RESUMEN
Sarcopenia is the loss of muscle mass and strength produced by aging or secondary to chronic diseases such as chronic liver disease (CLD). Although not all types of sarcopenia involve the same features, the most common are decreased fiber diameter and myosin heavy chain (MHC) levels, increased activity of ubiquitin-proteasome system (UPS) and reactive oxygen species (ROS). In this study, we aim to characterize the development of sarcopenia secondary to CLD induced by the hepatotoxin 5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). For this purpose, four-months-old male C57BL6 mice were fed with normal diet or DDC supplemented diet for 6 weeks. Functional tests to evaluate muscle strength, mobility, and motor skills were performed in alive mice. The muscle strength in isolated gastrocnemius was also assayed via electrophysiological measurements. Morphometric measures of fibers' diameter, total and ubiquitinated protein levels of myosin heavy chain (MHC), E3 ubiquitin ligases, ROS, and oxidation-dependent modified proteins in gastrocnemius tissue were also determined. Our results demonstrated that mice fed the DDC diet developed muscle wasting as evidenced by a loss of muscle mass and decreased muscle strength. The muscles of mice fed with DDC diet have a decreased diameter of fibers and MHC levels, also as increased MuRF-1 and atrogin-1 protein levels, ROS levels, and oxidation-modified protein levels. Additionally, control and DDC mice have the same food and water intake as well as mobility. Our results demonstrate mice with CLD develop sarcopenia involving decreased levels of myofibrillar proteins, increased UPS, and oxidative stress, but not for impaired caloric intake or immobility.
Asunto(s)
Hepatopatías/complicaciones , Músculo Esquelético/metabolismo , Estrés Oxidativo , Complejo de la Endopetidasa Proteasomal/metabolismo , Sarcopenia/metabolismo , Ubiquitinación , Animales , Línea Celular , Dicarbetoxidihidrocolidina/toxicidad , Hepatopatías/etiología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Musculares/metabolismo , Cadenas Pesadas de Miosina/metabolismo , Proteínas Ligasas SKP Cullina F-box/metabolismo , Sarcopenia/etiología , Proteínas de Motivos Tripartitos/metabolismo , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Mounting evidence indicates that structural and functional vascular changes associated with two-kidney, one-clip (2K-1C) hypertension result, at least in part, from altered activity of matrix metalloproteinases (MMPs). Because MMPs are upregulated by increased formation of reactive oxygen species (ROS), we hypothesized that antioxidant approaches could attenuate the increases in MMP-2 expression/activity and the vascular dysfunction and remodeling associated with 2K-1C hypertension. Sham-operated or 2K-1C hypertensive rats were treated with tempol 18 mg/kg/day or apocyanin 25 mg/kg/day (or vehicle). Systolic blood pressure was monitored weekly. After 8 weeks of treatment, aortic rings were isolated to assess endothelium-dependent and -independent relaxation. Quantitative morphometry of structural changes in the aortic wall was studied in hematoxylin/eosin sections. Aortic and systemic ROS levels were measured using dihydroethidine and thiobarbituric acid-reactive substances, respectively. Aortic MMP-2 levels and activity were determined by gelatin and in situ zymography, fluorimetry, and immunohistochemistry. Tempol and apocyanin attenuated 2K-1C hypertension (181+/-20.8 and 192+/-17.6 mm Hg, respectively, versus 213+/-18 mm Hg in hypertensive controls; both p<0.05) and prevented the reduction in endothelium-dependent vasorelaxation found in 2K-1C rats. Tempol, but not apocyanin (p>0.05), prevented the vascular remodeling found in 2K-1C rats (all p<0.01). Tempol was more effective than apocyanin in attenuating hypertension-induced increases in oxidative stress (both p<0.05), MMP-2 levels, and MMP-2 activity in hypertensive rats (all p<0.05). Our results suggest that antioxidant approaches decrease MMP-2 upregulation and attenuate the vascular dysfunction and remodeling during 2K-1C hypertension.
