Diabetes mellitus (DM) is a crucial metabolic disease that leads to severe disorders. These include macrovascular complications such as myocardial infarction, stroke, and peripheral artery disease and microvascular complications including diabetic nephropathy, neuropathy, and retinopathy. Diabetes mellitus, along with its associated organ pathologies, is one of the key problems in today’s medicine. Zebrafish is an upcoming disease model organism in diabetes research. Its glucose metabolism and the pathways of reactive metabolite formation are very similar to those of humans. Moreover, several physiological and pathophysiological pathways that also exist in humans and other mammals have been identified in this species or are currently under intense investigation. Zebrafish offer sophisticated imaging techniques and allow simple and fast genetic and pharmacological approaches with a high throughput. In this review, we highlight achievements and mechanisms concerning microvascular complications discovered in zebrafish, and we discuss the advantages and disadvantages of zebrafish as a model for studying diabetic complications.

The increased formation of methylglyoxal (MG) under hyperglycemia is associated with the development of microvascular complications in patients with diabetes mellitus; however, the effects of elevated MG levels in vivo are poorly understood. In zebrafish, a transient knockdown of glyoxalase 1, the main MG detoxifying system, led to the elevation of endogenous MG levels and blood vessel alterations. To evaluate effects of a permanent knockout of glyoxalase 1 in vivo, glo1 ^–/– zebrafish mutants were generated using CRISPR/Cas9. In addition, a diet-induced–obesity zebrafish model was used to analyze glo1 ^–/– zebrafish under high nutrient intake. Glo1 ^–/– zebrafish survived until adulthood without growth deficit and showed increased tissue MG concentrations. Impaired glucose tolerance developed in adult glo1 ^–/– zebrafish and was indicated by increased postprandial blood glucose levels and postprandial S6 kinase activation. Challenged by an overfeeding period, fasting blood glucose levels in glo1 ^–/– zebrafish were increased which translated into retinal blood vessel alterations. Thus, the data have identified a defective MG detoxification as a metabolic prerequisite and glyoxalase 1 alterations as a genetic susceptibility to the development of type 2 diabetes mellitus under high nutrition intake.

Increased methylglyoxal (MG) formation is associated with diabetes and its complications. In zebrafish, knockout of the main MG detoxifying system Glyoxalase 1, led to limited MG elevation but significantly elevated aldehyde dehydrogenases (ALDH) activity and aldh3a1 expression, suggesting the compensatory role of Aldh3a1 in diabetes. To evaluate the function of Aldh3a1 in glucose homeostasis and diabetes, aldh3a1−/− zebrafish mutants were generated using CRISPR-Cas9. Vasculature and pancreas morphology were analysed by zebrafish transgenic reporter lines. Corresponding reactive carbonyl species (RCS), glucose, transcriptome and metabolomics screenings were performed and ALDH activity was measured for further verification. Aldh3a1−/− zebrafish larvae displayed retinal vasodilatory alterations, impaired glucose homeostasis, which can be aggravated via pdx1 silencing induced hyperglycaemia. Unexpectedly, MG was not altered, but 4-hydroxynonenal (4-HNE), another prominent lipid peroxidation RCS exhibited high affinity with Aldh3a1, was increased in aldh3a1 mutants. 4-HNE was responsible for the retinal phenotype via pancreas disruption induced hyperglycaemia and can be rescued via l-Carnosine treatment. Furthermore, in type 2 diabetic patients, serum 4-HNE was increased and correlated with disease progression. Thus, our data suggest impaired 4-HNE detoxification and elevated 4-HNE concentration as biomarkers but also the possible inducers for diabetes, from genetic susceptibility to the pathological progression.

Hyperglycemia causes micro- and macrovascular complications in diabetic patients. Elevated glucose concentrations lead to increased formation of the highly reactive dicarbonyl methylglyoxal (MG), yet the early consequences of MG for development of vascular complications in vivo are poorly understood. In this study, zebrafish were used as a model organism to analyze early vascular effects and mechanisms of MG in vivo. High tissue glucose increased MG concentrations in tg(fli:EGFP) zebrafish embryos and rapidly induced several additional malformed and uncoordinated blood vessel structures that originated out of existing intersomitic blood vessels (ISVs). However, larger blood vessels, including the dorsal aorta and common cardinal vein, were not affected. Expression silencing of MG-degrading enzyme glyoxalase (glo) 1 elevated MG concentrations and induced a similar vascular hyperbranching phenotype in zebrafish. MG enhanced phosphorylation of vascular endothelial growth factor (VEGF) receptor 2 and its downstream target Akt/protein kinase B (PKB). Pharmacological inhibitors for VEGF receptor 2 and Akt/PKB as well as MG scavenger aminoguanidine and glo1 activation prevented MG-induced hyperbranching of ISVs. Taken together, MG acts on smaller blood vessels in zebrafish via the VEGF receptor signaling cascade, thereby describing a new mechanism that can explain vascular complications under hyperglycemia and elevated MG concentrations.

Engulfment and cell motility 1 (ELMO1) functions as a guanine exchange factor for Rac1 and was recently found to protect endothelial cells from apoptosis. Genome wide association studies suggest that polymorphisms within human elmo1 act as a potential contributing factor for the development of diabetic nephropathy. Yet, the function of ELMO1 with respect to the glomerulus and how this protein contributes to renal pathology was unknown. Thus, this study aimed to identify the role played by ELMO1 in renal development in zebrafish, under hyperglycaemic conditions, and in diabetic nephropathy patients. In zebrafish, hyperglycaemia did not alter renal ELMO1 expression. However, hyperglycaemia leads to pathophysiological and functional alterations within the pronephros, which could be rescued via ELMO1 overexpression. Zebrafish ELMO1 crispants exhibited a renal pathophysiology due to increased apoptosis which could be rescued by the inhibition of apoptosis. In human samples, immunohistochemical staining of ELMO1 in nondiabetic, diabetic and polycystic kidneys localized ELMO1 in glomerular podocytes and in the tubules. However, ELMO1 was not specifically or distinctly regulated under either one of the disease conditions. Collectively, these results highlight ELMO1 as an important factor for glomerular protection and renal cell survival via decreasing apoptosis, especially under diabetic conditions.