Resveratrol Inhibits β-Amyloid-Induced Neuronal Apoptosis through Regulation of SIRT1-ROCK1 Signaling Pathway

Alzheimer’s disease (AD) is characterized by the accumulation of β-amyloid peptide (Aβ) and loss of neurons. Recently, a growing body of evidences have indicated that as a herbal compound naturally derived from grapes, resveratrol modulates the pathophysiology of AD, however, with a largely unclear mechanism. Therefore, we aimed to investigate the protection of resveratrol against the neurotoxicity of β-amyloid peptide 25–35 (Aβ_25–35) and further explore its underlying mechanism in the present study. PC12 cells were injuried by Aβ_25–35, and resveratrol at different concentrations was added into the culture medium. We observed that resveratrol increased cell viability through the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) colorimetric assays. Flow cytometry indicated the reduction of cell apoptosis by resveratrol. Moreover, resveratrol also stabilized the intercellular Ca²⁺ homeostasis and attenuated Aβ_25–35 neurotoxicity. Additionally, Aβ_25–35-suppressed silent information regulator 1 (SIRT1) activity was significantly reversed by resveratrol, resulting in the downregulation of Rho-associated kinase 1 (ROCK1). Our results clearly revealed that resveratrol significantly protected PC12 cells and inhibited the β-amyloid-induced cell apoptosis through the upregulation of SIRT1. Moreover, as a downstream signal molecule, ROCK1 was negatively regulated by SIRT1. Taken together, our study demonstrated that SIRT1-ROCK1 pathway played a critical role in the pathomechanism of AD.     

FLZ Alleviates the Memory Deficits in Transgenic Mouse Model of Alzheimer’s Disease via Decreasing Beta-Amyloid Production and Tau Hyperphosphorylation

Alzheimer’s disease (AD) is the most common cause of dementia worldwide and mainly characterized by the aggregated β-amyloid (Aβ) and hyperphosphorylated tau. FLZ is a novel synthetic derivative of natural squamosamide and has been proved to improve memory deficits in dementia animal models. In this study, we aimed to investigate the mechanisms of FLZ’s neuroprotective effect in APP/PS1 double transgenic mice and SH-SY5Y (APPwt/swe) cells. The results showed that treatment with FLZ significantly improved the memory deficits of APP/PS1 transgenic mice and decreased apoptosis of SH-SY5Y (APPwt/swe) cells. FLZ markedly attenuated Aβ accumulation and tau phosphorylation both in vivo and in vitro. Mechanistic study showed that FLZ interfered APP processing, i.e., FLZ decreased β-amyloid precursor protein (APP) phosphorylation, APP-carboxy-terminal fragment (APP-CTF) production and β-amyloid precursor protein cleaving enzyme 1 (BACE1) expression. These results indicated that FLZ reduced Aβ production through inhibiting amyloidogenic pathway. The mechanistic study about FLZ’s inhibitory effect on tau phosphorylation revealed t the involvement of Akt/glycogen synthase kinase 3β (GSK3β) pathway. FLZ treatment increased Akt activity and inhibited GSK3β activity both in vivo and in vitro. The inhibitory effect of FLZ on GSK3β activity and tau phosphorylation was suppressed by inhibiting Akt activity, indicating that Akt/GSK3β pathway might be the possible mechanism involved in the inhibitory effect of FLZ on tau hyperphosphorylation. These results suggested FLZ might be a potential anti-AD drug as it not only reduced Aβ production via inhibition amyloidogenic APP processing pathway, but also attenuated tau hyperphosphoylation mediated by Akt/GSK3β.     

