with similar kinetics was unexpected. The mechanism mediating the TPA response appears to be the activation of protein kinase C (PKC), which subsequently results in phospholipase A2 activation. This is suggested by the observation that TPA-induced arachidonate release is inhibited 65% by 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7), an inhibitor of PKC and that TPA completely down-regulates PKC. In addition, down-regulation or depletion of PKC by prior treatment with TPA results in a 75% loss of response to a second TPA treatment. In vitro activation of partially purified PKC could be demonstrated for TPA but not 4 alpha TPA. 4 alpha TPA thus appears to induce the release of arachidonate by a different but unknown mechanism. The 4 alpha TPA effect is not significantly reduced by the PKC inhibitor H-7, and no evidence of PKC activation or down-regulation was observed. Additionally, 4 alpha TPA is unable to "down-regulate" arachidonate release when the two-treatment protocol is used and the down-regulation of PKC by TPA has little effect on 4 alpha TPA-induced arachidonate release. Cycloheximide inhibited TPA-induced arachidonate release by 80% and 4 alpha TPA-induced release by 50%, indicating a partial requirement for protein synthesis for both phorbol esters. Actinomycin D, on the other hand, inhibited the TPA response by 70%, but enhanced the 4 alpha TPA response by 169%. When used at 10- or 100-micrograms doses, 4 alpha TPA was found to lack activity with respect to ornithine decarboxylase induction, oxidant production, hyperplasia, inflammation, and tumor promotion, suggesting that arachidonate release is not sufficient to induce these events. This may be related to the observation that with TPA the extent of arachidonate metabolism to prostaglandin E2 is four- to fivefold greater than occurred with 4 alpha TPA, even under conditions of equivalent arachidonate release.
Assuntos
Ácidos Araquidônicos/metabolismo , Epiderme/metabolismo , Acetato de Tetradecanoilforbol/farmacologia , Animais , Ácido Araquidônico , Cicloeximida/farmacologia , Dactinomicina/farmacologia , Dinoprostona/metabolismo , Regulação para Baixo , Epiderme/efeitos dos fármacos , Camundongos , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismoRESUMO
Several responses suggested to be critical components of phorbol ester tumor promotion were compared in 12-O-tetradecanoylphorbol-13-acetate (TPA) promotion-sensitive SSIN and TPA promotion-resistant C57BL/6J mice. SSIN mice treated topically with 2 micrograms of TPA showed extensive hyperplasia accompanied by edema, measured as a 26% increase in water content of the skin. Only a very slight hyperplasia and 7% increased water content occurred after TPA treatment of C57BL/6J mice. The induction of ornithine decarboxylase was determined to be the same both in vivo and in vitro for SSIN and C57BL/6J mice, which does not correlate with the histological observations. Because hyperplasia and inflammation can be mediated by arachidonic acid metabolites, it was hypothesized that differences in this metabolic pathway would correlate with the histological responses. No significant qualitative or quantitative differences, however, were observed in the profiles of the major cyclooxygenase products between the strains of mice. Prostaglandin E2, the principal prostaglandin, was synthesized at a 3-fold greater level than prostaglandins D2 or F2 alpha in response to TPA. The most abundant lipoxygenase product was 12-hydroxyeicosatetraenoic acid followed by 8-, 15-, and 5-hydroxyeicosatetraenoic acid. 8-Lipoxygenase activity is elevated 24 h after TPA treatment in the SSIN mice by approximately 4-fold; no elevation is seen in C57BL/6J mice. A comparison of the oxidant response to TPA as well as to phospholipase C showed that SSIN epidermal cells generated a higher level, measured by chemiluminescence, than C57BL/6J cells. This suggests that oxidant generation or possibly 8-lipoxygenase activity may be the basis for the sensitivity or resistance to TPA as a hyperplasiogen and as a tumor promoter.
