RESUMEN
Amitriptyline, a tricyclic antidepressant, has been proposed as an antitumoral drug in oxidative therapy. Its pro-apoptotic effects, mediated by high reactive oxygen species generation, have been already described. In this study we analysed the effect of amitriptyline on the biosynthesis of coenzyme Q10 (CoQ), an essential component for electron transport and a potent membrane antioxidant involved in redox signaling. We treated H460 cells, a non-small-cell lung cancer cell line, with amitriptyline and we analysed CoQ levels by HPLC and CoQ biosynthesis rate, as well as the enzymes involved in CoQ biosynthesis by real-time PCR and Western blot. Amitriptyline treatment induced a dose-dependent decrease in CoQ levels in tumor cells. CoQ decreased levels were associated with down-regulation of the expression of COQ4 gene, as well as decreased Coq4 and Coq6 protein levels. Our findings suggest that the effect of amitriptyline on CoQ biosynthesis highlights the potential of this drug for antitumoral oxidative therapy.
Asunto(s)
Amitriptilina/farmacología , Neoplasias Pulmonares/patología , Ubiquinona/análogos & derivados , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Humanos , Cinética , Especies Reactivas de Oxígeno/metabolismo , Ubiquinona/biosíntesis , Ubiquinona/metabolismoRESUMEN
INTRODUCTION: Leptin, an inflammation-related adipokine, and its receptor (LEPR) are expressed in human dental pulp. Dentin sialophosphoprotein (DSPP) is involved in dentinogenesis and the dental pulp reparative response. The cell type expressing LEPR in dental human pulp and the resultant effect of the binding of leptin to LEPR remain unknown. This study describes the immmunohistochemical localization of LEPR and the effect of leptin on DSPP expression in human dental pulp. METHODS: Twenty-five dental pulp specimens were obtained from freshly extracted caries-free and restoration-free human third molars. LEPR localization was examined by immunohistochemistry using the antihuman LEPR monoclonal antibody. The effect of leptin on DSPP expression was determined by immunoblot analysis and quantitative real-time polymerase chain reaction. RESULTS: Immunoreactivity for LEPR concentrated in the odontoblast layer but was not evident in the central zone of the dental pulp. Leptin dose dependently stimulated DSPP expression. Western blot analysis revealed the presence of a protein with an apparent molecular weight of â¼00 kDa, the estimated molecular weight of DSPP. The expression of DSPP messenger RNA was confirmed by quantitative real-time polymerase chain reaction, and the size of the amplified fragments (298 bp) was confirmed by agarose gel electrophoresis. CONCLUSIONS: The present study shows that human dental pulp is immunoreactive for LEPR, with the immunoreactivity concentrated in the odontoblast layer, and that leptin stimulates, in a dose-dependent manner, DSPP protein and messenger RNA (mRNA) expression in human dental pulp. These findings further support the functional role of leptin in the dentin mineralization process and/or in dental pulp reparative and immune responses.
Asunto(s)
Pulpa Dental/metabolismo , Proteínas de la Matriz Extracelular/biosíntesis , Leptina/fisiología , Fosfoproteínas/biosíntesis , Sialoglicoproteínas/biosíntesis , Adulto , Humanos , Persona de Mediana Edad , Odontoblastos/metabolismo , Adulto JovenRESUMEN
AIMS: Fibromyalgia (FM) is a prevalent chronic pain syndrome characterized by generalized hyperalgesia associated with a wide spectrum of symptoms such as fatigue and joint stiffness. Diagnosis of FM is difficult due to the lack of reliable diagnostic biomarkers, while treatment is largely inadequate. We have investigated the role of coenzyme Q10 (CoQ10) deficiency and mitochondrial dysfunction in inflammasome activation in blood cells from FM patients, and in vitro and in vivo CoQ10 deficiency models. RESULTS: Mitochondrial dysfunction was accompanied by increased protein expression of interleukin (IL)-1ß, NLRP3 (NOD-like receptor family, pyrin domain containing 3) and caspase-1 activation, and an increase of serum levels of proinflammatory cytokines (IL-1ß and IL-18). CoQ10 deficiency induced by p-aminobenzoate treatment in blood mononuclear cells and mice showed NLRP3 inflammasome activation with marked algesia. A placebo-controlled trial of CoQ10 in FM patients has shown a reduced NLRP3 inflammasome activation and IL-1ß and IL-18 serum levels. INNOVATION: These results show an important role for the NLRP3 inflammasome in the pathogenesis of FM, and the capacity of CoQ10 in the control of inflammasome. CONCLUSION: These findings provide new insights into the pathogenesis of FM and suggest that NLRP3 inflammasome inhibition represents a new therapeutic intervention for the disease.
