Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2014 Apr 1.
Published in final edited form as: Exp Mol Pathol. 2013 Jan 11;94(2):366–371. doi: 10.1016/j.yexmp.2013.01.002

Ablation of the Microglial Protein DOCK2 Reduces Amyloid Burden in a Mouse Model of Alzheimer’s Disease

Patrick J Cimino 1,*, Yue Yang 1, Xianwu Li 1, Jake F Hemingway 1, Makenzie K Cherne 1, Shawn B Khademi 1, Yoshinori Fukui 2, Kathleen S Montine 1, Thomas J Montine 1, C Dirk Keene 1
PMCID: PMC3602334  NIHMSID: NIHMS435574  PMID: 23318649

Abstract

Alzheimer’s disease (AD) neuropathology is characterized by innate immune activation primarily through prostaglandin E2 (PGE2) signaling. Dedicator of cytokinesis 2 (DOCK2) is a guanyl nucleotide exchange factor expressed exclusively in microglia in the brain and is regulated by PGE2 receptor EP2. DOCK2 modulates microglia cytokine secretion, phagocytosis, and paracrine neurotoxicity. EP2 ablation in experimental AD results in reduced oxidative damage and amyloid beta (Aβ) burden. This discovery led us to hypothesize that genetic ablation of DOCK2 would replicate the anti-Aβ effects of loss of EP2 in experimental AD. To test this hypothesis, we crossed mice that lacked DOCK2 (DOCK2−/−), were hemizygous for DOCK2 (DOCK2+/−), or that expressed two DOCK2 genes (DOCK2+/+) with APPswe-PS1Δe9 mice (a model of AD). While we found no DOCK2-dependent differences in cortex or in hippocampal microglia density or morphology in APPswe-PS1Δe9 mice, cerebral cortical and hippocampal Aβ plaque area and size were significantly reduced in 10-month-old APPswe-PS1Δe9/DOCK2−/− mice compared with APPswe-PS1Δe9/DOCK2+/+ controls. DOCK2 hemizygous APPswe-PS1Δe9 mice had intermediate Aβ plaque levels. Interestingly, soluble Aβ42 was not significantly different among the three genotypes, suggesting the effects were mediated specifically in fibrillar Aβ. In combination with earlier cell culture results, our in vivo results presented here suggest DOCK2 contributes to Aβ plaque burden via regulation of microglial innate immune function and may represent a novel therapeutic target for AD.

Keywords: innate immunity, microglia, amyloid beta, PGE2 receptor EP2, Alzheimer’s disease

INTRODUCTION

Neuroinflammation is associated with most neurodegenerative conditions, including Alzheimer’s disease (AD) (Cimino, et al., 2008; Lobsiger and Cleveland, 2007; Wyss-Coray, 2006; Zipp and Aktas, 2006). However, the mechanisms by which neuroinflammation may enhance or suppress key pathogenic steps in AD remain unclear (Halle, et al., 2008; Hickman, et al., 2008; Liang, et al., 2005; Liang, et al., 2008; Miller and Streit, 2007; Shie, et al., 2005a; Shie, et al., 2005b; Wyss-Coray, 2006). In some AD mouse models, microglia appear to act in a feed-forward fashion to promote the signature lesion of AD, Aβ plaque formation (Wyss-Coray, 2006), a process regulated in part by prostaglandin (PG) signaling (Xiang, et al., 2002). Indeed, activation of the PG pathway has been linked to neurotoxicity in a number of cell culture and in vivo models using nonsteroidal anti-inflammatory drugs (NSAIDs) to suppress cyclooxygenase (COX) isozymes (Ahmad, et al., 2008; Cimino, Keene, Breyer, Montine and Montine, 2008; Jin, et al., 2007; Manabe, et al., 2004; Shie, Breyer and Montine, 2005a; Wu, et al., 2007), and compelling epidemiological evidence supports the efficacy of NSAIDs to prevent or delay the onset of the dementia stage of AD (McGeer and McGeer, 2007). Unfortunately, results from NSAID treatment trials of symptomatic subjects were disappointing, and the only NSAIDs prevention trial was suspended because of toxicity (Meinert, et al., 2009). In combination, these results have led academic and industry laboratories to pursue more specific targets within the PG pathway that are downstream of COX (Adapt Research Group, et al., 2007; Konstantinopoulos and Lehmann, 2005; Martin, et al., 2008). One such target is the PGE2 receptor subtype 2 (EP2) that has pro-inflammatory, pro-oxidative, and anti-phagocytic activity in mouse brain or primary microglial cultures (Cimino, Keene, Breyer, Montine and Montine, 2008; Echeverria, et al., 2005; Liang, Wang, Hand, Wu, Breyer, Montine and Andreasson, 2005; Liang, Wang, Shi, Lokteva, Breyer, Montine and Andreasson, 2008; Montine, et al., 2002; Shie, et al., 2005c; Wyss-Coray, 2006). However, EP2 has widespread cellular expression, including excitatory synapses, which may lead to untoward effects when using a small molecule inhibitor (Savonenko, et al., 2009). Moreover, EP2 signaling regulates COX2 expression, at least in microglia, and may thereby lead to a similar toxicity profile as COX2-selective NSAIDs (Funk and FitzGerald, 2007; Shie, Montine, Breyer and Montine, 2005c; Yu, et al., 2006; Yu, et al., 2007).