Assuntos
Araquidonato Lipoxigenases/biossíntese , Camundongos Endogâmicos/fisiologia , Ornitina Descarboxilase/biossíntese , Ésteres de Forbol/farmacologia , Dermatopatias/induzido quimicamente , Acetato de Tetradecanoilforbol/farmacologia , Animais , Cromatografia Líquida de Alta Pressão , Edema/induzido quimicamente , Indução Enzimática/efeitos dos fármacos , Ácidos Hidroxieicosatetraenoicos/metabolismo , Hiperplasia/induzido quimicamente , Camundongos , Camundongos Endogâmicos C57BL , Prostaglandinas/biossíntese , Dermatopatias/patologiaRESUMO
The ability of the hyperplasiogenic irritant ethyl phenylpropiolate (EPP) to act as a tumor promoter in two-stage carcinogenesis and to stimulate cellular events commonly cited as markers of tumor promoter action was evaluated. Treatment of adult, inbred SENCAR (SSIN) mice, initiated with 7,12-dimethylbenz(a)anthracene, with 5 mg of EPP twice weekly resulted in 100% of the mice developing tumors (4.8 tumors/mouse) after 40 weeks of promotion. Treatment with 3 mg EPP (twice weekly) resulted in 52% of the mice developing tumors (0.9 tumor/mouse). This treatment regimen with EPP produces a sustained epidermal hyperplasia without being overtly toxic. In addition, a 5-mg dose of EPP induced ornithine decarboxylase activity to a level comparable to that induced by the tumor promoter phorbol 12-myristate 13-acetate (PMA): 2.3 nmol CO2/mg protein/h for EPP versus 4.5 nmol CO2/mg protein/h for PMA versus 0.04 nmol CO2/mg protein/h for acetone control. Likewise, the time course of ornithine decarboxylase induction by EPP was the same as that seen with PMA (maximum induction at approximately 6 h). Vascular permeability of the dorsal skin increased significantly in response to EPP (8 times that seen in acetone controls) and exhibited the same kinetics as that seen after exposure to PMA. Activity of protein kinase C (PKC), the cellular receptor for PMA, decreased by 75 to 95% 48 h after treatment with PMA. In contrast, EPP treatment resulted in less than a 20% decrease in PKC activity 48 h after treatment. This slight decrease in PKC activity is thought to be an indirect effect caused by the hyperproliferative and inflammatory reactions, because EPP was found to be inactive as an in vitro activator of PKC. These results indicate not only that EPP is a good tumor promoter that causes morphological and biochemical responses similar to those induced by PMA, but also that the action of EPP is apparently mediated via a mechanism that does not involve direct interaction with PKC.
Assuntos
Alcinos/toxicidade , Carcinógenos/toxicidade , Neoplasias Cutâneas/induzido quimicamente , 9,10-Dimetil-1,2-benzantraceno , Animais , Regulação para Baixo , Indução Enzimática/efeitos dos fármacos , Feminino , Camundongos , Ornitina Descarboxilase/biossíntese , Proteína Quinase C/metabolismo , Neoplasias Cutâneas/enzimologia , Acetato de Tetradecanoilforbol , Fatores de TempoRESUMO
The activation of protein kinase C, induction of ornithine decarboxylase (ODC), and hyperplasia have been suggested to be linked, sequential processes resulting from phorbol ester application to mouse skin. However, evidence is presented indicating that these events are not necessarily linked or dependent on one another and that significant differences exist in these responses between phorbol ester promotion sensitive (SSIN) and resistant (C57BL/6J) mice. The epidermis from SSIN mice treated with a single application of 12-O-tetradecanoylphorbol-13-acetate (TPA) displayed a large induction of ODC and a subsequent extensive hyperplasia. A second TPA treatment at 24 or 48 h after the first did not result in ODC induction (refractory state), and protein kinase C was shown to be down-regulated at these times. By 72 h, however, a responsive state had returned even through protein kinase C remained down-regulated. The epidermis of C57BL/6J responds to a single application of TPA with a level of ODC induction similar to that of the SSIN mice. Protein kinase C was down-regulated by approximately 75% after 24 h and was virtually completely down-regulated at 48 and 72 h (95-97%). In contrast to the above findings for the sensitive mice, however, little, if any, hyperplasia was produced. In addition, while a second TPA treatment at 24 h did not result in ODC induction (refractory state), hyperplasia did occur within 24 to 48 h. When the second TPA application was given 48 h after the first, at a time when protein kinase C was down-regulated, an overinduction of ODC occurred, as well as subsequent hyperplasia. Furthermore, a significant number of papillomas resulted when these increased treatment frequencies, i.e., once a day or every other day, were used to promote dimethylbenz(a)anthracene-initiated C57BL/6J mice. It is concluded that, while hyperplasia remains an apparent requirement for tumor promotion, the ODC induction following an initial TPA treatment is insufficient for or not causally related to this hyperplasia. In addition, subsequent ODC induction, at least in the C57BL/6J mouse, is probably not mediated by protein kinase C.
