Jaume Folch

Early alterations in energy metabolism in the hippocampus of APPswe/PS1dE9 mouse model of Alzheimer's disease

The present study had focused on the behavioral phenotype and gene expression profile of molecules related to insulin receptor signaling in the hippocampus of 3 and 6 month-old APPswe/PS1dE9 (APP/PS1) transgenic mouse model of Alzheimers disease (AD). Elevated levels of the insoluble Aβ (1-42) were detected in the brain extracts of the transgenic animals as early as 3 months of age, prior to the Aβ plaque formation (pre-plaque stage). By the early plaque stage (6 months) both the soluble and insoluble Aβ (1-40) and Aβ (1-42) peptides were detectable. We studied the expression of genes related to memory function (Arc, Fos), insulin signaling, including insulin receptor (Insr), Irs1 and Irs2, as well as genes involved in insulin growth factor pathways, such as Igf1, Igf2, Igfr and Igfbp2. We also examined the expression and protein levels of key molecules related to energy metabolism (PGC1-α, and AMPK) and mitochondrial functionality (OXPHOS, TFAM, NRF1 and NRF2). 6 month-old APP/PS1 mice demonstrated impaired cognitive ability, were glucose intolerant and showed a significant reduction in hippocampal Insr and Irs2 transcripts. Further observations also suggest alterations in key cellular energy sensors that regulate the activities of a number of metabolic enzymes through phosphorylation, such as a decrease in the Prkaa2 mRNA levels and in the pAMPK (Thr172)/Total APMK ratio. Moreover, mRNA and protein analysis reveals a significant downregulation of genes essential for mitochondrial replication and respiratory function, including PGC-1α in hippocampal extracts of APP/PS1 mice, compared to age-matched wild-type controls at 3 and 6 months of age. Overall, the findings of this study show early alterations in genes involved in insulin and energy metabolism pathways in an APP/PS1 model of AD. These changes affect the activity of key molecules like NRF1 and PGC-1α, which are involved in mitochondrial biogenesis. Our results reinforce the hypothesis that the impairments in both insulin signaling and energy metabolism precede the development of AD amyloidogenesis.

Modulation of SIRT1 expression in different neurodegenerative models and human pathologies

We examined the expression of SIRT1 in several experimental paradigms of human pathologies. We used a neuroblastoma cell line (B65), neuronal primary cultures (hippocampus and cerebellar granule cells) and in vivo approaches in rat and senescence murine models (SAM). Cell cultures and rats were treated with several well-know neurotoxins, i.e. rotenone, MPP(+), kainate and 3-nitropropionic acid. Subsequently, SIRT1 expression was compared in these different paradigms of neurotoxicity. The pattern of expression of SIRT1 in proliferating cell cultures (B65) was different to that in quiescent cell cultures. In the murine model of senescence (senescence-accelerated mice prone, SAMP8), SIRT1 expression progressively decreased, while in the control strain (senescence-accelerated mice resistant, SAMR1) it increased. Finally, we studied human samples of Parkinsons disease (PD), dementia with Lewy bodies (DLB) and Huntingtons diseases (HD). SIRT1 expression decreased dramatically in HD, but there were no significant changes in Parkinson-related illnesses. In conclusion, SIRT1 expression may be a good sensor of toxic neuronal processes.

Role of cell cycle re-entry in neurons: a common apoptotic mechanism of neuronal cell death.

Currently, there is no effective treatment for neurodegenerative disorders such as Alzheimer’s disease and Parkinson’s disease. Thus, a major focus of neuroscience research is to examine the mechanisms involved in neuronal loss in order to identify potential drug targets. Recent results indicate that DNA damage and re-entry into the cell cycle may constitute a common pathway in apoptosis in neurological diseases. The role of the cell cycle in such disorders is supported by data on the brain of patients who showed an increase in cell-cycle protein expression. Indeed, studies performed in neuronal cell preparations indicate that oxidative stress could be the main mechanism responsible for cell cycle re-entry. DNA damage and repair after oxidative stress may activate the enzyme ataxia telangiectasia mutated, which is a cell-cycle regulator. Once the cell cycle is activated, the increase in the expression of transcription factor E2F-1 could induce neuronal apoptosis. Furthermore, the potential routes involved in E2F-1 induced apoptosis could be p53-dependent or p53-independent. Under this E2F-1 hypothesis of cell death, multiple mitochondria-dependent pathways may be activated, including caspase and caspase-independent signaling cascades. Finally, given that cyclin-dependent kinase inhibitory drugs have neuroprotective and anti-apoptotic effects in experimental models, their potential application for the treatment of neurological disorders should be taken into account.