Asunto(s)
Acetofenonas/administración & dosificación , Antioxidantes/administración & dosificación , Óxidos N-Cíclicos/administración & dosificación , Metaloproteinasa 2 de la Matriz/metabolismo , Arteria Renal/metabolismo , Animales , Determinación de la Presión Sanguínea , Dicarbetoxidihidrocolidina/análogos & derivados , Dicarbetoxidihidrocolidina/metabolismo , Hipertensión Renovascular/enzimología , Hipertensión Renovascular/patología , Hipertensión Renovascular/fisiopatología , Inmunohistoquímica , Masculino , Metaloproteinasa 2 de la Matriz/genética , Técnicas de Cultivo de Órganos , Estrés Oxidativo/fisiología , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Arteria Renal/patología , Arteria Renal/cirugía , Marcadores de Spin , Tiobarbitúricos/metabolismo , Vasodilatación/efectos de los fármacos , Vasodilatación/fisiologíaRESUMEN
This study focuses on the alterations suffered by the serotoninergic and kinurenergic routes of tryptophan (TRP) metabolism in liver, and their relation with gluconeogenic phosphoenolpyruvate-carboxykinase (PEPCK) blockage in experimental acute porphyria. This porphyria was induced in rats by a combined treatment of 2-allyl-2-isopropylacetamide (100, 250, 500 mg/kg bw) and 3,5-dietoxicarbonil 1,4-dihydrocollidine (constant 50 mg/kg bw dose). Results showed a marked dose-dependent increase of all TRP pyrrolase (TRPp) forms, active (holo, total) and inactive (apo), and a decrease in the degree of enzyme saturation by heme. Increases for holo, total, and apo-TRPp were 90, 150, and 230%, respectively, at the highest dose assayed (H). The treatment also impaired the serotoninergic route of TRP metabolism in liver, causing a decrease in serotonin level (H, 38%), and a concomitant enhancement in TRP content (H, 23%). The porphyrinogenic treatment promoted a blockage in PEPCK activity (H, 30%). This occurred in correlation to the development of porphyria, to TRPp alterations and to the production of hepatic microsomal thiobarbituric acid reactive substances. Porphyria was estimated through increases in 5-aminolevulinic acid-synthase (ALA-S) activity, ALA and porphobilinogen contents, and a decrease in ferrochelatase activity. Thus, the TRP kynurenine route was augmented whereas the serotoninergic route was reduced. PEPCK blockage could be partly attributed to quinolinate generated from TRP by the increase of TRPp activity, which would be due to the effect of porphyrinogenic drugs on TRP. The contribution of ROS to PEPCK blockage is analyzed. Likewise, the implication of these results in the control of porphyrias by glucose is discussed.
