Sirtuin 1 overexpression mice show a reference memory deficit, but not neuroprotection

Sirtuin 1 (SIRT1) is the closest mammalian homologue of yeast silent information regulator 2 (Sir2) and has a role in lifespan modulation. Reportedly, SIRT1 is also linked to neurodegenerative diseases. However, there are limited studies that report the relation between SIRT1 and neurodegenerative diseases using in vivo transgenic (Tg) methods. In the present study, we generated neuron-specific enolase (NSE) SIRT1 Tg mice that overexpress human SIRT1 in neurons. We examined possible neuroprotective effects of SIRT1 overexpression and compared their higher brain functions with those of wild-type (WT) mice. Overexpression of SIRT1 did not have any neuroprotective effects against the neuronal damage induced by ischemia or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). However, SIRT1 Tg mice exhibited a reference memory deficit. These findings suggest that an excessive expression of SIRT1 might induce the memory deficit in mice, but not neuroprotective effects.     

<Emphasis Type="Italic">N</Emphasis>-palmitoylethanolamide Prevents Parkinsonian Phenotypes in Aged Mice

Parkinson’s disease (PD) is a neurodegenerative disease characterized by degeneration of dopaminergic neurons. Aging is a major risk factor for idiopathic PD. Several prior studies examined the neuroprotective effects of palmitoylethanolamide (PEA), alone or combined with antioxidants, in a model of PD induced by the dopaminergic toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Here, we analyzed the pretreatment effect of micronized PEA (PEAm) on neuroinflammation and neuronal cell death in the MPTP model. Male CD mice (21 months of age) were pre-treated for 60 days with PEAm. After this time, they received four intraperitoneal injections of MPTP over a 24-h period and were killed 7 days later. On the 8th day, brains were processed. Pretreatment with PEAm ameliorated behavioral deficits and the reductions in expression of tyrosine hydroxylase and dopamine transporter, while blunting the upregulation of α-synuclein and β3-tubulin in the substantia nigra after MPTP induction. Moreover, PEAm reduced proinflammatory cytokine expression and showed a pro-neurogenic effect in hippocampus. These findings propose this strategy as a valid approach to prevent neurodegenerative diseases associated with old age.     

Sirtuin 2 (SIRT2) Enhances 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Nigrostriatal Damage via Deacetylating Forkhead Box O3a (Foxo3a) and Activating Bim Protein

Sirtuins are NAD-dependent protein deacetylases that were shown to have beneficial effects against age-related diseases. SIRT2 is a strong deacetylase that is highly expressed in brain. It has been associated with neurodegenerative diseases. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a dopaminergic neurotoxin that replicates most of the clinical features of Parkinson disease (PD) and produces a reliable and reproducible lesion of the nigrostriatal dopaminergic pathway and neurodegeneration after its systemic administration. Chronic administration of MPTP induces lesion via apoptosis. We show here that SIRT2 deacetylates Foxo3a, increases RNA and protein levels of Bim, and as a result, enhances apoptosis in the MPTP model of PD. We also show that neurodegeneration induced by chronic MPTP regimen is prevented by genetic deletion of SIRT2 in mouse. Deletion of SIRT2 leads to the reduction of apoptosis due to an increase in acetylation of Foxo3a and a decrease in Bim levels. We demonstrate that SIRT2 deacetylates Foxo3a, activates Bim, and induces apoptosis only in 1-methyl-4-phenylpyridinium-treated cells. Therefore, designing SIRT2 inhibitors might be helpful to develop effective treatments for PD.     

