Mitochondrial Lysates Induce Inflammation and Alzheimer's Disease-Relevant Changes in Microglial and Neuronal Cells

Abstract

Neuroinflammation occurs in AD. While AD genetic studies implicate inflammation-relevant genes and fibrillar amyloid β protein promotes inflammation, our understanding of AD neuroinflammation nevertheless remains incomplete. In this study we hypothesized damage-associated molecular pattern (DAMP) molecules arising from mitochondria, intracellular organelles that resemble bacteria, could contribute to AD neuroinflammation. To preliminarily test this possibility, we exposed neuronal and microglial cell lines to enriched mitochondrial lysates. BV2 microglial cells treated with mitochondrial lysates showed decreased TREM2 mRNA, increased TNFα mRNA, increased MMP-8 mRNA, increased IL-8 mRNA, redistribution of NFκB to the nucleus, and increased p38 MAPK phosphorylation. SH-SY5Y neuronal cells treated with mitochondrial lysates showed increased TNFα mRNA, increased NFκB protein, decreased IκBα protein, increased AβPP mRNA, and increased AβPP protein. Enriched mitochondrial lysates from SH-SY5Y cells lacking detectable mitochondrial DNA (ρ0 cells) failed to induce any of these changes, while mtDNA obtained directly from mitochondria (but not PCR-amplified mtDNA) increased BV2 cell TNFα mRNA. These results indicate at least one mitochondrial-derived DAMP molecule, mtDNA, can induce inflammatory changes in microglial and neuronal cell lines. Our data are consistent with the hypothesis that a mitochondrial-derived DAMP molecule or molecules could contribute to AD neuroinflammation.

Extracellular Mitochondria and Mitochondrial Components Act as Damage-Associated Molecular Pattern Molecules in the Mouse Brain

“…Multiple mitochondrial components are already known to function as DAMP molecules in other tissues and experimental models (Galluzzi et al 2012; Nakahira et al 2015). We previously showed mitochondrial lysates that contain mtDNA, but not mitochondrial lysates from mtDNA-depleted ρ0 cells, activate inflammatory pathways in cultured neuronal and microglial cells (Wilkins et al 2015). Findings from our current in vivo study are consistent with those from the in vitro study, and allow us to extend those observations to the mammalian brain.…”

“…The amount of directly injected mtDNA likely exceeded the amount that was introduced when disrupted whole mitochondria were injected. Exposure to disrupted whole mitochondria, though, introduces other known mitochondrial DAMP molecules (Wilkins et al 2015). Based on these experiments it is difficult to know which mitochondrial components contributed to the observed neuroinflammation response, how much a given component contributed, or what amount of a component exceeds its threshold for activating a neuroinflammatory response.…”

“…Mitochondria were rapidly freeze-thawed three times. Alternatively, mtDNA was isolated from the mitochondrial fractions as previously described (Wilkins et al 2015). …”

“…We recently hypothesized mitochondrial DAMP molecules may contribute to neuroinflammation in Alzheimer’s disease (AD), and in a proof-of-principle in vitro study found mitochondrial lysates activated inflammation pathways in neuronal and microglial cell lines (Wilkins et al 2015). Here, we considered this hypothesis in an in vivo setting.…”

Mitochondrial Signaling and Neurodegeneration

“…TLRs are present in dendritic cells, macrophages, and microglia in the CNS, as well as nonimmune cells such as neurons and epithelial cells, where they activate the infl ammasome and pro-infl ammatory gene expression [ 175 ]. In microglia and neuronal cultures, mtDAMPs cause dopaminergic cell death [ 176 ] and increased amyloidosis with AD-associated infl ammation [ 177 ]. In animal models, mtDAMP infl ammasome activation is associated with cognitive impairment [ 178 ] and neurodegenerative disease [ 176 , 179 , 180 ].…”