Patrick Pompl

Diet-induced insulin resistance promotes amyloidosis in a transgenic mouse model of Alzheimer's disease

Recent epidemiological evidence indicates that insulin resistance, a proximal cause of Type II diabetes [a non‐insulin dependent form of diabetes mellitus (NIDDM)], is associated with an increased relative risk for Alzheimers disease (AD). In this study we examined the role of dietary conditions leading to NIDDM‐like insulin resistance on amyloidosis in Tg2576 mice, which model AD‐like neuropathology. We found that diet‐induced insulin resistance promoted amyloidogenic β‐amyloid (Aβ) Aβ1–40 and Aβ1–42 peptide generation in the brain that corresponded with increased γ‐secretase activities and decreased insulin degrading enzyme (IDE) activities. Moreover, increased Aβ production also coincided with increased AD‐type amyloid plaque burden in the brain and impaired performance in a spatial water maze task. Further exploration of the apparent interrelationship of insulin resistance to brain amyloidosis revealed a functional decrease in insulin receptor (IR)‐mediated signal transduction in the brain, as suggested by decreased IR β‐subunit (IRβ) Y1162/1163 autophosphorylation and reduced phosphatidylinositol 3 (PI3)‐kinase/pS473‐AKT/Protein kinase (PK)‐B in these same brain regions. This latter finding is of particular interest given the known inhibitory role of AKT/PKB on glycogen synthase kinase (GSK)‐3α activity, which has previously been shown to promote Aβ peptide generation. Most interestingly, we found that decreased pS21‐GSK‐3α and pS9‐GSK‐ 3β phosphorylation, which is an index of GSK activation, positively correlated with the generation of brain C‐terminal fragment (CTF)‐γ cleavage product of amyloid precursor protein, an index of γ‐secretase activity, in the brain of insulin‐resistant relative to normoglycemic Tg2576 mice. Our study is consistent with the hypothesis that insulin resistance may be an underlying mechanism responsible for the observed increased relative risk for AD neuropathology, and presents the first evidence to suggest that IR signaling can influence Aβ production in the brain.

A therapeutic role for cyclooxygenase-2 inhibitors in a transgenic mouse model of amyotrophic lateral sclerosis

Recent studies indicate that the proinflammatory enzyme cyclooxygenase (COX)‐2, an enzyme involved in inflammatory cascades but also normal neuronal activities, is elevated in the brain and spinal cord of amyotrophic lateral sclerosis (ALS) patients and ALS mouse model systems. On the basis of this evidence, we explored the impact of COX‐2 inhibition on the onset and progression of ALS‐like disease in the G93A human superoxide dismutase (SOD)1 mouse model of ALS. We found that prophylactic administration of nimesulide, a preferential COX‐2 inhibitor, in the feed resulted in a significant delay in the onset of ALS type motor impairment. This delay of ALS symptomatology temporally overlapped with the inhibition of prostaglandin E2 elevation in the spinal cord of SOD1‐G93A transgenic mice relative to untreated SOD1‐G93A controls. This study strongly supports a role for COX‐2 in the pathophysiology of ALS and provides the first experimental evidence that prophylactic treatment with COX‐2 inhibitors can significantly delay the onset of motor dysfunction in the SOD1‐G93A transgenic mouse model of ALS.

Cyclooxygenase (COX)-2 and COX-1 potentiate β-amyloid peptide generation through mechanisms that involve γ-secretase activity

