Exogenous administration of mitochondrial DNA promotes ischemia reperfusion injury via TLR9-p38 MAPK pathway.

Previous studies have shown a role of mitochondrial DNA (mtDNA) in innate immunity. However, the specific role of mtDNA in acute myocardial infarction remains elusive. This study was designed to examine the damaging effect of mtDNA on cardiomyocytes. H9c2s cells were incubated with purified mtDNA or nuclear DNA with or without pretreatment by chloroquine, an inhibitor of Toll-like receptor 9(TLR9). The cell viability was tested by MTT. To demonstrate the toxicity of mtDNA, mtDNA fragments were injected into rats 10 min before ischemia for 30 min and reperfusion for 24 h. Infarct size was measured by TTC staining. Apoptosis of myocardium was detected by TUNEL staining and caspase-3 activity. The levels of TLR9, p-p38 MAPK, and p38 MAPK were detected by western blotting. The results showed that exogenous mtDNA reduced the viability of H9c2s cells and induced TLR9 expression, caspase 3 activation and p38 mitogen-activated protein kinase (MAPK) phosphorylation. However, these effects were inhibited by chloroquine. In contrast, nuclear DNA did not have these effects. Intravenous injection of mtDNA into rats aggravated ischemia-reperfusion injury and increased infarction area through TLR9-p38 MAPK activation. We concluded that mtDNA released into the circulation by AMI may has detrimental effect on myocardium through aggravating ischemia-reperfusion injury via TLR9-p38 MAPK pathway.

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

Mitochondria and mitochondrial debris are found in the brain's extracellular space, and extracellular mitochondrial components can act as damage associated molecular pattern (DAMP) molecules. To characterize the effects of potential mitochondrial DAMP molecules on neuroinflammation, we injected either isolated mitochondria or mitochondrial DNA (mtDNA) into hippocampi of C57BL/6 mice and seven days later measured markers of inflammation. Brains injected with whole mitochondria showed increased Tnfα and decreased Trem2 mRNA, increased GFAP protein, and increased NFκB phosphorylation. Some of these effects were also observed in brains injected with mtDNA (decreased Trem2 mRNA, increased GFAP protein, and increased NFκB phosphorylation), and mtDNA injection also caused several unique changes including increased CSF1R protein and AKT phosphorylation. To further establish the potential relevance of this response to Alzheimer's disease (AD), a brain disorder characterized by neurodegeneration, mitochondrial dysfunction, and neuroinflammation we also measured App mRNA, APP protein, and Aß1-42 levels. We found mitochondria (but not mtDNA) injections increased these parameters. Our data show that in the mouse brain extracellular mitochondria and its components can induce neuroinflammation, extracellular mtDNA or mtDNA-associated proteins can contribute to this effect, and mitochondria derived-DAMP molecules can influence AD-associated biomarkers.

Mitochondrial DNA toxicity in forebrain neurons causes apoptosis, neurodegeneration, and impaired behavior.

Mitochondrial dysfunction underlying changes in neurodegenerative diseases is often associated with apoptosis and a progressive loss of neurons, and damage to the mitochondrial genome is proposed to be involved in such pathologies. In the present study we designed a mouse model that allows us to specifically induce mitochondrial DNA toxicity in the forebrain neurons of adult mice. This is achieved by CaMKIIalpha-regulated inducible expression of a mutated version of the mitochondrial UNG DNA repair enzyme (mutUNG1). This enzyme is capable of removing thymine from the mitochondrial genome. We demonstrate that a continual generation of apyrimidinic sites causes apoptosis and neuronal death. These defects are associated with behavioral alterations characterized by increased locomotor activity, impaired cognitive abilities, and lack of anxietylike responses. In summary, whereas mitochondrial base substitution and deletions previously have been shown to correlate with premature and natural aging, respectively, we show that a high level of apyrimidinic sites lead to mitochondrial DNA cytotoxicity, which causes apoptosis, followed by neurodegeneration.