| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1991: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1990: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
We tried to develop a noble serum-free cell culture medium specially designed for rabbit corneal epithelium. First, when the effect of Ca^<2+> concentration was investigated in a basal medium tested, the Ca^<2+> concentrate : Lon of 0.03mM showed maxiTmnn growth in cultured corneal epithelial cells. Therefore, using this Ca^<2+> concentration, we compared the growth ability of basal medium among MEM, MCISBI53, F-10, MCDB153 and F-10(1 : 1), MCDB153 and MEM(1 : 1). The results indicated that the medium consisting of MCDBI53 and F-10(1 : 1)showed the highest cell growth. Third, using this basal medium supplemented with HEGF, hydrocortisone, insulin, BPE(bovine pituitary extract), primary cultured cells frozen at -70 C were successfully subcultured up to the 15th-generation. The medium reported here is presixmed to be far superior to any other available basal medium for culturing rabbit corneal epithelium.
We tried to establish in vitro rabbit corneal epithelial cell layer on type IV collagen sheet. Since two to three layers of cuboidal epithelial cells were successfully made one week after initial culture, this epithelial sheet can be transplanted into in vivo ocular surface.
We investigated whether biological alteration truly occurs in in vivo conjunctival epithelium in a certain environment, the biological characteristics of conjunctival epithelium implanted in a central corneal stromal pocket were immunohistochemically examined using 64Kd keratin expression, a specific differentiation marker. for corneal epithelium. The results suggest that the biological characteristics of conjunctival epithelium can be influenced by the external environment, becoming similar to those of corneal epithelium.
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