Susceptibility to acute cognitive dysfunction in aged mice is underpinned by reduced white matter integrity and microgliosis.

Age is a significant but heterogeneous risk factor for acute neuropsychiatric disturbances such as delirium. Neuroinflammation increases with aging but the determinants of underlying risk for acute dysfunction upon systemic inflammation are not clear. We hypothesised that, with advancing age, mice would become progressively more vulnerable to acute cognitive dysfunction and that neuroinflammation and neuronal integrity might predict heterogeneity in such vulnerability. Here we show region-dependent differential expression of microglial transcripts, but a ubiquitously observed primed signature: chronic Clec7a expression and exaggerated Il1b responses to systemic bacterial LPS. Cognitive frailty (vulnerability to acute disruption under acute stressors LPS and double stranded RNA; poly I:C) was increased in aged animals but showed heterogeneity and was significantly correlated with reduced myelin density, synaptic loss and severity of white matter microgliosis. The data indicate that white matter disruption and neuroinflammation may be key substrates of the progressive but heterogeneous risk for delirium in aged individuals.

Hyperthermia elevates brain temperature and improves behavioural signs in animal models of autism spectrum disorder.

BACKGROUND: Autism spectrum disorders (ASD) are predominantly neurodevelopmental and largely genetically determined. However, there are human data supporting the idea that fever can improve symptoms in some individuals, but those data are limited and there are almost no data to support this from animal models. We aimed to test the hypothesis that elevated body temperature would improve function in two animal models of ASD. METHODS: We used a 4 h whole-body hyperthermia (WBH) protocol and, separately, systemic inflammation induced by bacterial endotoxin (LPS) at 250 µg/kg, to dissociate temperature and inflammatory elements of fever in two ASD animal models: C58/J and Shank3B- mice. We used one- or two-way ANOVA and t-tests with normally distributed data and Kruskal-Wallis or Mann-Whitney with nonparametric data. Post hoc comparisons were made with a level of significance set at p < 0.05. For correlation analyses, data were adjusted by a linear regression model. RESULTS: Only LPS induced inflammatory signatures in the brain while only WBH produced fever-range hyperthermia. WBH reduced repetitive behaviours and improved social interaction in C58/J mice and significantly reduced compulsive grooming in Shank3B- mice. LPS significantly suppressed most activities over 5-48 h. LIMITATIONS: We show behavioural, cellular and molecular changes, but provide no specific mechanistic explanation for the observed behavioural improvements. CONCLUSIONS: The data are the first, to our knowledge, to demonstrate that elevated body temperature can improve behavioural signs in 2 distinct ASD models. Given the developmental nature of ASD, evidence that symptoms may be improved by environmental perturbations indicates possibilities for improving function in these individuals. Since experimental hyperthermia in patients would carry significant risks, it is now essential to pursue molecular mechanisms through which hyperthermia might bring about the observed benefits.

Redox Regulation of Microglial Inflammatory Response: Fine Control of NLRP3 Inflammasome through Nrf2 and NOX4.

The role of inflammation and immunity in the pathomechanism of neurodegenerative diseases has become increasingly relevant within the past few years. In this context, the NOD-like receptor protein 3 (NLRP3) inflammasome plays a crucial role in the activation of inflammatory responses by promoting the maturation and secretion of pro-inflammatory cytokines such as interleukin-1ß and interleukin-18. We hypothesized that the interplay between nuclear factor erythroid 2-related factor 2 (Nrf2) and NADPH oxidase 4 (NOX4) may play a critical role in the activation of the NLRP3 inflammasome and subsequent inflammatory responses. After priming mixed glial cultures with lipopolysaccharide (LPS), cells were stimulated with ATP, showing a significant reduction of IL1-ß release in NOX4 and Nrf2 KO mice. Importantly, NOX4 inhibition using GKT136901 also reduced IL-1ß release, as in NOX4 KO mixed glial cultures. Moreover, we measured NOX4 and NLRP3 expression in wild-type mixed glial cultures following LPS treatment, observing that both increased after TLR4 activation, while 24 h treatment with tert-butylhydroquinone, a potent Nrf2 inducer, significantly reduced NLRP3 expression. LPS administration resulted in significant cognitive impairment compared to the control group. Indeed, LPS also modified the expression of NLRP3 and NOX4 in mouse hippocampus. However, mice treated with GKT136901 after LPS impairment showed a significantly improved discrimination index and recovered the expression of inflammatory genes to normal levels compared with wild-type animals. Hence, we here validate NOX4 as a key player in NLRP3 inflammasome activation, suggesting NOX4 pharmacological inhibition as a potent therapeutic approach in neurodegenerative diseases.

