RESUMEN
The overall metabolic/energetic stress that occurs during an acute bout of exercise is proposed to be the main driving force for long-term training adaptations. Continuous and high-intensity interval exercise protocols (HIIE) are currently prescribed to acquire the muscular and metabolic benefits of aerobic training. We applied 1H NMR-based metabonomics to compare the overall metabolic perturbation and activation of individual bioenergetic pathways of three popular aerobic exercises matched for effort/strain. Nine men performed continuous, long-interval (3 min), and short-interval (30 s) bouts of exercise under isoeffort conditions. Blood was collected before and after exercise. The multivariate PCA and OPLS-DA models showed a distinct separation of pre- and postexercise samples in three protocols. The two models did not discriminate the postexercise overall metabolic profiles of the three exercise types. Analysis focused on muscle bioenergetic pathways revealed an extensive upregulation of carbohydrate-lipid metabolism and the TCA cycle in all three protocols; there were only a few differences among protocols in the postexercise abundance of molecules when long-interval bouts were performed. In conclusion, continuous and HIIE exercise protocols, when performed with similar effort/strain, induce comparable global metabolic response/stress despite their marked differences in work-bout intensities. This study highlights the importance of NMR metabonomics in comprehensive monitoring of metabolic consequences of exercise training in the blood of athletes and exercising individuals.
Asunto(s)
Ejercicio Físico , Metabolómica , Estrés Fisiológico , Adulto , Sangre/metabolismo , Ciclo del Ácido Cítrico/genética , Metabolismo Energético/genética , Humanos , Metabolismo de los Lípidos/genética , Espectroscopía de Resonancia Magnética , Masculino , Análisis Multivariante , Factores de Tiempo , Regulación hacia Arriba , Adulto JovenRESUMEN
Spinal cord injury (SCI) results in permanent loss of motor functions. A significant aspect of the tissue damage and functional loss may be preventable as it occurs, secondary to the trauma. We show that the phospholipase A(2) (PLA(2)) superfamily plays important roles in SCI. PLA(2) enzymes hydrolyze membrane glycerophospholipids to yield a free fatty acid and lysophospholipid. Some free fatty acids (arachidonic acid) give rise to eicosanoids that promote inflammation, while some lysophospholipids (lysophosphatidylcholine) cause demyelination. We show in a mouse model of SCI that two cytosolic forms [calcium-dependent PLA(2) group IVA (cPLA(2) GIVA) and calcium-independent PLA(2) group VIA (iPLA(2) GVIA)], and a secreted form [secreted PLA(2) group IIA (sPLA(2) GIIA)] are up-regulated. Using selective inhibitors and null mice, we show that these PLA(2)s play differing roles. cPLA(2) GIVA mediates protection, whereas sPLA(2) GIIA and, to a lesser extent, iPLA(2) GVIA are detrimental. Furthermore, completely blocking all three PLA(2)s worsens outcome, while the most beneficial effects are seen by partial inhibition of all three. The partial inhibitor enhances expression of cPLA(2) and mediates its beneficial effects via the prostaglandin EP1 receptor. These findings indicate that drugs that inhibit detrimental forms of PLA(2) (sPLA(2) and iPLA2) and up-regulate the protective form (cPLA2) may be useful for the treatment of SCI.