Assuntos
Ornitina Descarboxilase/metabolismo , Proteína Quinase C/metabolismo , Pele/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Animais , Carcinoma/induzido quimicamente , Indução Enzimática/efeitos dos fármacos , Hiperplasia , Camundongos , Camundongos Endogâmicos C57BL , Papiloma/induzido quimicamente , Pele/enzimologia , Pele/patologia , Neoplasias Cutâneas/induzido quimicamente , Fatores de TempoRESUMO
Epidermal cells were isolated from adult inbred SENCAR (SSIN) mice and separated by density-gradient centrifugation. The cells were pooled into three fractions shown by previous work to differ in their state of differentiation and proliferative potential. The three fractions were examined for their capacity to metabolize exogenous 14C-arachidonic acid (AA) into prostaglandins (PG) and hydroxyeicosatetraenoic acids (HETE). Cells found in the upper two fractions, which are less dense, have less proliferative potential in vitro, and are more differentiated than cells in the lower more dense fraction, are much more active in producing PG from exogenous AA than are the more dense cells. This was observed in intact cells as well as cells disrupted by freeze-thawing following density separation. The same relationship was found for HETE production in that cytoplasmic preparations from the two fractions containing the less dense cells were much more active in the production of HETE than cytoplasmic preparations from the more dense fraction. The two upper fractions differed little from each other in the production of PG or HETE. These results indicate the presence of higher levels of active cyclooxygenase and lipoxygenases in fractions containing the less dense, more differentiated cells than in the fraction containing the more dense, less differentiated cells which are highly enriched for basal keratinocytes.
Assuntos
Ácidos Araquidônicos/metabolismo , Epiderme/metabolismo , Animais , Ácido Araquidônico , Diferenciação Celular , Separação Celular , Centrifugação com Gradiente de Concentração , Células Epidérmicas , Ácidos Hidroxieicosatetraenoicos/biossíntese , Camundongos , Prostaglandinas/biossínteseRESUMO
Because arachidonic acid-derived eicosanoids are potent modulators of hyperproliferation and inflammation during skin tumor promotion with the phorbol ester, 12-0-tetradecanoylphorbol-13-acetate (TPA) (17, 18), it was hypothesized that dietary modification of epidermal fatty acids might modulate TPA-induced biochemical events in mouse skin. Semipurified diets containing 10% total fat composed of corn oil (CO) or a combination of CO and menhaden oil (MO) or coconut oil (CT) were fed to SENCAR mice for 4 weeks. Fatty acid composition of epidermal phospholipids generally reflected fatty acid composition of dietary oils fed to the mice. Since fatty acid-derived eicosanoids are thought to be essential in tumorigenesis, we compared the effects of dietary fats on prostaglandin E (PGE) production in epidermis treated with a single dose of TPA. TPA-induced PGE production in mouse epidermis from mice fed the MO diet was significantly reduced compared to PGE production in epidermal homogenates from mice fed the CO or CT diets. Type of dietary fats did not appear to modulate TPA-induced vascular permeability, however hyperplasia was slightly elevated in skins of mice fed MO. The subcellular distribution of protein kinase C, the plasma membrane receptor for TPA predominantly located in the cytosol (80%), was altered in epidermis from mice fed the MO diet compared to preparations from mice fed CO or CT diets which exhibited normal protein kinase C distribution. Our results suggest that n-3 rich dietary lipids modulate TPA-elicited events in mouse skin to a greater extent than diets containing higher proportions of saturated or n-6 fatty acids.
Assuntos
Gorduras Insaturadas na Dieta/farmacologia , Epiderme/efeitos dos fármacos , Óleos de Plantas , Acetato de Tetradecanoilforbol/farmacologia , Animais , Permeabilidade Capilar , Óleo de Coco , Cocos , Óleo de Milho/administração & dosagem , Gorduras na Dieta/administração & dosagem , Epiderme/química , Epiderme/metabolismo , Feminino , Óleos de Peixe/administração & dosagem , Hiperplasia/patologia , Camundongos , Fosfolipídeos/análise , Prostaglandinas E/biossíntese , Proteína Quinase C/metabolismoRESUMO
In the present studies we have examined the effects of a new calcium channel blocker, LY393615 ((N-Butyl-[5,5-bis-(4-fluorophenyl)tetrahydrofuran-2-yl]methylamine hydrochloride, NCC1048) in a model of hypoxia-hypoglycaemia in vitro and in a gerbil model of global and in two rat models of focal cerebral ischaemia in vivo. Results indicated that LY393615 protected against hypoxia-hypoglycaemic insults in brain slices and also provided significant protection against ischaemia-induced hippocampal damage in gerbil global cerebral ischaemia when dosed at 10, 12.5 (P<0.05) or 15 mg/kg i.p. (P<0.01) 30 min before and 2 h 30 min after occlusion. The compound penetrated the brain well after a 15 mg/kg i.p. dose and had a half-life of 2.5 h. In further studies LY393615 was protective 1 h post-occlusion when administered at 15 mg/kg i.p. followed by 2 doses of 5 mg/kg i.p. 2 and 3 h later. LY393615 dosed at 15 mg/kg i.p. followed by 2 further doses of 5 mg/kg i.p. (2 and 3 h later) also produced a significant reduction in the infarct volume following Endothelin-1 (Et-1) middle cerebral artery occlusion in the rat when administration was initiated immediately (P<0.01) or 1 h (P<0.05) after occlusion. The compound was also evaluated in the intraluminal monofilament model of focal ischaemia. The animals had the middle cerebral artery occluded for 2 h, and 15 min after reperfusion LY393615 was administered at 15 mg/kg i.p. followed by 2 mg/kg/h i.v. infusion for 6 h. There was no reduction in infarct volume using this dosing protocol. In conclusion, in the present studies we have reported that a novel calcium channel blocker, LY393615, with good bioavailability protects against neuronal damage caused by hypoxia-hypoglycaemia in vitro and both global and focal cerebral ischaemia in vivo. The compound is neuroprotective when administered post-occlusion and may therefore be a useful anti-ischaemic agent.
