RESUMO
The purpose of this study was to evaluate the effect of cyproterone acetate (CPA, A) compared with new synthetic steroids 3alpha-acetoxy-5,6-epoxy-16-pregnen-20-one (B) and 17alpha-hydroxy-16beta-methyl-1,4,6-pregnatriene-3,20-dione (C) in rat prostate and brain. Groups of animals were treated either with A, B or C (4 mg kg(-1) day(-1)) by the intraperitoneal route for 5 days. Levels of reduced glutathione (GSH), 5-hydroxy-indole acetic acid (5-HIAA), lipid peroxidation (as thiobarbituric acid reactive substances, TBARS) and the activities of Na(+), K(+)- and total ATPases were assayed in prostate and brain for each group of animals including a control group. No appreciable changes were shown in Na(+), K(+)-ATPase and total ATPases and TBARS on prostate and brain of rats that received A, B and C steroids. However, the levels of GSH and 5-HIAA decreased significantly (P < 0.05) in both tissues for the steroids assayed. It is concluded that CPA and the homologues B and C steroids induce changes in the levels of GSH and serotonin in rat prostate and brain.
Assuntos
Biomarcadores/análise , Encéfalo/metabolismo , Acetato de Ciproterona/farmacologia , Estresse Oxidativo/fisiologia , Pregnatrienos/farmacologia , Pregnenolona/análogos & derivados , Próstata/metabolismo , Serotonina/fisiologia , Animais , Encéfalo/efeitos dos fármacos , Glutationa/metabolismo , Ácido Hidroxi-Indolacético/metabolismo , Masculino , Estresse Oxidativo/efeitos dos fármacos , Pregnenolona/farmacologia , Próstata/efeitos dos fármacos , Ratos , ATPase Trocadora de Sódio-Potássio/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismoRESUMO
1. The Wobbler mouse suffers an autosomal recessive mutation producing severe motoneuron degeneration and astrogliosis in the spinal cord. It has been considered a suitable model of human motoneuron disease, including the sporadic form of amyotrophic lateral sclerosis (ALS). 2. Evidences exist demonstrating increased oxidative stress in the spinal cord of Wobbler mice, whereas antioxidant therapy delayed neurodegeneration and improved muscle trophism. 21-Aminosteroids are glucocorticoid-derived hydrophobic compounds with antioxidant potency 3 times higher than vitamin E and 100 times higher than methylprednisolone. They do not bind to intracellular receptors, and prevent lipid peroxidation by insertion into membrane lipid bilayers. 3. In common with the spinal cord of ALS patients, Wobbler mice present astrocytosis with hyperexpression of glial fibrillary acidic protein (GFAP), and increased expression of nitric oxide synthase (NOS) and growth-associated protein (GAP-43) in motoneurons. Here, we review our studies on the effects of a 21-aminosteroid on GFAP, NOS, and GAP-43. 4. First, we showed that 21-aminosteroid treatment further increased GFAP-expressing astrocytes in gray matter of the Wobbler spinal cord. This effect may provide neuroprotection if one considers a trophic and beneficial function of astrocytes during the course of degeneration. Other neuroprotectans used in Wobbler mice (T-588) also increased pre-existing astrocytosis. 5. Second, histochemical determination of NADPH-diaphorase, a parameter indicative of neuronal NOS activity, showed that the 21-aminosteroid down-regulated the high activity of this enzyme in ventral horn motoneurons. Therefore, suppression of nitric oxide by decreasing NADPH-diaphorase (NOS) activity may provide neuroprotection considering that excess NO is highly toxic to motoneurons. 6. Finally, 21-aminosteroid treatment significantly attenuated the aberrant expression of both GAP-43 protein and mRNA in Wobbler motoneurons. Hyperexpression of GAP-43 possibly indicated abnormal synaptogenesis, denervation, and muscle atrophy, parameters which may return to normal following antioxidant steroid treatment. 7. Besides 21-aminosteroids, other steroids also behave as neuroprotectans. In this regard, degenerative diseases may constitute potential targets of these hormones, based on the fact that the spinal cord expresses in a regional and cell-specific fashion, receptors for androgens. progesterone, adrenal steroids, and estrogens.
