title: Age-related loss of phospholipid asymmetry in APP(NLh)/APP(NLh) x PS-1(P264L)/PS-1(P264L) human double mutant knock-in mice: relevance to Alzheimer disease.
abstract: Using APP(NLh)/APP(NLh) x PS-1(P246L)/PS-1(P246L) human double knock-in (APP/PS-1) mice, we examined whether phosphatidylserine (PtdSer) asymmetry is significantly altered in brain of this familial Alzheimer disease mouse model in an age-dependent manner as a result of oxidative stress, toxic Abeta(1-42) oligomer production, and/or apoptosis. Annexin V (AV) and NBD-PS fluorescence in synaptosomes of wild-type (WT) and APP/PS-1 mice were used to determine PtdSer exposure with age, while Mg(2+) ATPase activity was determined to correlate PtdSer asymmetry changes with PtdSer translocase, flippase, activity. AV and NBD-PS results demonstrated significant PtdSer exposure beginning at 9 months compared to 1-month-old WT controls for both assays, a trend that was exacerbated in synaptosomes of APP/PS-1 mice. Decreasing Mg(2+) ATPase activity confirms that the age-related loss of PtdSer asymmetry is likely due to loss of flippase activity, more prominent in APP/PS-1 brain. Two-site sandwich ELISA on SDS- and FA-soluble APP/PS-1 brain fractions were conducted to correlate Abeta(1-40) and Abeta(1-42) levels with age-related trends determined from the AV, NBD-PS, and Mg(2+) ATPase assays. ELISA revealed a significant increase in both SDS- and FA-soluble Abeta(1-40) and Abeta(1-42) with age, consistent with PtdSer and flippase assay trends. Lastly, because PtdSer exposure is affected by pro-apoptotic caspase-3, levels of both latent and active forms were measured. Western blotting results demonstrated an increase in both active fragments of caspase-3 with age, while levels of pro-caspase-3 decrease. These results are discussed with relevance to loss of lipid asymmetry and consequent neurotoxicity in brain of subjects with Alzheimer disease.
authors.affiliation: Department of Chemistry, University of Kentucky, Lexington, KY 40506-0055, USA.
authors.forename: Miranda L
authors.initials: ML
authors.lastname: Bader Lange
authors.forename: Daret
authors.initials: D
authors.lastname: St Clair
authors.forename: William R
authors.initials: WR
authors.lastname: Markesbery
authors.forename: Christa M
authors.initials: CM
authors.lastname: Studzinski
authors.forename: M Paul
authors.initials: MP
authors.lastname: Murphy
authors.forename: D Allan
authors.initials: DA
authors.lastname: Butterfield
mesh_terms: D000375:Aging / Q000235:genetics / Q000378:metabolism*; D000544:Alzheimer Disease / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D016229:Amyloid beta-Peptides / Q000378:metabolism / Q000633:toxicity; D016564:Amyloid beta-Protein Precursor / Q000235:genetics; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000235:genetics; D001921:Brain / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D001923:Brain Chemistry / Q000235:genetics; D053148:Caspase 3 / Q000378:metabolism; D004195:Disease Models, Animal; D055879:Gene Knock-In Techniques; D006801:Humans; D050356:Lipid Metabolism / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D009410:Nerve Degeneration / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D018384:Oxidative Stress / Q000235:genetics; D010446:Peptide Fragments / Q000378:metabolism / Q000633:toxicity; D010718:Phosphatidylserines / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism*; D053764:Presenilin-1 / Q000235:genetics; D013574:Synaptosomes / Q000378:metabolism
chemical_list: D016229:Amyloid beta-Peptides; D016564:Amyloid beta-Protein Precursor; D017304:Annexin A5; D010446:Peptide Fragments; D010718:Phosphatidylserines; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids; D053764:Presenilin-1; C075222:amyloid beta-protein (1-42); D053148:Caspase 3