Abstract TP305: microRNA-182 Preserves Oxidative Phosphorylation During Substrate Limitation in Both Male and Female Primary Astrocyte Cultures

Abstract

Introduction: Stroke remains the leading cause of long-term disability in the United States, with therapy remaining limited to early reperfusion interventions. Adjuvants that aim to preserve neuronal survival could provide functional benefits to stroke survivors. Astrocytes promote neuronal survival by providing metabolic support to neurons in the absence of glucose availability, a critical substrate required for neuronal metabolism and survival. MicroRNAs (miRs) are non-coding RNAs that suppress translation of target genes. Our prior work identified miR-182 as a critical modulator of stroke by targeting genes that preserve astrocyte mitochondrial function. In the present study we assessed the role of miR-182 inhibition on oxygen consumption rate (OCR) during glucose deprivation (GD) injury. As stroke-induced mitochondrial dysfunction occurs mechanistically differently between males and females, we compared the effect of miR-182 inhibition between male and female astrocytes. Methods: Primary male and female cortical astrocyte cultures were generated from neonatal male and female C57BL/6 mice. Cultures were maintained at ambient O 2 tension and 5% CO 2 until day-in-vitro 12. Cultures were randomized to transfection with miR-182 inhibitor, or mismatch control RNA, 24h prior to 72h of GD injury. OCR was measured using the Lucid Scientific Recipher TM . Cell plating density was assessed using the nuclear stain DAPI. Results: No significant differences between treatments or between sexes in cell density were observed. GD injury resulted in a progressive decrease in OCR, requiring 12.5+7.8h to become significantly (p<0.05) lower in male astrocyte cultures, and 12.5+6.4h in females, with no difference between sexes. Comparatively, time-to-reach a lower OCR with miR-182 inhibitor required 38+7.1h in male cultures and 47+12.7h in females. The time-to-reach a lower OCR with miR-182 inhibition was significantly greater in both sexes versus control transfection, but not between sexes with miR-182 inhibition. Conclusions: These results suggest that inhibition of miR-182 represents a novel target to indirectly support neuronal metabolism during stroke that could be applied to both men and women survivors of stroke to improve functional outcomes.