β-Amyloid Precursor Protein Mutants Respond to γ-Secretase Modulators

Pathogenic generation of the 42-amino acid variant of the amyloid β-peptide (Aβ) by β- and γ-secretase cleavage of the β-amyloid precursor protein (APP) is believed to be causative for Alzheimer disease (AD). Lowering of Aβ₄₂ production by γ-secretase modulators (GSMs) is a hopeful approach toward AD treatment. The mechanism of GSM action is not fully understood. Moreover, whether GSMs target the Aβ domain is controversial. To further our understanding of the mode of action of GSMs and the cleavage mechanism of γ-secretase, we analyzed mutations located at different positions of the APP transmembrane domain around or within the Aβ domain regarding their response to GSMs. We found that Aβ₄₂-increasing familial AD mutations of the γ-secretase cleavage site domain responded robustly to Aβ₄₂-lowering GSMs, especially to the potent compound GSM-1, irrespective of the amount of Aβ₄₂ produced. We thus expect that familial AD patients carrying mutations at the γ-secretase cleavage sites of APP should respond to GSM-based therapeutic approaches. Systematic phenylalanine-scanning mutagenesis of this region revealed a high permissiveness to GSM-1 and demonstrated a complex mechanism of GSM action as other Aβ species (Aβ₄₁, Aβ₃₉) could also be lowered besides Aβ₄₂. Moreover, certain mutations simultaneously increased Aβ₄₂ and the shorter peptide Aβ₃₈, arguing that the proposed precursor-product relationship of these Aβ species is not general. Finally, mutations of residues in the proposed GSM-binding site implicated in Aβ₄₂ generation (Gly-29, Gly-33) and potentially in GSM-binding (Lys-28) were also responsive to GSMs, a finding that may question APP substrate targeting of GSMs.     

Involvement of Intracellular and Mitochondrial Aβ in the Ameliorative Effects of Huperzine A against Oligomeric Aβ42-Induced Injury in Primary Rat Neurons

Considerable studies indicate huperzine A is a promising natural product to suppress neuronal damages induced by β-amyloid (Aβ), a key pathogenic event in the Alzheimer’s disease (AD). As an extension, the present study for the first time explored whether the beneficial profiles of huperzine A against oligomeric Aβ₄₂ induced neurotoxicity are associated with the accumulation and detrimental function of intraneuronal/mitochondrial Aβ, on the basis of the emerging evidence that intracellular Aβ is more relevant to AD progression as compared with extracellular Aβ. Huperzine A treatment was shown to significantly attenuate the neurotoxicity of oligomeric Aβ₄₂, as demonstrated by increased neuronal viability. Interestingly, our results proved that exogenous Aβ₄₂ could accumulate intraneuronally in a dose- and time-dependent manner, while huperzine A treatment markedly reduced the level of intracellular Aβ₄₂. Moreover, huperzine A treatment rescued mitochondrial dysfunction induced by oligomeric Aβ₄₂, including adenosine triphosphate (ATP) reduction, reactive oxygen species (ROS) overproduction and membrane potential depolarization. Further study demonstrated that huperzine A also significantly reduced the level of Aβ₄₂ in the mitochondria-enriched subcellular fractions, as well as the Aβ₄₂ fluorescent signals colocalized with mitochondrial marker. This study indicates that interfering intracellular Aβ especially mitochondrial Aβ accumulation, together with ameliorating Aβ-associated mitochondrial dysfunction, may contribute to the protective effects of huperzine A against Aβ neurotoxicity. Above results may shed more light on the pharmacological mechanisms of huperzine A and provide important clues for discovering novel therapeutic strategies for AD.     

Synthesis and biological evaluation of radioiodinated 2,5-diphenyl-1,3,4-oxadiazoles for detecting β-amyloid plaques in the brain

This paper describes the synthesis and biological evaluation of a new series of 2,5-diphenyl-1,3,4-oxadiazole (1,3,4-DPOD) derivatives for detecting β-amyloid plaques in Alzheimer’s brains. The affinity for β-amyloid plaques was assessed by an in vitro binding assay using pre-formed synthetic Aβ42 aggregates. The new series of 1,3,4-DPOD derivatives showed affinity for Aβ42 aggregates with K_i values ranging from 20 to 349 nM. The 1,3,4-DPOD derivatives clearly stained β-amyloid plaques in an animal model of Alzheimer’s disease, reflecting the affinity for Aβ42 aggregates in vitro. Compared to 3,5-diphenyl-1,2,4-oxadiazole (1,2,4-DPOD) derivatives, they displayed good penetration of and fast washout from the brain in biodistribution experiments using normal mice. The novel radioiodinated 1,3,4-DPOD derivatives may be useful probes for detecting β-amyloid plaques in the Alzheimer’s brain.     

