A sublethal concentration of nApoE4_1-151 leads to morphological abnormalities in zebrafish embryos at the hatching phase.

Representative light phase contrast microscopic images following live imaging of embryos at 48 hpf following a 24-hour period with respective treatments (Control, 25 μg/ml of nApoE3_1-151, or 25 μg/ml of nApoE4_1-151. A: Arrows point to consistent morphological changes as compared to untreated controls that are healthy and categorized as having a developmental abnormality score of <1.0 (Panel A). The blue arrows designate pigmentation changes; orange arrows designate cerebellar primordium junction differences in the hindbrain. B: Exogenous treatment with nApoE3_1-151 impacted pigmentation pattern in otherwise healthy embryos. Embryos in this category were most likely to receive a score of <1. C: 24-hour incubation of a sublethal concentration of nApoE4_1-151 resulted in developmental abnormality scores >3. Embryos in this category were typically observed to be delayed in development with limited hindbrain folding (orange arrow), limited or lacking pigmentation (blue arrow), as well as enlargement of the cardiac cavity (red arrow). D. Quantitative developmental abnormality scores for each treatment group following treatment of embryos for 24 hours with respective fragments at low E3 (orange bar), E4 (pink bar) concentrations (25 μg/ml) or at high concentrations (50 μg/ml) E3 (gray bar), E4 (yellow bar). The nApoE4_1-151 25 μg/ml-treated groups were significantly different from controls (H(4) = -2.43, p = 0.0074). At 50 μg/ml both nApoE4_1-151 (H(4) = -3.32, p = 0.0004) and nApoE3_1-151-treatment groups (H(4) = -1.77, p = 0.037) were significantly different from controls. Errors bars represent ± S.E.M. *p<0.05, **p<0.01, ***p<0.001 E. Heart rate data obtained from live microscope analyses in 25 μg/ml and 50 μg/ml treatment groups nApoE3_1-151 and nApoE4_1-151 compared to non-treated controls. nApoE4_1-151 (pink bar, 25 μg/ml) was significantly different from controls (H(4) = 1.77, p = 0.038). nApoE4_1-151 (yellow bar, 50 μg/ml) was significantly different from nApoE3_1-151 25 μg/ml (H(4) = 1.94, p = 0.026) and controls (H(4) = 2.665, p = 0.0036). Errors bars represent ±S.E.M. All other comparisons were insignificant. *p<0.05, **p<0.01, ***p<0.001.