Assuntos
Doença dos Neurônios Motores/tratamento farmacológico , Doença dos Neurônios Motores/genética , Fármacos Neuroprotetores/farmacologia , Pregnatrienos/farmacologia , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Mutantes Neurológicos , Doença dos Neurônios Motores/patologiaAssuntos
Adenosina , Alopurinol , Dissacarídeos , Eletrólitos , Glutamatos , Glutationa , Histidina , Insulina , Intestino Delgado , Manitol , Soluções para Preservação de Órgãos , Preservação de Órgãos/efeitos adversos , Pregnatrienos/farmacologia , Rafinose , Animais , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/patologia , Ratos , Ratos Wistar , Fatores de TempoRESUMO
The Wobbler mouse, a model of amyotrophic lateral sclerosis (ALS), presents motorneuron degeneration and pronounced astrogliosis in the spinal cord. We have studied factors controlling astrocyte proliferation in cultures derived from Wobbler and control mice spinal cord. Basal rate of [3H]thymidine incorporation was 15 times lower in Wobbler astrocytes. While in control cultured cells interleukin-1alpha (IL-1) and corticosterone (CORT) significantly increased proliferation, both agents were inactive in Wobbler astrocytes. The lack of response to CORT was not due to the absence of glucocorticoid receptors, because similar receptor amounts were found in Wobbler and control astrocytes. In contrast to IL-1 and CORT, transforming growth factor-beta1 (TGF-beta1) substantially increased proliferation of Wobbler astrocytes but not of control cells. Differences in response to TGF-beta1 were also obtained by measuring glial fibrillary acidic protein (GFAP) immunoreaction intensity, which was substantially higher in Wobbler astrocytes. Thus, abnormal responses to different mitogens characterized Wobbler astrocytes in culture. We suggest that TGF-beta1 may play a role in the reactive gliosis and GFAP hyperexpression found in the degenerating spinal cord of this model of ALS.
Assuntos
Astrócitos/metabolismo , Receptores de Glucocorticoides/metabolismo , Medula Espinal/metabolismo , Animais , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Corticosterona/farmacologia , Feminino , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/genética , Gliose/patologia , Interleucina-1/farmacologia , Cinética , Masculino , Camundongos , Camundongos Endogâmicos , Camundongos Mutantes Neurológicos , Doença dos Neurônios Motores/genética , Doença dos Neurônios Motores/patologia , Fármacos Neuroprotetores/farmacologia , Pregnatrienos/farmacologia , Valores de Referência , Medula Espinal/citologia , Medula Espinal/efeitos dos fármacos , Timidina/metabolismo , Fator de Crescimento Transformador beta/farmacologiaRESUMO
Expression of the growth-associated protein GAP-43 is increased in the spinal cord of ALS patients and Wobbler (wr) mice, murine models of the disease. In this work we examined if expression of GAP-43 mRNA in control and wr mice was sensitive to steroid treatment. A group of control and wr mice received s.c. a 50 mg pellet of the natural hormone corticosterone (CORT) or the antioxidant 21-aminosteroid U-74389F during 4 days. Basal levels of GAP-43 mRNA were 10-fold elevated in ventral horn motoneurons of untreated wr mice, compared to the low levels in controls. The high expression of GAP-43 mRNA in wr was attenuated by treatment with CORT (41%, p < 0.001) and U-74389F (36%, p < 0.001). Although specific GAP-43 mRNA labelling was present in some neurons around the central canal, its cellular expression was similar in controls and wr. Also, steroid treatment was ineffective in neurons around the central canal. Other regions of the spinal cord (i.e., dorsal horn neurons) expressed GAP-43 mRNA slightly above background levels. It is possible that attenuation of GAP-43 expression due to the natural hormone and the antioxidant steroid resulted from reversal of motoneuron degeneration or aberrant sprouting. Therefore, steroid therapy may be of value to prevent denervation and/or muscular atrophy in this animal model.
Assuntos
Antioxidantes/farmacologia , Corticosterona/farmacologia , Proteína GAP-43/genética , Regulação da Expressão Gênica/fisiologia , Neurônios Motores/metabolismo , Pregnatrienos/farmacologia , Medula Espinal/metabolismo , Animais , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hibridização In Situ , Masculino , Camundongos , Camundongos Mutantes Neurológicos , Células do Corno Posterior/efeitos dos fármacos , Células do Corno Posterior/metabolismo , RNA Mensageiro/genética , Transcrição Gênica/efeitos dos fármacosRESUMO
The wobbler mouse suffers an autosomal recessive mutation producing severe neurodegeneration and astrogliosis in spinal cord. It has been considered a model for amyotrophic lateral sclerosis. We have studied in these animals the expression of two proteins, the growth-associated protein (GAP-43) and the NADPH-diaphorase, the nitric oxide synthesizing enzyme, employing immunocytochemistry and histochemistry. We found higher expression of GAP-43 immunoreactivity in dorsal horn, Lamina X, corticospinal tract and ventral horn motoneurons in wobbler mice compared to controls. Weak NADPH-diaphorase activity was present in control motoneurons, in contrast to intense labeling of the wobbler group. No differences in diaphorase activity was measured in the rest of the spinal cord between control and mutant mice. A group of animals received subcutaneously for 4 days a 50 mg pellet of U-74389F, a glucocorticoid-derived 21-aminosteroid with antioxidant properties but without glucocorticoid activity. U-74389F slightly attenuated GAP-43 immunostaining in dorsal regions of the spinal cord from wobblers but not in controls. However, in motoneurons of wobbler mice number of GAP-43 immunopositive neurons, cell processes and reaction intensity were reduced by U-74389F. The aminosteroid reduced by 50% motoneuron NADPH-diaphorase activity. Hyperexpression of GAP-43 immunoreactivity in wobbler mice may represent an exaggerated neuronal response to advancing degeneration or muscle denervation. It may also be linked to increased nitric oxide levels. U-74389F may stop neurodegeneration and/or increase muscle trophism and stop oxidative stress, consequently GAP-43 hyperexpression was attenuated. Wobbler mice may be important models to evaluate the use of antioxidant steroid therapy with a view to its use in human motoneuron disease.