Hyperphosphorylation of Tau Induced by Naturally Secreted Amyloid-β at Nanomolar Concentrations Is Modulated by Insulin-dependent Akt-GSK3β Signaling Pathway

Alzheimer disease (AD) is neuropathologically characterized by the formation of senile plaques from amyloid-β (Aβ) and neurofibrillary tangles composed of phosphorylated Tau. Although there is growing evidence for the pathogenic role of soluble Aβ species in AD, the major question of how Aβ induces hyperphosphorylation of Tau remains unanswered. To address this question, we here developed a novel cell coculture system to assess the effect of extracellular Aβ at physiologically relevant levels naturally secreted from donor cells on the phosphorylation of Tau in recipient cells. Using this assay, we demonstrated that physiologically relevant levels of secreted Aβ are sufficient to cause hyperphosphorylation of Tau in recipient N2a cells expressing human Tau and in primary culture neurons. This hyperphosphorylation of Tau is inhibited by blocking Aβ production in donor cells. The expression of familial AD-linked PSEN1 mutants and APP ΔE693 mutant that induce the production of oligomeric Aβ in donor cells results in a similar hyperphosphorylation of Tau in recipient cells. The mechanism underlying the Aβ-induced Tau hyperphosphorylation is mediated by the impaired insulin signal transduction because we demonstrated that the phosphorylation of Akt and GSK3β upon insulin stimulation is less activated under this condition. Treating cells with the insulin-sensitizing drug rosiglitazone, a peroxisome proliferator-activated receptor γ agonist, attenuates the Aβ-dependent hyperphosphorylation of Tau. These findings suggest that the disturbed insulin signaling cascade may be implicated in the pathways through which soluble Aβ induces Tau phosphorylation and further support the notion that correcting insulin signal dysregulation in AD may offer a potential therapeutic approach.     

Peptides of Presenilin-1 Bind the Amyloid Precursor Protein Ectodomain and Offer a Novel and Specific Therapeutic Approach to Reduce ?-Amyloid in Alzheimer’s Disease

β-Amyloid (Aβ) accumulation in the brain is widely accepted to be critical to the development of Alzheimer’s disease (AD). Current efforts at reducing toxic Aβ40 or 42 have largely focused on modulating γ-secretase activity to produce shorter, less toxic Aβ, while attempting to spare other secretase functions. In this paper we provide data that offer the potential for a new approach for the treatment of AD. The method is based on our previous findings that the production of Aβ from the interaction between the β-amyloid precursor protein (APP) and Presenilin (PS), as part of the γ-secretase complex, in cell culture is largely inhibited if the entire water-soluble NH₂-terminal domain of PS is first added to the culture. Here we demonstrate that two small, non-overlapping water-soluble peptides from the PS-1 NH₂-terminal domain can substantially and specifically inhibit the production of total Aβ as well as Aβ40 and 42 in vitro and in vivo in the brains of APP transgenic mice. These results suggest that the inhibitory activity of the entire amino terminal domain of PS-1 on Aβ production is largely focused in a few smaller sequences within that domain. Using biolayer interferometry and confocal microscopy we provide evidence that peptides effective in reducing Aβ give a strong, specific and biologically relevant binding with the purified ectodomain of APP 695. Finally, we demonstrate that the reduction of Aβ by the peptides does not affect the catalytic activities of β- or γ-secretase, or the level of APP. P4 and P8 are the first reported protein site-specific small peptides to reduce Aβ production in model systems of AD. These peptides and their derivatives offer new potential drug candidates for the treatment of AD.     

Advances on the Understanding of the Origins of Synaptic Pathology in AD

Although Alzheimer’s disease (AD) was first discovered a century ago, we are still facing a lack of definitive diagnosis during the patient’s lifetime and are unable to prescribe a curative treatment. However, the past 10 years have seen a “revamping” of the main hypothesis about AD pathogenesis and the hope to foresee possible treatment. AD is no longer considered an irreversible disease. A major refinement of the classic β-amyloid cascade describing amyloid fibrils as neurotoxins has been made to integrate the key scientific evidences demonstrating that the first pathological event occurring in AD early stages affects synaptic function and maintenance. A concept fully compatible with synapse loss being the best pathological correlate of AD rather than other described neuropathological hallmarks (amyloid plaques, neurofibrillary tangles or neuronal death). The notion that synaptic alterations might be reverted, thus offering a potential curability, was confirmed by immunotherapy experiments targeting β-amyloid protein in transgenic AD mice in which cognitive functions were improved despite no reduction in the amyloid plaques burden. The updated amyloid cascade now integrates the synapse failure triggered by soluble Aβ-oligomers. Still no consensus has been reached on the most toxic Aβ conformations, neither on their site of production nor on their extra- versus intra-cellular actions. Evidence shows that soluble Aβ oligomers or ADDLs bind selectively to neurons at their synaptic loci, and trigger major changes in synapse composition and morphology, which ultimately leads to dendritic spine loss. However, the exact mechanism is not yet fully understood but is suspected to involve some membrane receptor(s).     

