Endothelin‐converting enzyme‐1 in Alzheimer's disease and vascular dementia

J. C. Palmer, P. G. Kehoe and S. Love (2010) Neuropathology and Applied Neurobiology 36, 487–497
Endothelin‐converting enzyme‐1 in Alzheimer's disease and vascular dementia Aims: Alzheimer's disease (AD) is believed to be caused by the accumulation of amyloid beta (Aβ) peptide within the brain. Endothelin‐converting enzyme‐1 and 2 (ECE‐1 and ECE‐2) are expressed in endothelial cells and neurones, respectively, and both cleave ‘big endothelin’ to produce the vasoconstrictor endothelin‐1 (ET‐1). ECE‐1 and ECE‐2 also degrade Aβ. AD patients have regionally reduced microvascular blood flow in the brain, with impaired endothelium‐dependent relaxation and cerebrovascular autoregulation, and abnormal production of ET‐1 has been demonstrated in mice overexpressing amyloid precursor protein. We recently found ECE‐2 mRNA and protein to be elevated in the brain in AD. In vitro, expression of ECE‐2 was upregulated by Aβ. Our aims for this study were to examine expression of ECE‐1 (which has 57% homology with ECE‐2) in temporal cortex from patients with AD, vascular dementia (VaD) and controls. Methods: We examined the distribution of ECE‐1 with immunohistochemistry, and measured ECE‐1 mRNA by real‐time polymerase chain reaction (PCR). ECE‐1 protein levels were measured by western blot, and results analysed before and after adjustment for factor VIII‐related antigen. Results: We showed ECE‐1 to be in vascular endothelial cells. We did not find significant differences in ECE‐1 mRNA or protein levels (either full‐length ECE‐1 or the soluble spliced variant, ECE‐1sv) in AD or VaD compared with controls. Conclusions: Our findings suggest that any disease‐specific contribution of ECE‐1 to the accumulation of Aβ or reduction in local microvascular blood flow in AD or VaD is probably small, with abnormal production of ET‐1 being more likely to reflect Aβ‐mediated upregulation of ECE‐2.     

Chronic thrombin exposure results in an increase in apolipoprotein-E levels

Studies have shown that individuals with both a history of traumatic brain injury and inheritance of apolipoprotein E-4 (ApoE4) allele are associated with a poor neurologic outcome and an increased risk for Alzheimer's disease. We assessed the hypothesis that thrombin released during brain injury causes an increase in apolipoprotein-E levels and such increase in the levels of apolipoprotein-E4 isoform may have amyloidogenic effects. Rats received either thrombin (100 nm, 0.25 microl/hr, 28 days) or vehicle via intracerebroventricular (i.c.v.) infusion. Thrombin treatment increased apolipoprotein-E levels in hippocampus as compared to vehicle treatment (P < 0.001). Infusion of human apolipoprotein-E4 (0.6 ng/hr, i.c.v., 56 days) into rats resulted in beta-amyloid deposition and increased the number of GFAP-positive astrocytes. ApoE4 infusion also resulted in significant spatial memory deficits. These findings suggest that thrombin released during brain injury may contribute to an increase in apolipoprotein-E levels. Such increase in Apolipoprotein-E4 isoform facilitates beta-amyloid deposition and cognitive deficits.     

Metabolic approach of absence seizures in a genetic model of absence epilepsy, the GAERS: study of the leucine-glutamate cycle.

