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Structurally distinct ?-synuclein fibrils induce robust parkinsonian pathology.

ABSTRACT: OBJECTIVE:Alpha-synuclein (?-syn) is a major component of Lewy bodies, which are the pathological hallmark in Parkinson's disease, and its genetic mutations cause familial forms of Parkinson's disease. Patients with ?-syn G51D mutation exhibit severe clinical symptoms. However, in vitro studies showed low propensity for ?-syn with the G51D mutation. We studied the mechanisms associated with severe neurotoxicity of ?-syn G51D mutation using a murine model generated by G51D ?-syn fibril injection into the brain. METHODS:Structural analysis of wild-type and G51D ?-syn-fibrils were performed using Fourier transform infrared spectroscopy. The ability of ?-syn fibrils forming aggregates was first assessed in in vitro mammalian cells. An in vivo mouse model with an intranigral injection of ?-syn fibrils was then used to evaluate the propagation pattern of ?-syn and related cellular changes. RESULTS:We found that G51D ?-syn fibrils have higher ?-sheet contents than wild-type ?-syn fibrils. The addition of G51D ?-syn fibrils to mammalian cells overexpressing ?-syn resulted in the formation of phosphorylated ?-syn inclusions at a higher rate. Similarly, an injection of G51D ?-syn fibrils into the substantia nigra of a mouse brain induced more widespread phosphorylated ?-syn pathology. Notably, the mice injected with G51D ?-syn fibrils exhibited progressive nigral neuronal loss accompanied with mitochondrial abnormalities and motor impairment. CONCLUSION:Our findings indicate that the structural difference of G51D ?-syn fibrils plays an important role in the rapidly developed and more severe neurotoxicity of G51D mutation-linked Parkinson's disease. © 2019 International Parkinson and Movement Disorder Society.

SUBMITTER: Hayakawa H

PROVIDER: S-EPMC7202333 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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Project description:Previous studies demonstrate that intrastriatal injections of fibrillar alpha-synuclein (?-syn) into mice induce Parkinson's disease (PD)-like Lewy body (LB) pathology formed by aggregated ?-syn in anatomically interconnected regions and significant nigrostriatal degeneration. The aim of the current study was to evaluate whether exogenous mouse ?-syn pre-formed fibrils (PFF) injected into the striatum of rats would result in accumulation of LB-like intracellular inclusions and nigrostriatal degeneration. Sprague-Dawley rats received unilateral intrastriatal injections of either non-fibrillized recombinant ?-syn or PFF mouse ?-syn in 1- or 2- sites and were euthanized at 30, 60 or 180 days post-injection (pi). Both non-fibrillized recombinant ?-syn and PFF ?-syn injections resulted in phosphorylated ?-syn intraneuronal accumulations (i.e., diffuse Lewy neurite (LN)- and LB-like inclusions) with significantly greater accumulations following PFF injection. LB-like inclusions were observed in several areas that innervate the striatum, most prominently the frontal and insular cortices, the amygdala, and the substantia nigra pars compacta (SNpc). ?-Syn accumulations co-localized with ubiquitin, p62, and were thioflavin-S-positive and proteinase-k resistant, suggesting that PFF-induced pathology exhibits properties similar to human LBs. Although ?-syn inclusions within the SNpc remained ipsilateral to striatal injection, we observed bilateral reductions in nigral dopamine neurons at the 180-day time-point in both the 1- and 2-site PFF injection paradigms. PFF injected rats exhibited bilateral reductions in striatal dopaminergic innervation at 60 and 180 days and bilateral decreases in homovanillic acid; however, dopamine reduction was observed only in the striatum ipsilateral to PFF injection. Although the level of dopamine asymmetry in PFF injected rats at 180 days was insufficient to elicit motor deficits in amphetamine-induced rotations or forelimb use in the cylinder task, significant disruption of ultrasonic vocalizations was observed. Taken together, our findings demonstrate that ?-syn PFF are sufficient to seed the pathological conversion and propagation of endogenous ?-syn to induce a progressive, neurodegenerative model of ?-synucleinopathy in rats.

Project description:Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a central regulator of cellular stress responses and its transcriptional activation promotes multiple cellular defense and survival mechanisms. The loss of NRF2 has been shown to increase oxidative and proteotoxic stress, two key pathological features of neurodegenerative disorders such as Parkinson's disease (PD). Moreover, compromised redox homeostasis and protein quality control can cause the accumulation of pathogenic proteins, including alpha-synuclein (α-Syn) which plays a key role in PD. However, despite this link, the precise mechanisms by which NRF2 may regulate PD pathology is not clear. In this study, we generated a humanized mouse model to study the importance of NRF2 in the context of α-Syn-driven neuropathology in PD. Specifically, we developed NRF2 knockout and wild-type mice that overexpress human α-Syn (hα-Syn+/Nrf2-/- and hα-Syn+/Nrf2+/+ respectively) and tested changes in their behavior through nest building, challenging beam, and open field tests at three months of age. Cellular and molecular alterations in α-Syn, including phosphorylation and subsequent oligomerization, as well as changes in oxidative stress, inflammation, and autophagy were also assessed across multiple brain regions. It was observed that although monomeric α-Syn levels did not change, compared to their wild-type counterparts, hα-Syn+/Nrf2-/- mice exhibited increased phosphorylation and oligomerization of α-Syn. This was associated with a loss of tyrosine hydroxylase expressing dopaminergic neurons in the substantia nigra, and more pronounced behavioral deficits reminiscent of early-stage PD, in the hα-Syn+/Nrf2-/- mice. Furthermore, hα-Syn+/Nrf2-/- mice showed significantly amplified oxidative stress, greater expression of inflammatory markers, and signs of increased autophagic burden, especially in the midbrain, striatum and cortical brain regions. These results support an important role for NRF2, early in PD progression. More broadly, it indicates NRF2 biology as fundamental to PD pathogenesis and suggests that targeting NRF2 activation may delay the onset and progression of PD.

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Structurally distinct ?-synuclein fibrils induce robust parkinsonian pathology.

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