Proteome analysis of ventral midbrain in MPTP-treated normal and L1cam transgenic mice

Treatment of mice by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridene hydrochloride (MPTP) is a well established animal model for Parkinson's disease (PD), while overexpression of L1 cell adhesion molecule (L1cam) has been proposed to attenuate the degeneration of dopaminergic neurons induced by MPTP. To gain insight into the role of L1cam in the pathomechanism of PD, we investigated protein expression patterns after MPTP-treatment in both C57BL/6 (wild-type) and transgenic mice overexpressing L1cam in astrocytes. Our results showed that during the acute phase, proteins in functional complexes responsible for mitochondrial, glycolysis, and cytoskeletal function were down-regulated in MPTP-treated wild-type mice. After a recovery phase, proteins that were down-regulated in the acute phase reverted to normal levels. In L1cam transgenic mice, a much higher number of proteins was altered during the acute phase and this number even increased after the recovery phase. Many proteins involved in oxidative phosphorylation were still down-regulated and glycolysis related protein were still up-regulated. This pattern indicates a lasting severely impaired energy production in L1cam mice after MPTP treatment.     

Inhibition of repulsive guidance molecule-a protects dopaminergic neurons in a mouse model of Parkinson’s disease

Repulsive guidance molecule-a (RGMa), a glycosylphosphatidylinositol-anchored membrane protein, has diverse functions in axon guidance, cell patterning, and cell survival. Inhibition of RGMa attenuates pathological dysfunction in animal models of central nervous system (CNS) diseases including spinal cord injury, multiple sclerosis, and neuromyelitis optica. Here, we examined whether antibody-based inhibition of RGMa had therapeutic effects in a mouse model of Parkinson’s disease (PD). We treated mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and found increased RGMa expression in the substantia nigra (SN). Intraventricular, as well as intravenous, administration of anti-RGMa antibodies reduced the loss of tyrosine hydroxylase (TH)-positive neurons and accumulation of Iba1-positive microglia/macrophages in the SN of MPTP-treated mice. Selective expression of RGMa in TH-positive neurons in the SN-induced neuronal loss/degeneration and inflammation, resulting in a progressive movement disorder. The pathogenic effects of RGMa overexpression were attenuated by treatment with minocycline, which inhibits microglia and macrophage activation. Increased RGMa expression upregulated pro-inflammatory cytokine expression in microglia. Our observations suggest that the upregulation of RGMa is associated with the PD pathology; furthermore, inhibitory RGMa antibodies are a potential therapeutic option.Subject terms: Cell death in the nervous system, Parkinson''s disease     

Deficiency of inducible nitric oxide synthase protects against MPTP toxicity in vivo

MPTP produces clinical, biochemical, and neuropathologic changes reminiscent of those that occur in idiopathic Parkinson's disease (PD). In the present study we show that MPTP treatment led to activation of microglia in the substantia nigra pars compacta (SNpc), which was associated and colocalized with an increase in inducible nitric oxide synthase (iNOS) expression. In iNOS-deficient mice the increase of iNOS expression but not the activation of microglia was blocked. Dopaminergic SNpc neurons of iNOS-deficient mice were almost completely protected from MPTP toxicity in a chronic paradigm of MPTP toxicity. Because the MPTP-induced decrease in striatal concentrations of dopamine and its metabolites did not differ between iNOS-deficient mice and their wild-type littermates, this protection was not associated with a preservation of nigrostriatal terminals. Our results suggest that iNOS-derived nitric oxide produced in microglia plays an important role in the death of dopaminergic neurons but that other mechanisms contribute to the loss of dopaminergic terminals in MPTP neurotoxicity. We conclude that inhibition of iNOS may be a promising target for the treatment of PD.     

