斑马鱼体内氧化损伤标志物筛选
Screening for markers of oxidative damage in zebrafish

目的:探讨以端粒为靶点的氧化损伤标志物的筛选,进而可能可以用来临床上早期发现或预报氧化损伤。方法:采用端粒重复序列结合因子2(TRF2)基因敲除(TRF2-/-)斑马鱼胚胎作为氧化损伤模型,RT-qPCR法检测野生型和TRF2-/-斑马鱼胚胎中TRF2、PUM3、PACSIN3 mRNA的表达情况。构建pCS2-PUM3、pCS2-PACSIN3质粒并体外转录后,将mRNA分别显微注射至TRF2-/-斑马鱼受精卵中,比较注射组与未注射组斑马鱼畸形率。结果:RT-qPCR结果表明,与野生型斑马鱼胚胎相比,TRF2-/-斑马鱼胚胎TRF2、PUM3、PACSIN3 mRNA的表达量下降(P<0.05)。显微注射PUM3 mRNA至TRF2-/-斑马鱼受精卵后斑马鱼胚胎畸形率下降(P<0.05)。结论:TRF2能影响氧化应激相关基因PUM3、PACSIN3的表达,且PUM3能补救TRF2缺失造成的氧化损伤相关器官畸形。
Aim: To screen oxidative damage markers targeting telomeres, which may then be used to clinically detect or predict oxidative damage early.Methods: The telomeric repeat binding factor 2(TRF2)knockout(TRF2-/-)zebrafish embryos were used as an oxidative damage model. RT-qPCR was used to detect the expression of TRF2, PUM3 and PACSIN3 mRNA in wild-type and TRF2-/-zebrafish embryos. After pCS2-PUM3 and pCS2-PACSIN3 plasmids were constructed and in vitro transcribed, their mRNA were into TRF2-/-zebrafish fertilized eggs, and the zebrafish malformation rates in the groups with injection and without injection were summarized and statistically analyzed.Results: RT-qPCR results showed that the TRF2, PUM3, and PACSIN3 mRNA expression levels in TRF2-/-zebrafish embryos were significantly lower than those in wild-type zebrafish embryos(P<0.05).After microinjection of zebrafish PUM3 mRNA into TRF2-/-zebrafish fertilized eggs, the malformation rates of zebrafish embryos decreased(P<0.05).Conclusion: TRF2 can affect the expressions of oxidative stress related genes PUM3 and PACSIN3, and PUM3 can remedy the oxidative damage-related organ malformation caused by TRF2 deficiency. Targeting these molecules is expected to detect or predict oxidative stress early in clinical field