Work with transgenic mouse models of AD has shown that NSAIDs or genetic ablation of EP2 reduces cerebral cortical and hippocampal Aβ plaque burden and the concentration of soluble Aβ peptides (Cole, et al., 2004; Liang, Wang, Hand, Wu, Breyer, Montine and Andreasson, 2005). However, these in vivo data are unclear on whether the modulation of Aβ plaques and soluble Aβ peptides in AD transgenic mice is by neuronal or microglial EP2 signaling. One approach to resolving this question would be to identify EP2-dependent microglia-specific signaling components. Indeed, we discovered that expression of dedicator of cytokinesis 2 (DOCK2) is critically dependent on EP2 activation in mouse microglia, and that DOCK2 expression in mouse and human brain localizes exclusively to microglia (Cimino, et al., 2009). Moreover, DOCK2-expressing microglia are increased in diseased regions of AD brain compared to controls, and DOCK2-expressing microglia are intimately associated with Aβ plaques (Cimino, Sokal, Leverenz, Fukui and Montine, 2009).

DOCK2 is a guanyl-nucleotide exchange factor (GEF), which positively regulates Rac-(a Rho family small GTPase) mediated cellular processes that are key elements in innate and adaptive immunity (Cimino, Sokal, Leverenz, Fukui and Montine, 2009; Gollmer, et al., 2009; Gotoh, et al., 2010; Lei, et al., 2009; Nishikimi, et al., 2009). Primary murine cerebral cortical microglia derived from DOCK2−/− or EP2−/− mice are similar in their suppressed inflammatory responses and resulting decreased paracrine neurotoxicity (Cimino, Sokal, Leverenz, Fukui and Montine, 2009; Shie, Montine, Breyer and Montine, 2005c). However, primary cerebral cortical microglia from these two knockout mice are dissimilar in at least two respects: (i) EP2−/− microglia have enhanced non-Fc-mediated phagocytosis, while DOCK2−/− microglia show approximately one-third reduction in phagocytosis, and (ii) expression of COX2 is unchanged in DOCK2−/− microglia compared to a significant reduction in EP2−/− microglia. Here we tested the hypothesis that genetic ablation of DOCK2 might replicate some of the anti-Aβ effects of ablating EP2 in the same transgenic mouse model of AD.

MATERIALS AND METHODS

Animals

DOCK2−/− mice on a C57Bl6 background (developed by Dr. Fukui (Fukui, et al., 2001)), APPswe-PS1Δe9 hemizygous mice on a C57Bl6 background (Jackson, Bar Harbor, Maine), and wild-type C57Bl6 mice were used with approval by the University of Washington Institutional Animal Care and Use Committee. In APPswe-PS1Δe9, the amyloid precursor protein (APP) Swedish mutations K594N/M595L (APPswe) and the exon-9-deleted human presenilin-1 (PS1Δe9) transgenes are coexpressed under the control of the mouse prion promoter with plaque deposition beginning at five months (Jankowsky, et al., 2001). DOCK2 and APPswe-PS1Δe9 genotypes have been previously described and were confirmed by PCR (Fukui, Hashimoto, Sanui, Oono, Koga, Abe, Inayoshi, Noda, Oike, Shirai and Sasazuki, 2001; Jankowsky, Slunt, Ratovitski, Jenkins, Copeland and Borchelt, 2001; Sanui, et al., 2003). The APPswe-PS1Δe9 hemizygous mice were crossed with DOCK2−/− mice as the parental generation to produce desired F1 generation APPswe-PS1Δe9+/−:DOCK2+/− mice. The F1 generation APPswe-PS1Δe9 +/−:DOCK2+/− mice were then bred to wild-type C57Bl6 or DOCK2−/− mice (which are also of C57Bl6 background) to produce the F2 generation genotypes of interest (verified by PCR) which included APPswe-PS1Δe9+/−:DOCK2+/+, APPswe-PS1Δe9 +/−:DOCK2+/−, and APPswe-PS1Δe9 +/−:DOCK2−/− mice. F2 generation mice were aged for a total of 10 months and then euthanized under anesthesia by transcardial perfusion of ice-cold phosphate buffered saline. The brains were removed and bisected along the parasagittal plane with one half immediately immersion-fixed in 4% paraformaldehyde. The other half was dissected on ice and the anterior and posterior cortex, hippocampus, striatum, and cerebellum removed and flash frozen in liquid nitrogen for biochemical analysis of detergent-soluble and detergent-insoluble/guanidine-soluble Aβ.