Assuntos
Isquemia Encefálica/tratamento farmacológico , Butilaminas/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Furanos/farmacologia , Fármacos Neuroprotetores/farmacologia , Bloqueadores dos Canais de Sódio , Animais , Isquemia Encefálica/patologia , Butilaminas/química , Bloqueadores dos Canais de Cálcio/química , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Furanos/química , Gerbillinae , Hipocampo/irrigação sanguínea , Hipocampo/patologia , Técnicas In Vitro , Infarto da Artéria Cerebral Média/tratamento farmacológico , Infarto da Artéria Cerebral Média/patologia , Masculino , Fármacos Neuroprotetores/química , Células de Purkinje/efeitos dos fármacos , Células de Purkinje/patologia , Ratos , Ratos WistarRESUMO
In the present studies, we have evaluated the effects of N-[4-(2-¿[(3-Chlorophenyl)methyl]amino¿ethyl)phenyl]-2-thiophenecarbo ximidamide dihydrochloride (ARL 17477) on recombinant human neuronal NOS (nNOS) and endothelial NOS (eNOS). We then carried out pharmacokinetic studies and measured cortical nitric oxide synthase (NOS) inhibition to determine that the compound crossed the blood brain barrier. Finally, the compound was evaluated in a model of global ischaemia in the gerbil and two models of transient focal ischaemia in the rat. The IC(50) values for ARL 17477 on human recombinant human nNOS and eNOS were 1 and 17 microM, respectively. ARL 17477 (50 mg/kg i.p.) produced a significant reduction in the ischaemia-induced hippocampal damage following global ischaemia when administered immediately post-occlusion, but failed to protect when administration was delayed until 30 min post-occlusion. In the endothelin-1 model of focal ischaemia, ARL 17477 (1 mg/kg i.v.) significantly attenuated the infarct volume when administered at either 0, 1 or 2 h post-endothelin-1 (P<0.05). In the intraluminal suture model, ARL 17477 at both 1 and 3 mg/kg i.v. failed to reduce the infarct volume measured at 1, 3 or 7 days post-occlusion. These results demonstrate that ARL 17477 protects against global ischaemia in gerbils and provides some reduction in infarct volume following transient middle cerebral artery occlusion in rats, indicating that nNOS inhibition may be a useful treatment of ischaemic conditions.
Assuntos
Amidinas/farmacocinética , Isquemia Encefálica/tratamento farmacológico , Inibidores Enzimáticos/farmacocinética , Fármacos Neuroprotetores/farmacocinética , Óxido Nítrico Sintase/antagonistas & inibidores , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Isquemia Encefálica/enzimologia , Isquemia Encefálica/fisiopatologia , Linhagem Celular , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Interações Medicamentosas/fisiologia , Endotelina-1/farmacologia , Gerbillinae , Humanos , Masculino , Óxido Nítrico Sintase/efeitos dos fármacos , Óxido Nítrico Sintase/metabolismo , Ratos , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/fisiopatologia , Fatores de TempoRESUMO
In sickness-conditioned learning, animals become ill after sampling a new substance and develop an aversion that is expressed as avoidance of that substance in subsequent presentations. We examined the parameters of a one-trial, nongustatory, sickness-conditioned learning task in day-old chicks. Chicks pecked a bead and were made ill by i.p. injection of lithium chloride (LiCl). Both 0.5 and 1.0 M LiCl (0.1 ml) produced reliable avoidance at test. Chicks injected with LiCl between 15 and 45 min after training avoided the bead at test, whereas those injected within 5 or 10 min or more than 45 min after training did not. Avoidance was present until 24 h posttraining and absent after 48 h. Therefore, robust learning of the sickness-conditioned learning task occurs in one trial without the need for gustatory cues, and memory for the task lasts at least 24 h. Uses of this task to study memory formation in the day-old chick are discussed.