The Native Copper- and Zinc- Binding Protein Metallothionein Blocks Copper-Mediated Aβ Aggregation and Toxicity in Rat Cortical Neurons

Background

A major pathological hallmark of AD is the deposition of insoluble extracellular β-amyloid (Aβ) plaques. There are compelling data suggesting that Aβ aggregation is catalysed by reaction with the metals zinc and copper.

Methodology/Principal Findings

We now report that the major human-expressed metallothionein (MT) subtype, MT-2A, is capable of preventing the in vitro copper-mediated aggregation of Aβ_1–40 and Aβ_1–42. This action of MT-2A appears to involve a metal-swap between Zn₇MT-2A and Cu(II)-Aβ, since neither Cu₁₀MT-2A or carboxymethylated MT-2A blocked Cu(II)-Aβ aggregation. Furthermore, Zn₇MT-2A blocked Cu(II)-Aβ induced changes in ionic homeostasis and subsequent neurotoxicity of cultured cortical neurons.

Conclusions/Significance

These results indicate that MTs of the type represented by MT-2A are capable of protecting against Aβ aggregation and toxicity. Given the recent interest in metal-chelation therapies for AD that remove metal from Aβ leaving a metal-free Aβ that can readily bind metals again, we believe that MT-2A might represent a different therapeutic approach as the metal exchange between MT and Aβ leaves the Aβ in a Zn-bound, relatively inert form.     

Protection of TGF-β1 against Neuroinflammation and Neurodegeneration in Aβ1–42-Induced Alzheimer’s Disease Model Rats

Neuroinflammation has been reported to be associated with Alzheimer’s disease (AD) pathogenesis. Neuroinflammation is generally considered as an outcome of glial activation; however, we recently demonstrated that T helper (Th)17 cells, a subpopulation of proinflammatory CD4+ T cells, are also involved in AD pathogenesis. Transforming growth factor (TGF)-β1, a cytokine that can be expressed in the brain, can be immunosuppressive, but its effects on lymphocyte-mediated neuroinflammation in AD pathogenesis have not been well addressed. In the current study we administered TGF-β1 via intracerebroventricle (ICV) and intranasal (IN) routes in AD model rats to investigate its antiinflammatory and neuroprotective effects. The AD rat model was prepared by bilateral hippocampal injection of amyloid-β (Aβ)_1–42. TGF-β1 was administered via ICV one hour prior to Aβ_1–42 injection or via both nares seven days after Aβ_1–42 injection. ICV administration of TGF-β1 before Aβ_1–42 injection remarkably ameliorated Aβ_1–42-induced neurodegeneration and prevented Aβ_1–42-induced increases in glia-derived proinflammatory mediators (TNF-α, IL-1β and iNOS), as well as T cell-derived proinflammatory cytokines (IFN-γ, IL-2, IL-17 and IL-22), in the hypothalamus, serum or cerebrospinal fluid (CSF) in a concentration-dependent manner. TGF-β1 pretreatment also prevented Aβ_1–42-induced decreases in the neurotrophic factors, IGF-1, GDNF and BDNF, and in the antiinflammatory cytokine, IL-10. Similarly, IN administration of TGF-β1 after Aβ_1–42 injection reduced neurodegeneration, elevation of proinflammatory mediators and cytokines, and reduction of neurotrophic and antiinflammatory factors, in the hypothalamus, serum or CSF. These findings suggest that TGF-β1 suppresses glial and T cell-mediated neuroinflammation and thereby alleviates AD-related neurodegeneration. The effectiveness of IN administered TGF-β1 in reducing Aβ_1–42 neurotoxicity suggests a possible therapeutic approach in patients with AD.     

Increased Levels of Antigen-Bound β-Amyloid Autoantibodies in Serum and Cerebrospinal Fluid of Alzheimer’s Disease Patients

Recent studies have suggested a protective role of physiological β-amyloid autoantibodies (Aβ-autoantibodies) in Alzheimer’s disease (AD). However, the determination of both free and dissociated Aβ-autoantibodies in serum hitherto has yielded inconsistent results regarding their function and possible biomarker value. Here we report the application of a new sandwich enzyme-linked immunosorbent assay (ELISA) for the determination of antigen-bound Aβ-autoantibodies (intact Aβ-IgG immune complexes) in serum and cerebrospinal fluid (CSF) of a total number of 112 AD patients and age- and gender-matched control subjects. Both serum and CSF levels of Aβ-IgG immune complexes were found to be significantly higher in AD patients compared to control subjects. Moreover, the levels of Aβ-IgG complexes were negatively correlated with the cognitive status across the groups, increasing with declining cognitive test performance of the subjects. Our results suggest a contribution of IgG-type autoantibodies to Aβ clearance in vivo and an increased immune response in AD, which may be associated with deficient Aβ-IgG removal. These findings may contribute to elucidating the role of Aβ-autoantibodies in AD pathophysiology and their potential application in AD diagnosis.     

