Degraded mitochondrial DNA is a newly identified subtype of the damage associated molecular pattern (DAMP) family and possible trigger of neurodegeneration.

We previously showed a preferential degradation and down-regulation of mitochondrial DNA and RNA in hamster fibroblasts in response to hydrogen peroxide. Subsequent studies by others demonstrated that mitochondrial DNA can stimulate immune cells as a DAMP (damage associated molecular patterns) family member. However, the actual physical structure of this mitochondrial DNA DAMP and its importance in non-immune cell types are poorly understood. Here we report that transfected oxidant-initiated degraded mitochondrial polynucleotides, which we term "DeMPs", strongly induce the proinflammatory cytokines interleukin 6, monocyte chemotactic protein-1, and tumor necrosis factor α in mouse primary astrocytes. Additionally, proinflammatory IL1ß was induced, implicating DeMPs in inflammasome activation. Furthermore, human cerebrospinal fluid (CSF) and plasma were found to contain detectable DeMP signal. Finally, significant degradation of mitochondrial DNA was observed in response to either a bolus or steady state hydrogen peroxide. Combined, these studies demonstrate, all for the first time, that a pathophysiologically relevant form of mitochondrial DNA (degraded) can elicit a proinflammatory cytokine induction; that a brain cell type (astrocytes) elicits a proinflammatory cytokine induction in response to these DeMPs; that this induction includes the inflammasome; that astrocytes are capable of inflammasome activation by DeMPs; that DeMPs are detectable in CSF and plasma; and that hydrogen peroxide can stimulate an early stage cellular degradation of mitochondrial DNA. These results provide new insights and are supportive of our hypothesis that DeMPs are a newly identified trigger of neurodegenerative diseases such as Alzheimer's disease, which are known to be associated with early stage inflammation and oxidation.

Ethanol alters BDNF-induced Rho GTPase activation in axonal growth cones.

BACKGROUND: The effects of ethanol on development of postmitotic neurons include altered neurite outgrowth and differentiation, which may contribute to neuropathology associated with fetal alcohol spectrum disorders. We previously reported that ethanol exposure alters axon growth dynamics in dissociated cultures of rat hippocampal pyramidal neurons. Given the important regulatory role of small Rho guanosine triphosphatases (GTPases) in cytoskeletal reorganization associated with axon growth, and reports that ethanol alters whole cell Rho GTPase activity in other cell types, this study explored the hypothesis that ethanol alters Rho GTPase activity specifically in axonal growth cones. METHODS: Fetal rat hippocampal pyramidal neurons were maintained in dissociated cultures for 1 day in control medium or medium containing 11 to 43 mM ethanol. Some cultures were also treated with brain-derived neurotrophic factor (BDNF), an activator of Rac1 and Cdc42 GTPases that promotes axon extension. Levels of active Rho GTPases in growth cones were measured using in situ binding assays for GTP-bound Rac1, Cdc42, and RhoA. Axon length, growth cone area, and growth cone surface expression of tyrosine kinase B (TrkB), the receptor for BDNF, were assessed by digital morphometry and immunocytochemistry. RESULTS: Although ethanol increased the surface area of growth cones, the levels of active Rho GTPases in axonal growth cones were not affected in the absence of exogenous BDNF. In contrast, ethanol exposure inhibited BDNF-induced Rac1/Cdc42 activation in a dose-dependent manner and increased RhoA activation at the highest concentration tested. Similar TrkB expression was observed on the surface of axonal growth cones of control and ethanol-treated neurons. CONCLUSIONS: These results reveal an inhibitory effect of ethanol on growth cone signaling via small Rho GTPases during early stages of hippocampal development in vitro, and suggest a mechanism whereby ethanol may disrupt neurotrophic factor regulation of axon growth and guidance.