In previous studies we found that overexpression of the inducible form of cyclooxygenase, COX-2, in the brain exacerbated β-amyloid (Aβ) neuropathology in a transgenic mouse model of Alzheimers disease. To explore the mechanism through which COX may influence Aβ amyloidosis, we used an adenoviral gene transfer system to study the effects of human (h)COX-1 and hCOX-2 isoform expression on Aβ peptide generation. We found that expression of hCOXs in human amyloid precursor protein (APP)-overexpressing (Chinese hamster ovary (CHO)-APPswe) cells or human neuroglioma (H4-APP751) cells resulting in 10-25 nm prostaglandin (PG)-E2 concentration in the conditioned medium coincided with an ∼1.8-fold elevation of Aβ-(1-40) and Aβ-(1-42) peptide generation and an ∼1.8-fold induction of the C-terminal fragment (CTF)-γ cleavage product of the APP, an index of γ-secretase activity. Treatment of APP-overexpressing cells with the non-selective COX inhibitor ibuprofen (1 μm, 48 h) or with the specific γ-secretase inhibitor L-685,458 significantly attenuated hCOX-1- and hCOX-2-mediated induction of Aβ peptide generation and CTF-γ cleavage product formation. Based on this evidence, we next tested the hypothesis that COX expression might promote Aβ peptide generation via a PG-E2-mediated mechanism. We found that exposure of CHO-APPswe or human embryonic kidney (HEK-APPswe) cells to PG-E2 (11-deoxy-PG-E2) at a concentration (10 nm) within the range of PG-E2 found in hCOX-expressing cells similarly promoted (∼1.8-fold) the generation of the CTF-γ cleavage product of APP and commensurate Aβ-(1-40) and Aβ-(1-42) peptide elevation. The study suggests that expression of COXs may influence Aβ peptide generation through mechanisms that involve PG-E2-mediated potentiation of γ-secretase activity, further supporting a role for COX-2 and COX-1 in Alzheimers disease neuropathology.

A role of P301L tau mutant in anti-apoptotic gene expression, cell cycle and apoptosis

In exploring the causative role of the most common Pro(301)-to-Leu (TauP301L) tau missense mutation associated with neurodegenerative tauopathies, we examined TauP301L-mediated apoptotic cell death and the expression of a cluster of genes involved in the inhibition of apoptosis (IAPs) in human neuroblastoma SH-SY5Y cells. Our research found that the expression of TauP301L, but not wild-type tau, down regulated the expression of IAPs, including survivin, which plays a role in the mitotic spindle checkpoint. The inhibition of IAPs coincided with the activation of the pro-apoptotic caspase 3, but preceded apoptotic cell death by TUNEL. Furthermore, TauP301L altered the expression of the cell cycle regulatory proteins and induced the cell cycle arrest at G(2)/M phase. Our studies demonstrate that TauP301L downregulates the expression of genes that protect against apoptosis and regulate cell cycle progression. These results suggest a novel mechanism of apoptotic cell death in TauP301L-expressing cells that involves survivin-mediated activation of cell cycle checkpoint.

High-throughput Proteomics and Protein Biomarker Discovery in an Experimental Model of Inflammatory Hyperalgesia: Effects of Nimesulide

AbstractObjective: The goal of this study was to derive a systematic approach for the identification of protein species profiles that selectively identify subgroups of similar but distinct pain models and correlate with the therapeutic efficacy (or lack thereof) of drug intervention. Methods: Using high-throughput surface-enhanced laser desorption ionization (SELDI) mass spectrometry proteomic technology based on ProteinChip arrays, we conducted a comparative analysis of the profile of protein expression in cerebral spinal fluid (CSF) from rats exposed to either injection of complete Freunds adjuvant (CFA) [a model of inflammatory pain] or chronic constriction injury (CCI) [a model of neuropathic pain]. The CFA model was then further studied for the effects of treatment with the NSAID, nimesulide (a preferential cyclo-oxygenase [COX]-2 inhibitor). Results: Among other observations, we found that the content of two metal (copper) binding protein species (2.9 and 3.2kDa) and three anionic protein species (4.0, 6.9 and 8.2kDa) were increased in the CSF of rats with inflammatory pain in a time-dependent fashion, at 7 and 14 days after CFA injection. These changes were highly selective for the CFA model, as no detectable increase in these protein biomarkers was found in the CCI neuropathic pain model. Further, we found that most of the changes in the biomarker protein species induced by the inflammatory pain were prevented by treatment with nimesulide and correlated with the antihyperalgesic effect of this drug. Conclusion: This study demonstrates that CSF biomarker profiles, as detected by SELDI technology, can consistently and reproducibly differentiate inflammatory from neuropathic pain, and reflect the analgesic action produced by the preferential COX-2 inhibitor, nimesulide. The characterisation and identification of these biomarkers will provide invaluable insight into the pathophysiology of pain mechanisms, in addition to further understanding of the value of nimesulide in the treatment of inflammatory pain.

Amyloid immunization in Alzheimer's disease: do we promote amyloid scavenging at the cost of inflammatory degeneration?