Sirtuin activators: Designing molecules to extend life span

Resveratrol (RESV) exerts important pharmacological effects on human health: in addition to its beneficial effects on type 2 diabetes and cardiovascular diseases, it also modulates neuronal energy homeostasis and shows antiaging properties. Although it clearly has free radical scavenger properties, the mechanisms involved in these beneficial effects are not fully understood. In this regard, one area of major interest concerns the effects of RESV on the activity of sirtuin 1 (SIRT1), an NAD(+)-dependent histone deacetylase that has been implicated in aging. Indeed, the role of SIRT1 is currently the subject of intense research due to the antiaging properties of RESV, which increases life span in various organisms ranging from yeast to rodents. In addition, when RESV is administered in experimental animal models of neurological disorders, it has similar beneficial effects to caloric restriction. SIRT1 activation could thus constitute a potential strategic target in neurodegenerative diseases and in disorders involving disturbances in glucose homeostasis, as well as in dyslipidaemias or cardiovascular diseases. Therefore, small SIRT1 activators such as SRT501, SRT2104, and SRT2379, which are currently undergoing clinical trials, could be potential drugs for the treatment of type 2 diabetes, obesity, and metabolic syndrome, among other disorders. This review summarises current knowledge about the biological functions of SIRT1 in aging and aging-associated diseases and discusses its potential as a pharmacological target.

Inhibition of the cdk5/p25 fragment formation may explain the antiapoptotic effects of melatonin in an experimental model of Parkinson's disease

Abstract:  In this study, the effects of melatonin on MPP+‐treated cerebellar granule neurons (CGNs) in culture were investigated. Results showed that MPP+ treatment significantly decreased cell viability and increased the apoptotic cell population at 24 and 48 hr. Calpain and caspase‐3 activation was also determined, with results showing a strong increase in calpain (74%) and caspase 3 activity (70%), as measured by α‐spectrin cleavage and fluorometric and colorimetric analysis, respectively. There are several studies suggesting that the activation of the cdk5/p35 pathway at its cleavage to cdk5/p25 may play a role in neuronal cell death in neurodegenerative diseases. Moreover, these studies indicate that this cleavage is mediated by calpains, and that MPP+ prompted an increase in cdk5 expression, as well as the cleavage of p35–p25, in a time‐dependent manner. 1 mm Melatonin not only reduced the neurotoxic effects of MPP+ on cell viability, but also prevented apoptosis mediated by this Parkinsonian toxin in CGNs. 1 mm Melatonin reduced cdk5 expression, as well as the cleavage of p35–p25. These data indicate that melatonin possesses some neuro‐protective properties against MPP+‐induced apoptosis. Moreover, these data suggest that the calpain/cdk5 signaling cascade has a potential role in the MPP+‐mediated apoptotic process in CGNs.

High-fat diet-induced deregulation of hippocampal insulin signaling and mitochondrial homeostasis deficiences contribute to Alzheimer disease pathology in rodents

Global obesity is a pandemic status, estimated to affect over 2 billion people, that has resulted in an enormous strain on healthcare systems worldwide. The situation is compounded by the fact that apart from the direct costs associated with overweight pathology, obesity presents itself with a number of comorbidities, including an increased risk for the development of neurodegenerative disorders. Alzheimer disease (AD), the main cause of senile dementia, is no exception. Spectacular failure of the pharmaceutical industry to come up with effective AD treatment strategies is forcing the broader scientific community to rethink the underlying molecular mechanisms leading to cognitive decline. To this end, the emphasis is once again placed on the experimental animal models of the disease. In the current study, we have focused on the effects of a high-fat diet (HFD) on hippocampal-dependent memory in C57/Bl6 Wild-type (WT) and APPswe/PS1dE9 (APP/PS1) mice, a well-established mouse model of familial AD. Our results indicate that the continuous HFD administration starting at the time of weaning is sufficient to produce β-amyloid-independent, hippocampal-dependent memory deficits measured by a 2-object novel-object recognition test (NOR) in mice as early as 6months of age. Furthermore, the resulting metabolic syndrome appears to have direct effects on brain insulin regulation and mitochondrial function. We have observed pathological changes related to both the proximal and distal insulin signaling pathway in the brains of HFD-fed WT and APP/PS1 mice. These changes are accompanied by a significantly reduced OXPHOS metabolism, suggesting that mitochondria play an important role in hippocampus-dependent memory formation and retention in both the HFD-treated and AD-like rodents at a relatively young age.

Lithium treatment decreases activities of tau kinases in a murine model of senescence.

