Buss Dylan G, Giuliano Elizabeth A, Sharma Ajay, Mohan Rajiv R
College of Veterinary Medicine, University of Missouri, Columbia, MO, USA.
Vet Ophthalmol. 2010 Jan;13(1):37-42. doi: 10.1111/j.1463-5224.2009.00755.x.
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.
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.
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.
Equine corneal stromal keratocytes, fibroblasts and myofibroblasts can be predictably isolated and cultured in vitro using this protocol.
建立一个用于研究马角膜伤口愈合的体外模型。为实现这一目标，制定了一种分离和培养马角膜角质形成细胞、成纤维细胞和平滑肌肌成纤维细胞的方案。
从因与本研究无关的原因接受人道安乐死的健康研究用马身上无菌采集马角膜纽扣组织。在安乐死之前，由一位获得委员会认证的兽医眼科医生进行裂隙灯生物显微镜检查，以确保所有样本均取自无前段疾病的马。
采用机械技术分离马角膜基质，然后培养基质亚切片。使用不同培养条件下的定制培养基来促进角膜基质细胞生长并分化为角质形成细胞、成纤维细胞和平滑肌肌成纤维细胞。
细胞培养技术成功用于建立一种分离和培养马角膜角质形成细胞、成纤维细胞和平滑肌肌成纤维细胞的方法。使用α-平滑肌和F-肌动蛋白的免疫组织化学染色来明确区分这三种细胞类型。
使用该方案可在体外可预测地分离和培养马角膜基质角质形成细胞、成纤维细胞和平滑肌肌成纤维细胞。
