Antiviral Drug Ganciclovir Is a Potent Inhibitor of the Proliferation of Müller Glia-Derived Progenitors During Zebrafish Retinal Regeneration.
Antiviral Drug Ganciclovir Is a Potent Inhibitor of the Proliferation of Müller Glia-Derived Progenitors During Zebrafish Retinal Regeneration.
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抗病毒药物更昔洛韦是斑马鱼视网膜再生过程中穆勒胶质细胞衍生祖细胞增殖的有效抑制剂
DOI：
10.1167/iovs.15-18669
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发表时间：
2016-04-01
影响因子：
4.4
通讯作者：
Xu H
中科院分区：
文献类型：
--
作者：
Zhang S;Mu Z;He C;Zhou M;Liu D;Zhao XF;Goldman D;Xu H
本研究旨在探讨抗病毒药物更昔洛韦（GCV）对成年斑马鱼Müller胶质细胞去分化和增殖的影响及其细胞和分子机制。建立了转Tg（1016 tuba 1a：GFP）基因的Müller胶质细胞损伤诱导的去分化系。用针刺法对成年斑马鱼眼后部造成机械性视网膜损伤。在受伤时或通过角膜将磷酸盐缓冲盐水或GCV注射到眼睛的玻璃体中。通过反相HPLC法测定GCV从眼的清除率。采用绿色荧光蛋白（GFP）和溴脱氧尿苷（BrdU）免疫荧光法检测GCV对视网膜再生的影响。TUNEL法检测细胞凋亡。通过玻璃体注射异凝集素IB 4缀合物标记小胶质细胞。定量（q）PCR和Western印迹分析用于确定视网膜中的基因表达。更昔洛韦治疗在损伤后4天显著减少BrdU+ Müller胶质源性祖细胞（MGPC）的数量。进一步分析表明，GCV对Müller胶质细胞的去分化和MGPCs的初始形成没有影响。我们的数据表明GCV可能通过p53-p21 cip 1依赖性途径不可逆地抑制MGPC增殖。有趣的是，与对照细胞不同，GCV处理的Müller神经胶质细胞被“锁定”在长期的去分化状态。我们的研究揭示了一种新的抑制作用GCV对MGPC增殖，并建议其潜在的使用作为一种工具，以揭示视网膜再生的分子机制在斑马鱼。
The purpose of this study was to investigate the effect of the antiviral drug ganciclovir (GCV) on Müller glia dedifferentiation and proliferation and the underlying cellular and molecular mechanisms in adult zebrafish. A Tg(1016tuba1a:GFP) transgenic line was generated to identify injury-induced dedifferentiation of Müller glia. Mechanical retinal damage was induced by a needle-poke injury on the back of the eyes in adult zebrafish. Phosphate-buffered saline or GCV was injected into the vitreous of the eye at the time of injury or through the cornea. The GCV clearance rate from the eye was determined by a reversed-phase HPLC method. Green fluorescent protein (GFP) and bromodeoxyuridine (BrdU) immunofluorescence were used to determine the effect of GCV on retinal regeneration. Cell apoptosis was evaluated by TUNEL staining. Microglia were labeled by vitreous injection of isolectin IB4 conjugates. Quantitative (q)PCR and Western blot analysis were used to determine gene expression in the retina. Ganciclovir treatment significantly reduced the number of BrdU+ Müller glia–derived progenitor cells (MGPCs) at 4 days post injury. Further analysis showed that GCV had no impact on Müller glia dedifferentiation and the initial formation of MGPCs. Our data indicate that GCV irreversibly inhibited MGPC proliferation likely through a p53-p21cip1–dependent pathway. Interestingly, unlike control cells, GCV-treated Müller glia cells were “locked” in a prolonged dedifferentiated state. Our study uncovered a novel inhibitory effect of GCV on MGPC proliferation and suggests its potential use as a tool to uncover molecular mechanisms underlying retinal regeneration in zebrafish.