METHODS: Primary rabbit CSCs was cultured in vitro and identified by immunohistochemical staining. using lentivirus(LV)with marker gene enhanced green fluorescent protein(EGFP)transfection rabbit CSCs, the growth status and fluorescence intensity of the transfected cells were observed under an inverted fluorescence microscope. The in vitro animal experiments were randomly divided into 2 groups. experimental group lines of LV-EGFP tag of rabbit CSCs suspension stromal injection, control group amount of normal saline injection corneal stroma, Frozen sections were taken 1wk and 1mo after surgery to observe the fluorescence of transplanted CSCs, and hematoxylin-eosin(HE)was used to observe the tissue morphology of paraffin sections.

RESULTS: LV-EGFP transfected rabbit CSCs showed a small amount of fluorescence after 24h under an inverted fluorescence microscope, with the strongest at 96h and 110h. There was no significant difference in the morphology of the transfected CSCs and normal CSCs. Green fluorescence can be seen in the stromal layer of the cornea in the experimental group at 1wk and 1mo, while there is no green fluorescence in the control group. Paraffin section for 1wk showed obvious epithelial cell hyperplasia and slight corneal edema in the experimental group, and a small amount of inflammatory cell infiltration. 1mo after surgery, the epithelial cell hyperplasia was weakened in the experimental group, and no corneal layer edema was observed. No obvious abnormality was found in the control group for 1wk and 1mo.

CONCLUSION: Extracorporeal corneal stroma transplantation of LV-EGFP labeled rabbit CSCs can survive at least 1mo in the corneal and is compatible with adjacent tissues.