Asunto(s)
Gluconeogénesis , Hígado/metabolismo , Fosfoenolpiruvato Carboxiquinasa (ATP)/antagonistas & inhibidores , Porfirias/metabolismo , Triptófano/metabolismo , 5-Aminolevulinato Sintetasa/metabolismo , Enfermedad Aguda , Alilisopropilacetamida/toxicidad , Animales , Dicarbetoxidihidrocolidina/toxicidad , Relación Dosis-Respuesta a Droga , Femenino , Porfirias/inducido químicamente , Ratas , Ratas WistarRESUMEN
Acute hepatic porphyrias are human metabolic diseases characterized by the accumulation of heme precursors, such as 5-aminolevulinic acid (ALA). The administration of glucose can prevent the symptomatology of these diseases. The aim of this work was to study the relationship between glucose metabolism disturbances and the development of experimental acute hepatic porphyria, as well as the role of reactive oxygen species (ROS) through assays on hepatic key gluconeogenic and glycogenolytic enzymes; phosphoenolpyruvate carboxykinase (PEPCK) and glycogen phosphorylase (GP), respectively. Female Wistar rats were treated with three different doses of the porphyrinogenic drug 2-allyl-2-isopropylacetamide (AIA) and with a single dose of 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). Thus, rats were divided into the following groups: group L (100 mg AIA + 50 mg DDC/kg body wt.); group M (250 mg AIA + 50 mg DDC/kg body wt.) and group H (500 mg AIA + 50 mg DDC/kg body wt.). The control group (group C) only received vehicles (saline solution and corn oil). Acute hepatic porphyria markers ALA-synthase (ALA-S) and ferrochelatase, heme precursors ALA and porphobilinogen (PBG), and oxidative stress markers superoxide dismutase (SOD) and catalase (CAT) were also measured in hepatic tissue. On the other hand, hepatic cytosolic protein carbonyl content, lipid peroxidation and urinary chemiluminescence were determined as in vivo oxidative damage markers. All these parameters were studied in relation to the different doses of AIA/DDC. Results showed that enzymes were affected in a drug-dose-dependent way. PEPCK activity decreased about 30% in group H with respect to groups C and L, whereas GP activity decreased 53 and 38% in group H when compared to groups C and L, respectively. On the other hand, cytosolic protein carbonyl content increased three-fold in group H with respect to group C. A marked increase in urinary chemiluminescence and a definite increase in lipid peroxidation were also detected. The activity of liver antioxidant enzyme SOD showed an induction of about 235% in group H when compared to group C, whereas CAT activity diminished due to heme depletion caused by both drugs. Based on these results, we can speculate that the alterations observed in glucose metabolism enzymes could be partly related to the damage caused by ROS on their enzymatic protein structures, suggesting that they could be also linked to the beneficial role of glucose administration in acute hepatic porphyria cases.
Asunto(s)
Glucosa/metabolismo , Hígado/enzimología , Porfiria Intermitente Aguda/enzimología , Especies Reactivas de Oxígeno/metabolismo , Alilisopropilacetamida/toxicidad , Animales , Dicarbetoxidihidrocolidina/toxicidad , Modelos Animales de Enfermedad , Femenino , Hemo/biosíntesis , Peroxidación de Lípido , Hígado/metabolismo , Mediciones Luminiscentes , Porfiria Intermitente Aguda/inducido químicamente , Porfiria Intermitente Aguda/metabolismo , Ratas , Ratas Wistar , Orina/químicaRESUMEN
The naturally occurring polyamines--putrescine, spermidine and spermine--are organic cations present in all living cells and essential for cell growth and differentiation. The aim of the present study was to extend the investigations on the effects of porphyrinogenic compounds on polyamine metabolism. This was achieved by studying putrescine, spermidine and spermine levels in a model of acute porphyria, i.e. 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-induced porphyria, and in a model of non-acute porphyria, i.e. hexachlorobenzene (HCB)-induced porphyria. HCB administration to female Wistar rats for 7, 14, 21, 28 and 56 days did not alter polyamine levels in liver, even though rats presented clear signs of HCB-induced porphyria. In contrast to HCB, DDC treatment resulted in a remarkable increase in putrescine levels in the liver of female and male Sprague-Dawley rats. This increase was due, at least in part, to ornithine decarboxylase (ODC) activation. DDC induction of putrescine levels did not show organ specificity, since it could also be seen in adrenal gland. Interestingly, the deregulation of polyamine biosynthesis occurred concomitantly with the deregulation of the heme biosynthetic pathway. In addition to porphyria, it is known that DDC intoxication affects several proteins of the hepatocyte cytoskeleton. It is suggested that DDC-induced increase in ODC activity and putrescine levels may be an early event contributing to alter the cytoskeleton.
