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BACKGROUND: Type 2 diabetes (T2DM) is an established risk factor for developing Parkinson's disease (PD), but its effect on disease progression is not well understood. OBJECTIVE: The aim of this study was to investigate the influence of T2DM on aspects of disease progression in PD. METHODS: We analyzed data from the Tracking Parkinson's study to examine the effects of comorbid T2DM on PD progression and quality of life by comparing symptom severity scores assessing a range of motor and nonmotor symptoms. RESULTS: We identified 167 (8.7%) patients with PD and T2DM (PD + T2DM) and 1763 (91.3%) patients with PD without T2DM (PD). After controlling for confounders, patients with T2DM had more severe motor symptoms, as assessed by Movement Disorder Society Unified Parkinson's Disease Rating Scale, Part III (25.8 [0.9] vs. 22.5 [0.3] P = 0.002), and nonmotor symptoms, as assessed by Non-Motor Symptoms Scale total (38.4 [2.5] vs. 31.8 [0.7] P < 0.001), and were significantly more likely to report loss of independence (odds ratio, 2.08; 95% confidence interval [CI]: 1.34-3.25; P = 0.001) and depression (odds ratio, 1.62; CI: 1.10-2.39; P = 0.015). Furthermore, over time, patients with T2DM had significantly faster motor symptom progression (P = 0.012), developed worse mood symptoms (P = 0.041), and were more likely to develop substantial gait impairment (hazard ratio, 1.55; CI: 1.07-2.23; P = 0.020) and mild cognitive impairment (hazard ratio, 1.7; CI: 1.24-2.51; P = 0.002) compared with the PD group. CONCLUSIONS: In the largest study to date, T2DM is associated with faster disease progression in Parkinson's, highlighting an interaction between these two diseases. Because it is a potentially modifiable metabolic state, with multiple peripheral and central targets for intervention, it may represent a target for alleviating parkinsonian symptoms and slowing progression to disability and dementia. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Mutations in theSNCAgene cause autosomal dominant Parkinson’s disease (PD), with progressive loss of dopaminergic neurons in the substantia nigra, and accumulation of aggregates of α-synuclein. However, the sequence of molecular events that proceed from theSNCAmutation during development, to its end stage pathology is unknown. Utilising human induced pluripotent stem cells (hiPSCs) withSNCAmutations, we resolved the temporal sequence of pathophysiological events that occur during neuronal differentiation in order to discover the early, and likely causative, events in synucleinopathies. We adapted a small molecule-based protocol that generates highly enriched midbrain dopaminergic (mDA) neurons (>80%). We characterised their molecular identity using single-cell RNA sequencing and their functional identity through the synthesis and secretion of dopamine, the ability to generate action potentials, and form functional synapses and networks. RNA velocity analyses confirmed the developmental transcriptomic trajectory of midbrain neural precursors into mDA neurons using our approach, and identified key driver genes in mDA neuronal development. To characterise the synucleinopathy, we adopted super-resolution methods to determine the number, size and structure of aggregates inSNCA-mutant mDA neurons. At one week of differentiation, prior to maturation to mDA neurons of molecular and functional identity, we demonstrate the formation of small aggregates; specifically, β-sheet rich oligomeric aggregates, inSNCA-mutant midbrain immature neurons. The aggregation progresses over time to accumulate phosphorylated aggregates, and later fibrillar aggregates. When the midbrain neurons were functional, we observed evidence of impaired physiological calcium signalling, with raised basal calcium, and impairments in cytosolic and mitochondrial calcium efflux. Once midbrain identity fully developed,SNCA-mutant neurons exhibited bioenergetic impairments, mitochondrial dysfunction and oxidative stress. During the maturation of mDA neurons, upregulation of mitophagy and autophagy occured, and ultimately these multiple cellular stresses lead to an increase in cell death by six weeks post-differentiation. Our differentiation paradigm generates an efficient model for studying disease mechanisms in PD, and highlights that protein misfolding to generate intraneuronal oligomers is one of the earliest critical events driving disease in human neurons, rather than a late-stage hallmark of the disease.