Evaluation of TorsinA as a Target for Parkinson Disease Therapy in Mouse Models

Parkinson disease (PD) is a common and disabling disorder. No current therapy can slow or reverse disease progression. An important aspect of research in this field is target validation, a systematic approach to evaluating the likelihood that modification of a certain molecule, mechanism or biological pathway may be useful for the development of pharmacological or molecular treatments for the disease. TorsinA, a member of the AAA+ family of chaperone proteins, has been proposed as a potential target of neuroprotective therapy. TorsinA is found in Lewy bodies in human PD, and can suppress toxicity in cellular and invertebrate models of PD. Here, we evaluated the neuroprotective properties of torsinA in mouse models of PD based on intoxication with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) as well as recombinant adeno associated virus (rAAV) induced overexpression of alpha-synuclein (α-syn). Using either transgenic mice with overexpression of human torsinA (hWT mice) or mice in which torsinA expression was induced using an rAAV vector, we found no evidence for protection against acute MPTP intoxication. Similarly, genetic deletion of the endogenous mouse gene for torsinA (Dyt1) using an rAAV delivered Cre recombinase did not enhance the vulnerability of dopaminergic neurons to MPTP. Overexpression of α-syn using rAAV in the mouse substantia nigra lead to a loss of TH positive neurons six months after administration, and no difference in the degree of loss was observed between transgenic animals expressing forms of torsinA and wild type controls. Collectively, we did not observe evidence for a protective effect of torsinA in the mouse models we examined. Each of these models has limitations, and there is no single model with established predictive value with respect to the human disease. Nevertheless, these data do seem to support the view that torsinA is unlikely to be successfully translated as a target of therapy for human PD.     

Aquaporin‐4 deficiency reduces TGF‐β1 in mouse midbrains and exacerbates pathology in experimental Parkinson's disease

Aquaporin‐4 (AQP4), the main water‐selective membrane transport protein in the brain, is localized to the astrocyte plasma membrane. Following the establishment of a 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐induced Parkinson's disease (PD) model, AQP4‐deficient (AQP4^?/?) mice displayed significantly stronger microglial inflammatory responses and remarkably greater losses of tyrosine hydroxylase (TH⁺)‐positive neurons than did wild‐type AQP4 (AQP4^+/+) controls. Microglia are the most important immune cells that mediate immune inflammation in PD. However, recently, few studies have reported why AQP4 deficiency results in more severe hypermicrogliosis and neuronal damage after MPTP treatment. In this study, transforming growth factor‐β1 (TGF‐β1), a key suppressive cytokine in PD onset and development, failed to increase in the midbrain and peripheral blood of AQP4^?/? mice after MPTP treatment. Furthermore, the lower level of TGF‐β1 in AQP4^?/? mice partially resulted from impairment of its generation by astrocytes; reduced TGF‐β1 may partially contribute to the uncontrolled microglial inflammatory responses and subsequent severe loss of TH⁺ neurons in AQP4^?/? mice after MPTP treatment. Our study provides not only a better understanding of both aetiological and pathogenical factors implicated in the neurodegenerative mechanism of PD but also a possible approach to developing new treatments for PD via intervention in AQP4‐mediated immune regulation.     

HSPA12B promotes functional recovery after ischaemic stroke through an eNOS‐dependent mechanism

Stroke is the leading cause of disability worldwide. HSPA12B, a heat‐shock protein recently identified expression specifically in endothelial cells, is able to promote angiogenesis. Here, we have investigated its effects on functional recovery at chronic phase of ischaemic stroke. Ischaemic stroke was induced by 60 min. of middle cerebral artery occlusion in transgenic mice with overexpression of HSPA12B (HSPA12B Tg) and wild‐type littermates (WT). HSPA12B Tg mice demonstrated a significant higher survival rate than WT mice within 28 days post‐stroke. Significant improved neurological functions, increased spontaneous locomotor activity and decreased anxiety were detected inHSPA12B Tg mice compared with WT controls within 21 days post‐stroke. Stroke‐induced hippocampal degeneration was attenuated in HSPA12B Tg mice examined at day 28 post‐stroke. Interestingly, HSPA12B Tg mice showed enhanced peri‐infarct angiogenesis (examined 28 days post‐stroke) and hippocampal neurogenesis (examined 7 days post‐stroke), respectively, compared to WT mice. The stroke‐induced eNOS phosphorylation and TGF‐β1 expression were augmented in HSPA12B Tg mice. However, administration with eNOS inhibitor L‐NAME diminished the HSPA12B‐induced protection in neurological functional recovery and mice survival post‐stroke. The data suggest that HSPA12B promoted functional recovery and survival after stroke in an eNOS‐dependent mechanism. Targeting HSPA12B expression may have a therapeutic potential for the stroke‐evoked functional disability and mortality.     