We suggest that a dysregulation of energy metabolism in the brain of genetic absence epilepsy rats from Strasbourg (GAERS) could create a specific cerebral environment that would favor the expression of spike-and-wave discharges (SWD) in the thalamocortical loop, largely dependent on glutamatergic and gamma-aminobutyric acid (GABA)-ergic neurotransmissions. We tested several aspects of metabolic activity in the brain of GAERS compared to a genetic strain of nonepileptic (NE) rats. Glucose metabolism was higher in all brain regions of GAERS compared to those of NE rats along the whole glycolytic and aerobic pathways, as assessed by regional histochemical measurement of lactate dehydrogenase and cytochrome oxidase activities. Branched-chain amino acids (BCAA) and alpha-ketoisocaproate (alpha-KIC), the ketoacid of leucine, when injected intraperitoneally, increased the number of SWD in GAERS but had only a slight effect on their duration. These data speak in favor of a BCAA- or alpha-KIC-induced change in neuronal excitability. Leucine and alpha-KIC decreased the concentration of glutamate in thalamus and cortex without affecting GABA concentrations. Thus, BCAA and alpha-KIC, by decreasing glutamatergic neurotransmission, could favor GABAergic neurotransmission, which is known to increase the occurrence of seizures in GAERS. Finally, the transport of [1-(14)C]alpha-KIC in freshly isolated cortical neurons was lower in GAERS than in NE rats, and this difference was shown to be of metabolic origin. The addition of gabapentin, a specific inhibitor of BCAA transaminase (BCAT), reduced the transport of [1-(14)C]alpha-KIC in GAERS and NE rats to a level that became identical in both strains. This strain-dependent change was not related to a difference in the activity of BCAT, which was identical in GAERS and NE rats. The exact origin of this apparent metabolic dysregulation of energy metabolism in GAERS that could underlie the origin of seizures in that strain remains to be explored further.     

Long‐term effects of cognitive rehabilitation on brain,functional outcome and cognition in Parkinson's disease

Background and purpose

Cognitive rehabilitation has demonstrated efficacy in producing short‐term cognitive and brain changes in patients with Parkinson's disease (PD). To date, no study has assessed the long‐term effects of cognitive rehabilitation using neuroimaging techniques in PD. The aim was to assess the longitudinal effects of a 3‐month cognitive rehabilitation programme evaluating the cognitive, behavioural and neuroimaging changes after 18 months.

Methods

Fifteen patients with PD underwent a cognitive, behavioural and neuroimaging assessment at pre‐treatment (T₀), post‐treatment (T₁) and after 18 months (T₂). This study examined the long‐term effects (from T₀ to T₂) and the maintenance of the changes (from T₁ to T₂). T1‐weighted, diffusion‐weighted, functional magnetic resonance imaging during both a resting‐state and a memory paradigm were acquired. Voxel‐based morphometry and tract‐based spatial statistics were used for grey and white matter analyses. A region‐of‐interest‐to‐region‐of‐interest approach was used for resting‐state functional connectivity (FC) and a model‐based approach was used for brain activation during the memory paradigm.

Results

Patients with PD showed increased cognitive performance, decreased functional disability, increased brain FC and activation at T₂ compared with T₀ (P < 0.05, FDR). Moreover, patients showed maintenance of the improvements in cognition and functionality, and maintenance of the increased brain FC and activation at T₂ compared with T₁. However, significant grey matter reduction and alterations of white matter integrity were found at T₂ (P < 0.05, FWE).

Conclusions

Findings suggest that the improved cognitive performance and increased brain FC and activation after cognitive rehabilitation were significantly maintained after 18 months in patients with PD, despite the structural brain changes, consistent with a progression of neurodegenerative processes.     

In utero bacterial endotoxin exposure causes loss of tyrosine hydroxylase neurons in the postnatal rat midbrain.

We investigated whether in utero exposure to the Gram(-) bacteriotoxin lipopolysaccharide (LPS) induces dopamine (DA) neuron loss in rats. The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) kills DA neurons and is elevated in the brains of patients with Parkinson's disease (PD). LPS is a potent inducer of TNF-alpha, and both are increased in the chorioamniotic environment of women who have bacterial vaginosis (BV) during pregnancy, suggesting that BV might interfere with the normal development of fetal DA neurons. Gravid female rats were injected intraperitoneally with either LPS or normal saline at embryonic day 10.5 and their pups were killed at postnatal day 21. The brains of the pups were assessed for DA and TNF-alpha levels and DA cell counts in the mesencephalon using tyrosine hydroxylase immunoreactive (THir) cells as a DA neuron marker. Prenatal LPS exposure significantly reduced striatal DA (29%) and increased DA activity (72%) as well as TNF-alpha (101%). Stereological cell counts in the mesencephalon were also significantly reduced (27%) by prenatal LPS exposure. Prenatal exposure to LPS, as might occur in humans with BV, produces a significant loss of THir cells in rats that is still present 33 days following a single injection of LPS. Since this cell loss is well past the normal phase of DA neuron apoptosis that occurs in early postnatal life, rats so exposed may have a permanent loss of DA neurons, suggesting that prenatal infections may represent risk factors for PD.     