Aggregation of α-Synuclein (α-Syn) drives Parkinson’s disease, although the initial stages of self-assembly and structural conversion have not been captured inside neurons. We track the intracellular conformational states of α-Syn utilizing a single-molecule FRET biosensor, and show that α-Syn converts from its monomeric state to form two distinct oligomeric states in neurons in a concentration dependent, and sequence specific manner. 3D FRET-CLEM reveals the structural organization, and location of aggregation hotspots inside the cell. Notably multiple intracellular seeding events occur preferentially on membrane surfaces, especially mitochondrial membranes. The mitochondrial lipid, cardiolipin triggers rapid oligomerization of A53T α-Syn, and cardiolipin is sequestered within aggregating lipid-protein complexes. Mitochondrial aggregates impair complex I activity and increase mitochondrial ROS generation, which accelerates the oligomerization of A53T α-Syn, and ultimately causes permeabilization of mitochondrial membranes, and cell death. Patient iPSC derived neurons harboring A53T mutations exhibit accelerated oligomerization that is dependent on mitochondrial ROS, early mitochondrial permeabilization and neuronal death. Our study highlights a mechanism of de novo oligomerization at the mitochondria and its induction of neuronal toxicity.

Protein aggregation and abnormal lipid homeostasis are both implicated in neurodegeneration through unknown mechanisms. Here we demonstrate that aggregate-membrane interaction is critical to induce a form of cell death called ferroptosis. Importantly, the aggregate-membrane interaction that drives ferroptosis depends both on the conformational structure of the aggregate, as well as the oxidation state of the lipid membrane. We generated human stem cell-derived models of synucleinopathy, characterized by the intracellular formation of α-synuclein aggregates that bind to membranes. In human iPSC-derived neurons with SNCA triplication, physiological concentrations of glutamate and dopamine induce abnormal calcium signaling owing to the incorporation of excess α-synuclein oligomers into membranes, leading to altered membrane conductance and abnormal calcium influx. α-synuclein oligomers further induce lipid peroxidation. Targeted inhibition of lipid peroxidation prevents the aggregate-membrane interaction, abolishes aberrant calcium fluxes, and restores physiological calcium signaling. Inhibition of lipid peroxidation, and reduction of iron-dependent accumulation of free radicals, further prevents oligomer-induced toxicity in human neurons. In summary, we report that peroxidation of polyunsaturated fatty acids underlies the incorporation of β-sheet-rich aggregates into the membranes, and that additionally induces neuronal death. This suggests a role for ferroptosis in Parkinson’s disease, and highlights a new mechanism by which lipid peroxidation causes cell death.

Aggregation of alpha-synuclein (α-Syn) drives Parkinson’s disease (PD), although the initial stages of self-assembly and structural conversion have not been directly observed inside neurons. In this study, we tracked the intracellular conformational states of α-Syn using a single-molecule Förster resonance energy transfer (smFRET) biosensor, and we show here that α-Syn converts from a monomeric state into two distinct oligomeric states in neurons in a concentration-dependent and sequence-specific manner. Three-dimensional FRET-correlative light and electron microscopy (FRET-CLEM) revealed that intracellular seeding events occur preferentially on membrane surfaces, especially at mitochondrial membranes. The mitochondrial lipid cardiolipin triggers rapid oligomerization of A53T α-Syn, and cardiolipin is sequestered within aggregating lipid–protein complexes. Mitochondrial aggregates impair complex I activity and increase mitochondrial reactive oxygen species (ROS) generation, which accelerates the oligomerization of A53T α-Syn and causes permeabilization of mitochondrial membranes and cell death. These processes were also observed in induced pluripotent stem cell (iPSC)–derived neurons harboring A53T mutations from patients with PD. Our study highlights a mechanism of de novo α-Syn oligomerization at mitochondrial membranes and subsequent neuronal toxicity.

The molecular events causing memory loss and neuronal cell death in Alzheimer’s disease (AD) over time are still unknown. Here we found that picomolar concentrations of soluble oligomers of synthetic beta amyloid (Aβ42) aggregates incubated with BV2 cells or rat astrocytes caused a sensitised response of Toll-like receptor 4 (TLR4) with time, leading to increased production of TNF-α. Aβ aggregates caused long term potentiation (LTP) deficit in hippocampal slices and predominantly neuronal cell death in co-cultures of astrocytes and neurons, which was blocked by TLR4 antagonists. Soluble Aβ aggregates cause LTP deficit and neuronal death via an autocrine/paracrine mechanism due to TLR4 signalling. These findings suggest that the TLR4-mediated inflammatory response may be a key pathophysiological process in AD., Hughes et al. investigate the TLR4-mediated inflammatory response in Alzheimer’s disease and show that picomolar concentrations of soluble amyloid beta aggregates lead to a sensitised response of TLR4 with time, resulting in increased TNF-α production. They suggest the use of near physiological concentration of soluble aggregates can cause long-term potentiation deficit and neuronal death through an autocrine/paracrine mechanism due to TLR4 signalling.

