Reduction of Unscheduled R-Loops Accumulation Improves the Spinal Cord Injury Microenvironment

Spinal cord injury (SCI) is a severe condition that leads to dysfunction in motor, sensory, and autonomic systems, significantly impairing quality of life and imposing a substantial burden on healthcare systems. The pathological processes following SCI involve complex biochemical responses, including inflammation, oxidative stress, and mitochondrial dysfunction, all of which are closely related to genomic stability and DNA damage responses. Although extensive research has explored the mechanisms of neuronal death, microglial activation, and astrocyte responses following SCI, the role of DNA-RNA hybrids (i.e., R-loops) accumulation in these processes remains insufficiently studied.In this study, we used DRIPc-Seq (DNA-RNA immunoprecipitation combined with high-throughput sequencing) technology to investigate the dynamic changes of R-loops during the progression of SCI. Our findings revealed that R-loops rapidly accumulated during the early stages (3 days and 7 days) after SCI and significantly decreased at later stages (14 days and 28 days), suggesting that R-loop dynamics are closely related to transcriptional activity in SCI. Additionally, we examined the impact of non-canonical R-loops on cell death pathways, specifically neuronal apoptosis and ferroptosis, and their role in exacerbating tissue damage. The accumulation of R-loops promoted neuronal death in primary neurons and aggravated inflammation and glial scar formation in microglia and astrocytes.We also found that overexpression of Rnase H1 significantly reduced R-loop accumulation, alleviated inflammation, and promoted functional recovery in SCI models. Our results suggest that modulating R-loop levels may serve as a novel therapeutic strategy to promote neural repair.This study not only expands our understanding of SCI pathophysiology but also provides a theoretical foundation for future research on targeting R-loop accumulation as a therapeutic strategy for SCI and other neurodegenerative diseases.