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[Studies on conditions for the culture of rabbit corneal epithelium]
P Skopiński1, J Jóźwiak, J Lamprecht
1Zakładu Mikroskopii Elektronowej Instytutu Biostruktury AM w Warszawie. pskopin@ib.amwaw.edu.pl
Purpose:
Research was aimed at comparison of isolation methods as well as determination of growth and differentiation dynamics of rabbit corneal epithelium (CE) in vitro. Adhesion, growth and differentiation of CE cells growing on collagen membranes were evaluated.
Material And Methods:
Research was performed on the cells of rabbit corneal epithelium isolated mechanically or enzymatically (Dispase II) from comeas excised at the edge of limbus. CE cells were cultured in media with high or low contents of calcium, with addition of FCS, insulin, cholera toxin and EGF.
Results:
In comparison with mechanical isolation, enzymatic isolation yielded 4-5 times more living undifferentiated (CE) cells. The highest dynamics of in vitro growth was observed in primary cultures in low-calcium medium supplemented with the above substances. After 20 population doublings cells were differentiated and died. Only few cells on collagen membranes adhered to the collagen but did not enter division.
Conclusions:
Current research allowed for determination of methodology for CE excision and isolation. Optimal conditions for in vitro growth have been established. Growth dynamics and proliferation of CE in vitro have been evaluated. Growth of CE on standard collagen membrane has not been observed.