Autophagy in microglia degrades extracellular β-amyloid fibrils and regulates the NLRP3 inflammasome

Accumulation of β-amyloid (Aβ) and resultant inflammation are critical pathological features of Alzheimer disease (AD). Microglia, a primary immune cell in brain, ingests and degrades extracellular Aβ fibrils via the lysosomal system. Autophagy is a catabolic process that degrades native cellular components, however, the role of autophagy in Aβ degradation by microglia and its effects on AD are unknown. Here we demonstrate a novel role for autophagy in the clearance of extracellular Aβ fibrils by microglia and in the regulation of the Aβ-induced NLRP3 (NLR family, pyrin domain containing 3) inflammasome using microglia specific atg7 knockout mice and cell cultures. We found in microglial cultures that Aβ interacts with MAP1LC3B-II via OPTN/optineurin and is degraded by an autophagic process mediated by the PRKAA1 pathway. We anticipate that enhancing microglial autophagy may be a promising new therapeutic strategy for AD.     

Abeta42-Induced Neurodegeneration via an Age-Dependent Autophagic-Lysosomal Injury in Drosophila

The mechanism of widespread neuronal death occurring in Alzheimer''s disease (AD) remains enigmatic even after extensive investigation during the last two decades. Amyloid beta 42 peptide (Aβ_1–42) is believed to play a causative role in the development of AD. Here we expressed human Aβ_1–42 and amyloid beta 40 (Aβ_1–40) in Drosophila neurons. Aβ_1–42 but not Aβ_1–40 causes an extensive accumulation of autophagic vesicles that become increasingly dysfunctional with age. Aβ_1–42-induced impairment of the degradative function, as well as the structural integrity, of post-lysosomal autophagic vesicles triggers a neurodegenerative cascade that can be enhanced by autophagy activation or partially rescued by autophagy inhibition. Compromise and leakage from post-lysosomal vesicles result in cytosolic acidification, additional damage to membranes and organelles, and erosive destruction of cytoplasm leading to eventual neuron death. Neuronal autophagy initially appears to play a pro-survival role that changes in an age-dependent way to a pro-death role in the context of Aβ_1–42 expression. Our in vivo observations provide a mechanistic understanding for the differential neurotoxicity of Aβ_1–42 and Aβ_1–40, and reveal an Aβ_1–42-induced death execution pathway mediated by an age-dependent autophagic-lysosomal injury.     

CCL2 Accelerates Microglia-Mediated Aβ Oligomer Formation and Progression of Neurocognitive Dysfunction

Background

The linkages between neuroinflammation and Alzheimer''s disease (AD) pathogenesis are well established. What is not, however, is how specific immune pathways and proteins affect the disease. To this end, we previously demonstrated that transgenic over-expression of CCL2 enhanced microgliosis and induced diffuse amyloid plaque deposition in Tg2576 mice. This rodent model of AD expresses a Swedish β-amyloid (Aβ) precursor protein mutant.

Methodology/Principal Findings

We now report that CCL2 transgene expression accelerates deficits in spatial and working memory and hippocampal synaptic transmission in β-amyloid precursor protein (APP) mice as early as 2–3 months of age. This is followed by increased numbers of microglia that are seen surrounding Aβ oligomers. CCL2 does not suppress Aβ degradation. Rather, CCL2 and tumor necrosis factor-α directly facilitated Aβ uptake, intracellular Aβ oligomerization, and protein secretion.

Conclusions/Significance

We posit that CCL2 facilitates Aβ oligomer formation in microglia and propose that such events accelerate memory dysfunction by affecting Aβ seeding in the brain.     

