| Co-Investigator(Kenkyū-buntansha) |
WAKAKURA Masato Kitasato University School of Medicine, Ophthalmology, Associate Professor, 医学部, 助教授 (50137931)
ARAKI Masasuke Kyoto Prefectural University, Biology, Professor, 生物学科, 教授 (00118449)
TAUCHI Masaki Okayama Prefectural University, Welfare System and Health Science, Professor, 保健福祉学部, 教授 (00075425)
SAITO Takehiko Tsukuba University, Biology, Professor, 生物学系, 教授 (20081630)
FUKUDA Yutaka Osaka University School of Medicine, Physiology, Professor, 医学部, 教授 (90028598)
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| Research Abstract |
1) Degeneration process of retinal ganglion cells : By means of confocal laser scanning microscope, both dendritic and axonal processes located in different focal planes can be observed at a single image without any blur. Without sectioning flat-mounted retinas vertically, confocal laser scanning microscope enabled us to identify either on-type cells or off-type cells clearly and easily on the basis of their reconstructed vertical image.
2) Axonal regeneration of retinal ganglion cells : We concluded that even adult aged retinal ganglion cells had the ability of axonal regeneration after injury and that neurotrophic factors might enhanced these abilities. Therefore neurotrophic factors might have practicable appiocations in drug treatments for intractable disease of the neural retina and optic nerve.
3) Observation of possible cases for transplantation : Using a scanning laser ophthalmoscope (SLO) , the ocular fundus of a patient with Best's vitelliform maculardystrophywas examined. With
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the argon blue laser, a dark area was seen in the fovea. Areas of weakly bright reflex were seen corresponding to the scrambled-egg lesion observed by ophthalmoscope. With the argon green laser, highly bright spots were observed at the foveola. Intense bright reflex was seen over the scrambled-egg lesion at theposterior pole. The helium-neon laser revealed highly bright patches corresponding to the orderly ring distribution of the yellowish deposits at the posterior pole. The diode laser revealed a mottled appearance with dark and bright patches in the scrambled-egg lesion. SLO provided a morphologic enhancement in the specifically affected layrs of the maculs in Best's vitelliform macular dystrophy.
4) Analysis of retinal regeneration using avian embryo : The avian pineal is a photoreceptive organ and is believed to function as a circadian clock. Avian pineal cells are secretory rudimentary photoreceptors, and previous studies have demonstrated that there are two types of photoreceptors in developing quail pineals, one of which is rhodopsin-like immunoreactive and the other iodopsin-like immunoreactive. Much larger number of rhodopsin-like immunoreactive cells than of iodopsin-like immunoreactive cells were found in quail pineals, both in vivo and in vitro. In the present study we co-cultured pineal cells of embryonic quails with retinal cells but separated the two with a biomembrane filter We found that diffusible material produced by the cultured retinal cells intensely promotes the appearance of pineal iodopsin-like immunoreactive cells in vitro. This effect of retina-derived factors) is cell-type specific, since there is no effect on the differentiation of pineal rhodopsin-like immunoreactive cells. Retinal cell cultures had much more intensive iodopsin-promoting effect than other embryonic brain cultures such as cerebral cell cultures. The production of the retinal factors) seems to be developmentally regulated, since retiual cells from older embryos (E13 and older) did not have such effects. The factors) possibly act on pineal precursor cells by stimulating the expression of the iodopsin-like immunoreactive phenotype. Preliminary characterization of conditioned medium obtained from cultured retinal cells shows that the factor is a stable polypeptide, probably of low molecular weight. The pineal-retina culture system win provide a good experimental system to analyze the effect of extrinsic environments on cell differentiation.
5) Physiology of the retinal regeneration : Retinal cells dissociated from adult newt eyes were seeded onto plastic dishes which were coated with various cell adhesion substances. They were cultured in Leibovitz's L-15 medium with no serum. Concanavalin A (ConA) and poly-L-lysine (PLL) caused relatively good cell attachment, regardless of retinal cell types, in comparison with uncoated dishes. On the other hand, extracellular matrix (ECM) components such as collagen, fibronectin and laminin tended to inhibit the cell attachment. In both ConA-and PLL-coated dishes, ganglion cells survived for longer periods than other retinal cells and extended neurites with time in culture. The outgrowth of neurites by ganglion cells was found to be promoted to a far greater degree by ConA than PLL.Branching of neurites also appeared to occur more frequently on ConA.From these observations, ConA may be an useful tool for investigating retinal regeneration in vitro. Less
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