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Use of laser capture microdissection for the assessment of equine lamellar basal epithelial cell signalling in the early stages of laminitis
Author(s) -
Leise B. S.,
Watts M. R.,
Roy S.,
Yilmaz A. S.,
Alder H.,
Belknap J. K.
Publication year - 2015
Publication title -
equine veterinary journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.82
H-Index - 87
eISSN - 2042-3306
pISSN - 0425-1644
DOI - 10.1111/evj.12283
Subject(s) - laminitis , laser capture microdissection , biology , transcriptome , microdissection , basal (medicine) , pathology , gene expression , microbiology and biotechnology , gene , endocrinology , medicine , horse , genetics , diabetes mellitus , paleontology
Summary Reasons for performing study Dysadhesion of laminar basal epithelial cells ( LBECs ) from the underlying dermis is the central event leading to structural failure in equine laminitis. Although many studies of sepsis‐related laminitis have reported multiple events occurring throughout the lamellar tissue, there is minimal information regarding signalling events occurring specifically in LBECs . Objectives To determine signalling events in the LBECs during the early stages of carbohydrate‐induced laminitis. Study design Experimental study. Methods Eight horses were given an overload of carbohydrate ( CHO ) consisting of corn starch mixture via nasogastric tube. Prior to administration of CHO , lamellar biopsies were taken from the left forefoot (control [ CON ]). Biopsies were taken from the left hind foot at the onset of fever (developmental [ DEV ]) and from the right forefoot at the onset of Obel grade 1 lameness ( OG1 ). Laminar basal epithelial cells were isolated from cryosections using a laser capture microdissection ( LCM ) microscope. Next generation sequencing ( RNA ‐seq) was used to identify transcripts expressed in the LBECs for each time point and bioinformatic analysis was performed with thresholds for between group comparisons set at a greater than 2‐fold change and P value ≤0.05. Results Forty genes (22 increased/18 decreased) were significantly different from DEV time vs. CON and 107 genes (57 increased/50 decreased) were significantly different from OG1 time vs. CON . Significant increases in inflammatory genes were present in addition to significantly altered expression of genes related to extracellular matrix composition, stability and turnover. Conclusions Signalling related to inflammatory response and extracellular matrix regulation was strongly represented at the DEV and OG1 times. These results indicate that the LBEC is not only a casualty but also an active participant in lamellar events leading to structural failure of the digital lamellae in equine laminitis.