Unique expression of the atypical mitochondrial subunit NDUFA4L2 in cerebral pericytes fine tunes HIF activity in response to hypoxia.

A central response to insufficient cerebral oxygen delivery is a profound reprograming of metabolism, which is mainly regulated by the Hypoxia Inducible Factor (HIF). Among other responses, HIF induces the expression of the atypical mitochondrial subunit NDUFA4L2. Surprisingly, NDUFA4L2 is constitutively expressed in the brain in non-hypoxic conditions. Analysis of publicly available single cell transcriptomic (scRNA-seq) data sets coupled with high-resolution multiplexed fluorescence RNA in situ hybridization (RNA F.I.S.H.) revealed that in the murine and human brain NDUFA4L2 is exclusively expressed in mural cells with the highest levels found in pericytes and declining along the arteriole-arterial smooth muscle cell axis. This pattern was mirrored by COX4I2, another atypical mitochondrial subunit. High NDUFA4L2 expression was also observed in human brain pericytes in vitro, decreasing when pericytes are muscularized and further induced by HIF stabilization in a PHD2/PHD3 dependent manner. In vivo, Vhl conditional inactivation in pericyte targeting Ng2-cre transgenic mice dramatically induced NDUFA4L2 expression. Finally NDUFA4L2 inactivation in pericytes increased oxygen consumption and therefore the degree of HIF pathway induction in hypoxia. In conclusion our work reveals that NDUFA4L2 together with COX4I2 is a key hypoxic-induced metabolic marker constitutively expressed in pericytes coupling mitochondrial oxygen consumption and cellular hypoxia response.

Activation of NLRP3 Is Required for a Functional and Beneficial Microglia Response after Brain Trauma.

Despite the numerous research studies on traumatic brain injury (TBI), many physiopathologic mechanisms remain unknown. TBI is a complex process, in which neuroinflammation and glial cells play an important role in exerting a functional immune and damage-repair response. The activation of the NLRP3 inflammasome is one of the first steps to initiate neuroinflammation and so its regulation is essential. Using a closed-head injury model and a pharmacological (MCC950; 3 mg/kg, pre- and post-injury) and genetical approach (NLRP3 knockout (KO) mice), we defined the transcriptional and behavioral profiles 24 h after TBI. Wild-type (WT) mice showed a strong pro-inflammatory response, with increased expression of inflammasome components, microglia and astrocytes markers, and cytokines. There was no difference in the IL1ß production between WT and KO, nor compensatory mechanisms of other inflammasomes. However, some microglia and astrocyte markers were overexpressed in KO mice, resulting in an exacerbated cytokine expression. Pretreatment with MCC950 replicated the behavioral and blood-brain barrier results observed in KO mice and its administration 1 h after the lesion improved the damage. These findings highlight the importance of NLRP3 time-dependent activation and its role in the fine regulation of glial response.

Synthesis and Pharmacological Evaluation of New N-Sulfonylureas as NLRP3 Inflammasome Inhibitors: Identification of a Hit Compound to Treat Gout.

NLRP3 is involved in the pathophysiology of several inflammatory diseases. Therefore, there is high current interest in the clinical development of new NLRP3 inflammasome small inhibitors to treat these diseases. Novel N-sulfonylureas were obtained by the replacement of the hexahydroindacene moiety of the previously described NLRP3 inhibitor MCC950. These new derivatives show moderate to high potency in inhibiting IL-1ß release in vitro. The greatest effect was observed for compound 4b, which was similar to MCC950. Moreover, compound 4b was able to reduce caspase-1 activation, oligomerization of ASC, and therefore, IL-1ß processing. Additional in silico predictions confirmed the safety profile of compound 4b, and in vitro studies in AML12 hepatic cells confirmed the absence of toxicological effects. Finally, we evaluated in vivo anti-inflammatory properties of compound 4b, which showed a significant anti-inflammatory effect and reduced mechanical hyperalgesia at 3 and 10 mg/kg (i.p.) in an in vivo mouse model of gout.