Immunofluorescence Histology and Plaque Assessment

Mouse cerebral cortex and hippocampus were prepared, stained, and analyzed as previously described (Keene, et al., 2009; Keene, et al., 2010; Quinn, et al., 2007). Briefly, after immersion fixation in 4% paraformaldehyde and cryoprotection in sucrose/glycerol, hemibrains were sectioned coronally from frontal to occipital poles into 40 μm sections using a cryostat (Leica, Buffalo Grove, IL). Every sixth section (240 μm) was used for each immunostaining paradigm. Primary antibodies included anti-Iba-1 (Wako, Richmond, VA; 1:400) and anti-β-amyloid 22-35 (Sigma, St. Louis, MO); species-appropriate secondary antibodies were conjugated to Cy3 (1:400) or Cy2 (1:400; Jackson Immunoresearch, West Grove, PA.). Prolong-Gold Anti-Fade with 4′,6-diaminido-2-phenylindole (DAPI; Invitrogen, Grand Island, NY) was used for coverslipping and nuclear counterstain. To quantify Iba-1-immunopositive microglia, sections were analyzed by using systematic random sampling (West, 1993). Every sixth brain section (240 μm apart) was analyzed by a blinded observer at 400x magnification by using a Nikon fluorescence microscope (Mellville, NY) and StereoInvestigator software (MicroBrightfield, Williston, VT). A fractionator was used to assess Iba1 immunopositive cells with a counting frame measuring 100 μm × 100 μm applied every 500 μm in hippocampus and in cortex. For plaque assessment, stained sections were scanned (Nikon Super Coolscan 4000 ED) using the same exposure and the images were analyzed separately for cerebral cortex and hippocampus using Image J software (NIH, Bethesda, MD) by a blinded observer. The percent area occupied by Aβ-immunoreactive plaques, plaque frequency, and mean plaque size were averaged over all sections for hippocampus and cortex for each mouse, and averaged values from each mouse were used in statistical analyses.

Soluble Aβ Quantitation

Quantification of detergent-soluble Aβ42 was performed using followed published procedures (Keene, Chang, Lopez-Yglesias, Shalloway, Sokal, Li, Reed, Keene, Montine, Breyer, Rockhill and Montine, 2010; Woltjer, et al., 2005; Yang, et al., 2007). Mouse brain tissue was homogenized in 10 volume of extraction buffer (10 mM Tris, 1 mM DTT, 10% Sucrose, 1 mM Ethylene glycol-bis(2-aminoethyl-ether)-N,N,N,’N,’-tetraacetic acid, 1% Triton X-100, pH7.5) by sonication. Homogenates were spun at 13, 000 rpm for 15 min, and supernatants were collected and stored at −80°C. Triton-insoluble material (pellets remaining after supernatant was removed) was then extracted with 70% formic acid. Formic acid extracts of triton-insoluble proteins were dried by vacuum centrifugation and resolubilized by sonication in 20 vol of 5 mol/L guanidine HCl, 100 mmol/L Tris, pH 7.4, with 0.002% bromphenol blue added to confirm elimination of formic acid. Aβ42 levels were determined by ELISA kit following the manufacturer’s protocol (Invitrogen) and normalized to total protein. Samples were diluted 1:20 for ELISA. Standards were provided by the manufacturer, and standard curves were generated from 15.63 to 1000 pg/ml Aβ species with a detection limit of 15.63 pg/ml. All assessments were performed blinded.

Statistical Methods

Statistical analyses were performed as described in each results section using GraphPad Prism software (Graph-Pad Software Inc., San Diego CA). All experiments were performed with at least n ≥ 5 unless otherwise specified.

RESULTS

Since PGE2 receptor EP2 ablation results in reduced Aβ in APPswe-PS1Δe9 mice, and since DOCK2 is an upstream regulator of EP2 receptor function, we hypothesized that Aβ levels would be reduced in APPswe-PS1Δe9 mice in which DOCK2 is partially or completely ablated.

DOCK2 expression localizes exclusively to microglia in brain, and since DOCK2 regulates microglia, we first determined whether DOCK2−/− mice had altered morphologic features or levels of cerebral cortical or hippocampal microglia (Figure 1). As we reported previously in wild-type (wt) mouse brain (Cimino, Sokal, Leverenz, Fukui and Montine, 2009), DOCK2−/− microglia in APPswe-PS1Δe9 mice presented usual morphologic features with predominantly ramified cells that were relatively evenly distributed in cerebral cortex and hippocampus except in areas with Aβ plaques (confirmed with Aβ co-immunofluorescence). As expected, microglia density was increased around the periphery of amyloid plaques in cerebral cortex and hippocampus in all samples, but there was no apparent difference in frequency of plaque-associated microglia between DOCK2 genotypes. To assess total microglia, we used unbiased stereologic methodology to quantify Iba1 immunopositive microglia in cortex and hippocampus. Two-way analysis of variance of microglia/mm3 did not show a significant difference by brain region or across the three genotypes: DOCK2+/+, DOCK2−/+, and DOCK2−/− APPswe-PS1Δe9 mice.