Assuntos
Animais Recém-Nascidos/fisiologia , Aprendizagem da Esquiva/fisiologia , Galinhas/fisiologia , Aprendizagem/fisiologia , Reforço Psicológico , Animais , Depressores do Sistema Nervoso Central/administração & dosagem , Depressores do Sistema Nervoso Central/farmacologia , Relação Dose-Resposta a Droga , Injeções Intraperitoneais , Cloreto de Lítio/administração & dosagem , Cloreto de Lítio/farmacologia , Masculino , Memória/efeitos dos fármacos , Paladar/efeitos dos fármacosRESUMO
Aqueous solutions of CuSO(4) were shown to inhibit cytochrome c reduction by xanthine-xanthine oxidase. Such copper solutions also significantly inhibited oxidant production, induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), measured as chemiluminescence, in murine epidermal cells in vitro. At the non-toxic level of 250 mum, CuSO(4) inhibited by 40% the induction of ornithine decarboxylase by TPA in murine epidermal cultures. The superoxide generating system xanthine-xanthine oxidase was shown to induce ornithine decarboxylase by two- to threefold; such induction was partially inhibited by CuSO(4). Superoxide dismutase slightly inhibited TPA-induced, but not basal DNA synthesis in cultured epidermal cells. For unknown reasons, DNA synthesis was enhanced by CuSO(4) alone and was further enhanced in the presence of TPA. It appears that oxidants generated in response to TPA partially mediate the induction of ornithine decarboxylase and to a lesser extent DNA synthesis.
RESUMO
Murine epidermal cells have previously been shown to produce an oxidant response to the mouse skin tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA); however, the cellular source of these oxidants has not been well characterized. The demonstration that phospholipase C also elicits this oxidant response suggests that protein kinase C is the common mediator. In order to pursue this hypothesis, studies using protein kinase C inhibitors were carried out; both the induced oxidant response and the induction of ornithine decarboxylase (ODC), a known protein kinase C mediated event, were studied. At 100 µm-palmitoylcarnitine inhibited TPA-induced ODC by 50% and phospholipase C-induced ODC by 95%. At the same dose level, the negative analogue acetylcarnitine had no inhibitory effect on ODC with either inducer. The protein kinase C inhibitor 1-(5-isoquinolinesulphonyl)-2-methylpiperazine dihydrochloride (H-7) and its negative analogue N-(2-guanidinoethyl)-5-isoquinolinesulphonamide hydrochloride (HA-1004) were also used. At 100 µm, H-7 completely inhibited both TPA and phospholipase C-induced ODC; at the same dose HA-1004 had no effect. When these agents were included in the chemiluminescence assay for oxidant generation, similar results were seen: at 100 µm-palmitoylcarnitine and H-7, but not acetylcarnitine or HA-1004, suppressed the response by nearly 100%. These results suggest that the oxidant response to TPA in epidermal cells is mediated at least in part by protein kinase C.
RESUMO
Arachidonic acid (AA) release and metabolism to prostaglandins (PG) in response to the complete mouse skin tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and the first-stage promoter A23187 were compared in keratinocytes from normal adult human and two different strains of mouse with different sensitivities to TPA as a tumour promoter. The rate, extent and distribution of incorporated [(14)C]AA among the classes of phospholipids were similar in cultures of either CD-1, SSIN or human keratinocytes. Using prelabelled cultures, the amount of radiolabel released in control cultures was nearly identical for human and both strains of mice. Distinct species differences were observed, however, after TPA treatment. Cultures of human epidermal keratinocytes (HEK) released only half as much label by 6 hr compared with either strain of mouse. In addition, whereas the mouse epidermal keratinocytes (MEK) metabolized AA readily to PGE(2), very little PGE(2) could be detected in the human cultures. The extent of variability between individual human samples (n = 11) in response to 0.1 mug TPA/ml ranged from a 20% increase to a 200% increase in release of label, with a mean increase of 50%, whereas murine cells produced a mean increase of 400%. When MEK and HEK were stimulated with 10(-6) and 10(-5)m-A23187, an increase in the release of arachidonate by 200 and 400%, respectively, was observed for both species. Under these conditions of equal release, equivalent amounts of PGE(2) were produced. To compare further the ability of mouse and human cells to metabolize exogenous AA to PGE(2), freshly isolated, as well as cultured, cells from each species were incubated with [(14)C]arachidonate. Under both conditions, HEK have approximately the same ability as MEK to metabolize AA to PGE(2) (approx. 2% of the arachidonate for both species). The reduced ability of HEK, compared with MEK, to produce PGE(2) is specific to TPA and is due primarily to insufficient substrate, that is, low levels of arachidonic acid release.