Lysosomal trafficking defects link Parkinson's disease with Gaucher's disease

Lysosomal dysfunction has been implicated in multiple diseases, including lysosomal storage disorders such as Gaucher's disease, in which loss‐of‐function mutations in the GBA1 gene encoding the lysosomal hydrolase β‐glucocerebrosidase result in lipid substrate accumulation. In Parkinson's disease, α‐synuclein accumulates in Lewy bodies and neurites contributing to neuronal death. Previous clinical and genetic evidence has demonstrated an important link between Parkinson's and Gaucher's disease, as GBA1 mutations and variants increase the risk of Parkinson's and Parkinson's patients exhibit decreased β‐glucocerebrosidase activity. Using human midbrain neuron cultures, we have found that loss of β‐glucocerebrosidase activity promotes α‐synuclein accumulation and toxicity, whereas α‐synuclein accumulation further contributes to decreased lysosomal β‐glucocerebrosidase activity by disrupting β‐glucocerebrosidase trafficking to lysosomes. Moreover, α‐synuclein accumulation disrupts trafficking of additional lysosomal hydrolases, further contributing to lysosomal dysfunction and neuronal dyshomeostasis. Importantly, promoting β‐glucocerebrosidase activity reduces α‐synuclein accumulation and rescues lysosomal and neuronal dysfunction, suggesting that β‐glucocerebrosidase may be an important therapeutic target for advancing drug discovery in synucleinopathies including Parkinson's disease. © 2016 International Parkinson and Movement Disorder Society.     

Effect of age on the duration and extent of amyloid plaque reduction and microglial activation after injection of anti-Abeta antibody into the third ventricle of TgCRND8 mice

We have previously shown that anti beta-amyloid (Abeta) antibody injected into the third ventricle of mice is distributed throughout the brain within 24 hr and is completely washed out of brain within 36 hr after injection and that, in Tg2576 animals, a single injection of antibody reduces cerebral Abeta and restores presynaptic deficits 1 month after injection without producing hemorrhage or inflammation at an early plaque stage. Here we report the effects of a single ICV injection of anti-Abeta antibody on cerebral levels of immunoreactive Abeta and of microglial activation measured by immunoreactive interleukin-1beta (IL-1beta) at 1, 4, and 8 weeks after injections in TgCRND8 mice at two ages, 2 months (sparse plaques) and 8 months (abundant plaques). The data show that parenchymal amyloid accumulates before cerebral microvascular amyloid and that a single ICV injection reduces only parenchymal amyloid by about 70%, without affecting vascular amyloid, and reduces microglial activation by 46-60% at 1 week after injection. The reappearance of plaques after antibody injection takes 4-8 weeks, whereas plaque-associated focal microglial activation begins increasing between 1 and 4 weeks, suggesting that accumulation of nonfibrillar oligomeric Abeta may account for the earlier onset of microglial activation. No perivascular hemorrhage or inflammation was observed. These results suggest that periodic intraventricular administration of anti-Abeta is a potentially useful method for rapid reduction of both preexisting amyloid load and associated inflammation, providing a window of 4 weeks' duration for possible pharmacological cotreatment(s) to prevent de novo Abeta formation. This ICV method of passive immunization may be safer than active immunization, which has been known to produce encephalitis, or systemic passive immunization, which exposes amyloid-laden cerebral microvasculature to high levels of antibody in the blood and the potential of perivascular hemorrhages.     