Figure 1. Microglia frequency and morphology are not altered in APPswe-PS1Δe9 that lack DOCK2.

Figure 1

Open in a new tab

Cerebral cortical (A) and hippocampal (B) sections from APPswe-PS1Δe9;DOCK2−/− mice stained with anti-Iba1 (Cy3 – red) and anti-Aβ (Cy2 – green) with DAPI nuclear counterstain (blue) have relatively evenly distributed microglia with ramified morphology that are increased in association with Aβ plaque (insets, A and B). Unbiased stereologic quantification of microglia in cortex and hippocampus (C) demonstrated no significant difference in APPswe-PS1Δe9 mice that are DOCK2+/+, DOCK2+/−, or DOCK2−/− (one way ANOVA; P > 0.05). Scale Bar: 50 μm.

Our previously published studies demonstrated DOCK2-mediated microglia cytokine secretion, phagocytosis, and paracrine neurotoxicity. To test the role of DOCK2 in experimental Alzheimer’s disease, we assessed Aβ plaques in cerebral cortex (Figure 2) and hippocampus (Figure 3) of 10-month-old APPswe-PS1Δe9;DOCK2+/+, APPswe-PS1Δe9;DOCK2+/−, and APPswe-PS1Δe9;DOCK2−/− mice using Aβ immunofluorescence with stereological measurement of three different variables: fraction of brain area occupied by Aβ plaques, size of Aβ plaques, and number of Aβ plaques. DOCK2+/+ mice showed the expected Aβ plaque formation in these two regions of APPswe-PS1Δe9 mouse brain. There was a significant progressive reduction in the fraction of brain area occupied by Aβ plaque from DOCK2+/+ to DOCK2+/− to DOCK2−/− mice in both cerebral cortex (ANOVA P < 0.05) and hippocampus (ANOVA P < 0.05) with DOCK2−/− mice showing about half the amount of Aβ plaques as DOCK2+/+ mice; in both brain regions (Bonferroni-corrected multiple pair comparisons had P < 0.05 for DOCK2+/+ vs. DOCK2−/− but were non-significant for DOCK+/− mice). A stronger relationship was observed between DOCK2 genotype and Aβ plaque size in cerebral cortex (ANOVA P < 0.01) than in hippocampus (P < 0.05); again posttests showed significant difference between DOCK2+/+ and DOCK2−/− in cerebral cortex (P < 0.01) and hippocampus (P < 0.05) without significant difference for DOCK2+/− with either of the other groups. The number of Aβ plaques was reduced in both regions of mice lacking one or both DOCK2 alleles; however, this was significant only in the hippocampus (P < 0.05) and not cerebral cortex (P = 0.09). In summary, DOCK2−/− mice showed a 30% to 50% reduction in three different measures of Aβ plaque accumulation compared to DOCK2+/+ mice with DOCK2+/− mice displaying intermediate values.

Figure 2. Aβ plaques are reduced in cerebral cortex in APPswe-PS1Δe9 mice deficient in DOCK2.

Figure 2

Open in a new tab

Representative cortical sections of APPswe-PS1Δe9 with DOCK2+/+ (A), DOCK2+/− (B), or DOCK2−/− (C) stained with anti-Aβ (Cy3 – red) demonstrate a significant effect of DOCK2 genotype on Aβ plaque area (D), average plaque size (E), and plaque frequency (F) (Image J analysis; one way ANOVA; P < 0.05). Bonferroni post-hoc tests show significant reduction in Aβ plaque area, size and frequency (**P < 0.05) between APPswe-PS1Δe9 that were DOCK2+/+ and DOCK2−/−. No significant differences (ns) were identified between DOCK2+/− and the other groups although each measure for DOCK2+/− mice fell between those for DOCK2+/+ and knockout mice. Scale Bar: 500 μm.

Figure 3. Aβ plaques are reduced in hippocampus in APPswe-PS1Δe9 mice deficient in DOCK2.

Figure 3

Open in a new tab

Representative hippocampal sections of APPswe-PS1Δe9 with DOCK2+/+ (A), DOCK2+/− (B), or DOCK2−/− (C) stained with anti-Aβ (Cy3 – red) demonstrate a significant effect of DOCK2 genotype on total plaque area (D), average plaque size (E), and plaque frequency (F) (Image J analysis; one way ANOVA; P < 0.05). Bonferroni post-hoc tests show significant reduction in Aβ plaque area, size and frequency (**P < 0.05) between APPswe-PS1Δe9 that were DOCK2+/+ and DOCK2−/−. No significant differences (ns) were identified between DOCK2 hemizygous APPswe-PS1Δe9 mice and DOCK2 homozygotes or knockouts, although, as in cortex, each Aβ measurement for DOCK2+/− mice fell between those for DOCK2+/+ and DOCK2−/− mice. Scale Bar: 250 μm.

