Biomedical subjects

Chinatsu Kohno

Publications and source records attributed to Chinatsu Kohno.

4 recordsLinked to original sources

PubMed2006

Tissue engineered epithelial cell sheets for the creation of a bioartificial trachea.

To successfully engineer a bioartificial tracheal replacement, it is believed that the regeneration of a functional epithelial lining is a key requirement. In the present study, rabbit tracheal epithelial cells were cultured on temperature-responsive culture dishes, under normal culture conditions at 37 degrees C. By simple temperature reduction to 20 degrees C, the cultured epithelial cells were noninvasively harvested as intact sheets, without the use of any proteolytic enzymes. Support Dacron grafts that had been subcutaneously implanted for 4 weeks to allow for host tissue and vessel infiltration were then opened, and the tracheal epithelial cell sheets were transplanted to the luminal surface without sutures. These fabricated constructs were then used as tracheal replacements, in a rabbit model. Four weeks after transplantation, results showed that the tracheal grafts were covered by a mature, pseudostratified columnar epithelium. In contrast, control constructs that did not receive cell sheet transplantation demonstrated only a thin, immature epithelium at the center of the replacement graft. These results therefore demonstrate that these tracheal epithelial cell sheets can create an epithelial lining on the luminal surface of a bioartificial trachea.

Animals↗

PubMed2005

Functional human corneal endothelial cell sheets harvested from temperature-responsive culture surfaces.

This study reports a new method for fabricating bioengineered human corneal endothelial cell sheets suitable for ocular surgery and repair. We have initially cultured human corneal endothelial cells on type IV collagen-coated dishes and, after several passages, expanded cells were then seeded onto novel temperature-responsive culture dishes. Four weeks after reaching confluence, these cultured endothelial cells were harvested as intact monolayer cell sheets by simple temperature reduction without enzymatic treatment. Scanning electron microscopy indicated that these cells were primarily hexagonal with numerous microvilli and cilia, similar to the native corneal endothelium. The Na+, K+-ATPase pump sites were located at the cell borders as in vivo. Moreover, cell densities and numbers of pump sites were identical to those of in vivo human corneal endothelium under optimized conditions. A 3H-ouabain binding analysis demonstrated a linear proportionality for cell pump density between confluent cell densities of 575 cells/mm2 and 3070 cells/mm2. We also confirmed Na+, K+-ATPase activity in the sheets in vitro. Xenograft transplantation results showed that the fabricated sheets retain their function of maintaining proper stromal hydration in vivo. We have established a regimen to culture and proliferate human corneal endothelial cells and fabricate endothelial sheets ex vivo morphologically and functionally similar to the native corneal endothelium. Our results support the value of harvested cell sheets for clinical applications in ocular reconstructive surgery in patients with ocular endothelial decompensation.

Cell Culture Techniques↗

PubMed2005

Rat limbal epithelial side population cells exhibit a distinct expression of stem cell markers that are lacking in side population cells from the central cornea.

The side population (SP) phenotype is shared by stem cells in various tissues and species. Here we demonstrate SP cells with Hoechst dye efflux were surprisingly collected from the epithelia of both the rat limbus and central cornea, unlike in human and rabbit eyes. Our results show that rat limbal SP cells have a significantly higher expression of the stem cell markers ABCG2, nestin, and notch 1, compared to central corneal SP cells. Immunohistochemistry also revealed that ABCG2 and the epithelial stem/progenitor cell marker p63 were expressed only in basal limbal epithelial cells. These results demonstrate that ABCG2 expression is closely linked to the stem cell phenotype of SP cells.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMed2005

Cell sheet engineering: recreating tissues without biodegradable scaffolds.

While tissue engineering has long been thought to possess enormous potential, conventional applications using biodegradable scaffolds have limited the field's progress, demonstrating a need for new methods. We have previously developed cell sheet engineering using temperature-responsive culture dishes in order to avoid traditional tissue engineering approaches, and their related shortcomings. Using temperature-responsive dishes, cultured cells can be harvested as intact sheets by simple temperature changes, thereby avoiding the use of proteolytic enzymes. Cell sheet engineering therefore allows for tissue regeneration by either direct transplantation of cell sheets to host tissues or the creation of three-dimensional structures via the layering of individual cell sheets. By avoiding the use of any additional materials such as carrier substrates or scaffolds, the complications associated with traditional tissue engineering approaches such as host inflammatory responses to implanted polymer materials, can be avoided. Cell sheet engineering thus presents several significant advantages and can overcome many of the problems that have previously restricted tissue engineering with biodegradable scaffolds.

Animals↗