RESUMO
The biological activity, including metabolism and modulation of ornithine decarboxylase activity and DNA synthesis, of arachidonic acid (AA) and eicosapentaenoic acid (EPA) were compared in epidermal cells from SENCAR mice. Radiolabelled AA and EPA were found to be similarly incorporated into and released from membrane phospholipids of unstimulated cultures. However, when cells were stimulated with the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA), the release of AA was significantly higher than the release of EPA. The extent of metabolism of AA and EPA to prostaglandins was determined in both freeze-thawed cell preparations and in viable cultured cells. In the freeze-thawed preparations, use of AA as a substrate resulted in significantly more PGF than when EPA was used as the substrate. However, more PGE3 was formed than PGE2. PGD levels were the same for either fatty acid precursor. Prostaglandin production was also determined in viable cultured cells since other influences such as phospholipase A2 activity can modify prostaglandin production. Control cultures prelabelled with either AA or EPA produced similar amounts of the respective PGF, PGE, and PGD. However, TPA-stimulated cultures produced significantly higher amounts of each prostaglandin in cultures prelabelled with AA compared to cells prelabelled with EPA. HETE or HEPE production was the same both for cultured cells prelabelled with AA or EPA and for homogenates from uncultured cells incubated directly with the radiolabelled fatty acids. TPA-induced ornithine decarboxylase (ODC) was significantly higher in AA-treated cultures compared to EPA-treated cultures. AA supports DNA synthesis to a greater extent than EPA, either alone or in the presence of TPA.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Ácidos Araquidônicos/metabolismo , Ácido Eicosapentaenoico/metabolismo , Epiderme/metabolismo , Animais , DNA/biossíntese , Indução Enzimática , Células Epidérmicas , Congelamento , Lipoxigenase/metabolismo , Camundongos , Ornitina Descarboxilase/biossíntese , Fosfolipídeos/análise , Fosfolipídeos/isolamento & purificação , Prostaglandinas/biossínteseRESUMO
Exposure of isolated SENCAR mouse epidermal cells to the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) in vitro resulted in the production of oxidant species detected as chemiluminescence. This oxidant response can be inhibited by superoxide dismutase and copper complexes but not catalase or scavengers of hydroxyl radical or singlet oxygen, suggesting that the oxidant is superoxide anion. Inhibitors of various parts of the arachidonate cascade affect the TPA-induced oxidant response in a manner that corresponds to their effects on in vivo tumor promotion experiments. Agents that inhibit lipoxygenase activity, i.e. nordihydroguaiaretic acid, benoxaprofen, but not agents that are cyclooxygenase inhibitors, i.e. indomethacin, are effective in suppressing the oxidant response to TPA. Phospholipase C but not phospholipase A2 or D produced an oxidant response kinetically similar to that elicited by TPA. The inhibitors of TPA-induced oxidants inhibited the phospholipase C response to the same extent, suggesting that TPA and phospholipase C may produce an oxidant species through a common mechanism, via phospholipid turnover-protein kinase C activation. The relevance of oxidant production to the tumor promotion process is suggested by the ability of exogenous xanthine/xanthine oxidase, a superoxide anion-generating system, to induce ornithine decarboxylase, a characteristic of TPA-treated cells. In addition, oxidant production is significantly lower in cells from the TPA-promotion resistant C57BL/6J mouse. These studies provide further support for a role for reactive oxygens in the tumor promotion process.
