David Vergote

Proteolytic processing of SDF-1α reveals a change in receptor specificity mediating HIV-associated neurodegeneration

Proteolytic cleavage of constitutively expressed proteins can generate peptides with novel bioactive properties. Matrix metalloproteinase (MMP)-2 cleaves the 4 amino-terminal residues of the chemokine, stromal cell-derived factor (SDF)-1α, yielding a highly neurotoxic molecule, SDF(5-67), which fails to bind to its cognate receptor, CXCR4. Herein, we detected SDF(5-67) in brain monocytoid cells of HIV-infected persons, particularly in those with HIV-associated dementia. SDF(5-67) activated cell type-specific expression of proinflammatory genes including IL-1β, TNFα, indoleamine 2′,3′-dioxygenase (IDO), and IL-10 in both astrocytic and monocytoid cells (P < 0.05). Unlike SDF-1α, SDF(5-67) caused neuronal membrane perturbations with ensuing neurotoxicity and apoptosis (P < 0.05) through engagement of an inducible receptor. CXCR3 antagonists and siRNA-mediated knockdown of CXCR3 inhibited SDF(5-67)-stimulated neurophysiological changes, neuronal death, and neuroimmune activation (P < 0.05). Moreover SDF(5-67) bound directly to CXCR3 in a competitive manner, mediated by its amino terminus. In vivo neuroinflammation, neuronal loss, and neurobehavioral abnormalities caused by SDF(5-67) (P < 0.05) were prevented by a CXCR3 antagonist. These studies reveal additive neuropathogenic properties exerted by a proteolytically cleaved chemokine as consequences of a change in receptor specificity, culminating in neurodegeneration.

CXCR3 activation by lentivirus infection suppresses neuronal autophagy: neuroprotective effects of antiretroviral therapy

Previous studies have implicated CXCL12 in the neuropathogenesis of HIV infection. Proteolysis of CXCL12 generates a neurotoxic molecule, CXCL12(5‐67), which engages and activates CXCR3, in addition to exhibiting increased expression in the brains of patients with HIV‐associated dementia (HAD). Herein, we investigated CXCR3‐mediated neuronal injury, particularly, its contribution to autophagy suppression and the concomitant effects of antiretroviral therapy using human brain samples and models of HIV neuropathogenesis. Neurons in the brains of HAD patients and feline immunodeficiency virus (FlV)‐infected animals, as well as cultured human neurons, expressed CXCR3, which was modulated in a ligand‐specific manner. Exposure of human neurons to CXCL12(5‐67) caused a reduction in the autophagy‐associated molecule LC3 (P<0.05) and neuronal survival (P<0.05), which recapitulated findings in FIV‐ and HIV‐infected brains (P<0.05). Oral didanosine (ddI) treatment of FIV‐infected animals reduced neurobehavioral abnormalities in conjunction with diminished plasma viral load (P<0.05). F4/80 transcript abundance and CXCL12(5‐67) immunoreactivity were reduced with restored neuronal LC3 expression in the brains of FIV‐infected animals after ddI treatment (P<0.05). ddI treatment also prevented microglial activation and depletion of synaptic proteins in the cortex of FIV‐infected animals (P<0.05). These findings indicate that the beneficial effects of ddI might be a consequence of a reduced systemic viral burden and concurrent leukocyte activation, leading to diminished neuroinflammation with preservation of neuronal autophagy by regulating CXCR3 activation.—Zhu, Y., Vergote, D., Pardo, C., Noorbakhsh, F., McArthur, J. C., Hollenberg, M. D., Overall, C. M., Power, C. CXCR3 activation by lentivirus infection suppresses neuronal autophagy: neuroprotective effects of antiretroviral therapy. FASEB J. 23, 2928–2941 (2009). www.fasebj.org

Glucocorticoids regulate innate immunity in a model of multiple sclerosis: reciprocal interactions between the A1 adenosine receptor and β-arrestin-1 in monocytoid cells