Time-dependent dual effect of NLRP3 inflammasome in brain ischaemia.

BACKGROUND: Inflammasomes are cytosolic multiprotein complexes which, upon assembly, activate the maturation and secretion of the inflammatory cytokines IL-1ß and IL-18. However, participation of the NLRP3 inflammasome in ischaemic stroke remains controversial. Our aims were to determine the role of NLRP3 in brain ischaemia, and explore the mechanism involved in the potential protective effect of the neurovascular unit. METHODS: WT and NLRP3 knock-out mice were subjected to ischaemia by middle cerebral artery occlusion (60 min) with or without treatment with MCC950 at different time points post-stroke. Brain injury was measured histologically with 2,3,5-triphenyltetrazolium chloride (TTC) staining. RESULTS: We identified a time-dependent dual effect of NLRP3. While neither the pre-treatment with MCC950 nor the genetic approach (NLRP3 KO) proved to be neuroprotective, post-reperfusion treatment with MCC950 significantly reduced the infarct volume in a dose-dependent manner. Importantly, MCC950 improved the neuro-motor function and reduced the expression of different pro-inflammatory cytokines (IL-1ß and TNF-α), NLRP3 inflammasome components (NLRP3 and pro-caspase-1), protease expression (MMP9), and endothelial adhesion molecules (ICAM and VCAM). We observed a marked protection of the blood-brain barrier (BBB), which was also reflected in the recovery of the tight junction proteins (ZO-1 and Claudin-5). Additionally, MCC950 produced a reduction of the CCL2 chemokine in blood serum and in brain tissue, which lead to a reduction in the immune cell infiltration. CONCLUSIONS: These findings suggest that post-reperfusion NLRP3 inhibition may be an effective acute therapy for protecting the blood-brain barrier in cerebral ischaemia with potential clinical translation.

Serum Amyloid A1/Toll-Like Receptor-4 Axis, an Important Link between Inflammation and Outcome of TBI Patients.

Traumatic brain injury (TBI) is one of the leading causes of mortality and disability worldwide without any validated biomarker or set of biomarkers to help the diagnosis and evaluation of the evolution/prognosis of TBI patients. To achieve this aim, a deeper knowledge of the biochemical and pathophysiological processes triggered after the trauma is essential. Here, we identified the serum amyloid A1 protein-Toll-like receptor 4 (SAA1-TLR4) axis as an important link between inflammation and the outcome of TBI patients. Using serum and mRNA from white blood cells (WBC) of TBI patients, we found a positive correlation between serum SAA1 levels and injury severity, as well as with the 6-month outcome of TBI patients. SAA1 levels also correlate with the presence of TLR4 mRNA in WBC. In vitro, we found that SAA1 contributes to inflammation via TLR4 activation that releases inflammatory cytokines, which in turn increases SAA1 levels, establishing a positive proinflammatory loop. In vivo, post-TBI treatment with the TLR4-antagonist TAK242 reduces SAA1 levels, improves neurobehavioral outcome, and prevents blood-brain barrier disruption. Our data support further evaluation of (i) post-TBI treatment in the presence of TLR4 inhibition for limiting TBI-induced damage and (ii) SAA1-TLR4 as a biomarker of injury progression in TBI patients.

Acute systemic inflammation exacerbates neuroinflammation in Alzheimer's disease: IL-1ß drives amplified responses in primed astrocytes and neuronal network dysfunction.

Neuroinflammation contributes to Alzheimer's disease (AD) progression. Secondary inflammatory insults trigger delirium and can accelerate cognitive decline. Individual cellular contributors to this vulnerability require elucidation. Using APP/PS1 mice and AD brain, we studied secondary inflammatory insults to investigate hypersensitive responses in microglia, astrocytes, neurons, and human brain tissue. The NLRP3 inflammasome was assembled surrounding amyloid beta, and microglia were primed, facilitating exaggerated interleukin-1ß (IL-1ß) responses to subsequent LPS stimulation. Astrocytes were primed to produce exaggerated chemokine responses to intrahippocampal IL-1ß. Systemic LPS triggered microglial IL-1ß, astrocytic chemokines, IL-6, and acute cognitive dysfunction, whereas IL-1ß disrupted hippocampal gamma rhythm, all selectively in APP/PS1 mice. Brains from AD patients with infection showed elevated IL-1ß and IL-6 levels. Therefore, amyloid leaves the brain vulnerable to secondary inflammation at microglial, astrocytic, neuronal, and cognitive levels, and infection amplifies neuroinflammatory cytokine synthesis in humans. Exacerbation of neuroinflammation to produce deleterious outcomes like delirium and accelerated disease progression merits careful investigation in humans.