A large number of epidemiologic studies have indicated that the use of non-steroidal anti-inflammatory drugs (NSAIDs) may prevent or delay the clinical features of Alzheimer’s disease (AD) [3,8,12]. The pharmacological activity of NSAIDs is generally attributed to the inhibition of cyclooxygenase (COX), a rate-limiting enzyme in the production of prostaglandins (PGs). However, the mechanism by which these anti-inflammatory drugs, influence the clinical progression of AD dementia and/or AD neuropathology has yet to be defined. Further, evidence has indicated that COX inhibitors may also prevent AD-type amyloid (A ) neuropathology in experimental animals [6]. Interestingly, recent studies in murine models of AD have found that immunization with A peptides can mitigate A neuritic plaque pathology through the activation of the brains resident macrophages (microglia) [1,4,9,10]. Based on observations from these models, it has been demonstrated that either passive (intraperitoneal injection (IP) of pre-derived A antibodies) or active (IP injection of A peptides) immunization with A can activate pro-inflammatory microglial cells through immunoglobulin receptor signaling, and result in the clearance and degradation of A plaques [1,10]. This suggests a potential beneficial role for features of inflammation in AD. However, we note that the induction of pro-inflammatory cascades, e.g. microglial cytokine synthesis and generation of free radicals [5], is little understood and has been previously implicated in neuronal injury [13]. In humans, evidence has suggested that inflammation is integrally involved in the progression of AD dementia, and that separate segments of inflammatory cascades, may play important and possibly independent roles in different phases of the disease. For example, the induction of several cytokine inflammatory mediators, particularly those generated by microglial activation (such as IL-6 and TGF1, but not IL-1 , IL-1 , TNF and , among others), are

AN1792 vaccination immunotherapy in Alzheimer’s disease: the case of a therapy before its time

Due to increasing incidents of cerebral inflammation, Elan Corporation and American Home Products Corporation have recently announced the decision to terminate the clinical trial of AN1792 vaccine for Alzheimer’s disease (AD). Descriptive reports of the complications experienced in the Phase IIA trial of AN1792, following as little as one dose, have not been released. However, it has been reported that affected patients are recovering following anti-inflammatory drug treatment. Proponents of vaccination therapy attest that the unexpected events of the Phase IIA trial of AN1792 were part of the perceived risk in employing a novel therapeutic treatment. And in the case of treating a disorder as devastating as AD inevitably is, it may be that taking the “chance” with AN1792 was merited. This first open peer commentary to discuss vaccination therapy in AD reemphasizes the need for concerted efforts in approaching the design and initiation of AD therapeutics, and encouragingly, demonstrates the shared viewpoint that promise may lie in future therapeutic trials aimed at manipulating amyloid load in AD. However, there is evidence throughout the commentaries presented here (and in other forums), that there are many aspects of AD vaccination immunotherapy suggesting that we are not yet prepared to move forward with this therapeutic approach in humans. For example, it has been proposed that, although inflammation may be a component of AD vaccination immunotherapy, it is a short-lived phenomena and potentially integral to the eventual benefit of vaccination treatment. This theory is supported by experimental data [12], and human disease indicating, for example, that suppression of certain features of inflammation may unfavorably influence the progression of multiple sclerosis [1] (see Dr. Schenk’ commentary). This is very interesting, however, the role of inflammation associated with AD and with the vaccination-based clearance of amyloid from the

Inflammation and Cyclo-Oxygenase in Alzheimer’s Disease

There is clear evidence of inflammation in the degenerating Alzheimer’s disease (AD) brain, however, it is not presently known whether inflammatory mechanisms are a cause or consequence of this disease (Fig. 1) (1,2). The uncertainty stems from the fact that inflammatory events and pathological hallmarks of AD (such as senile plaques [SPs] and neurofibrillary tangles [NFTs]) occur in close temporal and physical proximity (3). For example, it has been shown that the majority of mature senile plaques are surrounded by reactive microglia that have the characteristics of antigen-presenting tissue macrophages, including HLA-DR surface markers, suggestive of an autoimmune response. It has been proposed that this is a remedial circumstance in which microglia are attempting to clear the amyloid debris from the neuropil. However, experimental studies show that suppression of microglia by treatment with ibuprofen (a nonsteroidal anti-inflammatory drug [NSAID]) can actually diminish the presence of amyloid deposits in the brain (4) and suggests a contradictive hypothesis. This latter hypothesis is further supported by retrospective epidemiological studies that have found decreased incidence of AD in populations who regularly take NSAIDs.