Department of Developmental and Cell Biology, University of California, Irvine, Natural Sciences II, Irvine, CA 92697, USA.
Department of Molecular Biology and Biochemistry, University of California, Irvine, Natural Sciences II, Irvine, CA 92697, USA.
Dis Model Mech. 2023 Aug 1;16(8). doi: 10.1242/dmm.050215. Epub 2023 Aug 4.
Transgene driven expression of Escherichia coli nitroreductase (NTR1.0) renders animal cells susceptible to the antibiotic metronidazole (MTZ). Many NTR1.0/MTZ ablation tools have been reported in zebrafish, which have significantly impacted regeneration studies. However, NTR1.0-based tools are not appropriate for modeling chronic cell loss as prolonged application of the required MTZ dose (10 mM) is deleterious to zebrafish health. We established that this dose corresponds to the median lethal dose (LD50) of MTZ in larval and adult zebrafish and that it induced intestinal pathology. NTR2.0 is a more active nitroreductase engineered from Vibrio vulnificus NfsB that requires substantially less MTZ to induce cell ablation. Here, we report on the generation of two new NTR2.0-based zebrafish lines in which acute β-cell ablation can be achieved without MTZ-associated intestinal pathology. For the first time, we were able to sustain β-cell loss and maintain elevated glucose levels (chronic hyperglycemia) in larvae and adults. Adult fish showed significant weight loss, consistent with the induction of a diabetic state, indicating that this paradigm will allow the modeling of diabetes and associated pathologies.
转基因表达大肠杆菌硝基还原酶（NTR1.0）使动物细胞对抗生素甲硝唑（MTZ）敏感。许多在斑马鱼中报道的 NTR1.0/MTZ 消融工具，对再生研究产生了重大影响。然而，基于 NTR1.0 的工具不适合用于模拟慢性细胞丢失，因为长期应用所需的 MTZ 剂量（10mM）对斑马鱼的健康有害。我们确定该剂量对应于 MTZ 在幼鱼和成鱼中的半数致死剂量（LD50），并且它诱导了肠道病理学。NTR2.0 是一种来自创伤弧菌 NfsB 的更活跃的硝基还原酶，需要的 MTZ 要少得多才能诱导细胞消融。在这里，我们报告了两种新的基于 NTR2.0 的斑马鱼系的产生，其中可以在没有 MTZ 相关肠道病理学的情况下实现急性β细胞消融。我们首次能够维持β细胞丢失并维持幼虫和成鱼中升高的葡萄糖水平（慢性高血糖症）。成年鱼表现出明显的体重减轻，与诱导糖尿病状态一致，表明该范例将允许糖尿病和相关病理学的建模。