TLR4 pathway impairs synaptic number and cerebrovascular functions through astrocyte activation following traumatic brain injury.

BACKGROUND AND PURPOSE: Activation of astrocytes contributes to synaptic remodelling, tissue repair and neuronal survival following traumatic brain injury (TBI). The mechanisms by which these cells interact to resident/infiltrated inflammatory cells to rewire neuronal networks and repair brain functions remain poorly understood. Here, we explored how TLR4-induced astrocyte activation modified synapses and cerebrovascular integrity following TBI. EXPERIMENTAL APPROACH: To determine how functional astrocyte alterations induced by activation of TLR4 pathway in inflammatory cells regulate synapses and neurovascular integrity after TBI, we used pharmacology, genetic approaches, live calcium imaging, immunofluorescence, flow cytometry, blood-brain barrier (BBB) integrity assessment and molecular and behavioural methods. KEY RESULTS: Shortly after a TBI, there is a recruitment of excitable and reactive astrocytes mediated by TLR4 pathway activation with detrimental effects on post-synaptic density-95 (PSD-95)/vesicular glutamate transporter 1 (VGLUT1) synaptic puncta, BBB integrity and neurological outcome. Pharmacological blockage of the TLR4 pathway with resatorvid (TAK-242) partially reversed many of the observed effects. Synapses and BBB recovery after resatorvid administration were not observed in IP3 R2-/- mice, indicating that effects of TLR4 inhibition depend on the subsequent astrocyte activation. In addition, TBI increased the astrocytic-protein thrombospondin-1 necessary to induce a synaptic recovery in a sub-acute phase. CONCLUSIONS AND IMPLICATIONS: Our data demonstrate that TLR4-mediated signalling, most probably through microglia and/or infiltrated monocyte-astrocyte communication, plays a crucial role in the TBI pathophysiology and that its inhibition prevents synaptic loss and BBB damage accelerating tissue recovery/repair, which might represent a therapeutic potential in CNS injuries and disorders.

Melatonin Reduces NLRP3 Inflammasome Activation by Increasing α7 nAChR-Mediated Autophagic Flux.

Microglia controls the immune system response in the brain. Specifically, the activation and dysregulation of the NLRP3 inflammasome is responsible for the initiation of the inflammatory process through IL-1ß and IL-18 release. In this work, we have focused on studying the effect of melatonin on the regulation of the NLRP3 inflammasome through α7 nicotinic receptor (nAChR) and its relationship with autophagy. For this purpose, we have used pharmacological and genetic approaches in lipopolysaccharide (LPS)-induced inflammation models in both in vitro and in vivo models. In the BV2 cell line, LPS inhibited autophagy, which increased NLRP3 protein levels. However, melatonin promoted an increase in the autophagic flux. Treatment of glial cultures from wild-type (WT) mice with LPS followed by extracellular adenosine triphosphate (ATP) produced the release of IL-1ß, which was reversed by melatonin pretreatment. In cultures from α7 nAChR knock-out (KO) mice, melatonin did not reduce IL-1ß release. Furthermore, melatonin decreased the expression of inflammasome components and reactive oxygen species (ROS) induced by LPS; co-incubation of melatonin with α-bungarotoxin (α-bgt) or luzindole abolished the anti-inflammatory and antioxidant effects. In vivo, melatonin reverted LPS-induced cognitive decline, reduced NLRP3 levels and promoted autophagic flux in the hippocampi of WT mice, whereas in α7 nAChR KO mice melatonin effect was not observed. These results suggest that melatonin may modulate the complex interplay between α7 nAChR and autophagy signaling.

Acute Inflammation Alters Brain Energy Metabolism in Mice and Humans: Role in Suppressed Spontaneous Activity, Impaired Cognition, and Delirium.