Asunto(s)
Fosfolipasas A2/metabolismo , Traumatismos de la Médula Espinal/enzimología , Animales , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/uso terapéutico , Femenino , Fosfolipasas A2 Grupo II/antagonistas & inhibidores , Fosfolipasas A2 Grupo II/deficiencia , Fosfolipasas A2 Grupo II/metabolismo , Fosfolipasas A2 Grupo IV/antagonistas & inhibidores , Fosfolipasas A2 Grupo IV/deficiencia , Fosfolipasas A2 Grupo IV/genética , Fosfolipasas A2 Grupo IV/metabolismo , Fosfolipasas A2 Grupo VI/antagonistas & inhibidores , Fosfolipasas A2 Grupo VI/deficiencia , Fosfolipasas A2 Grupo VI/metabolismo , Locomoción/efectos de los fármacos , Locomoción/fisiología , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Inhibidores de Fosfolipasa A2 , Fosfolipasas A2/clasificación , Fosfolipasas A2/deficiencia , Receptor Cross-Talk , Subtipo EP1 de Receptores de Prostaglandina E/metabolismo , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/patología , Traumatismos de la Médula Espinal/fisiopatologíaRESUMEN
The phospholipase A(2) (PLA(2)) superfamily hydrolyzes phospholipids to release free fatty acids and lysophospholipids, some of which can mediate inflammation and demyelination, hallmarks of the CNS autoimmune disease multiple sclerosis. The expression of two of the intracellular PLA(2)s (cPLA(2) GIVA and iPLA(2) GVIA) and two of the secreted PLA(2)s (sPLA(2) GIIA and sPLA(2) GV) are increased in different stages of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. We show using small molecule inhibitors, that cPLA(2) GIVA plays a role in the onset, and iPLA(2) GVIA in the onset and progression of EAE. We also show a potential role for sPLA(2) in the later remission phase. These studies demonstrate that selective inhibition of iPLA(2) can ameliorate disease progression when treatment is started before or after the onset of symptoms. The effects of these inhibitors on lesion burden, chemokine and cytokine expression as well as on the lipid profile provide insights into their potential modes of action. iPLA(2) is also expressed by macrophages and other immune cells in multiple sclerosis lesions. Our results therefore suggest that iPLA(2) might be an excellent target to block for the treatment of CNS autoimmune diseases, such as multiple sclerosis.
Asunto(s)
Encefalomielitis Autoinmune Experimental/enzimología , Fosfolipasas A2/fisiología , Isoformas de Proteínas/fisiología , Médula Espinal/enzimología , Adulto , Amidas/uso terapéutico , Animales , Citocinas/inmunología , Progresión de la Enfermedad , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/genética , Inhibidores Enzimáticos/uso terapéutico , Ácidos Grasos/análisis , Femenino , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Fluorocarburos , Expresión Génica , Humanos , Inmunohistoquímica , Cetonas/uso terapéutico , Macrófagos/enzimología , Masculino , Ratones , Ratones Endogámicos C57BL , Esclerosis Múltiple/enzimología , Esclerosis Múltiple/genética , Inhibidores de Fosfolipasa A2 , Fosfolipasas A2/química , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/química , ARN Mensajero/análisis , Linfocitos T/enzimología , Adulto JovenRESUMEN
We provide evidence that two members of the intracellular phospholipase A(2) family, namely calcium-dependent group IVA (cPLA(2) GIVA) and calcium-independent group VIA (iPLA(2) GVIA) may play important roles in Wallerian degeneration in the mouse sciatic nerve. We assessed the roles of these PLA(2)s in cPLA(2) GIVA(-/-) mice, and mice treated with a selective inhibitor of iPLA(2) GVIA (FKGK11). Additionally, the effects of both these PLA(2)s were assessed by treating cPLA(2) GIVA(-/-) mice with the iPLA(2) inhibitor. Our data suggest that iPLA(2) GVIA may play more of a role in the early stages of myelin breakdown, while cPLA(2) GIVA may play a greater role in myelin clearance by macrophages. Our results also show that the delayed myelin clearance and Wallerian degeneration after sciatic nerve crush injury in mice lacking cPLA(2) and iPLA(2) activities is accompanied by a delay in axon regeneration, target re-innervation and functional recovery. These results indicate that the intracellular PLA(2)s (cPLA(2) GIVA and iPLA(2) GVIA) contribute significantly to various aspects of Wallerian degeneration in injured peripheral nerves, which is then essential for successful axon regeneration. This work has implications for injury responses and recovery after peripheral nerve injuries in humans, as well as for understanding the slow clearance of myelin after CNS injury and its potential consequences for axon regeneration.
