Rottlerin Inhibits ROS Formation and Prevents NFκB Activation in MCF-7 and HT-29 Cells

Rottlerin, a polyphenol isolated from Mallotus Philippinensis, has been recently used as a selective inhibitor of PKC δ, although it can inhibit many kinases and has several biological effects. Among them, we recently found that Rottlerin inhibits the Nuclear Factor κB (NFκB), activated by either phorbol esters or H2O2. Because of the redox sensitivity of NFκB and on the basis of Rottlerin antioxidant property, we hypothesized that Rottlerin could prevent NFκB activation acting as a free radicals scavenger, as other natural polyphenols. The current study confirms the antioxidant property of Rottlerin against the 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) in vitro and against oxidative stress induced by H2O2 and by menadione in culture cells. We also demonstrate that Rottlerin prevents TNFα-dependent NFκB activation in MCF-7 cells and in HT-29 cells transfected with the NFκB-driven plasmid pBIIX-LUC, suggesting that Rottlerin can inhibit NFκB via several pathways and in several cell types

Comparison of pollutants effect on cutaneous inflammasomes activation

The skin is the outermost layer of the body and, therefore, is exposed to a variety of stressors, such as environmental pollutants, known to cause oxinflammatory reactions involved in the exacerbation of several skin conditions. Today, inflammasomes are recognized as important modulators of the cutaneous inflammatory status in response to air pollutants and ultraviolet (UV) light exposure. In this study, human skin explants were exposed to the best-recognized air pollutants, such as microplastics (MP), cigarette smoke (CS), diesel engine exhaust (DEE), ozone (O3), and UV, for 1 or 4 days, to explore how each pollutant can differently modulate markers of cutaneous oxinflammation. Exposure to environmental pollutants caused an altered oxidative stress response, accompanied by increased DNA damage and signs of premature skin aging. The effect of specific pollutants being able to exert different inflammasomes pathways (NLRP1, NLRP3, NLRP6, and NLRC4) was also investigated in terms of scaffold formation and cell pyroptosis. Among all environmental pollutants, O3, MP, and UV represented the main pollutants affecting cutaneous redox homeostasis; of note, the NLRP1 and NLRP6 inflammasomes were the main ones modulated by these outdoor stressors, suggesting their role as possible molecular targets in preventing skin disorders and the inflammaging events associated with environmental pollutant exposure

Ubiquitination as a key regulatory mechanism for O3-induced cutaneous redox inflammasome activation

NLRP1 is one of the major inflammasomes modulating the cutaneous inflammatory responses and therefore linked to a variety of cutaneous conditions. Although NLRP1 has been the first inflammasome to be discovered, only in the past years a significant progress was achieved in understanding the molecular mechanism and the stimuli behind its activation. In the past decades a crescent number of studies have highlighted the role of air pollutants as Particulate Matter (PM), Cigarette Smoke (CS) and Ozone (O3) as trigger stimuli for inflammasomes activation, especially via Reactive Oxygen Species (ROS) mediators. However, whether NLRP1 can be modulated by air pollutants via oxidative stress and the mechanism behind its activation is still poorly understood. Here we report for the first time that O3, one of the most toxic pollutants, activates the NLRP1 inflammasome in human keratinocytes via oxidative stress mediators as hydrogen peroxide (H2O2) and 4-hydroxy-nonenal (4HNE). Our data suggest that NLRP1 represents a target protein for 4HNE adduction that possibly leads to its proteasomal degradation and activation via the possible involvement of E3 ubiquitin ligase UBR2. Of note, Catalase (Cat) treatment prevented inflammasome assemble and inflammatory cytokines release as well as NLRP1 ubiquitination in human keratinocytes upon O3 exposure. The present work is a mechanistic study that follows our previous work where we have showed the ability of O3 to induce cutaneous inflammasome activation in humans exposed to this pollutant. In conclusion, our results suggest that O3 triggers the cutaneous NLRP1 inflammasome activation by ubiquitination and redox mechanism

Cigarette Smoke Affects ABCAl Expression via Liver X Receptor Nuclear Translocation in Human Keratinocytes

Cutaneous tissue is the first barrier against outdoor insults. The outer most layer of the skin, the stratum corneum (SC), is formed by corneocytes embedded in a lipid matrix (cholesterol, ceramide and fatty acids). Therefore, the regulation of lipids and, in particular, of cholesterol homeostasis in the skin is of great importance. ABCA1 is a membrane transporter responsible for cholesterol efflux and plays a key role in maintaining cellular cholesterol levels. Among the many factors that have been associated with skin diseases, the environmental stressor cigarette smoke has been recently studied. In the present study, we demonstrate that ABCA1 expression in human cells (HaCaT) was increased (both mRNA and protein levels) after CS exposure. This effect was mediated by the inhibition of NFkB (aldehydes adducts formation) that allows the translocation of liver X receptor (LXR). These findings suggest that passive smoking may play a role in skin cholesterol levels and thus affect cutaneous tissues functions