Systemic infection triggers a spectrum of metabolic and behavioral changes, collectively termed sickness behavior, which while adaptive, can affect mood and cognition. In vulnerable individuals, acute illness can also produce profound, maladaptive, cognitive dysfunction including delirium, but our understanding of delirium pathophysiology remains limited. Here, we used bacterial lipopolysaccharide (LPS) in female C57BL/6J mice and acute hip fracture in humans to address whether disrupted energy metabolism contributes to inflammation-induced behavioral and cognitive changes. LPS (250 µg/kg) induced hypoglycemia, which was mimicked by interleukin (IL)-1ß (25 µg/kg) but not prevented in IL-1RI-/- mice, nor by IL-1 receptor antagonist (IL-1RA; 10 mg/kg). LPS suppression of locomotor activity correlated with blood glucose concentrations, was mitigated by exogenous glucose (2 g/kg), and was exacerbated by 2-deoxyglucose (2-DG) glycolytic inhibition, despite preventing IL-1ß synthesis. Using the ME7 model of chronic neurodegeneration in female mice, to examine vulnerability of the diseased brain to acute stressors, we showed that LPS (100 µg/kg) produced acute cognitive dysfunction, selectively in those animals. These acute cognitive impairments were mimicked by insulin (11.5 IU/kg) and mitigated by glucose, demonstrating that acutely reduced glucose metabolism impairs cognition selectively in the vulnerable brain. To test whether these acute changes might predict altered carbohydrate metabolism during delirium, we assessed glycolytic metabolite levels in CSF in humans during inflammatory trauma-induced delirium. Hip fracture patients showed elevated CSF lactate and pyruvate during delirium, consistent with acutely altered brain energy metabolism. Collectively, the data suggest that disruption of energy metabolism drives behavioral and cognitive consequences of acute systemic inflammation.SIGNIFICANCE STATEMENT Acute systemic inflammation alters behavior and produces disproportionate effects, such as delirium, in vulnerable individuals. Delirium has serious short and long-term sequelae but mechanisms remain unclear. Here, we show that both LPS and interleukin (IL)-1ß trigger hypoglycemia, reduce CSF glucose, and suppress spontaneous activity. Exogenous glucose mitigates these outcomes. Equivalent hypoglycemia, induced by lipopolysaccharide (LPS) or insulin, was sufficient to trigger cognitive impairment selectively in animals with existing neurodegeneration and glucose also mitigated those impairments. Patient CSF from inflammatory trauma-induced delirium also shows altered brain carbohydrate metabolism. The data suggest that the degenerating brain is exquisitely sensitive to acute behavioral and cognitive consequences of disrupted energy metabolism. Thus "bioenergetic stress" drives systemic inflammation-induced dysfunction. Elucidating this may offer routes to mitigating delirium.

Effects of cannabis exposure in the prenatal and adolescent periods: Preclinical and clinical studies in both sexes.

Cannabis is the most commonly used illicit drug among adolescents and young adults, including pregnant women. There is substantial evidence for a significant association between prenatal cannabis exposure and lower birth weight in offspring, and mixed results regarding later behavioural outcomes in the offspring. Adolescent cannabis use, especially heavy use, has been associated with altered executive function, depression, psychosis and use of other drugs later in life. Human studies have limitations due to several confounding factors and have provided scarce information about sex differences. In general, animal studies support behavioural alterations reported in humans and have revealed diverse sex differences and potential underlying mechanisms (altered mesolimbic dopaminergic and hippocampal glutamatergic systems and interference with prefrontal cortex maturation). More studies are needed that analyse sex and gender influences on cannabis-induced effects with great clinical relevance such as psychosis, cannabis use disorder and associated comorbidities, to achieve more personalized and accurate treatments.

Bath salts and polyconsumption: in search of drug-drug interactions.

