Effect of clodronate on gene expression in the peripheral blood of horses.

There are two FDA-approved bisphosphonate products, clodronate (Osphos®) and tiludronate (Tildren®), for use in horses. It is hypothesized that bisphosphonates can produce analgesic effects and prevent proper healing of microcracks in bone. Therefore, bisphosphonate use is banned in racehorses. However, bisphosphonates have a short detection window in the blood before sequestration in the skeleton, making the reliability of current drug tests questionable. Seven exercising Thoroughbred horses were administered clodronate (1.8 mg/kg i.m.), and four were administered saline. RNA was isolated from peripheral blood mononuclear cells (PBMCs) collected immediately before a single dose of clodronate or saline and then on Days 1, 6, 28, 56 and 182 post-dose. mRNA was sequenced and analysed for differentially expressed transcripts. While no single transcripts were differentially expressed, pathway analysis revealed that p38 MAPK (p = .04) and Ras (p = .04) pathways were upregulated, and cadherin signalling (p = .02) was downregulated on Day 1. Previously investigated biomarkers, cathepsin K (CTSK) and type 5 acid phosphatase (ACP5), were analysed with RT-qPCR in a targeted gene approach, with no significant difference observed. A significant effect of time on gene expression for ACP5 (p = .03) and CTSK (p < .0001) was observed. Thus, these genes warrant further investigation for detecting clodronate use over time.

Previously Identified Genetic Variants in ADGRL3 Are not Associated with Risk for Equine Degenerative Myeloencephalopathy across Breeds.

Equine neuroaxonal dystrophy/equine degenerative myeloencephalopathy (eNAD/EDM) is a neurologic disease that has been reported in young horses from a wide range of breeds. The disease is inherited and associated with vitamin E deficiency during the first two years of life, resulting in bilateral symmetric ataxia. A missense mutation (chr3:71,917,591 C > T) within adhesion G protein-coupled receptor L3 (ADGRL3) was recently associated with risk for EDM in the Caspian breed. In order to confirm these findings, genotyping of this missense mutation, along with the three other associated single nucleotide polymorphisms (SNPs) in the genomic region, was carried out on 31 postmortem-confirmed eNAD/EDM cases and 43 clinically phenotyped controls from various breeds. No significant association was found between eNAD/EDM confirmed cases and genotype at any of the four identified SNPs (P > 0.05), including the nonsynonymous variant (EquCab2.0 chr3:71,917,591; allelic P = 0.85). These findings suggest that the four SNPs, including the missense variant in the ADGRL3 region, are not associated with risk for eNAD/EDM across multiple breeds of horses.