Finally, we quantified the concentrations of triton-soluble and triton-insoluble/guanidine-soluble Aβ42 in cerebral cortex and hippocampus of mice having none (n = 9), one (n = 8), or both (n = 5) copies of DOCK2. Triton-soluble Aβ42 averaged 208 + 28 pg/mg protein and guanidine-soluble Aβ42 averaged 537 + 16 pg/mg protein in DOCK2+/+ mice; there was no significant difference (p > 0.05) in concentrations of detergent-soluble Aβ42 among DOCK2+/+, DOCK2+/−, and DOCK2−/− mice (data not shown).

DISCUSSION

Observational data support suppression of the PG pathway with NSAIDs as a potential means to prevent or delay the onset of symptoms from AD. Clinical trials to treat dementia or prodromal stages of AD with NSAIDs failed, and the only prevention trial of NSAIDs was terminated due to toxicity. Thus, the efficacy of NSAIDs to prevent AD is supported by epidemiologic observations but has never been rigorously tested in people. These data have spurred several laboratories to pursue more specific targets within the PG signaling cascade that retain efficacy while averting toxicity. One promising pathway is EP2 receptor activation by PGE2.

We sought a microglia-specific EP2-dependent signaling component as a cell-specific target for suppressing the PG pathway in AD. Previous work led us to focus on DOCK2, whose exclusive microglial expression in brain is critically regulated by EP2 (Cimino, Sokal, Leverenz, Fukui and Montine, 2009). Similar to NSAIDs or ablation of EP2, we have shown previously that ablation of DOCK2 suppresses microglial inflammatory response and paracrine neurotoxicity; here we sought to determine whether ablation of DOCK2 might also replicate the anti-Aβ effects of NSAIDs and EP2 ablation in a transgenic mouse model of AD.

Our results showed that processes dependent on the expression of DOCK2, an exclusively microglial protein in brain, contributed to Aβ plaque accumulation but not the level of soluble Aβ42 in cerebral cortex and hippocampus. This argues that our data is not a result of APP processing and Aβ production, but rather a response to plaque deposition or plaque clearance. It is important to note that DOCK2−/− primary microglia have decreased phagocytic activity in vitro as assessed by fluorescent beads in primary cultures, yet DOCK2−/− mice accumulated less Aβ plaques in cerebral cortex and hippocampus. These data argue against microglial phagocytosis, and therefore plaque Aβ clearance by phagocytosis, as a significant regulator of Aβ plaques in mouse brain. Rather, our results align with those of others that have suggested that Aβ deposition, clearance, and plaque formation are regulated more by the inflammatory milieu (Farfara, et al., 2011; Lim, et al., 2000; Reed-Geaghan, et al., 2010; Town, et al., 2005; Town, et al., 2008; Wyss-Coray, 2006; Zhu, et al., 2011) than by altering phagocytosis (Reed-Geaghan, Reed, Cramer and Landreth, 2010). These results also are consistent with older experiments using NSAIDs that suppress immune response in brain but do not significantly alter microglial phagocytosis. Our results differ somewhat from those of some other models in that we observed a reduction in Aβ plaques but no change in concentrations of soluble Aβ42 concentration. This is an interesting distinction, one interpretation of which is that DOCK2-dependent processes are critical in the conversion of soluble Aβ peptides into insoluble plaque material but not in soluble Aβ production and clearance.

DOCK2 modulation of innate immune response does not appear to be related to microglia migration or proliferation, as we found no differences in microglia density or distribution in APPswe-PS1Δe9:DOCK2−/− mice compared with APPswe-PS1Δe9:DOCK2+/+ controls. This further supports the observation that microglia function, specifically cytokine production and subsequent paracrine neurotoxicity, is a critical regulator of Aβ plaque formation. By extension, microglia-mediated innate immune modulation and neurotoxicity are implicated in numerous neurodegenerative disorders making DOCK2 a logical potential therapeutic target for diverse neurological conditions.

In summary, our results show that the microglia-specific protein DOCK2 contributes significantly to cerebral cortical and hippocampal Aβ plaques in a transgenic mouse model of AD, add further evidence that suppression of the innate immune response may be more important than augmenting phagocytic capacity to in reduce Aβ plaques in vivo, and suggest that DOCK2 may provide a viable therapeutic target for AD and other neurodegenerative diseases characterized by innate immune activation.

Acknowledgments

This work was supported by F30 AG030914 (PJC), P50 AG05136, and the Nancy and Buster Alvord Endowment. The authors thank Ms. Meilany Wijaya for assistance with animal husbandry and Ms. Aimee Schantz for administrative support.

