Early laminar events involving endothelial activation in horses with black walnut- induced laminitis.
OBJECTIVE
To determine proinflammatory gene expression, endothelial adhesion molecule gene expression, and matrix metalloproteinase (MMP) concentrations in laminar specimens at 1.5 hours after administration of black walnut extract (BWE) and to compare these values with later time points.


METHODS
25 horses.


METHODS
After nasogastric administration of BWE, anesthesia was induced at 1.5 hours in early time point (ETP) horses (n = 5), between 3 and 4 hours in developmental time point horses (5), and between 9 and 10 hours in acute onset of lameness time point horses (5). Anesthesia was induced at 3 and 10 hours after nasogastric administration of water in 2 groups of control horses (3-hour control group, n = 5; 10-hour control group, 5). Real-time quantitative PCR assay was performed on laminar specimens from control and ETP horses for cyclooxygenase (COX)-1, COX-2, interleukin (IL)-1beta, tumor necrosis factor-alpha, IL-6, IL-8, IL-10, MMP-2, and MMP-9 gene expression; and on laminar specimens from all groups for endothelial adhesion molecules, intercellular adhesion molecule (ICAM)-1, and E-selectin gene expression. Leukocyte emigration was assessed via CD13 immunohistochemistry, and gelatinase accumulation was determined by gelatin zymography.


RESULTS
Laminar concentrations of IL-1beta, IL-6, IL-8, COX-2, ICAM-1, and E-selectin mRNA were significantly increased in ETP horses, compared with control horses. Concentrations of IL-1beta, IL-8, ICAM-1, and E-selectin mRNA peaked at 1.5 hours. In ETP horses, leukocyte emigration was present in 3 of 5 horses and pro-MMP-9 was detected in 2 of 5 horses.


CONCLUSIONS
Results indicated that endothelial activation and laminar inflammation are early events in laminitis; MMP accumulation likely is a downstream event.
Loftus
John P
JP
Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, MA 01003, USA.
Black
Samuel J
SJ
Pettigrew
Amanda
A
Abrahamsen
Eric J
EJ
Belknap
James K
JK
D000818:Animals; D051545:Cyclooxygenase 1 / Q000096:biosynthesis / Q000235:genetics; D051546:Cyclooxygenase 2 / Q000096:biosynthesis / Q000235:genetics; D019040:E-Selectin / Q000096:biosynthesis / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D004727:Endothelium / Q000201:enzymology / Q000276:immunology; D005534:Foot Diseases / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000662:veterinary*; D015870:Gene Expression; D006734:Horse Diseases / Q000201:enzymology / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006736:Horses; D007150:Immunohistochemistry / Q000662:veterinary; D018799:Intercellular Adhesion Molecule-1 / Q000096:biosynthesis / Q000235:genetics; D007378:Interleukins / Q000096:biosynthesis / Q000235:genetics; D031324:Juglans; D020778:Matrix Metalloproteinase 2 / Q000096:biosynthesis / Q000235:genetics; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis / Q000235:genetics; D010936:Plant Extracts; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics
D019040:E-Selectin; D007378:Interleukins; D010936:Plant Extracts; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9