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Antioxidantes/farmacologia , Di-Hidrolipoamida Desidrogenase/genética , Proteína GAP-43/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Neurônios Motores/metabolismo , Pregnatrienos/farmacologia , Medula Espinal/metabolismo , Esclerose Lateral Amiotrófica/genética , Animais , Di-Hidrolipoamida Desidrogenase/biossíntese , Modelos Animais de Doenças , Feminino , Proteína GAP-43/biossíntese , Humanos , Masculino , Camundongos , Camundongos Mutantes Neurológicos , Neurônios Motores/citologia , Neurônios Motores/patologia , Medula Espinal/citologia , Medula Espinal/patologiaRESUMO
1. Wobbler mice suffer an autosomal recessive mutation producing severe motoneuron degeneration and dense astrogliosis, with increased levels of glial fibrillary acidic protein (GFAP) in the spinal cord and brain stem. They have been considered animal models of amyotrophic lateral sclerosis and infantile spinal muscular atrophy. 2. Using Wobbler mice and normal littermates, we investigated the effects of the membrane-active steroid Lazaroid U-74389F on the number of GFAP-expressing astrocytes and glucocorticoid receptors (GR). Lazaroids are inhibitors of oxygen radical-induced lipid peroxidation, and proved beneficial in cases of CNS injury and ischemia. 3. Four days after pellet implantation of U-74389F into Wobbler mice, hyperplasia and hypertophy of GFAP-expressing astrocytes were apparent in the spinal cord ventral and dorsal horn, areas showing already intense astrogliosis in untreated Wobbler mice. In control mice, U-74389F also produced astrocyte hyperplasia and hypertophy in the dorsal horn and hyperplasia in the ventral-lateral funiculi of the cord. 4. Given in vivo U-74389F did not change GR in spinal cord of Wobbler or control mice, in line with the concept that it is active in membranes but does not bind to GR. Besides, U-74390F did not compete for [3H]dexamethasone binding when added in vitro. 5. The results suggest that stimulation of proliferation and size of GFAP-expressing astrocytes by U-74389F may be a novel mechanism of action of this compound. The Wobbler mouse may be a valuable animal model for further pharmacological testing of glucocorticoid and nonglucocorticoid steroids in neurodegenerative diseases.
Assuntos
Astrócitos/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/biossíntese , Pregnatrienos/farmacologia , Receptores de Glucocorticoides/biossíntese , Medula Espinal/efeitos dos fármacos , Adrenalectomia , Esclerose Lateral Amiotrófica , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Ligação Competitiva , Divisão Celular/efeitos dos fármacos , Corticosterona/sangue , Dexametasona/metabolismo , Modelos Animais de Doenças , Feminino , Genes Recessivos , Proteína Glial Fibrilar Ácida/análise , Homozigoto , Masculino , Camundongos , Camundongos Mutantes Neurológicos , Atrofia Muscular Espinal , Receptores de Glucocorticoides/análise , Valores de Referência , Medula Espinal/metabolismo , Medula Espinal/patologiaRESUMO
Lazaroids are a class of novel 21 aminosteroids. They have been reported to be potent inhibitors of lipid peroxidation, which is a major contributing factor to ischemia-reperfusion injury in the lung. A Lewis rat orthotopic left lung isotransplant model was used to investigate the effects of the lazaroid U74500A on pulmonary preservation. The heart-lung blocks of donor rats were flushed with and then stored in either standard University of Wisconsin solution or University of Wisconsin solution with 30 mumol/L of U74500A substituted for the dexamethasone. After 6 or 12 hours of cold storage at 0 degrees C, the left lungs were transplanted into recipient rats and reperfused for 1 hour. Pulmonary function was assessed by measuring oxygen and carbon dioxide tensions in arterial blood after removal of the right lung. Lipid peroxide concentrations were measured as a thiobarbituric acid-reactive substance. Although arterial oxygen and carbon dioxide pressures and water content after 6 hours of preservation followed by reperfusion were similar in both the lazaroid and dexamethasone groups, lipid peroxide concentration was significantly higher in the dexamethasone group (0.88 +/- 0.07 mumol/gm) than in the lazaroid group (0.54 +/- 0.07 mumol/gm) (p < 0.01). After 12 hours of preservation, there were significant differences between the lazaroid and dexamethasone groups in arterial oxygen pressure (339 +/- 70 vs 27 +/- 3 mm Hg, p < 0.01), arterial carbon dioxide pressure (24.3 +/- 2.7 vs 47.7 +/- 7.0 mm Hg, p < 0.001), and lipid peroxide concentrations (0.69 +/- 0.07 vs 1.30 +/- 0.09 mumol/gm, p < 0.001). We conclude that addition of U74500A to the flush and storage solution enhances the preservation of the pulmonary graft in this transplant model.