Objective: Reconstruct corneal stroma by tissue engineering.
Methods: Primary corneal stromal cells were isolated from newborn rabbit cornea. When the cultured cells reaching confluence, the stromal cells were mixed with polyglycolic acid (PGA) to form a cell-scaffold construct. After 1 week in vitro culture, the constructs were implanted into mother rabbit corneal stroma. Parts of corneal stromal cells were transfected with GFP gene as mark of transplanted cells. Tissues were harvested at 8 weeks for transmission electron microscopy (TEM), histology and Western blot evaluation. In control, PGA alone was implanted into the other cornea.
Results: The engineered corneal stroma became transparent gradually over a period of 8 weeks. The histology of engineered stromal lamellar was relatively similar to that of natural one, no significant differences were found in the diameter of cornea collagen fiber [(29.4 +/- 4.7) nm] in experimental rabbits compared with control [(28.5 +/- 3.5) nm], Student's t-test: P = 0.1316 > 0.05. TEM demonstrated that collagen fibrils deposited in engineered stroma had a similar diameter compared to that of normal counterpart. In addition, Western blot showed the positive expression of type I collagen in the collagen fibrils. In contrast, no new stroma tissue was formed when PGA alone implanted. A green colored stroma was observed when engineered with GFP-labeled cells under fluorescence light microscope.
Conclusion: The results demonstrate that nearly transparent corneal stroma can be obtained by the technique of cornea engineering.
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