Abstract
Altered levels of trace elements are associated with increased oxidative stress that is eventually responsible for pathologic conditions. Oxidative stress has been proposed to be involved in eye diseases, including cataract formation. We visualized the distribution of metals and other trace elements in the eye of zebrafish embryos by micro X-ray fluorescence (μ-XRF) imaging. Many elements showed highest accumulation in the retinal pigment epithelium (RPE) of the zebrafish embryo. Knockdown of the zebrafish brown locus homologues tyrp1a/b eliminated accumulation of these elements in the RPE, indicating that they are bound by mature melanosomes. Furthermore, albino (slc45a2) mutants, which completely lack melanosomes, developed abnormal lens reflections similar to the congenital cataract caused by mutation of the myosin chaperon Unc45b, and an in situ spin trapping assay revealed increased oxidative stress in the lens of albino mutants. Finally transplanting a wildtype lens into an albino mutant background resulted in cataract formation. These data suggest that melanosomes in pigment epithelial cells protect the lens from oxidative stress during embryonic development, likely by buffering trace elements.
Highlights
Of a fluorescence spectrum recorded from the sample overlaid with standard spectra from individual elements
An enhanced iron signal was detected in the pigment epithelial layer as well as in the highly vascularised choroidal and primordial hyaloid vasculature
The photoreceptor layer of the retina contains higher amounts of copper and iron than the other inner retinal layers. In light of these observations, we focused on the accumulation of elements in the RPE, which suggests the presence of a dedicated storage structure with potential biological function
Summary
Of a fluorescence spectrum recorded from the sample (black) overlaid with standard spectra from individual elements. To visualize sub-cellular element distributions in biological tissues, a number of chemical, or genetically encoded, fluorescent indicators have been developed for some elements such as calcium[10] and zinc[11] These fluorescent indicators have revealed dynamics of individual metal ions in many biological processes, the endogenous subcellular element distribution needs to be verified by direct visualization. In this context, hard X-ray fluorescence microscopy (μ -XRF) provides complementary and unique information. These results imply that melanosomes contribute to lens integrity by buffering elements that would otherwise cause deleterious lens protein modifications
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