SIRT1 protects against microglia-dependent amyloid-beta toxicity through inhibiting NF-kappaB signaling

Accumulating evidence suggests that neurodegeneration induced by pathogenic proteins depends on contributions from surrounding glia. Here we demonstrate that NF-kappaB signaling in microglia is critically involved in neuronal death induced by amyloid-beta (Abeta) peptides, which are widely presumed to cause Alzheimer disease. Constitutive inhibition of NF-kappaB signaling in microglia by expression of the nondegradable IkappaBalpha superrepressor blocked neurotoxicity, indicating a pivotal role for microglial NF-kappaB signaling in mediating Abeta toxicity. Stimulation of microglia with Abeta increased acetylation of RelA/p65 at lysine 310, which regulates the NF-kappaB pathway. Overexpression of SIRT1 deacetylase and the addition of the SIRT1 agonist resveratrol markedly reduced NF-kappaB signaling stimulated by Abeta and had strong neuroprotective effects. Our results support a glial loop hypothesis by demonstrating a critical role for microglial NF-kappaB signaling in Abeta-dependent neurodegeneration. They also implicate SIRT1 in this pathway and highlight the therapeutic potential of resveratrol and other sirtuin-activating compounds in Alzheimer disease.     

Partial depletion and repopulation of microglia have different effects in the acute MPTP mouse model of Parkinson’s disease

ObjectivesParkinson''s disease (PD) is a common neurodegenerative disorder characterized by the progressive and selective degeneration of dopaminergic neurons. Microglial activation and neuroinflammation are associated with the pathogenesis of PD. However, the relationship between microglial activation and PD pathology remains to be explored.Materials and MethodsAn acute regimen of MPTP was administered to adult C57BL/6J mice with normal, much reduced or repopulated microglial population. Damages of the dopaminergic system were comprehensively assessed. Inflammation‐related factors were assessed by quantitative PCR and Multiplex immunoassay. Behavioural tests were carried out to evaluate the motor deficits in MPTP‐challenged mice.ResultsThe receptor for colony‐stimulating factor 1 inhibitor PLX3397 could effectively deplete microglia in the nigrostriatal pathway of mice via feeding a PLX3397‐formulated diet for 21 days. Microglial depletion downregulated both pro‐inflammatory and anti‐inflammatory molecule expression at baseline and after MPTP administration. At 1d post‐MPTP injection, dopaminergic neurons showed a significant reduction in PLX3397‐fed mice, but not in control diet (CD)‐fed mice. However, partial microglial depletion in mice exerted little effect on MPTP‐induced dopaminergic injuries compared with CD mice at later time points. Interestingly, microglial repopulation brought about apparent resistance to MPTP intoxication.ConclusionsMicroglia can inhibit PD development at a very early stage; partial microglial depletion has little effect in terms of the whole process of the disease; and microglial replenishment elicits neuroprotection in PD mice.     

Inhibition of p38 pathway-dependent MPTP-induced dopaminergic neurodegeneration in estrogen receptor alpha knockout mice

Approximately, 7–10 million people in the world suffer from Parkinson's disease (PD). Recently, increasing evidence has suggested the protective effect of estrogens against nigrostriatal dopaminergic damage in PD. In this study, we investigated whether estrogen affects 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced behavioral impairment in estrogen receptor alpha (ERα)-deficient mice. MPTP (15 mg/kg, four times with 1.5-h interval)-induced dopaminergic neurodegeneration was evaluated in ERα wild-type (WT) and knockout (KO) mice. Larger dopamine depletion, behavioral impairments (Rotarod test, Pole test, and Gait test), activation of microglia and astrocytes, and neuroinflammation after MPTP injection were observed in ERα KO mice compared to those in WT mice. Immunostaining for tyrosine hydroxylase (TH) after MPTP injection showed fewer TH-positive neurons in ERα KO mice than WT mice. Levels of dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC, metabolite of dopamine) were also lowered in ERα KO mice after MPTP injection. Interestingly, a higher immunoreactivity for monoamine oxidase (MAO) B was found in the substantia nigra and striatum of ERα KO mice after MPTP injection. We also found an increased activation of p38 kinase (which positively regulates MAO B expression) in ERα KO mice. In vitro estrogen treatment inhibited neuroinflammation in 1-methyl-4-phenyl pyridium (MPP +)-treated cultured astrocyte cells; however, these inhibitory effects were removed by p38 inhibitor. These results indicate that ERα might be important for dopaminergic neuronal survival through inhibition of p38 pathway.     

