Shiro Amano1, Tatsuya Mimura2, Satoru Yamagami2, Yasuhiro Osakabe3, Kazunori Miyata3. 1. Department of Ophthalmology, University of Tokyo School of Medicine, Tokyo, Japan. amanos-tky@umin.ac.jp. 2. Department of Ophthalmology, University of Tokyo School of Medicine, Tokyo, Japan. 3. Meiwakai Medical Foundation, Miyata Eye Hospital, Miyakonojo, Miyazaki, Japan.
Abstract
PURPOSE: To examine the properties of corneas tissue-engineered with cultured human corneal endothelial cells (HCEC) and human corneal stroma. METHODS: Primary HCEC cultures were established from endothelial cell layer explants and propagated on culture dishes coated with bovine corneal endothelial extracellular matrix. A cell suspension of HCEC at the fifth passage was transferred onto human corneal stroma deprived of endothelial cells, and the corneas were gently centrifuged to enhance cell attachment. The cell density of the tissue-engineered corneas was examined after staining with alizarin red and trypan blue. The tissue-engineered corneas were histologically examined by light and electron microscopy. The pump function of the tissue-engineered corneas was measured using an Ussing chamber. RESULTS: The mean endothelial cell density of four tissue-engineered corneas was 2380 +/- 264 cells/mm(2) (mean +/- SD). HCEC on the tissue-engineered corneas had a morphology similar to HCEC in vivo. The pump function parameters of the tissue-engineered corneas were 55%-75% of those of normal corneas. CONCLUSIONS: HCEC on the tissue-engineered corneas have morphology and cellular density similar to HCEC in vivo, whereas the pump function of the tissue-engineered corneas was lower than in normal corneas.
PURPOSE: To examine the properties of corneas tissue-engineered with cultured human corneal endothelial cells (HCEC) and humancorneal stroma. METHODS: Primary HCEC cultures were established from endothelial cell layer explants and propagated on culture dishes coated with bovine corneal endothelial extracellular matrix. A cell suspension of HCEC at the fifth passage was transferred onto humancorneal stroma deprived of endothelial cells, and the corneas were gently centrifuged to enhance cell attachment. The cell density of the tissue-engineered corneas was examined after staining with alizarin red and trypan blue. The tissue-engineered corneas were histologically examined by light and electron microscopy. The pump function of the tissue-engineered corneas was measured using an Ussing chamber. RESULTS: The mean endothelial cell density of four tissue-engineered corneas was 2380 +/- 264 cells/mm(2) (mean +/- SD). HCEC on the tissue-engineered corneas had a morphology similar to HCEC in vivo. The pump function parameters of the tissue-engineered corneas were 55%-75% of those of normal corneas. CONCLUSIONS: HCEC on the tissue-engineered corneas have morphology and cellular density similar to HCEC in vivo, whereas the pump function of the tissue-engineered corneas was lower than in normal corneas.