| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2000: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1999: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1998: ¥1,900,000 (Direct Cost: ¥1,900,000)
|
|---|
| Research Abstract |
We have studied the cellular and molecular mechanism involved in transdifferentiation of retinal pigment epithelium (RPE) to neural retina (NR) both in adult newt eyes and in embryonic quail eyes carrying homozygous silver mutation.
Since no in vitro experimental system was available in the newt study, we have developed a new organ culture method in which adult newt RPEs were cultured with the surrounding choroidal tissue, on a filter membrane. Around day 7 in vitro, lightly pigmented "neuron-like cells" with neuritic processes were found migrating out from the explant onto the filter membrane. These cells were positively reacted to various antibodies against neuron-specific substances. Their number gradually increased day by day. BrdU-labeling study showed that RPE cells initiated to proliferate under the culture condition on day 4 in vitro, temporally correlating to the time course of retinal regeneration in vivo. Histological observations of cultured explants showed that proliferatin
… More
g RPE cells did not form the stratified structure typically observed in the NR but they rather migrated out from the explants. Fibroblast growth factor-2 (FGF-2), known as a potent factor for the transdifferentiation of ocular tissues in various vertebrates, substantially increased the numbers of both neuron-like cells. Interestingly, when purely isolated RPEs were cultured, no neuronal cells appeared after 30 days in vitro, but they were observed to differentiate from RPE cells in the presence of FGF-2. The results suggest FGF-2-like factors may trigger newt retinal regeneration and these factors will be derived from the choroidal tissue.
In the avian study, we developed an chimerical transplantation of optic vesicles from homozygous silver quails to wild-type chick embryos, and it was revealed that neural crest-derived mesenchymal cells have a crucial role in the RPE transdifferentiation. The migration of the cranial neural crest cells in the silver embryos appear to be affected by the mutation. In vitro study has shown that FGF-1 has a crucial role in this process. These results suggest that the neural crest cells have an essential role in the normal development of RPE cells, and mutation in the crest cells will cause a severe defect of eye development.
As a conclusion, mesenchymal tissues or connective tissues surrounding the optic cup or RPEs have a crucial role both in the development of avian RPE and retina and in the regeneration of the newt retina. Less
|
