Increased α-tocopherol metabolism in horses with equine neuroaxonal dystrophy.

Abstract

BACKGROUND: Equine neuroaxonal dystrophy/equine degenerative myeloencephalopathy (eNAD/EDM) is an inherited neurodegenerative disorder associated with a vitamin E deficiency within the first year of life. Vitamin E consists of 8 isoforms metabolized by the CYP4F2 enzyme. No antemortem diagnostic test currently exists for eNAD/EDM. HYPOTHESIS/OBJECTIVES: Based on the association of &#x3b1;-tocopherol deficiency with the development of eNAD/EDM, we hypothesized that the rate of &#x3b1;-tocopherol, but not &#x3b3;-tocopherol or tocotrienol metabolism, would be increased in eNAD/EDM-affected horses. ANIMALS: Vitamin E metabolism: Proof of concept (POC) study; eNAD/EDM-affected (n&#xa0;=&#xa0;5) and control (n&#xa0;=&#xa0;6) horses. Validation study: eNAD/EDM-affected Quarter Horses (QHs; n&#xa0;=&#xa0;6), cervical vertebral compressive myelopathy affected (n&#xa0;=&#xa0;6) horses and control (n&#xa0;=&#xa0;29) horses. CYP4F2 expression and copy number: eNAD/EDM-affected (n&#xa0;=&#xa0;12) and age- and sex-matched control (n&#xa0;=&#xa0;11-12) horses. METHODS: The rates of &#x3b1;-tocopherol/tocotrienol and &#x3b3;-tocopherol/tocotrienol metabolism were assessed in equine serum (POC and validation) and urine (POC only) using liquid chromatography tandem mass spectrometry (LC-MS/MS). Quantitative reverse-transcriptase PCR (qRT-PCR) and droplet digital (dd)-PCR were used to assay expression and genomic copy number of a CYP4F2 equine ortholog. RESULTS: Metabolic rate of &#x3b1;-tocopherol was increased in eNAD/EDM horses (POC,P&#x2009;<&#x2009;.0001; validation, P&#xa0;=&#xa0;.03), with no difference in the metabolic rate of &#x3b3;-tocopherol. Horses with eNAD/EDM had increased expression of the CYP4F2 equine orthologue (P&#xa0;=&#xa0;.02) but no differences in copy number. CONCLUSIONS AND CLINICAL IMPORTANCE: Increased &#x3b1;-tocopherol metabolism in eNAD/EDM-affected QHs provides novel insight into alterations in vitamin E processing in eNAD/EDM and highlights the need for high-dose supplementation to prevent the clinical phenotype in genetically susceptible horses.