下调lmna基因对斑马鱼胚胎髓系和红系造血干细胞发育的影响

摘要

目的：：探讨lmna基因下调对斑马鱼早期造血干细胞发育的影响。方法：显微注射法将lmna基因mRNA的吗啉代反义寡核苷酸注入单细胞期斑马鱼胚胎(实验组)，显微注射无意义的吗啉代寡核苷酸作为对照组。待胚胎发育至受精18、24、30、36 hpf后收集胚胎进行实验。 RT-PCR和全胚胎原位杂交方法检测斑马鱼髓系造血转录因子pu.1和红系造血转录因子gata1的变化。结果：RT-PCR结果显示，实验组pu.1和gata1表达均较对照组下降(均P<0．05)。原位杂交结果显示，实验组pu.1和gata1蓝黑色阳性杂交信号较对照组变浅。结论：lmna基因下调会阻碍斑马鱼髓系造血干细胞和红系造血干细胞的发育。%Objective: To investigate the effect of lmna gene down-regulation on early hematopoietic development of zebrafish. Methods: Phosphorodiamidate morpholino oligomer ( PMO) technology was used to downregulate lmna gene expression in Zebrafish. Zebrafish embryos injected phosphorodiamidate morpholino antisense oligonucleotide of lmna gene mRNA by microinjection at unicellular stage were taken as the experimental group, and those injected meaningless phosphorodiamidate morpholino antisense oligonucleotide were taken as the control. The embryos were collected at 18, 24, 30 and 36 hpf after the fertilization. The real-time fluorescent quantitative PCR ( RT-PCR) and whole embryo in situ hybridization methods were used to detect the expression of myeloid hematopoietic transcription factor pu. 1 and erythroid hematopoietic transcription factor gata1 in zebrafish. Results:RT-PCR showed that the expressions of pu. 1 and gata1 decreased in the experimental group compared with the control group ( all P<0. 05 ) . Whole embryo in situ hybridization showed that the blue-black positive hybridization signals of pu. 1 and gata1 in experimental group were shallow than those in the control group. Conclusion: Myeloid hematopoietic and erythroid hematopoietic of zebrafish are blocked with the downregulation of lmna gene.