Identification and immunohistochemical localization of annexin II in rat cornea.
OBJECTIVE
We have identified annexin II mRNA expression in the rat cornea and demonstrated immunolocalization of annexin II in normal and injured corneas. Furthermore, to investigate possible interaction between annexin II and its extracellular ligand tenascin during corneal wound healing, we also examined tenascin expression simultaneously.


METHODS
Total RNA was extracted from the corneal tissue of male Wistar rats as well as from the cell cultures of corneal epithelial cells and keratocytes. cDNA was obtained by reverse transcription (RT). Annexin II mRNA expression was examined by polymerase chain reaction (PCR). Western blot analysis of annexin II protein was performed with protein samples that were obtained from the corneal tissue and cell cultures. In addition, the localization of annexin II and that of tenascin were clarified by immunohistochemical analysis, using both uninjured and epithelial scraped corneas.


RESULTS
RT-PCR analysis revealed that annexin II mRNA was expressed in corneal tissues, epithelial cells and keratocytes. Western blot analysis of corneal epithelium and keratocytes showed a 38kDa band that corresponded to the molecular weight of annexin II. Immunohistochemical study showed that annexin II was present in keratocytes as well as the basal cells of corneal epithelium in the central cornea, and basal/suprabasal cells of the limbal epithelium. Positive annexin II immunoreactivity is translocated from cytoplasm to the cell periphery/extracellular position during the epithelial wound healing process. Annexin II and tenascin are coexpressed on the basal surface of limbal epithelium and the leading edge of the healing epithelium. The reappearance of cytoplasmic annexin II staining at the periphery of the cornea correlated with the epithelial cell proliferation.


CONCLUSIONS
Annexin II is abundantly expressed in corneal tissue. The translocalization of annexin II protein suggests its role in the corneal epithelial migration during wound healing. The colocalization of annexin II and tenascin in migrating and proliferating corneal epithelial cells suggests that annexin II-tenascin interaction may play a role in epithelial wound healing.
Matsuda
A
A
Department of Ophthalmology, Hokkaido University School of Medicine. Sapporo, Japan. akimatsu@med.hokudai.ac.jp
Tagawa
Y
Y
Yamamoto
K
K
Matsuda
H
H
Kusakabe
M
M
D000818:Animals; D017306:Annexin A2 / Q000096:biosynthesis / Q000235:genetics*; D015153:Blotting, Western; D003315:Cornea / Q000378:metabolism*; D065306:Corneal Injuries; D004847:Epithelial Cells / Q000378:metabolism; D005131:Eye Injuries / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D008297:Male; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019063:Tenascin / Q000096:biosynthesis / Q000235:genetics; D014945:Wound Healing
D017306:Annexin A2; D012333:RNA, Messenger; D019063:Tenascin