Desensitization of seven transmembrane receptors (7TMRs), which are modulated by the β‐arrestins, leads to altered G protein activation. The A1 adenosine receptor (A1AR) is an antiinflammatory 7TMR exhibiting reduced expression and activity in both multiple sclerosis (MS) and the murine MS model, experimental autoimmune en‐cephalomyelitis (EAE) in monocytoid cells. Herein, we report that β‐arrestin‐1 expression was increased in brains of MS patients relative to non‐MS brains, whereas A1AR expression was concomitantly reduced. This inverse relationship between β‐arrestin‐1 and A1AR was confirmed in cultured monocytoid cells as β‐arrestin‐1 overexpression resulted in a down‐regulation of A1AR together with the internal‐ization of the surface receptor. Moreover, a physical interaction between β‐arrestin‐1 and A1AR was demonstrated in monocytoid cells. Proinflammatory cytokines regulated the A1AR/β‐arrestin‐1 interactions, while A1AR activation also modulated proinflamma‐tory cytokines expression. During EAE, β‐arrestin‐1 and A1AR expression in the spinal cord displayed a similar pattern compared to that observed in MS brains. EAE‐induced neuroinflammation and neu‐robehavioral deficits were suppressed by glucocorti‐coid treatments, accompanied by concurrent reduced β ‐arrestin‐1 and enhanced A1AR expression. Thus, the interplay between β‐arrestin‐1 and A1AR in the central nervous system during neuroinflammation represents a reciprocal regulatory mechanism through which neuroprotective therapeutic strategies for neuroinflammatory diseases might be further developed.—Tsutsui, S., Vergote, D., Shariat, N., Warren, K., Ferguson, S. S. G., Power, C. Glucocor‐ticoids regulate innate immunity in a model of multiple sclerosis: reciprocal interactions between the A1 adenosine receptor and β‐arrestin‐1 in monocytoid cells. FASEB J. 22, 786–796 (2008)

Up-regulation of Neurohemerythrin Expression in the Central Nervous System of the Medicinal Leech, Hirudo medicinalis, following Septic Injury

We report here some results of a proteomic analysis of changes in protein expression in the leech Hirudo medicinalis in response to septic injury. Comparison of two-dimensional protein gels revealed several significant differences between normal and experimental tissues. One protein found to be up-regulated after septic shock was identified, through a combination of Edman degradation, mass spectrometry, and molecular cloning, as a novel member of the hemerythrin family, a group of non-heme-iron oxygen transport proteins found in four invertebrate phyla: sipunculids, priapulids, brachiopods, and annelids. We found by in situ hybridization and immunocytochemistry that the new leech protein, which we have called neurohemerythrin, is indeed expressed in the leech central nervous system. Both message and protein were detected in the pair of large glia within the ganglionic neuropile, in the six packet glia that surround neuronal somata in each central ganglion, and in the bilateral pair of glia that separate axonal fascicles in the interganglionic connective nerves. No expression was detected in central neurons or in central nervous system microglia. Expression was also observed in many other, non-neuronal tissues in the body wall. Real-time PCR experiments suggest that neurohemerythrin is up-regulated posttranscriptionaly. We consider potential roles of neurohemerythrin, associated with its ability to bind oxygen and iron, in the innate immune response of the leech nervous system to bacterial invasion.

Role of Cathepsin D in U18666A-induced Neuronal Cell Death POTENTIAL IMPLICATION IN NIEMANN-PICK TYPE C DISEASE PATHOGENESIS