BACKGROUND AND RATIONALE: Polydrug use is a widespread phenomenon, especially among adolescents and young adults. Synthetic cathinones are frequently consumed in combination with other drugs of abuse. However, there is very little information regarding the consequences of this specific consumption pattern. OBJECTIVES: The aim of this review is to introduce this topic and highlight the gaps in the existing literature. In three different sections, we focus on specific interactions of synthetic cathinones with alcohol, cannabinoids, and the stimulants nicotine and cocaine. We then dedicate a section to the existence of sex and gender differences in the effects of synthetic cathinones and the long-term psychophysiological consequences of adolescent and prenatal exposure to these drugs. MAJOR FINDINGS: Epidemiological studies, case reports, and results obtained in animal models point to the existence of pharmacological and pharmacokinetic interactions between synthetic cathinones and other drugs of abuse. This pattern of polyconsumption can cause the potentiation of negative effects, and the dissociation between objective and subjective effects can increase the combined use of the drugs and the risk of toxicity leading to serious health problems. Certain animal studies indicate a higher vulnerability and effect of cathinones in females. In humans, most of the users are men and case reports show long-term psychotic symptoms after repeated use. CONCLUSIONS: The co-use of synthetic cathinones and the other drugs of abuse analyzed indicates potentiation of diverse effects including dependence and addiction, neurotoxicity, and impaired cognition and emotional responses. The motivations for and effects of synthetic cathinone use appear to be influenced by sex/gender. The long-term consequences of their use by adolescents and pregnant women deserve further investigation.

Chronic neurodegeneration induces type I interferon synthesis via STING, shaping microglial phenotype and accelerating disease progression.

Type I interferons (IFN-I) are the principal antiviral molecules of the innate immune system and can be made by most cell types, including central nervous system cells. IFN-I has been implicated in neuroinflammation during neurodegeneration, but its mechanism of induction and its consequences remain unclear. In the current study, we assessed expression of IFN-I in murine prion disease (ME7) and examined the contribution of the IFN-I receptor IFNAR1 to disease progression. The data indicate a robust IFNß response, specifically in microglia, with evidence of IFN-dependent genes in both microglia and astrocytes. This IFN-I response was absent in stimulator of interferon genes (STING-/- ) mice. Microglia showed increased numbers and activated morphology independent of genotype, but transcriptional signatures indicated an IFNAR1-dependent neuroinflammatory phenotype. Isolation of microglia and astrocytes demonstrated disease-associated microglial induction of Tnfα, Tgfb1, and of phagolysosomal system transcripts including those for cathepsins, Cd68, C1qa, C3, and Trem2, which were diminished in IFNAR1 and STING deficient mice. Microglial increases in activated cathepsin D, and CD68 were significantly reduced in IFNAR1-/- mice, particularly in white matter, and increases in COX-1 expression, and prostaglandin synthesis were significantly mitigated. Disease progressed more slowly in IFNAR1-/- mice, with diminished synaptic and neuronal loss and delayed onset of neurological signs and death but without effect on proteinase K-resistant PrP levels. Therefore, STING-dependent IFN-I influences microglial phenotype and influences neurodegenerative progression despite occurring secondary to initial degenerative changes. These data expand our mechanistic understanding of IFN-I induction and its impact on microglial function during chronic neurodegeneration.

Astrocytes: Heterogeneous and Dynamic Phenotypes in Neurodegeneration and Innate Immunity.

Astrocytes are the most numerous cell type in the brain and perform several essential functions in supporting neuronal metabolism and actively participating in neural circuit and behavioral function. They also have essential roles as innate immune cells in responding to local neuropathology, and the manner in which they respond to brain injury and degeneration is the subject of increasing attention in neuroscience. Although activated astrocytes have long been thought of as a relatively homogenous population, which alter their phenotype in a relatively stereotyped way upon central nervous system injury, the last decade has revealed substantial heterogeneity in the basal state and significant heterogeneity of phenotype during reactive astrocytosis. Thus, phenotypic diversity occurs at two distinct levels: that determined by regionality and development and that determined by temporally dynamic changes to the environment of astrocytes during pathology. These inflammatory and pathological states shape the phenotype of these cells, with different consequences for destruction or recovery of the local tissue, and thus elucidating these phenotypic changes has significant therapeutic implications. In this review, we will focus on the phenotypic heterogeneity of astrocytes in health and disease and their propensity to change that phenotype upon subsequent stimuli.

Development and validation of a risk stratification model for prediction of disability and hospitalisation in patients with heart failure: a study protocol.

