An In Vivo Zebrafish Model for Interrogating ROS-Mediated Pancreatic β-Cell Injury, Response, and Prevention

Figure 1

Time-dependent metronidazole induction of β-cell-specific ROS. (a) Schematic of MTZ treatments and imaging. Zebrafish (NTR⁺) larvae were treated with MTZ or vehicle for 0, 1, 3, 6, 12, or 24 hours with a “staggered start” such that all treatments were completed simultaneously; larvae were then incubated with CellROX green stain at 105 hpf and fixed/analyzed at 106 hpf. (b) Representative immunofluorescence images of zebrafish pancreatic islets stained with insulin antibody and CellROX green after 7.5 mM MTZ treatments. Magnified insets (bounded by dashed boxes) highlight the dose-dependent increase in CellROX green signal in β-cells. (c) Quantification of CellROX green intensity in insulin-positive β-cells showing a significant increase in ROS generation after 1, 3, 6, and 12 hours of MTZ treatment as compared to vehicle-treated controls ( for each condition). (d) MTZ treatment caused a significant decrease in β-cell area after 12 or 24 hours of treatment as compared to untreated controls. (e) Representative immunofluorescence images of zebrafish pancreatic islets treated for 3 hours with 0 or 7.5 mM MTZ and stained with 5 μM DHE. Dotted lines demarcate the boundaries of the pancreas. Graphed data are presented as mean ± SEM (). Statistical significance was determined by one-way ANOVA followed by post hoc Holm-Sidak test. Scale bar indicates 10 μm.

(a)

(b)

(c)

(d)

(e)