Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Serdang 43400, Selangor, Malaysia.
Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur 50603, Selangor, Malaysia.
Int J Mol Sci. 2022 Jul 27;23(15):8290. doi: 10.3390/ijms23158290.
Background: Type 2 diabetes mellitus has recently been identified as a mediator of neurodegeneration. However, the molecular mechanisms have not been clearly elucidated. We aimed to investigate insulin resistance associated with neurodegenerative events in zebrafish larvae. Methods: Larvae aged 72 h-post-fertilization (hpf) were induced to insulin resistance by immersion in 250 nM insulin and were then reinduced with 100 nM insulin at 96 hpf. This model was validated by a glucose levels assay, qPCR analysis of selected genes (akt, pepck, zglut3 and claudin-5a) and Oil Red-O (ORO) staining of the yolk sac for lipid distribution. The association of insulin resistance and neurodegeneration was validated by malondialdehyde (MDA), glutathione (GSH) assays, and by integrating next-generation sequencing with database for annotation, visualization and integrated discovery (DAVID). Results: There was a significant increase in glucose levels at 180 min in the insulin-resistant group. However, it decreased at 400 min after the re-challenge. Insulin-signaling mediators, akt and pepck, were showed significantly downregulated up to 400 min after insulin immersion (p < 0.05). Meanwhile, claudin-5a assessed blood−brain barrier (BBB) integrity and showed significant deterioration after 400 min of post-insulin immersion. ORO staining remarked the increase in yolk sac size in the insulin-resistant group. After the confirmation of insulin resistance, MDA levels increased significantly in the insulin-resistant group compared to the control group in the following parameters. Furthermore, dysregulated MAPK- and Wnt/Ca2+-signaling pathways were observed in the insulin-resistant group, disrupting energy metabolism and causing BBB injury. Conclusions: We conclude that the insulin-resistant zebrafish larvae alter the metabolic physiology associated with neurodegeneration.
2 型糖尿病最近被认为是神经退行性变的中介。然而，其分子机制尚未阐明。我们旨在研究斑马鱼幼虫中与神经退行性事件相关的胰岛素抵抗。
将受精后 72 小时（hpf）的幼虫浸入 250 nM 胰岛素中诱导胰岛素抵抗，然后在 96 hpf 时用 100 nM 胰岛素再诱导。通过葡萄糖水平测定、选择基因（akt、pepck、zglut3 和 claudin-5a）的 qPCR 分析以及卵黄囊中油红 O（ORO）染色评估脂质分布来验证该模型。通过丙二醛（MDA）、谷胱甘肽（GSH）测定和下一代测序与数据库整合用于注释、可视化和综合发现（DAVID）来验证胰岛素抵抗和神经退行性变之间的关联。
在胰岛素抵抗组中，180 分钟时葡萄糖水平显著增加，但在再挑战后 400 分钟时降低。胰岛素信号转导介质 akt 和 pepck 在胰岛素浸泡后 400 分钟内显著下调（p<0.05）。同时，claudin-5a 评估血脑屏障（BBB）完整性，在胰岛素浸泡后 400 分钟后显示出明显恶化。ORO 染色标记胰岛素抵抗组卵黄囊大小增加。在确认胰岛素抵抗后，与对照组相比，胰岛素抵抗组的 MDA 水平在以下参数中显著增加。此外，在胰岛素抵抗组中观察到 MAPK 和 Wnt/Ca2+-信号通路失调，破坏能量代谢并导致 BBB 损伤。
我们得出结论，胰岛素抵抗的斑马鱼幼虫改变了与神经退行性变相关的代谢生理学。
