Development of equine models of podal keratinocytes and endothelial cells cultures for the study of equine laminitis

Laminitis is a painful systemic disease of which the principal manifestation is a diffuse aseptic pododermatitis. An ischaemia-reperfusion and/or an activation of matrix métalloproteinases following a proinflammatory status (mainly gastro-intestinal) or hormonal disorders involving metabolic dysfunctions related to the development of insulin resistance, are at the origin of laminitis. The presence of histological lesions of keratinocytes and endothelial cells, and the infiltration of polymorphonuclear neutrophils (PMNs) in laminar tissues, warranted the development of the two cellular culture models corresponding for the study of this disease. Therefore, keratinocytes and endothelial cells are isolated by enzymatic digestion and mechanical separation, respectively from coronary bands and carotids, and are put in culture under controlled atmosphere in DMEM supplemented with additives. Microscopic observations of phenotype, the immunocytochemistry, growth curves, and apoptosis stainings allowed to characterize the cells. The cells can be used, between passage 3 and 5 for the keratinocytes and 3 and 6 for the endothelial cells, in models of interaction with PMNs and their enzymes, in condition of anoxia/reoxygenation with variable glucose.concentrations.