Ethanol inhibits retinal and CNS differentiation due to failure of cell cycle exit via an apoptosis-independent pathway

• Main Authors: Hsin-Yu Chung, 鍾昕妤
• Other Authors: Chin-Hwa Hu
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/71998809542528488053

博士 === 國立臺灣海洋大學 === 生物科技研究所 === 101 === Alcohol exposure during embryogenesis results in a variety of developmental disorders. Here, we demonstrate that continuous exposure to 1.5% ethanol causes substantial apoptosis and abrogated retinal and CNS development in zebrafish embryos. Chronic exposure to ethanol for twenty four hours before hatching also induces apoptosis and retinal disorder. After the 2-day post-fertilization (dpf) stage, chronic exposure to ethanol continued to induce apoptosis, but did not block retinal differentiation. Although continuous ethanol exposure induces substantial accumulation of reactive oxygen species (ROS) and increases p53 expression, depletion of p53 did not eliminate ethanol-induced apoptosis. On the other hand, sequestering ROS with the antioxidant reagent N-acetylcysteine (NAC) successfully inhibited ethanol-associated apoptosis, suggesting that the ethanol-induced cell death primarily results from ROS accumulation. Continuous ethanol treatment of embryos reduced expression of the mature neural and photoreceptor markers elavl3/huC, rho, and crx, as well as sustained the expression pattern of the neural and retinal progenitor markers ascl1b and pax6b at undifferentiated stage, indicating that the retinal and CNS neural progenitor cells did not undergo further differentiation. Moreover, ethanol treatment enhanced BrdU incorporation, histone H3 phosphorylation, and pcna expression in the neural progenitor cells, thereby maintaining a high proliferation rate.Furthermore, ethanol treatment resulted in sustained transcription of ccnd1/cyclin D1 and ccne/cyclin E throughout development in neural progenitor cells without an appropriate increase of cdkn1b/p27 and cdkn1c/p57 expression, suggesting that these cells failed to exit from cell cycle. Although NAC was able to mitigate ethanol-mediated apoptosis, it was unable to ameliorate the defects in visual and CNS neural differentiation, suggesting that abrogated neural development in ethanol-exposed embryos is unlikely to arise from excessive apoptosis. In conclusion, we demonstrate that the pathological effect of ethanol on zebrafish embryos is partially attributable to cell death and inhibition of visual and CNS neuron differentiation. Excessive apoptosis largely results from the accumulation of ROS, whereas abrogated neural development is caused by failure of cell cycle arrest, which in turn prevents a successful transition from proliferation to differentiation.