Asunto(s)
Proteínas Portadoras/metabolismo , Fibromialgia/sangre , Inflamasomas/metabolismo , Ubiquinona/análogos & derivados , Vitaminas/administración & dosificación , Adulto , Animales , Proteínas Portadoras/genética , Estudios de Casos y Controles , Caspasa 1/metabolismo , Células Cultivadas , Citocinas/sangre , Suplementos Dietéticos , Método Doble Ciego , Activación Enzimática , Femenino , Fibromialgia/tratamiento farmacológico , Fibromialgia/inmunología , Expresión Génica , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Lipopolisacáridos/farmacología , Masculino , Ratones , Persona de Mediana Edad , Mitocondrias/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR , Estrés Oxidativo , Ubiquinona/administración & dosificación , Ubiquinona/metabolismoRESUMEN
Besides intervening in calcium homeostasis by means of calcitonin, C cells are also implicated in the synthesis of an increasing number of regulatory peptides that could exert a paracrine regulation on the neighbouring follicular cells. Among the latest peptides reported in C cells, there are several characteristic hypothalamic peptides, such as TRH, CART, and ghrelin, which are mainly involved in the regulation of the metabolism at hypothalamic-pituitary-thyroid axis. The main aim of the present work has been to study the synthesis of the referred hypothalamic peptides by normal and neoplastic C cells of different mammals as well as in C-cell lines of both rat (CA-77, 6-23) and human (TT) origins in order to elucidate whether this is a fact in this kind of vertebrates. With that objective, we have applied the immunoperoxidase technique to analyze the presence of TRH, CART, ghrelin, and somatostatin in thyroid tissues of different species, and immunofluorescence to study those same peptides in C-cell cultures. Furthermore, we have investigated their expression at mRNA level by RT-PCR analysis. Our results demonstrate immunocolocalization of CART, ghrelin, somatostatin and TRH with calcitonin in normal C cells of different mammals, as well as in rat and human neoplastic C cells. We also confirm the expression of those peptides in rat and human C-cell lines by RT-PCR. Consequently, we can conclude that the synthesis of those peptides by C cells is a general event characteristic of the thyroid gland in mammals.
Asunto(s)
Glándula Tiroides/citología , Glándula Tiroides/metabolismo , Animales , Ghrelina/metabolismo , Cobayas , Humanos , Técnicas In Vitro , Proteínas del Tejido Nervioso/metabolismo , Conejos , Ratas , Somatostatina/metabolismo , Porcinos , Hormona Liberadora de Tirotropina/metabolismoRESUMEN
Fibromyalgia (FM) is a complex disorder that affects up to 5% of the general population worldwide. Both mitochondrial dysfunction and inflammation have been implicated in the pathophysiology of FM. We have investigated the possible relationship between mitochondrial dysfunction, oxidative stress, and inflammation in FM. We studied 30 women diagnosed with FM and 20 healthy women. Blood mononuclear cells (BMCs) from FM patients showed reduced level of coenzyme Q10 (CoQ10) and mtDNA contents and high level of mitochondrial reactive oxygen species (ROS) and serum tumor necrosis factor (TNF)-alpha and transcript levels. A significant negative correlation between CoQ10 and TNF-alpha levels (r=-0.588; p<0.01), and a positive correlation between ROS and TNF-alpha levels (r=0.791; p<0.001) were observed accompanied by a significant correlation of visual analogical scale with serum TNF-alpha and transcript levels (r=0.4507; p<0.05 and r=0.7089; p<0.001, respectively). TNF-alpha release was observed in an in vitro (BMCs) and in vivo (mice) CoQ10 deficiency model. Oral CoQ10 supplementation restored biochemical parameters and induced a significant improvement in clinical symptoms (p<0.001). These results lead to the hypothesis that inflammation could be a mitochondrial dysfunction-dependent event implicated in the pathophysiology of FM in several patients indicating at mitochondria as a possible new therapeutic target.
Asunto(s)
Fibromialgia/patología , Inflamación/patología , Mitocondrias/patología , Animales , Modelos Animales de Enfermedad , Femenino , Fibromialgia/metabolismo , Humanos , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/metabolismo , Leucocitos Mononucleares/patología , Masculino , Ratones , Mitocondrias/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factor de Necrosis Tumoral alfa/sangre , Ubiquinona/análogos & derivados , Ubiquinona/sangre , Ubiquinona/deficienciaRESUMEN
Melatonin is an indoleamine with a wide spectrum of biological activities other than transmitting photoperiod information, including antioxidant, oncostatic, anti-aging and immunomodulatory properties. Although melatonin is synthesized mainly in the pineal gland, other tissues have the same capacity. In the present study, we examined whether two key enzymes in melatonin biosynthesis, arylalkylamine Nacetyltransferase (AANAT) and hydroxyindole-O-methyltransferase (HIOMT) and its receptor MT1 are expressed in the two endocrine thyroid cells of the rat, follicular cells and C cells. Reverse transcriptase polymerase chain reaction analyses demonstrated that both AANAT and HIOMT mRNAs are expressed in the rat thyroid C-cells, and MT1 expression has been detected in C cells and follicular cells. Immunofluorescence revealed that AANAT protein is localized in C-cell cytoplasm, and MT1 protein in both cell populations. These findings demonstrate that the rat thyroid expresses AANAT, HIOMT, and its receptor MT1, showing that C cells are the main melatonin-synthesizing sites in the thyroid. This local C-cell-secreted melatonin may protect follicular cells from the oxidative stress inherent to the thyroid gland, and could also have paracrine and autocrine functions.