Aggresome-related biogenesis of Lewy bodies

Neurodegenerative disorders such as Parkinson's disease (PD) and 'dementia with Lewy bodies' (DLB) are characterized pathologically by selective neuronal death and the appearance of intracytoplasmic protein aggregates (Lewy bodies). The process by which these inclusions are formed and their role in the neurodegenerative process remain elusive. In this study, we demonstrate a close relationship between Lewy bodies and aggresomes, which are cytoplasmic inclusions formed at the centrosome as a cytoprotective response to sequester and degrade excess levels of potentially toxic abnormal proteins within cells. We show that the centrosome/aggresome-related proteins gamma-tubulin and pericentrin display an aggresome-like distribution in Lewy bodies in PD and DLB. Lewy bodies also sequester the ubiquitin-activating enzyme (E1), the proteasome activators PA700 and PA28, and HSP70, all of which are recruited to aggresomes for enhanced proteolysis. Using novel antibodies that are specific and highly sensitive to ubiquitin-protein conjugates, we revealed the presence of numerous discrete ubiquitinated protein aggregates in neuronal soma and processes in PD and DLB. These aggregates appear to be being transported from peripheral sites to the centrosome where they are sequestered to form Lewy bodies in neurons. Finally, we have shown that inhibition of proteasomal function or generation of misfolded proteins cause the formation of aggresome/Lewy body-like inclusions and cytotoxicity in dopaminergic neurons in culture. These observations suggest that Lewy body formation may be an aggresome-related event in response to increasing levels of abnormal proteins in neurons. This phenomenon is consistent with growing evidence that altered protein handling underlies the etiopathogenesis of PD and related disorders.     

Human apolipoprotein E ?4 expression impairs cerebral vascularization and blood–brain barrier function in mice

Human apolipoprotein E (APOE) exists in three isoforms ɛ2, ɛ3, and ɛ4, of which APOE4 is the main genetic risk factor of Alzheimer''s disease (AD). As cerebrovascular defects are associated with AD, we tested whether APOE genotype has an impact on the integrity and function of the blood–brain barrier (BBB) in human APOE-targeted replacement mice. Using the quantitative in situ brain perfusion technique, we first found lower (13.0% and 17.0%) brain transport coefficient (Clup) of [³H]-diazepam in APOE4 mice at 4 and 12 months, compared with APOE2 and APOE3 mice, reflecting a decrease in cerebral vascularization. Accordingly, results from immunohistofluorescence experiments revealed a structurally reduced cerebral vascularization (26% and 38%) and thinner basement membranes (30% and 35%) in 12-month-old APOE4 mice compared with APOE2 and APOE3 mice, suggesting vascular atrophy. In addition, APOE4 mice displayed a 29% reduction in [³H]-d-glucose transport through the BBB compared with APOE2 mice without significant changes in the expression of its transporter GLUT1 in brain capillaries. However, an increase of 41.3% of receptor for advanced glycation end products (RAGE) was found in brain capillaries of 12-month-old APOE4 mice. In conclusion, profound divergences were observed between APOE genotypes at the cerebrovascular interface, suggesting that APOE4-induced BBB anomalies may contribute to AD development.     

Short‐term effect of combined drug therapy and cognitive stimulation therapy on the cognitive function of Alzheimer's disease

Background: Acetylcholinesterase inhibitors (i.e. donepezil) are known to benefit Alzheimer's disease (AD) patients. However, the combined effects of acetylcholinesterase and cognitive stimulation therapy (CST) are still debated. The present study examined their combined effects on the progression of cognitive decline in AD. Methods: The present study was a non‐randomized controlled study and included two groups of patients with AD (i.e. CST group and control group). The CST group consisted of 31 patients with AD who received donepezil and weekly, 30‐min CST sessions over the course of 7 weeks. The control group consisted of 18 patients who received only donepezil. Changes in cognitive abilities were assessed with Hasegawa's Dementia Scale‐Revised (HDS‐R) and were statistically analyzed by repeated‐measure analysis of variance (anova ). Results: ANOVA showed a significant group × time interaction effect on the HDS‐R score. HDS‐R scores for the CST group increased significantly during the intervention period, whereas the scores for the control group did not increase. Differences between the means of pre‐ and post‐test HDS‐R scores were significantly different between the groups; scores were significantly higher for the CST group than the control group. The groups differed significantly in the proportion of subjects whose score increased by more than four points on the HDS‐R (Fisher's exact test, P < 0.05; 8 patients (25.8%) in the CST group and none (0.0%) in the control group). Conclusions: These results suggest that CST is one of the important non‐pharmacological treatment strategies for patients with AD.     