Assuntos
Cocarcinogênese , Epiderme/metabolismo , Oxigênio/metabolismo , Neoplasias Cutâneas/induzido quimicamente , Acetato de Tetradecanoilforbol/farmacologia , Animais , Ácido Araquidônico , Ácidos Araquidônicos/metabolismo , Células Cultivadas , Epiderme/efeitos dos fármacos , Feminino , Medições Luminescentes , Masculino , Camundongos , Camundongos Endogâmicos , Ornitina Descarboxilase/metabolismo , Neoplasias Cutâneas/metabolismo , Superóxidos/metabolismoRESUMO
The type of dietary fat has been shown to modulate the initiation stage of mammary tumorigenesis, with saturated fat fed before and/or during carcinogen treatment resulting in increased tumor incidence. This study was designed to determine whether different types of dietary fat alter the initiation stage of skin carcinogenesis by use of the initiation-promotion mouse skin carcinogenesis model. Sencar mice were divided into three groups and maintained on one of the experimental diets. The AIN-76-based diets consisted of 10% total fat with various types of fat: 8.5% menhaden oil plus 1.5% corn oil, 8.5% coconut oil plus 1.5% corn oil, and 10% corn oil. After three weeks mice were initiated with 10 nmol dimethylbenz[a]anthracene (DMBA). Two weeks later, all mice were switched to a diet containing 5% corn oil. Promotion began four weeks after initiation with twice-weekly application of 1 microgram 12-O-tetradecanoylphorbol-13-acetate and continued for 12 weeks. No statistically significant differences in kilocalories of food consumed or body weights were observed between diet groups during the study. The final papilloma incidence, yield, and size were not significantly different among the diet groups. In a parallel study, [3H]DMBA binding to epidermal DNA showed no dietary differences. Unlike the mammary carcinogenesis model, these data suggest that the type of fat fed during DMBA initiation had minimal effects on this stage of skin carcinogenesis.
Assuntos
Gorduras na Dieta/administração & dosagem , Papiloma/induzido quimicamente , Neoplasias Cutâneas/induzido quimicamente , 9,10-Dimetil-1,2-benzantraceno , Animais , Peso Corporal , Óleo de Coco , Óleo de Milho/farmacologia , DNA/metabolismo , Ingestão de Alimentos , Ácidos Graxos/análise , Feminino , Óleos de Peixe/farmacologia , Camundongos , Fosfolipídeos/análise , Óleos de Plantas/administração & dosagem , Pele/químicaRESUMO
Fish oil has been shown to have a protective effect in some cancer models. To determine whether fish oil alters skin tumorigenesis, a study was designed using the initiation-promotion mouse skin carcinogenesis model, feeding mice during the promotion stage a constant overall amount of dietary fat (10%) in which the levels of menhaden oil (MO) varied from 0 to 8.5% or corn oil (CO) at 10%. SENCAR mice were initiated with 10 nmol dimethylbenz[a]anthracene. Two weeks later mice were divided into five groups and maintained on one of the following AIN-76 based diets consisting of: 8.5% coconut oil (CT)/1.5% CO (diet A); 1% MO/7.5% CT/1.5% CO (diet B); 4% MO/4.5% CT/1.5% CO (diet C); 8.5% MO/1.5% CO (diet D); or 10% CO (diet E). Two weeks later, promotion with twice weekly applications of 1 micrograms 12-O-tetradecanoylphorbol-13-acetate (TPA) was begun and continued for 24 weeks. No statistically significant differences in kcal food consumed or body wts were observed between diet groups during the study. The final papilloma and carcinoma incidence was not different among the diet groups. However, differences were seen in the rate of papilloma appearance with the group fed diet E (10% CO) being the slowest and diet B being the most rapid. In a parallel study, ornithine decarboxylase activity, a suggested marker of promotion, was greatly elevated in the epidermis of all TPA-treated mice and the effect of diet tended to reflect the different rates of tumor formation observed among the groups. These data indicate that the diets containing fish oil were not protective in the final incidence of tumor formation and suggest that a better understanding of the complex interactions is warranted before recommendations are made to alter the human diet for cancer prevention.
Assuntos
Antineoplásicos , Carcinoma/induzido quimicamente , Gorduras na Dieta/uso terapêutico , Óleos de Peixe/uso terapêutico , Neoplasias Cutâneas/induzido quimicamente , Acetato de Tetradecanoilforbol/toxicidade , Animais , Carcinoma/patologia , Carcinoma/prevenção & controle , Ingestão de Energia , Ácidos Graxos/análise , Feminino , Camundongos , Camundongos Endogâmicos , Ornitina Descarboxilase/metabolismo , Pele/efeitos dos fármacos , Pele/enzimologia , Pele/patologia , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/prevenção & controleRESUMO
Substantial evidence suggests that inflammation is an essential component of the phorbol ester tumor promotion stage of multistage carcinogenesis in mouse skin. In order to understand better the significance of this relationship, studies were directed at identifying the principal mediators of the vascular permeability component of inflammation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). Antihistamines and inhibitors of arachidonic acid metabolism were compared with respect to their anti-inflammatory activity and the correlation of this parameter with their effect on tumor promotion. The H1 histamine receptor antagonist, diphenhydramine, suppressed TPA-induced vascular leakage by 25 and 50% at topical doses of 0.342 mumol (100 micrograms) and 0.856 mumol (250 micrograms) respectively. In initiated mice, these same doses inhibited papilloma development by 40 and 75%. Inhibition of tumors was also observed when diphenhydramine was given orally. The H2 antagonist, cimetidine, which could only be supplied orally, had little effect on either TPA-induced vascular permeability or promotion. The lipoxygenase inhibitor nordihydroguaiaretic acid also suppressed vascular permeability and has been reported to inhibit papilloma development. The cyclooxygenase inhibitor indomethacin, however, has no effect on TPA-induced vascular permeability. Collectively, these data suggest that the increased vascular leakage observed with TPA contributes to tumor development and that this event is mediated by both the H1 histamine receptors and one or more of the lipoxygenase products of arachidonic acid.
