UOSD SAFU, IRCCS - Regina Elena National Cancer Institute, Via Elio Chianesi 53, 00144, Rome, Italy.
IFOM - FIRC Institute of Molecular Oncology, Milan, Italy.
Sci Rep. 2021 Jan 12;11(1):671. doi: 10.1038/s41598-020-79530-5.
We developed a novel reporter transgenic zebrafish model called MITO-Luc/GFP zebrafish in which GFP and luciferase expression are under the control of the master regulator of proliferation NF-Y. In MITO-Luc/GFP zebrafish it is possible to visualize cell proliferation in vivo by fluorescence and bioluminescence. In this animal model, GFP and luciferase expression occur in early living embryos, becoming tissue specific in juvenile and adult zebrafish. By in vitro and ex vivo experiments we demonstrate that luciferase activity in adult animals occurs in intestine, kidney and gonads, where detectable proliferating cells are located. Further, by time lapse experiments in live embryos, we observed a wave of GFP positive cells following fin clip. In adult zebrafish, in addition to a bright bioluminescence signal on the regenerating tail, an early unexpected signal coming from the kidney occurs indicating not only a fin cell proliferation, but also a systemic response to tissue damage. Finally, we observed that luciferase activity was inhibited by anti-proliferative interventions, i.e. 5FU, cell cycle inhibitors and X-Rays. In conclusion, MITO-Luc/GFP zebrafish is a novel animal model that may be crucial to assess the spatial and temporal evolution of cell proliferation in vivo.
我们开发了一种新型的报告基因转基因斑马鱼模型，称为 MITO-Luc/GFP 斑马鱼，其中 GFP 和荧光素酶的表达受增殖主调控因子 NF-Y 的控制。在 MITO-Luc/GFP 斑马鱼中，可以通过荧光和生物发光在体内可视化细胞增殖。在这种动物模型中，GFP 和荧光素酶的表达发生在早期胚胎中，在幼年和成年斑马鱼中成为组织特异性表达。通过体外和离体实验，我们证明成年动物中的荧光素酶活性存在于肠道、肾脏和性腺中，这些组织中存在可检测的增殖细胞。此外，通过活体胚胎的延时实验，我们观察到在鳍夹后 GFP 阳性细胞的波状出现。在成年斑马鱼中，除了再生尾巴上明亮的生物发光信号外，还会出现来自肾脏的早期意外信号，这不仅表明鳍细胞增殖，还表明对组织损伤的全身反应。最后，我们观察到，用抗增殖干预措施（如 5FU、细胞周期抑制剂和 X 射线）抑制了荧光素酶的活性。总之，MITO-Luc/GFP 斑马鱼是一种新型的动物模型，可能对评估体内细胞增殖的时空演变至关重要。