Background: Cathepsin D has been implicated in Niemann-Pick type C (NPC) disease, which is associated with intracellular cholesterol accumulation. Results: Increased cytosolic and extracellular levels of cathepsin D enhanced neuronal death via different mechanisms. Conclusion: Leakage of cathepsin D within and outside the cell can cause cell death. Significance: Cathepsin D may be involved in the degeneration of neurons in NPC pathology. Cathepsin D is an aspartyl protease that plays a crucial role in normal cellular functions and in a variety of neurodegenerative disorders, including Niemann-Pick type C (NPC) disease, which is characterized by intracellular accumulation of cholesterol and glycosphingolipids in many tissues, including the brain. There is evidence that the level and activity of cathepsin D increased markedly in vulnerable neurons in NPC pathology, but its involvement in neurodegeneration remains unclear. In the present study, using mouse hippocampal cultured neurons, we evaluated the significance of cathepsin D in toxicity induced by U18666A, a class II amphiphile, which triggers cell death by impairing the trafficking of cholesterol, as observed in NPC pathology. Our results showed that U18666A-mediated toxicity is accompanied by an increase in cathepsin D mRNA and enzyme activity but a decrease in the total peptide content. The cytosolic level of cathepsin D, on the other hand, was increased along with cytochrome c and activated caspase-3 in U18666A-treated neurons. The cathepsin D inhibitor, pepstatin A, partially protected neurons against toxicity by attenuating these signaling mechanisms. Additionally, down-regulation of cathepsin D level prevented, whereas overexpression of the protease increased, vulnerability of cultured N2a cells to U18666A-induced toxicity. We also showed that extracellular cathepsin D from U18666A-treated neurons or application of exogenous enzyme can induce neurotoxicity by activating the autophagic pathway. These results suggest that increased release/activation of cathepsin D can trigger neurodegeneration and possibly development of NPC pathology. Thus, targeting cathepsin D level/activity may provide a new therapeutic opportunity for the treatment of NPC pathology.

APOE and COMT polymorphisms are complementary biomarkers of status, stability, and transitions in normal aging and early mild cognitive impairment.

Objective: Research has reported associations among selected genetic susceptibility biomarkers and risk of (a) normal cognitive aging decrements, (b) established mild cognitive impairment (MCI), and (c) sporadic Alzheimers disease (AD). In focusing on the transitional normal-to-early MCI phase, we examine associations among three theoretically relevant polymorphisms (APOE [rs429358, rs7412], BDNF [rs6265], COMT [rs4680]) and both baseline cognitive status (MCI vs. normal aging) and two-wave (four-year) longitudinal stability or change profiles. The latter included three profiles: (a) stable as normal aging, (b) stable or chronic impairment (MCI-to-MCI), and (c) emergence of impairment (normal-to-MCI). Method: Genotyped older adults (n = 237 at baseline; age range = 64–91; 62% women) from the Victoria Longitudinal Study were examined for (a) independent and interactive associations of three genetic polymorphisms with (b) two objectively classified cognitive status groups (not-impaired controls (NIC) and MCI) at (c) both baseline and across a two-wave (four-year) longitudinal interval. Results: First, logistic regression revealed that the presence of at least one APOE ε4 allele (the risk factor for AD) was linked to greater baseline risk of objective MCI. Second, multinomial logistic regression revealed that (a) the presence of an APOE ε4 allele was associated with an increased risk of 4-year MCI status stability (chronicity), and (b) the COMT homozygous risk genotype (G/G or Val/Val) was associated with an increased risk of both MCI-to-MCI stability (chronicity) and emerging NIC-to-MCI conversion. Discussion: Both chronicity and emergence of objectively classified early cognitive impairment may be genetically heterogeneous phenomena, with influences from a panel of both normal cognitive aging (COMT) and AD-related (APOE) polymorphisms.

Synergistic associations of catechol-O-methyltransferase and brain-derived neurotrophic factor with executive function in aging are selective and modified by apolipoprotein E.

Genetic polymorphisms of catechol-O-methyltransferase (COMT) and brain-derived neurotrophic factor (BDNF) have shown promising but inconsistent linkages with executive function (EF) in normal aging. We tested (1) independent contributions of COMT and BDNF risk; (2) potential magnification by risk-related interactions or additive effects with age; and (3) effect modification through stratification by apolipoprotein E (APOE) (risk: ε4+). Multiple linear regression models were applied with nondemented older adults (N = 634; range: 53-95 years) for an EF latent variable. No independent effects of BDNF or COMT on EF were observed. Additive (but not interactive) effects of COMT, BDNF, and age showed that older adults with a high-risk allelic combination performed differentially worse. Of 2 tested models of synergistic effects, the additive approach selectively supported a magnification hypothesis, which was qualified by the presence or the absence of APOE ε4.