Assuntos
Permeabilidade Capilar/efeitos dos fármacos , Cimetidina/farmacologia , Difenidramina/farmacologia , Papiloma/induzido quimicamente , Neoplasias Cutâneas/induzido quimicamente , Pele/patologia , Acetato de Tetradecanoilforbol/toxicidade , 9,10-Dimetil-1,2-benzantraceno/toxicidade , Animais , Feminino , Indometacina/farmacologia , Inflamação , Masoprocol/farmacologia , Camundongos , Camundongos Endogâmicos , Papiloma/patologia , Pele/efeitos dos fármacos , Neoplasias Cutâneas/patologiaRESUMO
The F1 progeny of a cross between 12-O-tetradecanoyl-phorbol-13-acetate (TPA) tumor promotion-sensitive SSIN mice and TPA promotion-resistant C57BL/6J mice were found to be sensitive to TPA as a tumor promoter. The tumor response was substantial, with an average of 15 papillomas per mouse and a 100% incidence following initiation with 400 nmol dimethylbenz[a]anthracene and promotion with 6.5 nmol (4 micrograms) TPA. To determine whether tumor promotability of the F1 mice correlates with other parameters believed to be associated with TPA responsiveness, oxidant generation, epidermal hyperplasia and edema were compared in the parents and F1 hybrids. The SSIN produced a strong hyperplastic response to TPA, the C57BL/6J a negligible response and the F1 hybrids a moderate response. In the SSIN, 6.5 nmol (4 micrograms) TPA caused an 18% increase in the water content of the skin (edema) while no change was seen for either the C57BL/6J or F1 hybrids. The oxidant response of the F1 hybrids to either TPA or phospholipase C was markedly less than that observed for the SSIN and was similar to the response previously observed for the C57BL/6J mice. These findings suggest that the oxidant response may not be an essential aspect of TPA tumor promotion. It may be related to the edema response, suggesting that at least this aspect of inflammation is not necessary for promotion.
Assuntos
Transformação Celular Neoplásica/efeitos dos fármacos , Ésteres de Forbol/farmacologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Testes de Mutagenicidade , Oxirredução , Acetato de Tetradecanoilforbol/farmacologia , Fosfolipases Tipo C/metabolismoRESUMO
Fish or vegetable oils were fed during the promotion stage of a mouse skin carcinogenesis model in order to investigate the effects of dietary fat on tumor development. Two weeks after initiation with 10 nmol dimethylbenz[a]anthracene, SENCAR mice were divided into five groups and maintained on one of the following semipurified diets containing 10% total fat and varying the type of fat: 8.5% coconut oil (CT)/1.5% corn oil (CO); 1% menhaden oil (MO)/7.5% CT/1.5% CO; 4% MO/4.5% CT/1.5% CO; 8.5% MO/1.5% CO; or 10% CO. Promotion with twice-weekly applications of 40 mg benzoyl peroxide was begun 2 weeks later and continued for 52 weeks. No statistically significant differences in kcal food consumed or body weights were observed between diet groups. Papilloma latency, incidence and yield differed among the diet groups with the group fed the 8.5% CT/1.5% CO diet having the shortest latency and highest papilloma incidence and number. In addition, carcinoma latency and incidence was assessed and the first carcinoma appeared in the group fed 8.5% CT/1.5% CO after 20 weeks of benzoyl peroxide treatment; this group yielded the highest carcinoma incidence throughout the study. In comparison, the group fed the 10% CO diet had the longest latency period, and among the lowest papilloma and carcinoma incidence and fewest tumors. In parallel studies, ornithine decarboxylase activity, vascular permeability and hyperplasia were elevated in the epidermis of benzoyl peroxide-treated mice but the extent of the response did not correlate with the different rates of tumor formation observed among the diet groups. These data indicate that dietary fat modulates tumor promotion by benzoyl peroxide in this skin carcinogenesis model with the predominantly saturated fat diet producing the highest rates of papilloma and carcinogen formation and the polyunsaturated fat diet the lowest.