BACKGROUND: Chronic heart failure (CHF) reduces quality of life and causes hospitalisation and death. Identifying predictive factors of such events may help change the natural history of this condition. AIM: To develop and validate a stratification system for classifying patients with CHF, according to their degree of disability and need for hospitalisation due to any unscheduled cause, over a period of 1 year. METHODS AND ANALYSIS: Prospective, concurrent, cohort-type study in two towns in the Madrid autonomous region having a combined population of 1 32 851. The study will include patients aged over 18 years who meet the following diagnostic criteria: symptoms and typical signs of CHF (Framingham criteria) and left ventricular ejection fraction (EF)<50% or structural cardiac lesion and/or diastolic dysfunction in the presence of preserved EF (EF>50%).Outcome variables will be(a) Disability, as measured by the WHO Disability Assessment Schedule V.2.0 Questionnaire, and (b) unscheduled hospitalisations. The estimated sample size is 557 patients, 371 for predictive model development (development cohort) and 186 for validation purposes (validation cohort). Predictive models of disability or hospitalisation will be constructed using logistic regression techniques. The resulting model(s) will be validated by estimating the probability of outcomes of interest for each individual included in the validation cohort. ETHICS AND DISSEMINATION: The study protocol has been approved by the Clinical Research Ethics Committee of La Princesa University Teaching Hospital (PI-705). All results will be published in a peer-reviewed journal and shared with the medical community at conferences and scientific meetings.

Systemic TNF-α produces acute cognitive dysfunction and exaggerated sickness behavior when superimposed upon progressive neurodegeneration.

Inflammation influences chronic neurodegeneration but its precise roles are not yet clear. Systemic inflammation caused by infection, trauma or co-morbidity can alter the brain's inflammatory status, produce acute cognitive impairments, such as delirium, and drive new pathology and accelerated decline. Consistent with this, elevated systemic TNF-α is associated with more rapid cognitive decline over 6months in Alzheimer's disease patients. In the current study we challenged normal animals and those with existing progressive neurodegeneration (ME7 prion disease) with TNF-α (i.p.) to test the hypothesis that this cytokine has differential effects on cognitive function, sickness behavior and features of underlying pathology contingent on the animals' baseline condition. TNF-α (50µg/kg) had no impact on performance of normal animals (normal brain homogenate; NBH) on working memory (T-maze) but produced acute impairments in ME7 animals similarly challenged. Plasma TNF-α and CCL2 levels were equivalent in NBH and ME7 TNF-challenged animals but hippocampal and hypothalamic transcription of IL-1ß, TNF-α and CCL2 and translation of IL-1ß were higher in ME7+TNF-α than NBH+TNF-α animals. TNF-α produced an exaggerated sickness behavior response (hypothermia, weight loss, inactivity) in ME7 animals compared to that in NBH animals. However a single challenge with this dose was not sufficient to produce de novo neuronal death, synaptic loss or tau hyperphosphorylation that was distinguishable from that arising from ME7 alone. The data indicate that acutely elevated TNF-α has robust acute effects on brain function, selectively in the degenerating brain, but more sustained levels may be required to significantly impact on underlying neurodegeneration.

Blockage of neonatal leptin signaling induces changes in the hypothalamus associated with delayed pubertal onset and modifications in neuropeptide expression during adulthood in male rats.

The neonatal leptin surge, occurring from postnatal day (PND) 5 to 13 and peaking at PND9 in rodents, is important for the development of neuroendocrine circuits involved in metabolic control and reproductive function. We previously demonstrated that treatment with a leptin antagonist from PND 5 to 9, coincident with peak leptin levels in the neonatal surge, modified trophic factors and markers of cell turnover and neuronal maturation in the hypothalamus of peri-pubertal rats. The kisspeptin system and metabolic neuropeptide and hormone levels were also modified. Here our aim was to investigate if the timing of pubertal onset is altered by neonatal leptin antagonism and if the previously observed peripubertal modifications in hormones and neuropeptides persist into adulthood and affect male sexual behavior. To this end, male Wistar rats were treated with a pegylated super leptin antagonist (5mg/kg, s.c.) from PND 5 to 9 and killed at PND102-103. The appearance of external signs of pubertal onset was delayed. Hypothalamic kiss1 mRNA levels were decreased in adult animals, but sexual behavior was not significantly modified. Although there was no effect on body weight or food intake, circulating leptin, insulin and triglyceride levels were increased, while hypothalamic leptin receptor, POMC and AgRP mRNA levels were decreased. In conclusion, alteration of the neonatal leptin surge can modify the timing of pubertal onset and have long-term effects on hypothalamic expression of reproductive and metabolic neuropeptides.