Diabetologie und Stoffwechsel 2015; 10 - FV5
DOI: 10.1055/s-0035-1549511

Zebrafish as a model organism for studying late diabetic complications

K Jörgens 1, 2, E Lodd 1, F Schmöhl 1, SJ Stoll 1, 2, TH Fleming 3, PP Nawroth 3, HP Hammes 4, J Kroll 1
  • 1Department of Vascular Biology and Tumor Angiogenesis, Center for Biomedicine and Medical Technology Mannheim (CBTM), Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany
  • 2Division of Vascular Oncology and Metastasis, German Cancer Research Center (DKFZ-ZMBH Alliance), Heidelberg, Germany
  • 3Department of Medicine I and Clinical Chemistry, Heidelberg University, Heidelberg, Germany
  • 45th Medical Department, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany

Research Question: In addition to high blood glucose levels, oxidative stress and reactive metabolites can lead to microvascular complications in diabetic patients. To better understand formation of late diabetic complications, we analyze the effects of hyperglycemia, methylglyoxal (MG) as a toxic dicarbonyl, and the knockdown of aldo-keto reductase (Akr), an enzyme of the polyol pathway, upregulated during hyperglycemia, in zebrafish.

Methodology: Zebrafish (tg(fli:egfp)) embryos were incubated with glucose and MG and the MG degrading enzyme Glo1 was knocked down in a morpholino-based approach. Phenotypic analysis was carried out using a confocal microscope and MG measurements were determined via HPLC. To analyze long term effects of increased MG or lack of Akr in zebrafish we are generating a permanent knockout using the CRISPR-Cas9-technology.

Findings: Incubation experiments using zebrafish embryos with glucose and MG as well as the Glo1-knockdown, all leads to increased MG concentrations and to a hyperbranching of smaller blood vessels, while larger blood vessels were not affected. The CRISPR constructs were generated and we are currently characterizing mosaic mutant zebrafish embryos. After raising and outcrossing these zebrafish, analysis of adult hetero- and homozygous zebrafish mutants will be carried out with a specific focus on the retina and kidney.

Conclusion: Collectively, the hyperbranching vascular phenotype based on increased MG levels and the generation of different mutant lines highlights the zebrafish as an excellent model organism to analyze late diabetic complications.