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Antioxidant Nanoparticles for Concerted Inhibition of ?-Synuclein Fibrillization, and Attenuation of Microglial Intracellular Aggregation and Activation.

ABSTRACT: Parkinson's Disease is characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta, the extracellular accumulation of toxic ?-synuclein (?SYN) aggregates, and neuroinflammation. Microglia, resident macrophages of the brain, are one of the critical cell types involved in neuroinflammation. Upon sensing extracellular stimuli or experiencing oxidative stress, microglia become activated, which further exacerbates neuroinflammation. In addition, as the first line of defense in the central nervous system, microglia play a critical role in ?SYN clearance and degradation. While the role of microglia in neurodegenerative pathologies is widely recognized, few therapeutic approaches have been designed to target both microglial activation and ?SYN aggregation. Here, we designed nanoparticles (NPs) to deliver aggregation-inhibiting antioxidants to ameliorate ?SYN aggregation and attenuate activation of a pro-inflammatory microglial phenotype. Ferulic acid diacid with an adipic acid linker (FAA) and tannic acid (TA) were used as shell and core molecules to form NPs via flash nanoprecipitation. These NPs showed a strong inhibitory effect on ?SYN fibrillization, significantly diminishing ?SYN fibrillization in vitro compared to untreated ?SYN using a Thioflavin T assay. Treating microglia with NPs decreased overall ?SYN internalization and intracellular ?SYN oligomer formation. NP treatment additionally lowered the in vitro secretion of pro-inflammatory cytokines TNF-? and IL-6, and also attenuated nitric oxide and reactive oxygen species production induced by ?SYN. NP treatment also significantly decreased Iba-1 expression in ?SYN-challenged microglia and suppressed nuclear translocation of nuclear factor kappa B (NF-?B). Overall, this work lays the foundation for an antioxidant-based nanotherapeutic candidate to target pathological protein aggregation and neuroinflammation in neurodegenerative diseases.

SUBMITTER: Zhao N

PROVIDER: S-EPMC7046761 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Antioxidant Nanoparticles for Concerted Inhibition of α-Synuclein Fibrillization, and Attenuation of Microglial Intracellular Aggregation and Activation.

Zhao Nanxia N Yang Xue X Calvelli Hannah R HR Cao Yue Y Francis Nicola L NL Chmielowski Rebecca A RA Joseph Laurie B LB Pang Zhiping P ZP Uhrich Kathryn E KE Baum Jean J Moghe Prabhas V PV

Frontiers in bioengineering and biotechnology 20200221

Parkinson's Disease is characterized by the loss of dopaminergic neurons in the <i>substantia nigra pars compacta</i>, the extracellular accumulation of toxic α-synuclein (αSYN) aggregates, and neuroinflammation. Microglia, resident macrophages of the brain, are one of the critical cell types involved in neuroinflammation. Upon sensing extracellular stimuli or experiencing oxidative stress, microglia become activated, which further exacerbates neuroinflammation. In addition, as the first line of ...[more]

PMID: 32154238

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Project description:Prevention of abnormal misfolding and aggregation of alpha synuclein (syn) protein in vulnerable neurons should be viable therapeutic strategies for reducing pathogenesis in Parkinson's disease. The nonamyloid component (NAC) region of alpha-syn shows strong tendencies to form beta-sheet structures, and deletion of this region has been shown to reduce aggregation and toxicity in vitro and in vivo. The binding of a molecular species to this region may mimic the effects of such deletions. Single-chain variable fragment (scFv) antibodies retain the binding specificity of antibodies and, when genetically manipulated to create high-diversity libraries, allow in vitro selection against peptides. Accordingly, we used a yeast surface display library of an entire naive repertoire of human scFv antibodies to select for binding to a NAC peptide. Candidate scFv antibodies (after transfer to mammalian expression vectors) were screened for viability in a neuronal cell line by transient cotransfection with A53T mutant alpha-syn. This provided a ranking of the protective efficacies of the initial panel of intracellular antibodies (intrabodies). High steady-state expression levels and apparent conformational epitope binding appeared more important than in vitro affinity in these assays. None of the scFv antibodies selected matched the sequences of previously reported anti-alpha-syn scFv antibodies. A stable cell line expressing the most effective intrabody, NAC32, showed highly significant reductions in abnormal aggregation in two separate models. Recently, intrabodies have shown promising antiaggregation and neuroprotective effects against misfolded mutant huntingtin protein. The NAC32 study extends such work significantly by utilizing information about the pathogenic capacity of a specific alpha-syn region to offer a new generation of in vitro-derived antibody fragments, both for further engineering as direct therapeutics and as a tool for rational drug design for Parkinson's disease.

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Dataset Information

Antioxidant Nanoparticles for Concerted Inhibition of ?-Synuclein Fibrillization, and Attenuation of Microglial Intracellular Aggregation and Activation.

Publications

Antioxidant Nanoparticles for Concerted Inhibition of α-Synuclein Fibrillization, and Attenuation of Microglial Intracellular Aggregation and Activation.

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