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Isolation and cultivation of equine corneal keratocytes, fibroblasts and myofibroblasts
Author(s) -
Buss Dylan G.,
Giuliano Elizabeth A.,
Sharma Ajay,
Mohan Rajiv R.
Publication year - 2010
Publication title -
veterinary ophthalmology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.594
H-Index - 50
eISSN - 1463-5224
pISSN - 1463-5216
DOI - 10.1111/j.1463-5224.2009.00755.x
Subject(s) - myofibroblast , stromal cell , stroma , pathology , in vitro , cornea , biology , immunohistochemistry , cell culture , medicine , ophthalmology , fibrosis , biochemistry , genetics
Objective To establish an in vitro model for the investigation of equine corneal wound healing. To accomplish this goal, a protocol to isolate and culture equine corneal keratocytes, fibroblasts and myofibroblasts was developed. Animal material Equine corneal buttons were aseptically harvested from healthy research horses undergoing humane euthanasia for reasons unrelated to this study. Slit‐lamp biomicroscopy was performed prior to euthanasia by a board‐certified veterinary ophthalmologist to ensure that all samples were harvested from horses free of anterior segment disease. Procedure Equine corneal stroma was isolated using mechanical techniques and stromal sub‐sections were then cultured. Customized media at different culture conditions was used to promote growth and differentiation of corneal stromal cells into keratocytes, fibroblasts and myofibroblasts. Results Cell culture techniques were successfully used to establish a method for the isolation and culture of equine corneal keratocytes, fibroblasts and myofibroblasts. Immunohistochemical staining for alpha‐smooth muscle and F‐actin was used to definitively differentiate the three cell types. Conclusion Equine corneal stromal keratocytes, fibroblasts and myofibroblasts can be predictably isolated and cultured in vitro using this protocol.