Abbreviations

AD

Alzheimer’s disease

APP

amyloid precursor protein

Aβ

amyloid beta

COX

cyclooxygenase

DAPI

4′,6-diaminido-2-phenylindole

DOCK2

dedicator of cytokinesis 2

EP2

PGE2 receptor subtype 2

GEF

guanyl-nucleotide exchange factor

NSAIDs

nonsteroidal anti-inflammatory drugs

PG

prostaglandin

PGE2

prostaglandin E2

PS1Δe9

exon-9-deleted human presenilin-1

swe

Swedish mutations K594N/M595L

wt

wild-type

Footnotes

Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

References

  • 1.Adapt Research Group. Lyketsos CG, Breitner JC, et al. Naproxen and celecoxib do not prevent AD in early results from a randomized controlled trial. Neurology. 2007;68:1800–1808. doi: 10.1212/01.wnl.0000260269.93245.d2. [DOI] [PubMed] [Google Scholar]
  • 2.Ahmad AS, Yun YT, Ahmad M, Maruyama T, Dore S. Selective blockade of PGE2 EP1 receptor protects brain against experimental ischemia and excitotoxicity, and hippocampal slice cultures against oxygen-glucose deprivation. Neurotox Res. 2008;14:343–351. doi: 10.1007/BF03033858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Cimino PJ, Keene CD, Breyer RM, Montine KS, Montine TJ. Therapeutic targets in prostaglandin E2 signaling for neurologic disease. Curr Med Chem. 2008;15:1863–1869. doi: 10.2174/092986708785132915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Cimino PJ, Sokal I, Leverenz J, Fukui Y, Montine TJ. DOCK2 is a microglial specific regulator of central nervous system innate immunity found in normal and Alzheimer’s disease brain. Am J Pathol. 2009;175:1622–1630. doi: 10.2353/ajpath.2009.090443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Cole GM, Morihara T, Lim GP, Yang F, Begum A, Frautschy SA. NSAID and antioxidant prevention of Alzheimer’s disease: lessons from in vitro and animal models. Ann N Y Acad Sci. 2004;1035:68–84. doi: 10.1196/annals.1332.005. [DOI] [PubMed] [Google Scholar]
  • 6.Echeverria V, Clerman A, Dore S. Stimulation of PGE receptors EP2 and EP4 protects cultured neurons against oxidative stress and cell death following beta-amyloid exposure. Eur J Neurosci. 2005;22:2199–2206. doi: 10.1111/j.1460-9568.2005.04427.x. [DOI] [PubMed] [Google Scholar]
  • 7.Farfara D, Trudler D, Segev-Amzaleg N, Galron R, Stein R, Frenkel D. gamma-Secretase component presenilin is important for microglia beta-amyloid clearance. Ann Neurol. 2011;69:170–180. doi: 10.1002/ana.22191. [DOI] [PubMed] [Google Scholar]
  • 8.Fukui Y, Hashimoto O, Sanui T, et al. Haematopoietic cell-specific CDM family protein DOCK2 is essential for lymphocyte migration. Nature. 2001;412:826–831. doi: 10.1038/35090591. [DOI] [PubMed] [Google Scholar]
  • 9.Funk CD, FitzGerald GA. COX-2 inhibitors and cardiovascular risk. J Cardiovasc Pharmacol. 2007;50:470–479. doi: 10.1097/FJC.0b013e318157f72d. [DOI] [PubMed] [Google Scholar]
  • 10.Gollmer K, Asperti-Boursin F, Tanaka Y, et al. CCL21 mediates CD4+ T-cell costimulation via a DOCK2/Rac-dependent pathway. Blood. 2009;114:580–588. doi: 10.1182/blood-2009-01-200923. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Gotoh K, Tanaka Y, Nishikimi A, et al. Selective control of type I IFN induction by the Rac activator DOCK2 during TLR-mediated plasmacytoid dendritic cell activation. J Exp Med. 2010;207:721–730. doi: 10.1084/jem.20091776. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Halle A, Hornung V, Petzold GC, et al. The NALP3 inflammasome is involved in the innate immune response to amyloid-beta. Nat Immunol. 2008;9:857–865. doi: 10.1038/ni.1636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Hickman SE, Allison EK, El Khoury J. Microglial dysfunction and defective beta-amyloid clearance pathways in aging Alzheimer’s disease mice. J Neurosci. 2008;28:8354–8360. doi: 10.1523/JNEUROSCI.0616-08.2008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Jankowsky JL, Slunt HH, Ratovitski T, Jenkins NA, Copeland NG, Borchelt DR. Co-expression of multiple transgenes in mouse CNS: a comparison of strategies. Biomol Eng. 2001;17:157–165. doi: 10.1016/s1389-0344(01)00067-3. [DOI] [PubMed] [Google Scholar]
