Elevated MG-H1 in glo1-/-aldh3a1-/- zebrafish caused microvasculature alterations and glucose metabolism impairment via decreased proteasome activity. Elevated MG-H1 in glo1-/-aldh3a1-/- zebrafish caused microvasculature alterations and glucose metabolism impairment via decreased proteasome activity

Dicarbonyl stress is characterized by the abnormal accumulation of dicarbonyl reactive metabolites, leading to increased modification of proteins, DNA and lipids, thereby contributing to cellular and tissue dysfunction in diabetes, diabetic complications and other diseases. glo1 knockout zebrafish exhibited moderately increased MG levels, yet this was inadequate to induce trunk vessels alterations due to elevated ALDH activity and mRNA expression levels, which partially act as compensatory mechanism. Excess 4-HNE induced pancreas dysfunction in aldh3a1 knockout zebrafish larvae, inhibiting insulin expression and thereby facilitating hyperglycemia and hyaloid vasculature alterations. To evaluate the combined function of Glo1 and Aldh3a1 in glucose homeostasis and diabetic microvasculature, glo1-/-aldh3a1-/- zebrafish were generated using CRISPR/Cas9 technology. Multiple experiments are performed regarding vasculature alterations, glucose homeostasis, transcriptome, and metabolomics in Tg(fli1:EGFP) zebrafish. glo1-/-aldh3a1-/- zebrafish larvae displayed angiogenic hyaloid vasculature caused by the elevated MG-H1 and decreased proteasomal chymotrypsin-like activity, which could be rescued by the MG-H1 scavenger L-carnosine and proteasome activator Betulinic acid treatments. In adult glo1-/-aldh3a1-/- zebrafish, impaired glucose metabolism, angiogenic retina vasculature and thickened GBM were observed. Thus, our data suggested that an important upstream factor contributing to these phenomena in the context of lacking glo1 and aldh3a1 is MG-H1, possibly through reduced proteasome activity. Overall design: glo1-/-aldh3a1-/-zebrafish were generated using CRISPR/Cas9 technology. Multiple experiments are performed regarding vasculature alterations, glucose homeostasis, transcriptome, and metabolomics in Tg(fli1:EGFP) zebrafish.

二羰基应激（Dicarbonyl stress）以二羰基活性代谢物的异常蓄积为特征，可引发蛋白质、DNA及脂质的修饰水平升高，进而导致糖尿病、糖尿病并发症及其他疾病中的细胞与组织功能障碍。Glo1敲除斑马鱼的MG水平呈中度升高，但由于乙醛脱氢酶（ALDH）活性与mRNA表达水平上调（此为部分代偿机制），该变化不足以诱发躯干血管异常。过量的4-羟基壬烯醛（4-HNE）可在醛脱氢酶3a1（aldh3a1）敲除的斑马鱼幼体中诱发胰腺功能障碍，抑制胰岛素表达，进而促进高血糖症及玻璃体血管异常的发生。为探究Glo1与Aldh3a1在葡萄糖稳态（glucose homeostasis）及糖尿病微血管病变（diabetic microvasculature）中的协同功能，本研究借助CRISPR/Cas9基因编辑技术构建了glo1-/-aldh3a1-/-双敲除斑马鱼模型。本研究针对Tg(fli1:EGFP)转基因斑马鱼（Tg(fli1:EGFP)）开展了多项实验，涵盖血管异常、葡萄糖稳态、转录组学（transcriptome）及代谢组学（metabolomics）分析。glo1-/-aldh3a1-/-斑马鱼幼体呈现出血管生成性玻璃体血管异常，该表型由MG-H1水平升高及蛋白酶体糜蛋白酶样活性（proteasomal chymotrypsin-like activity）降低所介导，且可通过MG-H1清除剂L-肌肽（L-carnosine）与蛋白酶体激活剂白桦酸（Betulinic acid）处理得以挽救。在成年glo1-/-aldh3a1-/-斑马鱼中，可观察到葡萄糖代谢受损、血管生成性视网膜血管异常以及GBM增厚。综上，本研究数据表明，在Glo1与Aldh3a1缺失的背景下，引发上述表型的关键上游因子为MG-H1，其作用机制可能与蛋白酶体活性降低有关。整体实验设计：本研究借助CRISPR/Cas9基因编辑技术构建了glo1-/-aldh3a1-/-双敲除斑马鱼模型，并针对Tg(fli1:EGFP)转基因斑马鱼开展了涵盖血管异常、葡萄糖稳态、转录组学及代谢组学的多项实验。