Protein aggregation and abnormal lipid homeostasis are both implicated in neurodegeneration through unknown mechanisms. Here we demonstrate that aggregate-membrane interaction is critical to induce a form of cell death called ferroptosis. Importantly, the aggregate-membrane interaction that drives ferroptosis depends both on the conformational structure of the aggregate, as well as the oxidation state of the lipid membrane. We generated human stem cell-derived models of synucleinopathy, characterized by the intracellular formation of α-synuclein aggregates that bind to membranes. In human iPSC-derived neurons with SNCA triplication, physiological concentrations of glutamate and dopamine induce abnormal calcium signaling owing to the incorporation of excess α-synuclein oligomers into membranes, leading to altered membrane conductance and abnormal calcium influx. α-synuclein oligomers further induce lipid peroxidation. Targeted inhibition of lipid peroxidation prevents the aggregate-membrane interaction, abolishes aberrant calcium fluxes, and restores physiological calcium signaling. Inhibition of lipid peroxidation, and reduction of iron-dependent accumulation of free radicals, further prevents oligomer-induced toxicity in human neurons. In summary, we report that peroxidation of polyunsaturated fatty acids underlies the incorporation of β-sheet-rich aggregates into the membranes, and that additionally induces neuronal death. This suggests a role for ferroptosis in Parkinson's disease, and highlights a new mechanism by which lipid peroxidation causes cell death.

Misfolding and aggregation of the proteins amyloid-β, tau and alpha-synuclein is the predominant pathology underlying the neurodegenerative disorders, Alzheimer's and Parkinson's disease. While end stage insoluble products of aggregation have been well characterised in human and animal models of disease, accumulating evidence from biophysical, cellular and in vivo studies has shown that soluble intermediates of aggregation, or oligomers, may be the key species that mediate toxicity and underlie seeding and spreading in disease. Here, we review the process of protein misfolding, and the intrinsic and extrinsic processes that cause the native states of the key aggregating proteins to undergo conformational change to form oligomers and ultimately fibrils. We discuss the structural features of the key toxic intermediate, and describe the putative mechanisms by which oligomers may cause cell toxicity. Finally, we explore the potential therapeutic approaches raised by the oligomer hypothesis in neurodegenerative disease.

Adult hippocampal neurogenesis is a highly dynamic process in which new cells are born, but only some of which survive. Of late it has become clear that these surviving newborn neurons have functional roles, most notably in certain forms of memory. Conventional methods to look at adult neurogenesis are based on the quantification of the number of newly born neurons using a simple cell counting methodology. However, this type of approach fails to capture the dynamic aspects of the neurogenic process, where neural proliferation, death and differentiation take place continuously and simultaneously. In this paper, we propose a simple mathematical approach to better understand the adult neurogenic process in the hippocampus which in turn will allow for a better analysis of this process in disease states and following drug therapies.

Background The aggregation of the protein ɑ-synuclein (ɑS) underlies a range of increasingly common neurodegenerative disorders including Parkinson’s disease. One widely explored therapeutic strategy for these conditions is the use of antibodies to target aggregated ɑS, although a detailed molecular-level mechanism of the action of such species remains elusive. Here, we characterize ɑS aggregation in vitro in the presence of two ɑS-specific single-domain antibodies (nanobodies), NbSyn2 and NbSyn87, which bind to the highly accessible C-terminal region of ɑS. Results We show that both nanobodies inhibit the formation of ɑS fibrils. Furthermore, using single-molecule fluorescence techniques, we demonstrate that nanobody binding promotes a rapid conformational conversion from more stable oligomers to less stable oligomers of ɑS, leading to a dramatic reduction in oligomer-induced cellular toxicity. Conclusions The results indicate a novel mechanism by which diseases associated with protein aggregation can be inhibited, and suggest that NbSyn2 and NbSyn87 could have significant therapeutic potential. Electronic supplementary material The online version of this article (doi:10.1186/s12915-017-0390-6) contains supplementary material, which is available to authorized users.

Huntington's disease (HD) is an inherited neurodegenerative disorder that is classically defined by a triad of movement and cognitive and psychiatric abnormalities with a well-established pathology that affects the dopaminergic systems of the brain. This has classically been described in terms of an early loss of dopamine D2 receptors (D2R), although interestingly the treatments most effectively used to treat patients with HD block these same receptors. We therefore sought to examine the dopaminergic system in HD not only in terms of striatal function but also at extrastriatal sites especially the hippocampus, given that transgenic (Tg) mice also exhibit deficits in hippocampal-dependent cognitive tests and a reduction in adult hippocampal neurogenesis. We showed that there was an early reduction of D2R in both the striatum and dentate gyrus (DG) of the hippocampus in the R6/1 transgenic HD mouse ahead of any overt motor signs and before striatal neuronal loss. Despite downregulation of D2Rs in these sites, further reduction of the dopaminergic input to these sites by either medial forebrain bundle lesions or receptor blockade using sulpiride was able to improve both deficits in motor performance and adult hippocampal neurogenesis. In contrast, a reduction in dopaminergic innervation of the neurogenic niches resulted in impaired neurogenesis in healthy WT mice. This study therefore provides evidence that D2R blockade improves hippocampal and striatal deficits in HD mice although the underlying mechanism for this is unclear, and suggests that agents working within this network may have greater effects than previously thought.