Asunto(s)
Axones/fisiología , Fosfolipasas A2 Grupo IV/metabolismo , Fosfolipasas A2 Grupo VI/metabolismo , Regeneración Nerviosa/fisiología , Degeneración Walleriana/enzimología , Animales , Axones/enzimología , Técnica del Anticuerpo Fluorescente , Fluorocarburos , Fosfolipasas A2 Grupo IV/genética , Fosfolipasas A2 Grupo VI/antagonistas & inhibidores , Fosfolipasas A2 Grupo VI/genética , Cetonas/farmacología , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Electrónica , Modelos Animales , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Nervio Ciático/enzimología , Nervio Ciático/lesiones , Nervio Ciático/patología , Piel/inervación , Degeneración Walleriana/patologíaRESUMEN
The addition of (pentafluoroethyl)- and (heptafluoropropyl)lithium to various acyl derivatives was studied. Weinreb and morpholine amides led to polyfluoro ketones in high to quantitative yields in short reaction times. Derivatives of 2-fluorocarboxylic acids may produce 1,1,1,2,2,4-hexafluoro ketones and 1,1,1,2,2,3,3,5-octafluoro ketones. The methodology can provide inhibitors for various lipolytic enzymes, including phospholipase A2.
Asunto(s)
Fluorocarburos/síntesis química , Cetonas/síntesis química , Compuestos de Litio/química , Inhibidores Enzimáticos/síntesis química , Fluorocarburos/química , Indicadores y Reactivos , Métodos , Fosfolipasas/antagonistas & inhibidoresRESUMEN
Laetiporus sulphureus is an edible wood-rooting basidiomycete. The nutritional and medicinal properties of this mushroom have long been known by traditional practitioners. The aim of this study was to determine the proximate composition, total phenol antioxidant capacity and antimicrobial activities of different extracts of L. sulphureus. Different extraction methodologies, including high energy techniques, were employed and their effect was examined on the activity of the extracts. Optimum extraction methodologies (classical and ultrasound-assisted) provided one fraction containing neutral and polar lipids and the other fraction containing fungal carotenoids and pigments. Fatty acid analysis indicated a predominant level of polyunsaturated fatty acids followed by saturated and mono-unsaturated fatty acids. Both the aqueous methanolic and water extracts contained higher TPC and showed better antioxidant capacity than the ethanolic extract. Irrespective of the type of extraction applied, L. sulphureus showed good antimicrobial activity against all the tested bacteria and fungi, being in some cases stronger than the used antibiotics and mycotics. Therefore, this edible mushroom could be considered as a positive candidate to be utilised by the food industry, not only for obtaining bioactive compounds to be used as natural antioxidants/antimicrobial agents, but possibly also for its nutritional value and health benefits.
Asunto(s)
Agaricales/química , Coriolaceae/química , Antiinfecciosos/análisis , Antiinfecciosos/farmacología , Antioxidantes/análisis , Antioxidantes/farmacología , Productos Biológicos/análisis , Carotenoides/análisis , Carotenoides/farmacología , Cromatografía Liquida , Bacterias Gramnegativas/efectos de los fármacos , Bacterias Gramnegativas/aislamiento & purificación , Concentración 50 Inhibidora , Espectrometría de Masas , Valor Nutritivo , Fenoles/análisis , Fenoles/farmacologíaRESUMEN
Sodium potassium pump (Na(+)/K(+)ATPase) is a validated pharmacological target for the treatment of congestive heart failure. Recent data with inotropic drugs such as digoxin & digitoxin (digitalis) suggest a potent anti-cancer action of these drugs and promote Na(+)/K(+)ATPase as a novel therapeutic target in cancer. However, digitalis have narrow therapeutic indices, are pro-arrhythmic and are considered non-developable drugs by the pharmaceutical industry. On the contrary, a series of recently-developed steroidal inhibitors showed better pharmacological properties and clinical activities in cardiac patients. Their anti-cancer activity however, remained unknown. In this study, we synthesized seventeen steroidal cardiac inhibitors and explored for the first time their anti-cancer activity in vitro and in vivo. Our results indicate potent anti-cancer actions of steroidal cardiac inhibitors in multiple cell lines from different tumor panels including multi-drug resistant cells. Furthermore, the most potent compound identified in our studies, the 3-[(R)-3- pyrrolidinyl]oxime derivative 3, showed outstanding potencies (as measured by GI50, TGI and LC50 values) in most cells in vitro, was selectively cytotoxic in cancer versus normal cells showing a therapeutic index of 31.7 and exhibited significant tumor growth inhibition in prostate and lung xenografts in vivo. Collectively, our results suggest that previously described cardiac Na(+)/K(+)ATPase inhibitors have potent anti-cancer actions and may thus constitute strong re-purposing candidates for further cancer drug development.