  • 15.Jin J, Shie FS, Liu J, et al. Prostaglandin E2 receptor subtype 2 (EP2) regulates microglial activation and associated neurotoxicity induced by aggregated alpha-synuclein. J Neuroinflammation. 2007;4:2. doi: 10.1186/1742-2094-4-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Keene CD, Chang R, Stephen C, et al. Protection of hippocampal neurogenesis from toll-like receptor 4-dependent innate immune activation by ablation of prostaglandin E2 receptor subtype EP1 or EP2. Am J Pathol. 2009;174:2300–2309. doi: 10.2353/ajpath.2009.081153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Keene CD, Chang RC, Lopez-Yglesias AH, et al. Suppressed accumulation of cerebral amyloid beta peptides in aged transgenic Alzheimer’s disease mice by transplantation with wild-type or prostaglandin E2 receptor subtype 2-null bone marrow. Am J Pathol. 2010;177:346–354. doi: 10.2353/ajpath.2010.090840. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 18.Konstantinopoulos PA, Lehmann DF. The cardiovascular toxicity of selective and nonselective cyclooxygenase inhibitors: comparisons, contrasts, and aspirin confounding. J Clin Pharmacol. 2005;45:742–750. doi: 10.1177/0091270005278202. [DOI] [PubMed] [Google Scholar]
  • 19.Lei Y, Liu C, Saito F, Fukui Y, Takahama Y. Role of DOCK2 and DOCK180 in fetal thymus colonization. Eur J Immunol. 2009;39:2695–2702. doi: 10.1002/eji.200939630. [DOI] [PubMed] [Google Scholar]
  • 20.Liang X, Wang Q, Hand T, et al. Deletion of the prostaglandin E2 EP2 receptor reduces oxidative damage and amyloid burden in a model of Alzheimer’s disease. J Neurosci. 2005;25:10180–10187. doi: 10.1523/JNEUROSCI.3591-05.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Liang X, Wang Q, Shi J, et al. The prostaglandin E2 EP2 receptor accelerates disease progression and inflammation in a model of amyotrophic lateral sclerosis. Ann Neurol. 2008;64:304–314. doi: 10.1002/ana.21437. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Lim GP, Yang F, Chu T, et al. Ibuprofen suppresses plaque pathology and inflammation in a mouse model for Alzheimer’s disease. J Neurosci. 2000;20:5709–5714. doi: 10.1523/JNEUROSCI.20-15-05709.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Lobsiger CS, Cleveland DW. Glial cells as intrinsic components of non-cell-autonomous neurodegenerative disease. Nat Neurosci. 2007;10:1355–1360. doi: 10.1038/nn1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Manabe Y, Anrather J, Kawano T, et al. Prostanoids, not reactive oxygen species, mediate COX-2-dependent neurotoxicity. Ann Neurol. 2004;55:668–675. doi: 10.1002/ana.20078. [DOI] [PubMed] [Google Scholar]
  • 25.Martin BK, Szekely C, Brandt J, et al. Cognitive function over time in the Alzheimer’s Disease Anti-inflammatory Prevention Trial (ADAPT): results of a randomized, controlled trial of naproxen and celecoxib. Arch Neurol. 2008;65:896–905. doi: 10.1001/archneur.2008.65.7.nct70006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.McGeer PL, McGeer EG. NSAIDs and Alzheimer disease: epidemiological, animal model and clinical studies. Neurobiol Aging. 2007;28:639–647. doi: 10.1016/j.neurobiolaging.2006.03.013. [DOI] [PubMed] [Google Scholar]
  • 27.Meinert CL, McCaffrey LD, Breitner JC. Alzheimer’s Disease Anti-inflammatory Prevention Trial: design, methods, and baseline results. Alzheimers Dement. 2009;5:93–104. doi: 10.1016/j.jalz.2008.09.004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Miller KR, Streit WJ. The effects of aging, injury and disease on microglial function: a case for cellular senescence. Neuron Glia Biol. 2007;3:245–253. doi: 10.1017/S1740925X08000136. [DOI] [PubMed] [Google Scholar]
  • 29.Montine TJ, Milatovic D, Gupta RC, Valyi-Nagy T, Morrow JD, Breyer RM. Neuronal oxidative damage from activated innate immunity is EP2 receptor-dependent. J Neurochem. 2002;83:463–470. doi: 10.1046/j.1471-4159.2002.01157.x. [DOI] [PubMed] [Google Scholar]
  • 30.Nishikimi A, Fukuhara H, Su W, et al. Sequential regulation of DOCK2 dynamics by two phospholipids during neutrophil chemotaxis. Science. 2009;324:384–387. doi: 10.1126/science.1170179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Quinn JF, Bussiere JR, Hammond RS, et al. Chronic dietary alpha-lipoic acid reduces deficits in hippocampal memory of aged Tg2576 mice. Neurobiol Aging. 2007;28:213–225. doi: 10.1016/j.neurobiolaging.2005.12.014. [DOI] [PubMed] [Google Scholar]