The protein alpha-synuclein (αS) self-assembles into small oligomeric species and subsequently into amyloid fibrils that accumulate and proliferate during the development of Parkinson's disease. However, the quantitative characterization of the aggregation and spreading of αS remains challenging to achieve. Previously, we identified a conformational conversion step leading from the initially formed oligomers to more compact oligomers preceding fibril formation. Here, by a combination of single-molecule fluorescence measurements and kinetic analysis, we find that the reaction in solution involves two unimolecular structural conversion steps, from the disordered to more compact oligomers and then to fibrils, which can elongate by further monomer addition. We have obtained individual rate constants for these key microscopic steps by applying a global kinetic analysis to both the decrease in the concentration of monomeric protein molecules and the increase in oligomer concentrations over a 0.5-140-µM range of αS. The resulting explicit kinetic model of αS aggregation has been used to quantitatively explore seeding the reaction by either the compact oligomers or fibrils. Our predictions reveal that, although fibrils are more effective at seeding than oligomers, very high numbers of seeds of either type, of the order of 10(4), are required to achieve efficient seeding and bypass the slow generation of aggregates through primary nucleation. Complementary cellular experiments demonstrated that two orders of magnitude lower numbers of oligomers were sufficient to generate high levels of reactive oxygen species, suggesting that effective templated seeding is likely to require both the presence of template aggregates and conditions of cellular stress.

Presence of α-synuclein aggregates in the brain is one of the major pathological hallmarks of Parkinson's disease (PD) and a point mutation in the gene encoding a-synuclein has been reported in familial PD cases. Previously, we have shown that for wild-type α-synuclein aggregation into oligomeric status is required for mitochondrial regulated cell toxicity. However, there is little understanding of how variant strains (monomers, oligomers, fibrils) of the α-synuclein mutations implicate neuronal vulnerability associated with PD. Here using various α-synuclein strains, monomers, oligomers and fibrils, carrying A30P, A53T or E46K point mutation, we examined the effect of these different forms of the mutants on neuronal survival following 24 hours incubation and the immediate effect on mitochondrial membrane potential as well as reactive oxygen spices (ROS) production using live cell imaging. We also measured the cellular redox state following exposure to the mutants using NADH autofluorescence. Our results show that unlike wild-type α-synuclein, all three mutant forms of α-synuclein induce cell toxicity and loss of mitochondrial membrane potential in their unfolded monomeric forms, prior to their aggregation into oligomeric species. Importantly, the different mutations A30P and A53T lead to an increase in cellular ROS production through different mechanisms. This work suggests that point mutations in α-synuclein, in addition to misfolding and aggregation of α-synuclein, are sufficient to cause neuronal vulnerability including cell death through oxidative stress related to mitochondrial dysfunction and excessive ROS.

In the original publication of this article, the author Magarida Rodrigues was written incorrectly.

Parkinson's disease (PD) is a common neurodegenerative disease characterised initially loss of dopaminergic neurons in the substantia nigra (SN) and later widespread nondopaminergic neuronal loss including in the cortex. Genetic mutations can cause familial PD, but the sequence of molecular events that trigger neuronal vulnerability is poorly understood. Of the familial forms of PD, point mutations of gene encoding alpha-synuclein (SNCA), the major component of the predominant pathological hallmark called Lewy bodies/neurites, result in dominant PD. To characterize the cellular pathophysiology, we differentiated patient-specific induced pluripotent stem cells (iPSCs) from PD patient fibroblasts carrying alpha-synuclein point mutation, A53T, into functional cortical and midbrain neurons. We demonstrated that endogenous levels of SNCA-A53T reduce cell survival and the source of vulnerability is the excess reactive oxygen species (ROS) evidenced by increased superoxide production and lipid peroxidation levels compared to control. By utilizing the power of patient specific iPSC derived neurons, this study provides an insight into the primary pathological events leading to neuronal cell death and may furthermore be an important direct tool for therapeutic development.