Asunto(s)
Poliaminas Biogénicas/metabolismo , Dicarbetoxidihidrocolidina/farmacología , Hexaclorobenceno/farmacología , Porfirinas/biosíntesis , 5-Aminolevulinato Sintetasa/metabolismo , Animales , Femenino , Ferroquelatasa/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ornitina Descarboxilasa/metabolismo , Proteínas/metabolismo , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Caracteres Sexuales , Uroporfirinógeno Descarboxilasa/metabolismoRESUMEN
The use of antineoplastics is common in cancer therapy, and some of them have been associated with the development of porphyria in patients with cancer. However, knowledge of their effects on the haeme metabolic pathway is at present scarce and unclear. So, the present study evaluates the porphyrinogenic ability of nine antineoplastics (both alkylating and non-alkylating). These were tested either alone or in conjunction with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (latent porphyria model) in chick embryos and in mice. The results obtained suggest that the use of cyclophosphamide, azathioprine, 5-fluorouracil, busulphan, procarbazine and hexamethylmelamine be avoided in the treatment of porphyric patients. On the other hand, dacarbazine, chlorambucil and melphalan are non-porphyrinogenic. We also provide evidence showing that neither the presence of the mustard group in the structure of the antineoplastic nor alterations in ferrochelatase or protoporphyrinogen oxidase activities are responsible for the porphyrinogenic ability of cyclophosphamide.
Asunto(s)
Antineoplásicos/toxicidad , Dicarbetoxidihidrocolidina/toxicidad , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH , Porfirias/inducido químicamente , Alquilantes/toxicidad , Altretamina/toxicidad , Animales , Antineoplásicos/efectos adversos , Antineoplásicos/química , Azatioprina/toxicidad , Busulfano/toxicidad , Embrión de Pollo , Ciclofosfamida/efectos adversos , Ciclofosfamida/química , Ciclofosfamida/toxicidad , Dacarbazina/toxicidad , Femenino , Ferroquelatasa/efectos de los fármacos , Ferroquelatasa/metabolismo , Flavoproteínas , Fluorouracilo/toxicidad , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Proteínas Mitocondriales , Oxidorreductasas/efectos de los fármacos , Oxidorreductasas/metabolismo , Porfirinas/metabolismo , Procarbazina/toxicidad , Protoporfirinógeno-Oxidasa , Relación Estructura-ActividadRESUMEN
In this paper, we studied the effect of heme availability on corticosterone and aldosterone synthesis in rat adrenal. We found that hemin stimulated corticosterone and aldosterone production in adrenal homogenates in a dose-dependent fashion. Hemin administration to rats also provoked an increase in both corticosterone and aldosterone content in adrenal. 3,5-Diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC), an inhibitor of liver ferrochelatase activity, was able to inhibit this enzyme in rat adrenal. This resulted in an impairment of heme concentration and consequently adrenal ALA-synthase and porphyrin content were increased. Thus, it was proven that DDC inhibits heme biosynthesis in adrenal as it does in liver. In vivo experiments with rats showed that DDC was able to partially blocked ACTH-mediated corticosterone and aldosterone production while hemin administration was able to partially restore it. These data indicate that heme availability affects steroid biosynthesis in rat adrenal.
Asunto(s)
Glándulas Suprarrenales/metabolismo , Aldosterona/biosíntesis , Corticosterona/biosíntesis , Hemo/metabolismo , 5-Aminolevulinato Sintetasa/metabolismo , Glándulas Suprarrenales/efectos de los fármacos , Hormona Adrenocorticotrópica/farmacología , Animales , Desoxicorticosterona/metabolismo , Dicarbetoxidihidrocolidina/farmacología , Ferroquelatasa/efectos de los fármacos , Ferroquelatasa/metabolismo , Hemina/farmacología , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Porfirinas/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Aldosterone production from 11-deoxycorticosterone was stimulated by hemin in primary cultures and homogenates of calf adrenal zona glomerulosa, in a time- and dose-dependent fashion. The ferrochelatase inhibitor 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) blocked the stimulation of aldosterone mediated by adrenocorticotropin (ACTH). Addition of hemin after treatment with DDC partially restored ACTH action. These results suggest that hemin may play a role in regulation of aldosterone production.