Transcriptome diversity and differential expression in supporting limb laminitis.

Laminitis results in impaired tissue integrity and Inflammation of the epidermal and dermal lamellae connecting the hoof capsule to the underlying distal phalanx and causes loss-of-use, poor quality of life and euthanasia in horses. Historically, studies to better understand the etiology of laminitis by documenting changes in gene expression were hampered by the paucity of gene annotation specific to hoof tissues. Next-generation sequencing enables improvements to annotation by incorporating equine- and hoof-specific transcripts. Here we characterize the hoof lamellar tissue transcriptome of naturally occurring supporting limb laminitis (SLL) using archived lamellar tissue from Thoroughbred racehorses consisting of 13 SLL hospital cases and seven age-matched control horses. This was achieved using: 1) Applied transcriptome annotation by long-read sequencing to document transcript diversity and 2) short-read RNA sequencing to document changes in gene expression correlating to the developmental and acute stages of naturally occurring SLL. 1.99Gbp of long-read transcriptome sequencing deeply documented 5067 unique loci, while short read RNA-seq under very stringent quality filters described 66 differentially expressed loci. Functional analysis of these loci revealed alterations in cell replication and growth, stress response and leukocyte recruitment and activation pathways. Differential expression of the Ezrin and TIMP3 genes suggests they may have utility as biomarkers for laminitis disease, while NR1D1 and genes relevant to the inflammasome are promising targets for novel pharmacological treatments.