  • 32.Reed-Geaghan EG, Reed QW, Cramer PE, Landreth GE. Deletion of CD14 attenuates Alzheimer’s disease pathology by influencing the brain’s inflammatory milieu. J Neurosci. 2010;30:15369–15373. doi: 10.1523/JNEUROSCI.2637-10.2010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Sanui T, Inayoshi A, Noda M, et al. DOCK2 is essential for antigen-induced translocation of TCR and lipid rafts, but not PKC-theta and LFA-1, in T cells. Immunity. 2003;19:119–129. doi: 10.1016/s1074-7613(03)00169-9. [DOI] [PubMed] [Google Scholar]
  • 34.Savonenko A, Munoz P, Melnikova T, et al. Impaired cognition, sensorimotor gating, and hippocampal long-term depression in mice lacking the prostaglandin E2 EP2 receptor. Exp Neurol. 2009;217:63–73. doi: 10.1016/j.expneurol.2009.01.016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Shie FS, Breyer RM, Montine TJ. Microglia lacking E Prostanoid Receptor subtype 2 have enhanced Abeta phagocytosis yet lack Abeta-activated neurotoxicity. Am J Pathol. 2005a;166:1163–1172. doi: 10.1016/s0002-9440(10)62336-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Shie FS, Montine KS, Breyer RM, Montine TJ. Microglial EP2 as a new target to increase amyloid beta phagocytosis and decrease amyloid beta-induced damage to neurons. Brain Pathol. 2005b;15:134–138. doi: 10.1111/j.1750-3639.2005.tb00509.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Shie FS, Montine KS, Breyer RM, Montine TJ. Microglial EP2 is critical to neurotoxicity from activated cerebral innate immunity. Glia. 2005c;52:70–77. doi: 10.1002/glia.20220. [DOI] [PubMed] [Google Scholar]
  • 38.Town T, Nikolic V, Tan J. The microglial “activation” continuum: from innate to adaptive responses. J Neuroinflammation. 2005;2:24. doi: 10.1186/1742-2094-2-24. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Town T, Laouar Y, Pittenger C, et al. Blocking TGF-beta-Smad2/3 innate immune signaling mitigates Alzheimer-like pathology. Nat Med. 2008;14:681–687. doi: 10.1038/nm1781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40.West MJ. New stereological methods for counting neurons. Neurobiol Aging. 1993;14:275–285. doi: 10.1016/0197-4580(93)90112-o. [DOI] [PubMed] [Google Scholar]
  • 41.Woltjer RL, Cimino PJ, Boutte AM, et al. Proteomic determination of widespread detergent-insolubility including Abeta but not tau early in the pathogenesis of Alzheimer’s disease. Faseb J. 2005;19:1923–1925. doi: 10.1096/fj.05-4263fje. [DOI] [PubMed] [Google Scholar]
  • 42.Wu L, Wang Q, Liang X, Andreasson K. Divergent effects of prostaglandin receptor signaling on neuronal survival. Neurosci Lett. 2007;421:253–258. doi: 10.1016/j.neulet.2007.05.055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 43.Wyss-Coray T. Inflammation in Alzheimer disease: driving force, bystander or beneficial response? Nat Med. 2006;12:1005–1015. doi: 10.1038/nm1484. [DOI] [PubMed] [Google Scholar]
  • 44.Xiang Z, Ho L, Yemul S, et al. Cyclooxygenase-2 promotes amyloid plaque deposition in a mouse model of Alzheimer’s disease neuropathology. Gene Expr. 2002;10:271–278. doi: 10.3727/000000002783992352. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 45.Yang W, Woltjer RL, Sokal I, et al. Quantitative proteomics identifies surfactant-resistant alpha-synuclein in cerebral cortex of Parkinsonism-dementia complex of Guam but not Alzheimer’s disease or progressive supranuclear palsy. Am J Pathol. 2007;171:993–1002. doi: 10.2353/ajpath.2007.070015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 46.Yu Y, Fan J, Chen XS, et al. Genetic model of selective COX2 inhibition reveals novel heterodimer signaling. Nat Med. 2006;12:699–704. doi: 10.1038/nm1412. [DOI] [PubMed] [Google Scholar]
  • 47.Yu Y, Fan J, Hui Y, et al. Targeted cyclooxygenase gene (ptgs) exchange reveals discriminant isoform functionality. J Biol Chem. 2007;282:1498–1506. doi: 10.1074/jbc.M609930200. [DOI] [PubMed] [Google Scholar]
  • 48.Zhu Y, Hou H, Rezai-Zadeh K, et al. CD45 deficiency drives amyloid-beta peptide oligomers and neuronal loss in Alzheimer’s disease mice. J Neurosci. 2011;31:1355–1365. doi: 10.1523/JNEUROSCI.3268-10.2011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 49.Zipp F, Aktas O. The brain as a target of inflammation: common pathways link inflammatory and neurodegenerative diseases. Trends Neurosci. 2006;29:518–527. doi: 10.1016/j.tins.2006.07.006. [DOI] [PubMed] [Google Scholar]

RESOURCES