Garcia 2014 Abstract SECF
|Melatonin disrupts the NF-κB/NLRP3 connection in sepsis through RORα receptor.|
Overaggressive and excessive inflammatory response to noxious signals has been implicated in several acute and chronic inflammatory diseases, including sepsis and septic shock. Research efforts suggest that overstimulation of NF-κB, and subsequent high levels of NF-κB-dependent inflammatory mediators, causes a massive accumulation of ROS-generating mitochondria, whereas other data show that mitochondrial ROS induce NLRP3 inflammasome activation and IL-1β release. Thus, persistent activation and cooperation between NF-κB/NLRP3 signaling pathways amplifies and sustains a high-grade of inflammatory state. Melatonin has been proposed as pharmacologic support in inflammatory diseases due to its anti-inflammatory and antioxidant properties. Moreover, melatonin maintains mitochondrial homeostasis boosting its bioenergetic efficacy and reducing ROS production. Importantly, retinoid-related orphan receptor α (RORα) is not only a nuclear binding site of melatonin but also negatively regulates the inflammatory response. Therefore, this study was designed to analyze the NF-κB- and NLRP3-dependent molecular mechanisms involved in the development of sepsis, and to evaluate whether RORα is a key factor in anti-inflammatory actions of melatonin.
C57BL/6J and homozygous RORαsg/sg mice were grouped as follows: control group, septic groups, and septic mice treated with melatonin. Sepsis was induced by cecal ligation and puncture, and melatonin was given s.c. At 6, 8 or 24 hours after surgery or melatonin treatment, mice of both strains were sacrificed and heart was collected and used to evaluate the expression and level of key molecules involved in NF-κB and NLRP3 signaling pathway by qRT-PCR and Western-blot, respectively. Moreover, ELISA kits were used to determine the DNAbinding capacity NF-κB p65 subunit and total NAD+ levels. Finally, cytosolic oxidative stress was measured by spectrophotometric methods.
Parallel and transient activation of NF-κB and NLRP3 was observed in septic mice, leading to proinflammatory and prooxidant status during sepsis. At both, early and late septic stages, melatonin treatment modulated the nuclear translocation and DNA binding activity of NF-kB increasing the nuclear levels of Sirt-1 and its deacetylase activity. Therefore, melatonin decreased the level and expression of NF-κB-dependent proinflammatory cytokines and restored redox balance in septic mice. Moreover, melatonin inhibited NLRP3 inflammasome avoiding caspase-1 activation. Finally, we demonstrated that melatonin actions on NF-κB but not NLRP3 required a functional RORα transcription factor.
These results show by the first time the participation of the inflammasome NLRP3 in the developing of sepsis, and the molecular mechanisms underlying the anti-inflammatory activity of melatonin, revealing new molecular targets against sepsis. The results reinforce the utility of melatonin in the treatment of sepsis and other inflammatory disorders.
• Keywords: melatonin, NF-κB, NLRP3 inflammasome
• O2k-Network Lab: ES Granada Acuna-Castroviejo D
Labels: MiParea: Pharmacology;toxicology Pathology: Sepsis
Organism: Mouse Tissue;cell: Heart
1-Instituto de Biotecnología, Centro de Investigación Biomédica, Parque Tecnológico de Ciencias de la Salud, Universidad de Granada, Avda. Del Conocimiento s/n, 18100 Armilla, Granada, Spain. 2-Departamento de Fisiología, Facultad de Medicina,Universidad de Granada, Avenida de Madrid 11, 18012 Granada, Spain. 3-Unidad de Gestión Clínica de Laboratorios Clínicos, Hospital Universitario San Cecilio, Avenida Dr. Olóriz s/n, 18012 Granada, Spain.
Supported in part by grants # PI08-1664; P07-CTS-03135 and PI13-00981