Cdk5 suppression blocks SIRT1 degradation via the ubiquitin-proteasome pathway in Parkinson's disease models

The NAD⁺-dependent protein deacetylase sirtuin 1 (SIRT1), a member of the sirtuin family, may have a neuroprotective effect in multiple neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's disease (PD) and Amyotrophic lateral sclerosis (ALS). Many studies have suggested that overexpression-induced or resveratrol-treated activation of SIRT1 could significantly ameliorate several neurodegenerative diseases in mouse models. However, the type of SIRT1, protein expression levels and underlying mechanisms remain unclear, especially in PD. In this study, the results demonstrated that SIRT1 knockout markedly worsened the movement function in MPTP-lesioned animal model of PD. SIRT1 expression was found to be markedly decreased not only in environmental factor PD models, neurotoxin MPP⁺-treated primary culture neurons and MPTP-induced mice but also in genetic factor PD models, overexpressed α-synuclein-A30PA53T SH-SY5Y stable cell line and hm²α-SYN-39 transgenic mouse strain. Importantly, the degradation of SIRT1 during MPP⁺ treatment was mediated by the ubiquitin-proteasome pathway. Furthermore, the results indicated that cyclin-dependent kinase 5 (Cdk5) was also involved in the decrease of SIRT1 expression, which could be efficiently blocked by the inhibition of Cdk5. In conclusion, our findings revealed that the Cdk5-dependent ubiquitin-proteasome pathway mediated degradation of SIRT1 plays a vital role in the progression of PD.     

Preventive role of PD‐1 on MPTP‐induced dopamine depletion in mice

Many current studies of Parkinson's disease (PD) suggest that inflammation is involved in the neurodegenerative process. PD‐1, a traditional Korean medicine, used to treat various brain diseases in Korea. This study was designed to investigate the effect of PD‐1 extract in the Parkinson's model of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) lesioned mice. The MPTP administration caused the dopamine neuron loss in the striatum and substantia nigra pars compacta (SNpc), which was demonstrated by a depletion of tyrosine hydroxylase (TH). In addition, a reduction of bcl‐2 expression with elevation of bax expression, caspase‐3 activation, and release of cytochrome c into cytosol in dopaminergic neurons of SNpc were noted. Oral administration of PD‐1 extract (50 and 100 mg kg^?1) attenuated the MPTP‐induced depletion of TH proteins in the striatum and SNpc and prevented the apoptotic effects. These results indicate that PD‐1 extract is able to protect dopaminergic neurons from MPTP‐induced neuronal death, with important implications for the treatment of PD. Copyright © 2010 John Wiley & Sons, Ltd.     

Nei-like 1 inhibition results in motor dysfunction and promotes inflammation in Parkinson’s disease mice model

