Ischemia–Reperfusion Alters Retrotransposon Transcript Abundance in the Mouse Brain and Acidosis Conditions Are Associated with Transcript Changes in Neuro-2a Cells

Retrotransposons are repetitive DNA elements that are normally suppressed through epigenetic mechanisms. In aging and neurodegenerative diseases, abnormal retrotransposon activation occurs and can worsen neurotoxicity. However, it remains unclear whether ischemic stroke induces changes in retrotransposon transcript levels between infarct and peri-infarct brain tissue and whether the abundance of retrotransposon transcripts differs between these regions. To answer this question, we performed 45 min of transient middle cerebral artery occlusion (tMCAO) followed by reperfusion on adult wild-type male mice, then measured expression of retrotransposons LINE-1, IAP and SINE B2 in infarct and peri-infarct brain tissues using RT-qPCR. Next, we performed Western blotting to determine retroviral GAG protein expression. Since brain ischemia leads to tissue acidosis, which exhibits a positive correlation with ischemic brain injury, we further asked whether acidosis itself alters retrotransposon expression. We exposed neuro-2a (N2A) cells to acidic extracellular pH (6.4 or 6.0) and analyzed retrotransposon transcript levels. tMCAO significantly increased transcript abundance of LINE-1 Orf1, IAPEz-gag, IAPEY4 and B2 in ipsilateral ischemic brain tissue compared with sham controls at 24 h. Following correction of the four paired regional comparisons, only the difference in IAPEz-gag expression between infarct and corresponding contralateral region remained statistically significant. Western blotting analysis revealed changes in the abundance and patterns of GAG-immunoreactive proteins, including decreased precursor Pr65 abundance at 6 h of reperfusion, and reduced levels of both Pr65 and mature capsid p30 products at 24 h. In N2A cells, extracellular acidosis was associated with time-dependent increases in the abundance of LINE-1 Orf1, IAPEz-gag and B2 transcripts. Increased cellular injury was also observed under these conditions and may have contributed to these changes. These data suggest distinct retrotransposon transcript profiles between infarct and peri-infarct regions following focal ischemia–reperfusion. Although the in vitro results do not establish whether the transcript changes are attributable to extracellular acidosis, associated cellular injury, or both, exposure to acidic conditions was associated with a subset of the retrotransposon transcript changes observed in ischemic brain tissue. These findings provide a foundation for further investigation of retrotransposon dysregulation in stroke pathology.