Review Article Isoprostanes and 4-Hydroxy-2-nonenal: Markers or Mediators of Disease? Focus on Rett Syndrome as a Model of Autism Spectrum Disorder

Lipid peroxidation, a process known to induce oxidative damage to key cellular components, has been implicated in several diseases. Following three decades of explorations mainly on in vitro models reproducible in the laboratories, lipid peroxidation has become increasingly relevant for the interpretation of a wide range of pathophysiological mechanisms in the clinical setting. This cumulative effort has led to the identification of several lipid peroxidation end-products meeting the needs of the in vivo evaluation. Among these different molecules, isoprostanes and 4-hydroxy-2-nonenal protein adducts appear to be particularly interesting. This review shows how specific oxidation products, deriving from polyunsaturated fatty acids precursors, are strictly related to the clinical manifestations and the natural history of Rett syndrome, a genetically determined neurodevelopmental pathology, currently classified among the autism spectrum disorders. In our experience, Rett syndrome offers a unique setting for physicians, biologists, and chemists to explore the borders of the lipid mediators concept

Redox imbalance is associated to lung damage triggered by silver nanoparticles exposure

Along with the AgNP applications development, the concern about their possible toxicity has increasingly gained attention. As the respiratory system is one of the main exposure routes, the aim of this study was to evaluate the harmful effects developed in the lung after an acute AgNP exposure. In vivo studies using Balb/c mice intranasally instilled with 0.1 mg AgNP/kg b.w, were performed. 99mTc-AgNP showed the lung as the main organ of deposition, where, in turn, AgNP may exert barrier injury observed by increased protein content and total cell count in BAL samples. In vivo acute exposure showed altered lung tissue O2 consumption due to increased mitochondrial active respiration and NOX activity. Both O2 consumption processes release ROS triggering the antioxidant system as observed by the increased SOD, catalase and GPx activities and a decreased GSH/GSSG ratio. In addition, increased protein oxidation was observed after AgNP exposure. In A549 cells, exposure to 2.5 μg/mL AgNP during 1 h resulted in augment NOX activity, decreased mitochondrial ATP associated respiration and higher H2O2 production rate. Lung 3D tissue model showed AgNP-initiated barrier alterations as TEER values decreased and morphological alterations. Taken together, these results show that AgNP exposure alters O2 metabolism leading to alterations in oxygen metabolism lung toxicity. AgNP-triggered oxidative damage may be responsible for the impaired lung function observed due to alveolar epithelial injury.Fil: Garces, Mariana Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Magnani, Natalia Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Pecorelli, Alessandra. Università di Ferrara; ItaliaFil: Calabró López, María Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Marchini, Timoteo Oscar. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Caceres, Lourdes Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Pambianchi, Erika. Università di Ferrara; ItaliaFil: Galdopórpora, Juan Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Metabolismo del Fármaco; ArgentinaFil: Vico, Tamara Antonela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Salgueiro, María Jimena. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Zubillaga, Marcela Beatriz. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Moretton, Marcela Analía. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Desimone, Martín Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Metabolismo del Fármaco; ArgentinaFil: Alvarez, Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Valacchi, Giuseppe. Università di Ferrara; ItaliaFil: Evelson, Pablo Andrés. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; Argentin

Defective proteasome biogenesis into skin fibroblasts isolated from Rett syndrome subjects with {MeCP}2 non-sense mutations

Rett Syndrome (RTT) is a rare X-linked neurodevelopmental disorder which affects about 1: 10000 live births. In >95% of subjects RTT is caused by a mutation in Methyl-CpG binding protein-2 (MECP2) gene, which encodes for a transcription regulator with pleiotropic genetic/epigenetic activities. The molecular mechanisms underscoring the phenotypic alteration of RTT are largely unknown and this has impaired the development of therapeutic approaches to alleviate signs and symptoms during disease progression. A defective proteasome biogenesis into two skin primary fibroblasts isolated from RTT subjects harbouring non-sense (early-truncating) MeCP2 mutations (i.e., R190fs and R255X) is herewith reported. Proteasome is the proteolytic machinery of Ubiquitin Proteasome System (UPS), a pathway of overwhelming relevance for post-mitotic cells metabolism. Molecular, transcription and proteomic analyses indicate that MeCP2 mutations down-regulate the expression of one proteasome subunit, α7, and of two chaperones, PAC1 and PAC2, which bind each other in the earliest step of proteasome biogenesis. Furthermore, this molecular alteration recapitulates in neuron-like SH-SY5Y cells upon silencing of MeCP2 expression, envisaging a general significance of this transcription regulator in proteasome biogenesis