Adult hippocampal neurogenesis is a highly dynamic process in which new cells are born, but only some of which survive. Of late it has become clear that these surviving newborn neurons have functional roles, most notably in certain forms of memory. Conventional methods to look at adult neurogenesis are based on the quantification of the number of newly born neurons using a simple cell counting methodology. However, this type of approach fails to capture the dynamic aspects of the neurogenic process, where neural proliferation, death and differentiation take place continuously and simultaneously. In this paper, we propose a simple mathematical approach to better understand the adult neurogenic process in the hippocampus which in turn will allow for a better analysis of this process in disease states and following drug therapies.

Neurogenesis and Spatial Memory The dentate gyrus of the hippocampus is one of two sites in the brain where new neurons are produced throughout life. Adult-born neurons integrate into the dentate gyrus circuitry and are thought to play a role in learning and memory. However, their contribution to hippocampal function remains unclear. Clelland et al. (p. 210 ) disrupted neurogenesis in mice and used two behavioral tasks to test for impairment in the formation of uncorrelated episodic memory representations. In one task, two arms were presented and the mice were rewarded for choosing the most recently visited arm in an earlier sequence; in the second task, animals were rewarded for choosing a certain location on a touch screen. Ablation of neurogenesis affected discrimination performance in both tasks but only when the arms or screen locations were close to one another. Neurogenesis is thus necessary for spatial pattern separation in the dentate gyrus.

Despite the wealth of genomic and transcriptomic data in Parkinson's disease (PD), the initial molecular events are unknown. Using LD score regression analysis, we show significant enrichment in PD heritability within regulatory sites for LPS-activated monocytes and that TLR4 expression is highest within human substantia nigra, the most affected brain region, suggesting a role for TLR4 inflammatory responses. We then performed extended incubation of cells with physiological concentrations of small alpha-synuclein oligomers observing the development of a TLR4-dependent sensitized inflammatory response with time, including TNF-α production. ROS and cell death in primary neuronal cultures were significantly reduced by TLR4 antagonists revealing that an indirect inflammatory mechanism involving cytokines produced by glial cells makes a major contribution to neuronal death. Prolonged exposure to low levels of alpha-synuclein oligomers sensitizes TLR4 responsiveness in astrocytes and microglial, explaining how they become pro-inflammatory, and may be an early causative event in PD.

Despite the wealth of genomic and transcriptomic data in Parkinson's disease (PD), the initial molecular events are unknown. Using LD score regression analysis, we show significant enrichment in PD heritability within regulatory sites for LPS-activated monocytes and that TLR4 expression is highest within human substantia nigra, the most affected brain region, suggesting a role for TLR4 inflammatory responses. We then performed extended incubation of cells with physiological concentrations of small alpha-synuclein oligomers observing the development of a TLR4-dependent sensitized inflammatory response with time, including TNF-α production. ROS and cell death in primary neuronal cultures were significantly reduced by TLR4 antagonists revealing that an indirect inflammatory mechanism involving cytokines produced by glial cells makes a major contribution to neuronal death. Prolonged exposure to low levels of alpha-synuclein oligomers sensitizes TLR4 responsiveness in astrocytes and microglial, explaining how they become pro-inflammatory, and may be an early causative event in PD.

The molecular events causing memory loss and neuronal cell death in Alzheimer's disease (AD) over time are still unknown. Here we found that picomolar concentrations of soluble oligomers of synthetic beta amyloid (Aβ42) aggregates incubated with BV2 cells or rat astrocytes caused a sensitised response of Toll-like receptor 4 (TLR4) with time, leading to increased production of TNF-α. Aβ aggregates caused long term potentiation (LTP) deficit in hippocampal slices and predominantly neuronal cell death in co-cultures of astrocytes and neurons, which was blocked by TLR4 antagonists. Soluble Aβ aggregates cause LTP deficit and neuronal death via an autocrine/paracrine mechanism due to TLR4 signalling. These findings suggest that the TLR4-mediated inflammatory response may be a key pathophysiological process in AD.

The molecular events causing memory loss and neuronal cell death in Alzheimer's disease (AD) over time are still unknown. Here we found that picomolar concentrations of soluble oligomers of synthetic beta amyloid (Aβ42) aggregates incubated with BV2 cells or rat astrocytes caused a sensitised response of Toll-like receptor 4 (TLR4) with time, leading to increased production of TNF-α. Aβ aggregates caused long term potentiation (LTP) deficit in hippocampal slices and predominantly neuronal cell death in co-cultures of astrocytes and neurons, which was blocked by TLR4 antagonists. Soluble Aβ aggregates cause LTP deficit and neuronal death via an autocrine/paracrine mechanism due to TLR4 signalling. These findings suggest that the TLR4-mediated inflammatory response may be a key pathophysiological process in AD.