Microglial phagocytosis is enhanced by monomeric alpha-synuclein, not aggregated alpha-synuclein: implications for Parkinson's disease

Gathering evidence has associated activation of microglia with the pathogenesis of numerous neurodegenerative diseases of the central nervous system (CNS) such as Alzheimer's disease and Parkinson's disease. Microglia are the resident macrophages of the CNS whose functions include chemotaxis, phagocytosis, and secretion of a variety of cytokines and proteases. In this study, we examined the possibility that alpha-synuclein (alpha-syn), which is associated with the pathogenesis of Parkinson's disease, may affect the phagocytic function of microglia. We found that extracellular monomeric alpha-syn enhanced microglial phagocytosis in both a dose- and time-dependent manner, but beta- and gamma- syn did not. We also found that the N-terminal and NAC region of alpha-syn, especially the NAC region, might be responsible for the effect of alpha-syn on microglial phagocytosis. In contrast to monomeric alpha-syn, aggregated alpha-syn actually inhibited microglial phagocytosis. The different effects of monomeric and aggregated alpha-syn on phagocytosis might be related to their localization in cells. This study indicates that alpha-syn can modulate the function of microglia and influence inflammatory changes such as those seen in neurodegenerative disorders.     

Cell therapy for central nervous system disorders: Current obstacles to progress

Cell therapy for disorders of the central nervous system has progressed to a new level of clinical application. Various clinical studies are underway for Parkinson's disease, stroke, traumatic brain injury, and various other neurological diseases. Recent biotechnological developments in cell therapy have taken advantage of the technology of induced pluripotent stem (iPS) cells. The advent of iPS cells has provided a robust stem cell donor source for neurorestoration via transplantation. Additionally, iPS cells have served as a platform for the discovery of therapeutics drugs, allowing breakthroughs in our understanding of the pathology and treatment of neurological diseases. Despite these recent advances in iPS, adult tissue‐derived mesenchymal stem cells remain the widely used donor for cell transplantation. Mesenchymal stem cells are easily isolated and amplified toward the cells' unique trophic factor‐secretion property. In this review article, the milestone achievements of cell therapy for central nervous system disorders, with equal consideration on the present translational obstacles for clinic application, are described.     

Review: Parkinson's disease: from synaptic loss to connectome dysfunction

Parkinson's disease (PD) is a common neurodegenerative disorder with prominent loss of nigro‐striatal dopaminergic neurons. The resultant dopamine (DA) deficiency underlies the onset of typical motor symptoms (MS). Nonetheless, individuals affected by PD usually show a plethora of nonmotor symptoms (NMS), part of which may precede the onset of motor signs. Besides DA neuron degeneration, a key neuropathological alteration in the PD brain is Lewy pathology. This is characterized by abnormal intraneuronal (Lewy bodies) and intraneuritic (Lewy neurites) deposits of fibrillary aggregates mainly composed of α‐synuclein. Lewy pathology has been hypothesized to progress in a stereotypical pattern over the course of PD and α‐synuclein mutations and multiplications have been found to cause monogenic forms of the disease, thus raising the question as to whether this protein is pathogenic in this disorder. Findings showing that the majority of α‐synuclein aggregates in PD are located at presynapses and this underlies the onset of synaptic and axonal degeneration, coupled to the fact that functional connectivity changes correlate with disease progression, strengthen this idea. Indeed, by altering the proper action of key molecules involved in the control of neurotransmitter release and re‐cycling as well as synaptic and structural plasticity, α‐synuclein deposition may crucially impair axonal trafficking, resulting in a series of noxious events, whose pressure may inevitably degenerate into neuronal damage and death. Here, we provide a timely overview of the molecular features of synaptic loss in PD and disclose their possible translation into clinical symptoms through functional disconnection.     

Plasma α‐synuclein in patients with Parkinson's disease with and without treatment

Alpha‐synuclein (α‐syn) is an intracellular protein with a high tendency to aggregation. It is the major component of Lewy bodies and may play a key role in the pathogenesis of Parkinson's disease (PD). α‐Syn is also released by neurons and can be detected in biological fluids, such as plasma. The purpose of this study was to determine whether plasma α‐syn concentrations are elevated in newly diagnosed PD patients before treatment (nontreated PD group, ntPD; n = 53) and to compare them with concentrations in PD patients with at least 1 year of specific treatment (tPD; n = 42) and in healthy controls (n = 60). Plasma α‐syn concentrations in the ntPD and tPD groups were similar and significantly higher than in healthy controls. In conclusion, α‐syn was elevated early in the development of PD and specific PD treatment did not change plasma α‐syn levels. © 2010 Movement Disorder Society