Neary J T, Whittemore S R, Zhu Q, Norenberg M D
Laboratory of Neuropathology, Research Service, VA Medical Center, Miami, Florida.
J Neurochem. 1994 Dec;63(6):2021-7. doi: 10.1046/j.1471-4159.1994.63062021.x.
The expression of the astrocyte-specific intermediate filament protein, glial fibrillary acidic protein (GFAP), is decreased in hepatic encephalopathy and increased in numerous neurological conditions including brain injury. However, little is known about the molecular mechanisms that regulate GFAP expression. Here it is reported that treatment of cultured astrocytes with ammonium chloride reduces GFAP mRNA by up to 85% without inhibiting total RNA synthesis. The effect of NH4Cl was time and dose dependent. The reduction in GFAP mRNA was detected 3 h after initiation of ammonia treatment with a maximum effect observed at 24 h. Significant decreases in GFAP mRNA were observed at 2, 5, and 10 mM NH4Cl. Concurrent treatment with extracellular ATP prevented the loss of GFAP mRNA, possibly by activation of purinergic receptors. In addition, removal of ammonium chloride restored GFAP mRNA to normal levels. Nuclear runoff experiments indicated that NH4Cl did not inhibit GFAP mRNA transcription. Studies using alpha-amanitin, an inhibitor of RNA polymerase II, showed that NH4Cl decreased the stability of GFAP mRNA by approximately 50%. This destabilization of GFAP mRNA may be an important factor in the pathogenesis of hepatic encephalopathy. Because increased GFAP is an important component of reactive gliosis, understanding the mechanisms that destabilize GFAP mRNA may facilitate strategies to minimize the gliosis associated with brain injury.
星形胶质细胞特异性中间丝蛋白，即胶质纤维酸性蛋白（GFAP），在肝性脑病中表达降低，而在包括脑损伤在内的多种神经疾病中表达增加。然而，关于调节GFAP表达的分子机制却知之甚少。本文报道，用氯化铵处理培养的星形胶质细胞可使GFAP mRNA减少多达85%，而不抑制总RNA合成。NH4Cl的作用具有时间和剂量依赖性。在用氨处理3小时后可检测到GFAP mRNA减少，24小时时观察到最大效应。在2、5和10 mM NH4Cl时观察到GFAP mRNA显著减少。细胞外ATP的同时处理可防止GFAP mRNA的丢失，可能是通过嘌呤能受体的激活。此外，去除氯化铵可使GFAP mRNA恢复到正常水平。核转录实验表明，NH4Cl不抑制GFAP mRNA转录。使用RNA聚合酶II抑制剂α-鹅膏蕈碱的研究表明，NH4Cl使GFAP mRNA的稳定性降低了约50%。GFAP mRNA的这种不稳定可能是肝性脑病发病机制中的一个重要因素。由于GFAP增加是反应性胶质增生的一个重要组成部分，了解使GFAP mRNA不稳定的机制可能有助于制定策略，以尽量减少与脑损伤相关的胶质增生。
