Toll-like Receptor 4 Signaling is Critical for the Adaptive Cellular Stress Response Effects Induced by Intermittent Fasting in the Mouse Brain.

Among different kinds of dietary energy restriction, intermittent fasting (IF) has been considered a dietary regimen which causes a mild stress to the organism. IF can stimulate proteins and signaling pathways related to cell stress that can culminate in the increase of the body resistance to severe stress conditions. Energy intake reduction induced by IF can induce modulation of receptors, kinases, and phosphatases, which in turn can modulate the activation of transcription factors such as NF-E2-related factor 2 (NRF2) and cAMP response element-binding (CREB) which regulate the transcription of genes related to the translation of proteins such as growth factors: brain-derived neurotrophic factor (BDNF), chaperone proteins: heat shock proteins (HSP), and so on. It has been shown that toll-like receptors (TLRs) are important molecules in innate immune response which are present not only in the periphery but also in neurons and glial cells. In central nervous system, TLRs can exert functions related to set up responses to infection, as well as influence neural progenitor cell proliferation and differentiation, being involved in cognitive parameters such as learning and memory. Little is known about the involvement of TLR4 on the beneficial effects induced by IF protocol. The present work investigated the effects of IF on memory and on the signaling mechanisms associated with NRF2 and CREB in Tlr4 knockout mice. The results suggest that TLR4 participates in the modulatory effects of IF on oxidative stress levels, on the transcription factors CREB and NRF2, and on BDNF and HSP90 expressions in hippocampus.

Alpha 2 Na+,K+-ATPase silencing induces loss of inflammatory response and ouabain protection in glial cells.

Ouabain (OUA) is a cardiac glycoside that binds to Na+,K+-ATPase (NKA), a conserved membrane protein that controls cell transmembrane ionic concentrations and requires ATP hydrolysis. At nM concentrations, OUA activates signaling pathways that are not related to its typical inhibitory effect on the NKA pump. Activation of these signaling pathways protects against some types of injury of the kidneys and central nervous system. There are 4 isoforms of the alpha subunit of NKA, which are differentially distributed across tissues and may have different physiological roles. Glial cells are important regulators of injury and inflammation in the brain and express the α1 and α2 NKA isoforms. This study investigated the role of α2 NKA in OUA modulation of the neuroinflammatory response induced by lipopolysaccharide (LPS) in mouse primary glial cell cultures. LPS treatment increased lactate dehydrogenase release, while OUA did not decrease cell viability and blocked LPS-induced NF-κB activation. Silencing α2 NKA prevented ERK and NF-κB activation by LPS. α2 NKA also regulates TNF-α and IL-1ß levels. The data reported here indicate a significant role of α2 NKA in regulating central LPS effects, with implications in the associated neuroinflammatory processes.

Exercise training decreases NADPH oxidase activity and restores skeletal muscle mass in heart failure rats.

We have recently demonstrated that NADPH oxidase hyperactivity, NF-κB activation, and increased p38 phosphorylation lead to atrophy of glycolytic muscle in heart failure (HF). Aerobic exercise training (AET) is an efficient strategy to counteract skeletal muscle atrophy in this syndrome. Therefore, we tested whether AET would regulate muscle redox balance and protein degradation by decreasing NADPH oxidase hyperactivity and reestablishing NF-κB signaling, p38 phosphorylation, and proteasome activity in plantaris muscle of myocardial infarcted-induced HF (MI) rats. Thirty-two male Wistar rats underwent MI or fictitious surgery (SHAM) and were randomly assigned into untrained (UNT) and trained (T; 8 wk of AET on treadmill) groups. AET prevented HF signals and skeletal muscle atrophy in MI-T, which showed an improved exercise tolerance, attenuated cardiac dysfunction and increased plantaris fiber cross-sectional area. To verify the role of inflammation and redox imbalance in triggering protein degradation, circulating TNF-α levels, NADPH oxidase profile, NF-κB signaling, p38 protein levels, and proteasome activity were assessed. MI-T showed a reduced TNF-α levels, NADPH oxidase activity, and Nox2 mRNA expression toward SHAM-UNT levels. The rescue of NADPH oxidase activity induced by AET in MI rats was paralleled by reducing nuclear binding activity of the NF-κB, p38 phosphorylation, atrogin-1, mRNA levels, and 26S chymotrypsin-like proteasome activity. Taken together our data provide evidence for AET improving plantaris redox homeostasis in HF associated with a decreased NADPH oxidase, redox-sensitive proteins activation, and proteasome hyperactivity further preventing atrophy. These data reinforce the role of AET as an efficient therapy for muscle wasting in HF.NEW & NOTEWORTHY This study demonstrates, for the first time, the contribution of aerobic exercise training (AET) in decreasing muscle NADPH oxidase activity associated with reduced reactive oxygen species production and systemic inflammation, which diminish NF-κB overactivation, p38 phosphorylation, and ubiquitin proteasome system hyperactivity. These molecular changes counteract plantaris atrophy in trained myocardial infarction-induced heart failure rats. Our data provide new evidence into how AET may regulate protein degradation and thus prevent skeletal muscle atrophy.

The Influence of Na(+), K(+)-ATPase on Glutamate Signaling in Neurodegenerative Diseases and Senescence.