Lithium modulates glycogen synthase kinase 3β (GSK-3β), a kinase involved in Alzheimer disease-related tau pathology. To investigate mechanisms of aging and the potential therapy of lithium in neurodegenerative disease, we treated senescence-accelerated mouse (SAM)P8 mice, a murine model of senescence, and mice of the control SAMR1 strain with lithium. The treatment reduced hippocampal caspase 3 and calpain activation, indicating that it provides neuroprotection. Lithium also reduced both the levels and activity of GSK-3β and the activity of cyclin-dependent kinase 5 and reduced hyperphosphorylation of 3 different phosphoepitopes of tau: Ser199, Ser212, and Ser396. In lithium-treated primary cultures of SAMP8 and SAMR1 cerebellar neurons, there was a marked reduction in protease activity mediated by calpain and caspase 3. Both lithium and SB415286, a specific inhibitor of GSK-3β, reduced apoptosis in vitro. Taken together, these in vivo and in vitro findings of lithium-mediated reductions in GSK-3β and cyclin-dependent kinase 5 activities, tau phosphorylation, apoptotic activity, and cell death provide a strong rationale for the use of lithium as a potential treatment in neurodegenerative diseases.

Evidence in favour of a role for peripheral-type benzodiazepine receptor ligands in amplification of neuronal apoptosis.

The mitochondrial peripheral benzodiazepine receptor (PBR) is involved in a functional structure designated as the mitochondrial permeability transition (MPT) pore, which controls apoptosis. PBR expression in nervous system has been reported in glial and immune cells. We now show expression of both PBR mRNA and protein, and the appearance of binding of a synthetic ligand fluo-FGIN-1-27 in mitochondria of rat cerebellar granule cells (CGCs). Additionally, the effect of PBR ligands on colchicine-induced apoptosis was investigated. Colchicine-induced neurotoxicity in CGCs was measured at 24 h. We show that, in vitro, PBR ligands 1-(2-chlorophenyl-N-methylpropyl)-3-isoquinolinecarboxamide (PK11195), 7-chloro-5-(4-chlorophenyl)-1,3-dihydro-1-methyl-2H-1,4- benzodiazepin-2-one (Ro5-4864) and diazepam (25– 50 μM) enhanced apoptosis induced by colchicine, as demonstrated by viability experiments, flow cytometry and nuclear chromatin condensation. Enhancement of colchicine-induced apoptosis was characterized by an increase in mitochondrial release of cytochrome c and AIF proteins and an enhanced activation of caspase-3, suggesting mitochondrion dependent mechanism that is involved in apoptotic process. Our results indicate that exposure of neural cells to PBR ligands generates an amplification of apoptotic process induced by colchicine and that the MPT pore may be involved in this process.

The antiproliferative activity of melatonin in B65 rat dopaminergic neuroblastoma cells is related to the downregulation of cell cycle-related genes

Abstract:  A potential application of melatonin is its ability to rescue many cell types from cell death, because of its antioxidant properties. Likewise, recent studies suggest that melatonin may also be used as an anti‐tumor drug, due to its anti‐proliferative properties in tumor cells when administered at physiologic or pharmacologic doses. In the present study, we investigated the mechanisms involved in the apoptosis induced by acute exposure to melatonin and roscovitine in the rat dopaminergic neuroblastoma B65 cell line. Cell growth studies revealed that, at 24 hr of treatment, roscovitine blocked cell growth and induced apoptosis whereas melatonin delayed cell growth and induced a slight increase in the number of apoptotic nuclei. Melatonin also increased the percentage of cells in the G1‐phase of the cell cycle, whereas roscovitine blocked cells in the G2/M‐phase. Both compounds significantly downregulated the transcriptional activity of cdk4, while melatonin also downregulated cdk2 and cyclin D1. Taken together, our data show that melatonin at millimolar concentrations inhibits dopaminergic B65 proliferation, induces cell apoptosis, and modulates cell cycle progression by inhibiting the transcriptional activity of cyclins and cdks related to the progression of the G1‐phase.

Neuroprotective and anti-ageing role of leptin

Leptin (Lep), an adipose-derived hormone, exerts very important functions in the body mainly on energy storage and availability. The physiological effects of Lep controlling the body weight and suppressing appetite are mediated by the long form of Lep receptor in the hypothalamus. Lep receptor activates several downstream molecules involved in key pathways related to cell survival such as STAT3, PI3K, MAPK, AMPK, CDK5 and GSK3β. Collectively, these pathways act in a coordinated manner and form a network that is fully involved in Lep physiological response. Although the major interest in Lep is related to its role in the regulation of energy balance, and since resistance to Lep affects is the primary risk factor for obesity, the interest on their effects on brain cognition and neuroprotection is increasing. Thus, Lep and Lep mimetic compounds now await and deserve systematic exploration as the orchestrator of protective responses in the nervous system. Moreover, Lep might promote the activation of a cognitive process that may retard or even partially reverse selected aspects of Alzheimers disease or ageing memory loss.