Expression and activity of collagenases in the digital laminae of horses with carbohydrate overload-induced acute laminitis.
BACKGROUND
Matrix metalloproteinases (MMP) are hypothesized to degrade structurally important components of the laminar extracellular matrix (ECM) in horses with laminitis.


OBJECTIVE
To compare levels of expression of stromelysin-1 (MMP-3), collagenases (MMP-1, -13), and membrane type-MMPs (MMP-14, -15, -16), and the distribution of their ECM substrates, in laminae of healthy horses and horses with carbohydrate overload laminitis.


METHODS
Twenty-five adult horses.


METHODS
Gene and protein expression were determined in extracts of laminae using real-time quantitative polymerase chain reaction and Western blotting after sodium dodecylsulfate polyacrylamide gel electrophoresis. Distribution of MMP-13 and ECM components was determined using indirect immunofluorescent microscopy of nonfixed frozen sections. ECM morphology was assessed by hematoxylin and eosin staining.


RESULTS
Of the genes studied, only those encoding MMP-1 and -13 were upregulated in CHO-induced laminitis; MMP-1 at Obel grade (OG)1 lameness and MMP-13 at OG3 lameness. Laminar MMP-1 was present as 52 kDa proenzyme only. MMP-13 was present as pro- (61 kDa) and processed (48 kDa) enzyme. MMP-13 localized to the basal epithelium of the secondary epidermal laminae and its increased expression were accompanied by the appearance in secondary dermal laminae (SDL) of multiple foci that were devoid of collagen I, fibronectin, chondroitin and keratan sulfate glycosaminoglycans, and eosin-staining material.


CONCLUSIONS
MMP-13 is upregulated in laminae of horses with CHO-induced OG3 lameness and, by degrading components of the ECM, may contribute to the formation of ECM-free lesions (gaps or tears) that appear in the SDL with OG3 lameness.
Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, MA.
L
L
Wang
E A
EA
Pawlak
P J
PJ
Johnson
J K
JK
Belknap
D
D
Alfandari
S J
SJ
Black
D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D005109:Extracellular Matrix / Q000201:enzymology / Q000378:metabolism*; D005534:Foot Diseases / Q000201:enzymology / Q000378:metabolism / Q000662:veterinary*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D006724:Hoof and Claw / Q000201:enzymology / Q000378:metabolism*; D006734:Horse Diseases / Q000201:enzymology / Q000378:metabolism*; D006736:Horses; D007150:Immunohistochemistry / Q000662:veterinary; D020782:Matrix Metalloproteinases / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000662:veterinary; D012313:RNA / Q000737:chemistry / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D018709:Statistics, Nonparametric
D012313:RNA; D020782:Matrix Metalloproteinases
CHO‐laminitis; Equine; Matrix metalloproteinases