Ca2+ Influx through Store-operated Ca2+ Channels Reduces Alzheimer Disease β-Amyloid Peptide Secretion

Alzheimer disease (AD), the leading cause of dementia, is characterized by the accumulation of β-amyloid peptides (Aβ) in senile plaques in the brains of affected patients. Many cellular mechanisms are thought to play important roles in the development and progression of AD. Several lines of evidence point to the dysregulation of Ca²⁺ homeostasis as underlying aspects of AD pathogenesis. Moreover, direct roles in the regulation of Ca²⁺ homeostasis have been demonstrated for proteins encoded by familial AD-linked genes such as PSEN1, PSEN2, and APP, as well as Aβ peptides. Whereas these studies support the hypothesis that disruption of Ca²⁺ homeostasis contributes to AD, it is difficult to disentangle the effects of familial AD-linked genes on Aβ production from their effects on Ca²⁺ homeostasis. Here, we developed a system in which cellular Ca²⁺ homeostasis could be directly manipulated to study the effects on amyloid precursor protein metabolism and Aβ production. We overexpressed stromal interaction molecule 1 (STIM1) and Orai1, the components of the store-operated Ca²⁺ entry pathway, to generate cells with constitutive and store depletion-induced Ca²⁺ entry. We found striking effects of Ca²⁺ entry induced by overexpression of the constitutively active STIM1_D76A mutant on amyloid precursor protein metabolism. Specifically, constitutive activation of Ca²⁺ entry by expression of STIM1_D76A significantly reduced Aβ secretion. Our results suggest that disruptions in Ca²⁺ homeostasis may influence AD pathogenesis directly through the modulation of Aβ production.     

Cathepsin B Degrades Amyloid-β in Mice Expressing Wild-type Human Amyloid Precursor Protein

Accumulation of amyloid-β (Aβ), believed to be a key trigger of Alzheimer disease (AD), could result from impaired clearance mechanisms. Previously, we showed that the cysteine protease cathepsin B (CatB) degrades Aβ, most likely by C-terminal truncation, in mice expressing human amyloid precursor protein with familial AD-linked mutations (hAPP_FAD). In addition, the Aβ-degrading activity of CatB is inhibited by its endogenous inhibitor, cystatin C (CysC). Reducing CysC expression markedly lowers Aβ levels by enhancing CatB-mediated Aβ degradation in hAPP_FAD mice. However, because a vast majority of AD patients do not carry familial mutations, we investigated how the CysC-CatB axis affects Aβ levels in mice expressing wild-type hAPP (hAPP_WT). Enhancing CatB activity by CysC deletion significantly lowered total Aβ and Aβ42 levels in hAPP_WT mice, whereas CatB deletion increased Aβ levels. To determine whether neuron-derived CatB degrades Aβ in vivo, we generated transgenic mice overexpressing CatB under the control of a neuron-specific enolase promoter. Enhancing neuronal CatB activity in hAPP_WT mice significantly lowered Aβ42 levels. The processing of hAPP_WT was unaffected by increasing or ablating CatB activity. Thus, the CysC-CatB axis affects degradation of Aβ42 derived from hAPP lacking familial mutations. These findings support the notion that enhancing CatB activity could lower Aβ, especially Aβ42, in AD patients with or without familial mutations.     

Intracellular selection of peptide inhibitors that target disulphide-bridged Aβ42 oligomers

The β-amyloid (Aβ) peptide aggregates into a number of soluble and insoluble forms, with soluble oligomers thought to be the primary factor implicated in Alzheimer''s disease pathology. As a result, a wide range of potential aggregation inhibitors have been developed. However, in addition to problems with solubility and protease susceptibility, many have inadvertently raised the concentration of these soluble neurotoxic species. Sandberg et al. previously reported a β-hairpin stabilized variant of Aβ₄₂ that results from an intramolecular disulphide bridge (A21C/A31C; Aβ_42cc), which generates highly toxic oligomeric species incapable of converting into mature fibrils. Using an intracellular protein-fragment complementation (PCA) approach, we have screened peptide libraries using E. coli that harbor an oxidizing environment to permit cytoplasmic disulphide bond formation. Peptides designed to target either the first or second β-strand have been demonstrated to bind to Aβ_42cc, lower amyloid cytotoxicity, and confer bacterial cell survival. Peptides have consequently been tested using wild-type Aβ₄₂ via ThT binding assays, circular dichroism, MTT cytotoxicity assays, fluorescence microscopy, and atomic force microscopy. Results demonstrate that amyloid-PCA selected peptides function by both removing amyloid oligomers as well as inhibiting their formation. These data further support the use of semirational design combined with intracellular PCA methodology to develop Aβ antagonists as candidates for modification into drugs capable of slowing or even preventing the onset of AD.