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NPD1 shifts βAPP processing from the amyloidogenic to the non-amyloidogenic pathway.(A) Control or HNG cells over-expressing βAPPsw were treated with increasing concentrations (0, 50, 100, 500 nM) of NPD1 for 48 h and subjected to Western blot detection of holo-βAPP (βAPP holoenzyme), sAPPα, sAPPβsw, CTFα and CTFβ in comparison to β-actin levels in the same sample; (B) Quantification of gel bands in (A) analyzing βAPP fragments with increasing doses of NPD1. Results are means ± SEM (n = 4); *p<0.01 vs. βAPPsw control.
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pone-0015816-g006: NPD1 shifts βAPP processing from the amyloidogenic to the non-amyloidogenic pathway.(A) Control or HNG cells over-expressing βAPPsw were treated with increasing concentrations (0, 50, 100, 500 nM) of NPD1 for 48 h and subjected to Western blot detection of holo-βAPP (βAPP holoenzyme), sAPPα, sAPPβsw, CTFα and CTFβ in comparison to β-actin levels in the same sample; (B) Quantification of gel bands in (A) analyzing βAPP fragments with increasing doses of NPD1. Results are means ± SEM (n = 4); *p<0.01 vs. βAPPsw control.

Mentions: Aβ42-peptides are secreted from human brain cells as they age or in response to physiological stress [4], [9], [27], [47], [48]. The processing of βAPP holoenzyme and secretion of βAPP fragments is controlled in large part by alpha-, beta- and gamma- (α-,β- and γ-) secretases [3], [4]. To assess the effects of NPD1 on secretase-mediated Aβ42 peptide generation, we used HNG cells transiently-transfected with βAPPsw and assayed for the abundance of the α-secretase–generating enzymes precursor-ADAM10 (pro-ADAM10), mature-ADAM10 (m-ADAM10), β-amyloid cleavage enzyme (BACE1) and the gamma-secretase presenilin-1 (PS1) (Figure 6). Western blot analysis revealed that the steady-level of BACE1 was reduced by 500 nM of NPD1. Meanwhile, the active and mature form of ADAM10 (m-ADAM10), the putative α-secretase, was dose-dependently increased in response to NPD1. We did not find changes in the pro-ADAM10, the inactive precursor or in the mRNA abundance of ADAM10 (data not shown; Figure 6). The undergoing changes in these two secretases are in agreement with alterations in Aβ42 peptide abundance, and in other cleavage products of βAPP (Figure 7). Interestingly, both m-ADAM10 and BACE1 levels were elevated in βAPP-over-expressing cells (Figure 6). Presenilin 1 (PS1), the main catalytic component for γ-secretase, remains unchanged after different βAPPsw or NPD1 treatments (Figure 6). This same pattern was also seen in their C-terminal counterparts, CTFβ and CTFα; importantly, no change was observed in the steady-state level of the neural cell abundant βAPP (holo-βAPP; see Figure 7A). NPD1-mediated up-regulation of m-ADAM-10 and down-regulation of BACE1 was apparent with maximal effect at 500 nM, the highest concentration used in these experiments (Figures 6 and 7).

Docosahexaenoic Acid-Derived Neuroprotectin D1 Induces Neuronal Survival via Secretase- and PPARγ-Mediated Mechanisms in Alzheimer's Disease Models

Zhao Y, Calon F, Julien C, Winkler JW, Petasis NA, Lukiw WJ, Bazan NG - PLoS ONE (2011)

Bottom Line: Moreover, NPD1 suppresses Aβ42 peptide shedding by down-regulating β-secretase-1 (BACE1) while activating the α-secretase ADAM10 and up-regulating sAPPα, thus shifting the cleavage of βAPP holoenzyme from an amyloidogenic into the non-amyloidogenic pathway.Use of the thiazolidinedione peroxisome proliferator-activated receptor gamma (PPARγ) agonist rosiglitazone, the irreversible PPARγ antagonist GW9662, and overexpressing PPARγ suggests that the NPD1-mediated down-regulation of BACE1 and Aβ42 peptide release is PPARγ-dependent.In conclusion, NPD1 bioactivity potently down regulates inflammatory signaling, amyloidogenic APP cleavage and apoptosis, underscoring the potential of this lipid mediator to rescue human brain cells in early stages of neurodegenerations.

Affiliation: School of Medicine, Louisiana State University Health Sciences Center, New Orleans, Louisiana, United States of America.

ABSTRACT
Neuroprotectin D1 (NPD1) is a stereoselective mediator derived from the omega-3 essential fatty acid docosahexaenoic acid (DHA) with potent inflammatory resolving and neuroprotective bioactivity. NPD1 reduces Aβ42 peptide release from aging human brain cells and is severely depleted in Alzheimer's disease (AD) brain. Here we further characterize the mechanism of NPD1's neurogenic actions using 3xTg-AD mouse models and human neuronal-glial (HNG) cells in primary culture, either challenged with Aβ42 oligomeric peptide, or transfected with beta amyloid precursor protein (βAPP)(sw) (Swedish double mutation APP695(sw), K595N-M596L). We also show that NPD1 downregulates Aβ42-triggered expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2) and of B-94 (a TNF-α-inducible pro-inflammatory element) and apoptosis in HNG cells. Moreover, NPD1 suppresses Aβ42 peptide shedding by down-regulating β-secretase-1 (BACE1) while activating the α-secretase ADAM10 and up-regulating sAPPα, thus shifting the cleavage of βAPP holoenzyme from an amyloidogenic into the non-amyloidogenic pathway. Use of the thiazolidinedione peroxisome proliferator-activated receptor gamma (PPARγ) agonist rosiglitazone, the irreversible PPARγ antagonist GW9662, and overexpressing PPARγ suggests that the NPD1-mediated down-regulation of BACE1 and Aβ42 peptide release is PPARγ-dependent. In conclusion, NPD1 bioactivity potently down regulates inflammatory signaling, amyloidogenic APP cleavage and apoptosis, underscoring the potential of this lipid mediator to rescue human brain cells in early stages of neurodegenerations.

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