Single-Cell Multiome Analysis Reveals Neuroprotection by Elovanoids in Experimental Ischemic Stroke

Abstract

Ischemic stroke results in severe loss of neurological functions, and a large proportion of survivors display the onset of cognitive impairments. Administration of a novel class of very long-chain polyunsaturated fatty acids (VLC-PUFAs) termed elovanoids (ELV-N34) or its VLC-PUFA precursor (C34:6) are protective against both in vitro and in vivo models of uncompensated oxidative stress and ischemic stress. We found that ELV-N34 or C34:6 delivery attenuates sensorimotor deficits after ischemic stroke. Furthermore, we uncovered that ELV and its precursors protect against ischemic damage by modulating inflammatory signaling and increasing the expression of neuroprotective genes. We used 10X Single Cell Multiome (ATAC + Gene Expression) to compare genomic and epigenomic responses in naïve rats and after experimental ischemic treatment. Adult Sprague-Dawley rats underwent 2 hours of middle cerebral artery occlusion (MCAo) via intraluminal suture. Rats received vehicle, ELV-N34, or C34:6 delivered intranasally 1 hour, 1 day, and 2 days after 2 hours of MCAo. Brains were harvested on day 3 from the ipsilesional cortex (penumbra region) and subcortex. Nuclei were isolated and processed according to 10X Genomics Single Cell Multiome protocol. ELV-N34 or C34:6 treatment reduced neurological deficits after MCAo. Joint single-cell analysis revealed increases in microglia and leukocyte abundance as well as a loss of neuronal populations after MCAo. Differential gene expression revealed that ELV or its precursors protected against MCAo through complex mechanisms involving anti- inflammatory signaling and promoted neuroregeneration in microglia, astrocytes, neurons, and other neural cells. ELV/precursors exert their bioactivity by modulating conversion to neuroprotective cell phenotypes in neurodegenerative diseases such as stroke or Alzheimer's disease.