Asunto(s)
Androstenos/farmacología , Antineoplásicos/farmacología , Neoplasias Pulmonares/tratamiento farmacológico , Miocardio/enzimología , Oximas/farmacología , Pirrolidinas/farmacología , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Esteroides/farmacología , Androstenos/uso terapéutico , Animales , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Femenino , Xenoinjertos , Humanos , Masculino , Ratones SCID , Trasplante de Neoplasias , Oximas/química , Oximas/uso terapéutico , Neoplasias de la Próstata/tratamiento farmacológico , Pirrolidinas/química , Pirrolidinas/uso terapéutico , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Esteroides/química , Esteroides/uso terapéuticoRESUMEN
Group VIA calcium-independent phospholipase A(2) (GVIA iPLA(2)) has recently emerged as a novel pharmaceutical target. We have now explored the structure-activity relationship between fluoroketones and GVIA iPLA(2) inhibition. The presence of a naphthyl group proved to be of paramount importance. 1,1,1-Trifluoro-6-(naphthalen-2-yl)hexan-2-one (FKGK18) is the most potent inhibitor of GVIA iPLA(2) (X(I)(50) = 0.0002) ever reported. Being 195 and >455 times more potent for GVIA iPLA(2) than for GIVA cPLA(2) and GV sPLA(2), respectively, makes it a valuable tool to explore the role of GVIA iPLA(2) in cells and in vivo models. 1,1,1,2,2,3,3-Heptafluoro-8-(naphthalene-2-yl)octan-4-one inhibited GVIA iPLA(2) with a X(I)(50) value of 0.001 while inhibiting the other intracellular GIVA cPLA(2) and GV sPLA(2) at least 90 times less potently. Hexa- and octafluoro ketones were also found to be potent inhibitors of GVIA iPLA(2); however, they are not selective.
Asunto(s)
Flúor , Fosfolipasas A2 Grupo VI/antagonistas & inhibidores , Cetonas/farmacología , Naftalenos/farmacología , Línea Celular , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/química , Humanos , Cetonas/uso terapéutico , Naftalenos/uso terapéutico , Fosfolipasas A2 Calcio-Independiente/antagonistas & inhibidores , Relación Estructura-ActividadRESUMEN
The development of selective inhibitors for individual PLA(2) enzymes is necessary in order to target PLA(2)-specific signaling pathways, but it is challenging due to the observed promiscuity of known PLA(2) inhibitors. In the current work, we present the development and application of a variety of synthetic routes to produce pentafluoro, tetrafluoro, and trifluoro derivatives of activated carbonyl groups in order to screen for selective inhibitors and characterize the chemical properties that can lead to selective inhibition. Our results demonstrate that the pentafluoroethyl ketone functionality favors selective inhibition of the GVIA iPLA(2), a very important enzyme for which specific, potent, reversible inhibitors are needed. We find that 1,1,1,2,2-pentafluoro-7-phenyl-heptan-3-one (FKGK11) is a selective inhibitor of GVIA iPLA(2) (X(I)(50) = 0.0073). Furthermore, we conclude that the introduction of an additional fluorine atom at the alpha' position of a trifluoromethyl ketone constitutes an important strategy for the development of new potent GVIA iPLA(2) inhibitors.