CYP7A1 Is Significantly Up-Regulated in Spinal Cord from Horses Affected with Equine Neuroaxonal Dystrophy (NAD)

Equine neuroaxonal dystrophy (NAD) is an inherited neurodegenerative disease characterized by low α-tocopherol (α-TP) concentrations. Horses with NAD have histologic lesions identical to human spinocerebellar ataxias caused by mutations in genes involved in α-TP absorption and transport. Studies have been unable to map a chromosomal locus for equine NAD. The purpose of the present study was to determine if differential gene expression analysis would identify genes involved in α-TP metabolism and/or the pathogenesis of NAD. Cervical spinal cord (C1) rather than brainstem was selected for RNA-Seq analysis based on identification of primary segregation by disease phenotype on principal component analysis. RNA-Seq was performed on C1 from six NAD-affected and six sex-matched control yearlings at 20 million 100pb PE reads/horse. Differential gene expression analysis identified 481 significantly (FDR=0.05, minimum alignment count=10) down-regulated and 336 significantly up-regulated genes. Of the 817 significantly dysregulated genes, 75 were down-regulated with log₂ fold change<2 and 10 were up-regulated with log₂ fold change>2. Of these, only one highly up-regulated gene, CYP7A1 (log₂ fold change= 3.43), is potentially involved in α-TP bioactivity, lipid transport or metabolism. Quantitative RT-PCR demonstrated a 5.6 fold up-regulation (P=0.006) of CYP7A1 expression in the NAD-affected versus control horses (n = 11 and 9, respectively), validating results obtained with RNA-Seq. Within the CYP gene family, a CYP4f14-null mouse model has been shown to have clear disruption of α-TP metabolism. We therefore hypothesize that increased metabolism of α-TP due to overexpression of CYP7A1 in the spinal cord may underlie the pathophysiology of equine NAD.