Researchers have discovered how α-synuclein (α-syn), a key protein in Parkinson’s disease and Lewy body dementia, leads to inflammation and disruption of the axons in the brain.
Failure of the barrier
Unlike other organs, the brain is heavily protected from many compounds in the bloodstream in order to prevent damage, with a unique combination of cells and junction proteins involved in this layered defense: this is the blood-brain barrier (BBB) [1]. As expected, damage to the BBB is directly related to neurodegenerative diseases [2].
This relationship has been very heavily studied for Alzheimer’s disease [3], but only limited work has been done to tie together the BBB and α-synucleinopathies [4]. While most work on α-syn has focused on how it directly damages cells in the context of Parkinson’s [5], these researchers note that BBB damage is likely to have its own negative effects that need to be included for accurate drug development [6].
Aggregates cause damage to cells
For accurate results, the researchers investigated both the usual, monomeric form of α-syn along with an aggregate called preformed fibril α-syn (PFF). They then introduced each of these proteins to endothelial cells that line the BBB (HBMVECs) in vitro.
Both regular α-syn and PFF were taken up by the HBMVECs. While these cells did not react to the aggregate within the first hour, the PFF fibrils were carried to lysosomes for processing within a day. Before it could be properly processed, however, PFF was found to lead to disruption of vascular endothelial cahedrin (VE-cadherin), a core protein of the BBB. This disruption led to increased penetration of a form of dextran thaat does not normally penetrate the BBB. Unaggregated α-syn, which is associated with normal function, did not have such effects.
A gene expression analysis revealed a link to inflammation. While there were no interesting differences between the monomeric α-syn group and the control group, the PFF group had significant differences. At 24 hours, a gene cluster that is related to inflammatory factors, such as NF-κB and TNF-α, was strongly upregulated compared to controls. Cellular proliferation and growth were also impeded according to this analysis.
Because genes are strongly inter-related and the underlying biology is exceptionally complicated, the researchers used an AI algorithm to determine what upstream pathways were responsible for this change in gene expression; many of these genes were well-known in the literature for being related to inflammation. They also found that TNF-α was uniquely upregulated, becoming elevated by nearly 250-fold within one hour compared to controls.
Critically, inhibiting TNF-α in these cells, while it did not completely reverse the effects, led to lower permeability of dextran. This suggests potential benefits for the BBB.
Heavy BBB leakage in mice
The researchers used mice that were modified to aggregate α-syn (G2-3 mice), first confirming that these aggregates were indeed found in both the brain tissue and in the vasculature. They then tested for claudin 5, a core protein responsible for BBB integrity. Unsurprisingly, they found that claudin 5 in G2-3 mice was significantly decreased from that of wild-type mice, although it took 13 months of aging for this damage to appear to a statisticallly significant degree. An antibody for immunoglobulin G (IgG) demonstrated a tremendous amount of BBB leakage: at 13 months, approximately six times as much IgG had worked its way into the brains of the G2-3 mice than those of wild-type mice.
Pericytes, along with the endfeet of astrocytes, are also part of BBB maintenance. Compared to wild-type controls, the G2-3 mice had more astrocytic activity, showing that they were working harder to compensate for the porous BBB. However, aquaporin 4, a protein involved in disposing of potentially dangerous waste, was depleted, which suggests that the astrocytes were overwhelmed. Similarly, a marker of pericyte activity, PDGFRβ, was upregulated, suggesting that these cells were also working much harder to defend the leaky BBB.
Small vessel disease (SVD) is a BBB failure that leads to nerve damage. Microglial inflammation near the vasculature, which is found in SVD, was also found in the G2-3 mice. Degeneration of the extracellular matrix was discovered, and there was evidence of axonal damage near the vasculature as well.
A potential treatment may already exist
In another experiment, the researchers used a wild-type strain of mice and injected their brains with PFF. They then dosed some of the mice with etanercept, a TNF-α inhibitor that is used to treat arthritis and does not normally penetrate the BBB. Compared to the mice that received PFF but not etanercept, the treated mice had significantly less IgG infiltration, nearly to the levels of the control group. Very significant effects were also found when etanercept was given to G2-3 mice, including benefits against the effects of SVD.
Etanercept was also found to have downstream benefits in mice. α-syn aggregation was found to cause damage to both novel object recognition, which measures cognitive function, and the rotarod test, which measures balance ability. Both of these metrics were improved with etanercept.
While there is still no evidence that this could work as a treatment for Parkinson’s in human beings, it is clear that BBB disruption and the resulting inflammation are likely to strongly contribute to Parkinson’s pathology. A clinical trial could validate whether etanercept or another drug that disrupts TNF-α could blunt the effects of this debilitating disease.
Literature
[1] Jeon, M. T., Kim, K. S., Kim, E. S., Lee, S., Kim, J., Hoe, H. S., & Kim, D. G. (2021). Emerging pathogenic role of peripheral blood factors following BBB disruption in neurodegenerative disease. Ageing research reviews, 68, 101333.
[2] Baloyannis, S. J., & Baloyannis, I. S. (2012). The vascular factor in Alzheimer’s disease: a study in Golgi technique and electron microscopy. Journal of the neurological sciences, 322(1-2), 117-121.
[3] Ryu, J. K., & McLarnon, J. G. (2009). A leaky blood–brain barrier, fibrinogen infiltration and microglial reactivity in inflamed Alzheimer’s disease brain. Journal of cellular and molecular medicine, 13(9a), 2911-2925.
[4] Pediaditakis, I., Kodella, K. R., Manatakis, D. V., Le, C. Y., Hinojosa, C. D., Tien-Street, W., … & Karalis, K. (2021). Modeling alpha-synuclein pathology in a human brain-chip to assess blood-brain barrier disruption. Nature communications, 12(1), 5907.
[5] Michel, P. P., Hirsch, E. C., & Hunot, S. (2016). Understanding dopaminergic cell death pathways in Parkinson disease. Neuron, 90(4), 675-691.
[6] Elabi, O., Gaceb, A., Carlsson, R., Padel, T., Soylu-Kucharz, R., Cortijo, I., … & Paul, G. (2021). Human α-synuclein overexpression in a mouse model of Parkinson’s disease leads to vascular pathology, blood brain barrier leakage and pericyte activation. Scientific reports, 11(1), 1120.
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