Transcription Factor Brn-3b Overexpression Enhances Neurite Outgrowth in PC12 Cells Under Condition of Hypoxia.
Transcription Factor Brn-3b Overexpression Enhances Neurite Outgrowth in PC12 Cells Under Condition of Hypoxia.
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DOI：
10.1007/s10571-015-0171-0
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发表时间：
2015-08
影响因子：
4
通讯作者：
Krishnamoorthy RR
中科院分区：
文献类型：
--
作者：
Phatak NR;Stankowska DL;Krishnamoorthy RR
转录因子 Brn-3b 在发育过程中视网膜神经节细胞分化、存活和轴突生长中发挥关键作用。然而，Brn-3b 在正常成人视网膜以及神经变性过程中的确切作用尚不清楚。在当前的研究中，在常氧和缺氧条件下的 PC12 细胞中体外评估了 Brn-3b 过表达的影响。免疫印迹分析表明，PC12 细胞和 661W 细胞中 Brn-3b 的过表达导致生长锥标记物、生长相关蛋白 43 (GAP-43) 和乙酰化微管蛋白 (ac-TUBA) 显着增加。此外，与过表达空载体的 PC12 细胞相比，在过表达 Brn-3b 的 PC12 细胞中观察到 GAP-43 和 ac-TUBA 的免疫染色增加，同时伴随着神经突生长的显着增加。在单独的实验中，将一组用Brn-3b表达载体或空载体转染的PC12细胞置于缺氧条件下2小时，而另一组类似转染的PC12细胞维持在常氧条件下。结果发现，常氧条件下过表达Brn-3b的PC12细胞中GAP-43和ac-TUBA的上调在缺氧条件下持续存在。免疫细胞化学分析显示，与空载体转染的 PC12 细胞相比，在常氧和缺氧条件下，转染 Brn-3b 的 PC12 细胞不仅 GAP-43 和 ac-TUBA 上调，而且神经突生长增加。这些发现对 Brn-3b 在神经退行性疾病中的潜在作用具有影响，在神经退行性疾病中，缺氧/缺血会导致该疾病的病理生理学。
Transcription factor Brn-3b plays a key role in retinal ganglion cell differentiation, survival and axon outgrowth during development. However, the precise role of Brn-3b in the normal adult retina as well as during neurodegeneration is unclear. In the current study, the effect of overexpression of Brn-3b was assessed in vitro, in PC12 cells under conditions of normoxia and hypoxia. Immunoblot analysis showed that overexpression of Brn-3b in PC12 cells as well as 661W cells produced significant increase in the growth cone marker, growth associated protein-43 (GAP-43) and acetylated-tubulin (ac-TUBA). In addition, an increased immunostaining for GAP-43 and ac-TUBA was observed in PC12 cells overexpressing Brn-3b, which was accompanied by a marked increase in neurite outgrowth, compared to PC12 cells overexpressing empty vector. In separate experiments, one set of PC12 cells transfected either with a Brn-3b expression vector or an empty vector was subjected to conditions of hypoxia for 2h, while another set of similarly transfected PC12 cells was maintained in normoxic conditions. It was found that the upregulation of GAP-43 and ac-TUBA in PC12 cells overexpressing Brn-3b under conditions of normoxia was sustained under conditions of hypoxia. Immunocytochemical analysis revealed not only an upregulation of GAP-43 and ac-TUBA, but also increased neurite outgrowth in PC12 cells transfected with Brn-3b as compared to PC12 cells transfected with empty vector in both normoxia and hypoxia. The findings have implications for a potential role of Brn-3b in neurodegenerative diseases in which hypoxia/ischemia contribute to pathophysiology of the disease.