Decreased Na(+), K(+)-ATPase (NKA) activity causes energy deficiency, which is commonly observed in neurodegenerative diseases. The NKA is constituted of three subunits: α, ß, and γ, with four distinct isoforms of the catalytic α subunit (α1-4). Genetic mutations in the ATP1A2 gene and ATP1A3 gene, encoding the α2 and α3 subunit isoforms, respectively can cause distinct neurological disorders, concurrent to impaired NKA activity. Within the central nervous system (CNS), the α2 isoform is expressed mostly in glial cells and the α3 isoform is neuron-specific. Mutations in ATP1A2 gene can result in familial hemiplegic migraine (FHM2), while mutations in the ATP1A3 gene can cause Rapid-onset dystonia-Parkinsonism (RDP) and alternating hemiplegia of childhood (AHC), as well as the cerebellar ataxia, areflexia, pescavus, optic atrophy and sensorineural hearing loss (CAPOS) syndrome. Data indicates that the central glutamatergic system is affected by mutations in the α2 isoform, however further investigations are required to establish a connection to mutations in the α3 isoform, especially given the diagnostic confusion and overlap with glutamate transporter disease. The age-related decline in brain α2∕3 activity may arise from changes in the cyclic guanosine monophosphate (cGMP) and cGMP-dependent protein kinase (PKG) pathway. Glutamate, through nitric oxide synthase (NOS), cGMP and PKG, stimulates brain α2∕3 activity, with the glutamatergic N-methyl-D-aspartate (NMDA) receptor cascade able to drive an adaptive, neuroprotective response to inflammatory and challenging stimuli, including amyloid-ß. Here we review the NKA, both as an ion pump as well as a receptor that interacts with NMDA, including the role of NKA subunits mutations. Failure of the NKA-associated adaptive response mechanisms may render neurons more susceptible to degeneration over the course of aging.

Suppression of MAPK attenuates neuronal cell death induced by activated glia-conditioned medium in alpha-synuclein overexpressing SH-SY5Y cells.

BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disease with characteristics and symptoms that are well defined. Nevertheless, its aetiology remains unknown. PD is characterized by the presence of Lewy bodies inside neurons. α-Synuclein (α-syn) is a soluble protein present in the pre-synaptic terminal of neurons. Evidence suggests that α-syn has a fundamental role in PD pathogenesis, given that it is an important component of Lewy bodies localized in the dopaminergic neurons of PD patients. METHODS: In the present study, we investigated the influence of wild type (WT) and A30P α-syn overexpression on neuroblastoma SH-SY5Y toxicity induced by the conditioned medium (CM) from primary cultures of glia challenged with lipopolysaccharide (LPS) from Escherichia coli. RESULTS: We observed that SH-SY5Y cells transduced with α-syn (WT or A30P) and treated with CM from LPS-activated glia cells show evidence of cell death, which is not reverted by NF-κB inhibition by sodium salicylate or by blockage of P50 (NF-κB subunit). Furthermore, the expression of A30P α-syn in neuroblastoma SH-SY5Y decreases the cell death triggered by the CM of activated glia versus WT α-syn or control group. This effect of A30P α-syn may be due to the low MAPK42/44 phosphorylation. This finding is substantiated by MEK1 inhibition by PD98059, decreasing LDH release by CM in SH-SY5Y cells. CONCLUSION: Our results suggest that SH-SY5Y cells transduced with α-syn (WT or A30P) and treated with CM from LPS-activated glia cells show cell death, which is not reverted by NF-κB blockage. Additionally, the expression of A30P α-syn on neuroblastoma SH-SY5Y leads to decreased cell death triggered by the CM of activated glia, when compared to WT α-syn or control group. The mechanism underlying this process remains to be completely elucidated, but the present data suggest that MAPK42/44 phosphorylation plays an important role in this process. PROSPERO: CRD42015020829.

NADPH oxidase hyperactivity induces plantaris atrophy in heart failure rats.

BACKGROUND: Skeletal muscle wasting is associated with poor prognosis and increased mortality in heart failure (HF) patients. Glycolytic muscles are more susceptible to catabolic wasting than oxidative ones. This is particularly important in HF since glycolytic muscle wasting is associated with increased levels of reactive oxygen species (ROS). However, the main ROS sources involved in muscle redox imbalance in HF have not been characterized. Therefore, we hypothesized that NADPH oxidases would be hyperactivated in the plantaris muscle of infarcted rats, contributing to oxidative stress and hyperactivation of the ubiquitin-proteasome system (UPS), ultimately leading to atrophy. METHODS: Rats were submitted to myocardial infarction (MI) or Sham surgery. Four weeks after surgery, MI and Sham groups underwent eight weeks of treatment with apocynin, a NADPH oxidase inhibitor, or placebo. NADPH oxidase activity, oxidative stress markers, NF-κB activity, p38 MAPK phosphorylation, mRNA and sarcolemmal protein levels of NADPH oxidase components, UPS activation and fiber cross-sectional area were assessed in the plantaris muscle. RESULTS: The plantaris of MI rats displayed atrophy associated with increased Nox2 mRNA and sarcolemmal protein levels, NADPH oxidase activity, ROS production, lipid hydroperoxides levels, NF-κB activity, p38 MAPK phosphorylation and UPS activation. NADPH oxidase inhibition by apocynin prevented MI-induced skeletal muscle atrophy by reducing ROS production, NF-κB hyperactivation, p38 MAPK phosphorylation and proteasomal hyperactivity. CONCLUSION: Our data provide evidence for NADPH oxidase hyperactivation as an important source of ROS production leading to plantaris atrophy in heart failure rats, suggesting that this enzyme complex plays key role in skeletal muscle wasting in HF.