Parkinson’s disease (PD) is well known as a neurodegenerative disorder with progressive loss of dopaminergic (DA) neurons. Nei-like 1 (NEIL1) is one of four mammalian DNA glycosylases involved in the progression of various diseases, including neuroinflammation. However, it is still unknown if the expression changes of NEIL1 could contribute to PD progression. In the present study, we established mouse model with PD using 6-hydroxydopamine (6-OHDA) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to explore the effects of NEIL1 on PD development. Here, we found that NEIL1 deletion significantly promoted the motor dysfunction in the wild type mice treated with 6-OHDA. Furthermore, DA neuronal loss was further accelerated by NEIL1 deletion in 6-OHDA-injected mice, as evidenced by the significantly reduced expression of tyrosine hydroxylase (TH) and dopamine transporter (DAT). Furthermore, in PD mice induced by MPTP, remarkably reduced expression of NEIL1 was observed in nigra and striatum of mice. A strong positive correlation was detected in the expression of NEIL1 and the survival rate of DA neurons. Also, NEIL1 ablation further elevated the DA neuronal loss in MPTP-treated mice, accompanied with higher glial activation, as evidenced by the obvious up-regulation of glial fibrillary acidic protein (GFAP) and Ionized calcium-Binding Adapter molecule 1 (Iba1). Moreover, MPTP-triggered inflammation was highly aggravated by the loss of NEIL1 through inducing the expression of pro-inflammatory cytokines and chemokines. In contrast, promoting NEIL1 expression effectively reversedPD progression induced by MPTP in mice. Together, these results demonstrated that NEIL1 insufficiency might be a contributing factor for the progression of PD, which therefore could be considered as a novel candidate to develop effective treatments against PD progression.     

Loss of Allograft Inflammatory Factor-1 Ameliorates Experimental Autoimmune Encephalomyelitis by Limiting Encephalitogenic CD4 T-Cell Expansion

Experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis (MS), is mediated by myelin-specific autoreactive T cells that cause inflammation and demyelination in the central nervous system (CNS), with significant contributions from activated microglia and macrophages. The molecular bases for expansion and activation of these cells, plus trafficking to the CNS for peripheral cells, are not fully understood. Allograft inflammatory factor-1 (Aif-1) (also known as ionized Ca²⁺ binding adapter-1 [Iba-1]) is induced in leukocytes in MS and EAE; here we provide the first assessment of Aif-1 function in this setting. After myelin oligodendrocyte glycoprotein peptide (MOG_35–55) immunization, Aif-1–deficient mice were less likely than controls to develop EAE and had less CNS leukocyte infiltration and demyelination; their spinal cords contained fewer CD4 T cells and microglia and more CD8 T cells. These mice also showed significantly less splenic CD4 T-cell expansion and activation, plus decreased proinflammatory cytokine expression. These findings identify Aif-1 as a potent molecule that promotes expansion and activation of CD4 T cells, plus elaboration of a proinflammatory cytokine milieu, in MOG_35–55-induced EAE and as a potential therapeutic target in MS.     

Dexmedetomidine protects against degeneration of dopaminergic neurons and improves motor activity in Parkinson's disease mice model

Parkinson’s disease (PD) is the result of dopaminergic (DA) neuronal death in the substantianigra pars compacta (SNc). Current treatments for PD such as L-dopa are limited in effectiveness and fail to address the cause. Targeted anti-inflammatory therapies, particularly directed at nuclear factor kappa B (NF‐κB) activity in alleviating degeneration of DA-neurons is of evolving interest. In the present study, we hypothesised that dexmedetomidine (DEX), an alpha-2 receptor adrenergic agonist, suppress the inflammatory responses associated with PD and restores dopaminergic levels by alleviating substantia nigral degeneration. Male mice (C57Bl/10, 8–11 months old and of 34–40 g of weight) were divided into: the control, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and MPTP + dexmedetomidine (MPTP + DEX) (n = 26 each group). Dex restored dopamine levels in SNpc of MPTP-induced PD mice model. Results of immunohisto staining revealed that Dex treatment post-MPTP induction restored TH-positive cells, with only 12.37% increase (^##p < 0.01 vs MPTP) on the third day and a steep 55% increase (^###p < 0.001 vs MPTP) following the seventh day of Dex treatment. Moreover, the expressions of proinflammatory markers regulated by NF-κB were diminished in Dex + MPTP group. In addition, cylinder test revealed that Dex treatment improved asymmetric limb usage pattern in MPTP induced mice over the course of 7 days. Hence, in this study, we provided insight on the effect of Dex in the inhibition of NF-κB1 regulated proinflammatory mediators to improve dopamine levels and reduce SNpc dopaminergic neuronal degeneration.