IDE (rs6583817) Polymorphism and Pulse Pressure are Independently and Interactively Associated with Level and Change in Executive Function in Older Adults

OBJECTIVE We report a gene × environment (health) study focusing on concurrent performance and longitudinal change in a latent-variable executive function (EF) phenotype. Specifically, we tested the independent and interactive effects of a recently identified insulin degrading enzyme genetic polymorphism (IDE rs6583817) and pulse pressure (PP; one prominent aging-related vascular health indicator) across up to 9 years of EF data in a sample of older adults from the Victoria Longitudinal Study. Both factors vary across a continuum of risk-elevating to risk-reducing and have been recently linked to normal and impaired cognitive aging. METHOD We assembled a genotyped and typically aging group of older adults (n = 599, M age = 66 years at baseline), following them for up to 3 longitudinal waves (M interval = 4.4 years). We used confirmatory factor analyses, latent growth modeling, and path analyses to pursue 3 main research goals. RESULTS First, the EF single factor model was confirmed comprising 4 executive function tasks and it demonstrated measurement invariance across the waves. Second, older adults with the major IDE G allele exhibited better EF outcomes than homozygotes for the minor A allele at the centering age of 75 years. Adults with higher PP performed more poorly on EF tasks at age 75 years and exhibited greater EF longitudinal decline. Third, gene × health interaction analyses showed that worsening vascular health (higher PP) differentially affected EF performance in older adults with the IDE G allele. CONCLUSION Genetic interaction analyses can reveal differential and magnifying effects on cognitive phenotypes in aging. In the present case, pulse pressure is confirmed as a risk factor for concurrent and changing cognitive health in aging, but the effects operate differently across the risk and protective allelic distribution of this IDE gene.

IDE (rs6583817) polymorphism and type 2 diabetes differentially modify executive function in older adults

We tested independent and interactive contributions of a recently noted and promising insulin degrading enzyme polymorphism (IDE; rs6583817) and type 2 diabetes (T2D) to executive function performance, concurrently and longitudinally. Regarding normal neurocognitive decline and Alzheimers disease, T2D is a known risk factor and this IDE variant might contribute risk or risk reduction via the minor (A) or major (G) allele. We compared normal aging and T2D groups (baseline n = 574; ages 53-95 years) over 2 longitudinal waves (mean interval = 4.4 years). We used confirmatory factor analysis, latent growth curve modeling, and path analysis. A confirmed single-factor model of 4 executive function tasks established the cognitive phenotype. This IDE variant predicted concurrent group differences and differential change in cognitive performance. Furthermore, the IDE major allele reduced risk of cognitive decline. T2D predicted performance only concurrently. Both IDE and T2D are associated with executive function levels in older adults, but only IDE moderated 2-wave change. Previously linked to Alzheimers disease, this IDE variant should be further explored for its potential influence on cognitive phenotypes of normal aging.

ApoE and Pulse Pressure Interactively Influence Level and Change in the Aging of Episodic Memory: Protective Effects Among ε2 Carriers

OBJECTIVE We tested independent and interactive effects of Apolipoprotein E (ApoE) and pulse pressure (PP) concurrently and longitudinally across 9 years (3 waves) of episodic (EM) and semantic memory (SM) data from the Victoria Longitudinal Study. METHOD We assembled a sample of older adults (n = 570, baseline M age = 71, age range = 53-95) and used latent growth modeling to test 4 research goals. RESULTS First, the best fitting memory model was 2 single latent variables for EM and SM, each exhibiting configural, metric, and partial scalar invariance. This model was analyzed as a parallel process model. Second, baseline level of PP predicted EM performance at centering age (75) and rate of 9-year EM change. Third, we observed no main effects of ApoE on EM or SM. Fourth, EM was affected by higher PP but differentially less so for carriers of the ApoE ε2 allele than the ε3 or ε4 alleles. CONCLUSIONS PP is confirmed as a risk factor for concurrent and changing cognitive health in aging, but the effects operate differently across risk and protective allelic distribution of the ApoE gene.