Applied Tissue Engineering by Minoru Ueda (Ed.)

By Minoru Ueda (Ed.)

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To avoid cell damage from osmotic stress, 6 ml of chilled DMEM containing 10% FCS was gradually added, and the melted cryoprotectant solution was diluted to less than 2% of DMSO or glycerol concentration. 5 ml of diluted cryoprotectant solution. 5%. All of the diluted cryoprotectant solution was then decanted and replaced with new culture medium. The cell sheets were allowed to equilibrate for several more minutes and washed again with the same medium. Structural damage such as vacuolar degeneration was more clearly observed in the corneal epithelial cell sheets cryopreserved with DMSO than those with glycerol, especially at -80°C, whereas only minor morphologic changes were observed in the corneal epithelial cell sheets cryopreserved in glycerol at both temperatures.

A: The cells became a membranous structure with enough mechanical strength to handle with forceps. Cells can be obtained by a conventional explant culture of periosteal fragment. B: Phase-contrast photomicrograph showing the cultured CP. C: Photomicrograph showing hematoxylin and eosin staining of CP section. The CP is approximately 100–300 µm in thickness and consists of 20–30 cellular layers (From Mase et al. 2006. Reprinted with permission). A major disadvantage of using CP for clinical treatment might be the time period required for tissue culture.

G. VEGF) and bone regeneration emphasizes the important role of vasculature not only for survival but also for the proper formation of tissue-engineered bone. In 1997, Asahara et al. characterized endothelial progenitor cells (EPCs) in human peripheral blood using magnetic beads selection [10]. Since EPCs can give rise to endothelial cells (ECs) and are known to facilitate the neovascularization of an implanted site, EPCs may be used to facilitate collateral vessel growth into ischemic tissues through delivery of antiangiogenic or proangiogenic agents.

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