These results indicate that anti-inflammatory treatment of early stage laminitis (and the horse at risk of laminitis) should include not only therapeutic drugs that address prostanoid activity, but should also address the marked increases in lamellar cytokine expression.

Summary Reasons for performing study: The mediators and signalling cascades important in the initiation of laminitis remain unclear. We therefore wanted to explore the genes and overall signalling mechanisms that play an important role in the developmental stage of laminitis. Objective: To use a broad genomic screening technique to identify novel genes that are differentially regulated in the equine lamellae during the developmental period of laminitis. Methods: Differential mRNA display (DRD) was performed to discover regulated genes, and real‐time quantitative polymerase chain reaction (RT‐qPCR) was then used to evaluate lamellar mRNA levels of a regulated gene (MAIL) and mediators related to that gene (IL‐1β and IL‐6) in control horses (n = 5) and horses administered black walnut extract (BWE; n = 5). Results: Using DRD, MAIL was identified as a regulated gene. RT‐qPCR indicated a 4‐fold increase in expression of the MAIL mRNA in BWE lamellae compared to controls. A 30‐fold increase in IL‐1β, and a 160‐fold difference in IL‐6 mRNA expression was present in BWE lamellae. Differences in MAIL, IL‐1β and IL‐6 mRNA expression were statistically significant between groups (P<0.05). Conclusions and potential relevance: The data strongly support a role for inflammatory cytokines in the developmental stages of laminitis, possibly inducing the vascular and metabolic alterations reported to occur in the affected digit. These results potentially support the use of anti‐inflammatory drugs in horses at risk of laminitis, and warrant further investigation of the link between systemic disease processes associated with laminitis and the reported digital inflammation.

Background: Recent reports indicate increased amounts of mRNA from inflammation-related genes in the prodromal stage of laminitis.Hypothesis: Cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) undergo distinct patterns of expression in equine laminae in the developmental stage (DEV) and acute clinical stage (LAM) of laminitis.Animals: Horses selected from an outbred population were placed into 1 of 4 groups: DEV (n 5 5), CON-3h (control group for DEV, n 5 5), LAM (n 5 5) and CON-10h (control group for LAM, n 5 5).Methods: Laminar and skin samples were obtained from (1) animals either undergoing leukopenia (DEV) or the onset of clinical signs of laminitis (LAM) after black walnut extract (BWE) administration and (2) animals either 3 (CON-3h ) or 10 (CON-10h) hours after administration of water. Real-time quantitative polymerase chain reaction (RT-qPCR), immunoblotting, and immunohistochemical analysis were performed for COX-1 and COX-2.Results: Upon immunohistochemical analysis of all 4 groups, COX-2 was expressed by most viable epithelial cells in both laminae and skin. COX-1 exhibited similar epithelial expression to COX-2 in skin epidermis, but was expressed exclusively in the basal layer of laminar epidermis. COX-1 protein was not detectable in dermal vasculature of equine skin or laminae, whereas COX-2 was present in endothelial and vascular smooth muscle cells of dermal vasculature in both skin and laminae in all groups. A marked increase in laminar COX-2 protein concentrations was detected on immunoblotting in the DEV group, although a lesser increase was observed in the LAM group.Conclusions and Clinical Importance: COX-2 protein expression is markedly increased in the resident laminar cell types in the developmental stage of BWE-induced laminitis.

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Background: Recent reports indicate increased amounts of mRNA from inflammation-related genes in the prodromal stage of laminitis.Hypothesis: Cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) undergo distinct patterns of expression in equine laminae in the developmental stage (DEV) and acute clinical stage (LAM) of laminitis.Animals: Horses selected from an outbred population were placed into 1 of 4 groups: DEV (n 5 5), CON-3h (control group for DEV, n 5 5), LAM (n 5 5) and CON-10h (control group for LAM, n 5 5).Methods: Laminar and skin samples were obtained from (1) animals either undergoing leukopenia (DEV) or the onset of clinical signs of laminitis (LAM) after black walnut extract (BWE) administration and (2) animals either 3 (CON-3h ) or 10 (CON-10h) hours after administration of water. Real-time quantitative polymerase chain reaction (RT-qPCR), immunoblotting, and immunohistochemical analysis were performed for COX-1 and COX-2.Results: Upon immunohistochemical analysis of all 4 groups, COX-2 was expressed by most viable epithelial cells in both laminae and skin. COX-1 exhibited similar epithelial expression to COX-2 in skin epidermis, but was expressed exclusively in the basal layer of laminar epidermis. COX-1 protein was not detectable in dermal vasculature of equine skin or laminae, whereas COX-2 was present in endothelial and vascular smooth muscle cells of dermal vasculature in both skin and laminae in all groups. A marked increase in laminar COX-2 protein concentrations was detected on immunoblotting in the DEV group, although a lesser increase was observed in the LAM group.Conclusions and Clinical Importance: COX-2 protein expression is markedly increased in the resident laminar cell types in the developmental stage of BWE-induced laminitis.