Abstract
The majority of kidney diseases arise from the loss of podocytes and from morphological changes of their highly complex foot process architecture, which inevitably leads to a reduced kidney filtration and total loss of kidney function. It could have been shown that microRNAs (miRs) play a pivotal role in the pathogenesis of podocyte-associated kidney diseases. Due to their fully functioning pronephric kidney, larval zebrafish have become a popular vertebrate model, to study kidney diseases in vivo. Unfortunately, there is no consensus about a proper normalization strategy of RT-qPCR-based miRNA expression data in zebrafish. In this study we analyzed 9 preselected candidates dre-miR-92a-3p, dre-miR-206-3p, dre-miR-99-1, dre-miR-92b-3p, dre-miR-363-3p, dre-let-7e, dre-miR-454a, dre-miR-30c-5p, dre-miR-126a-5p for their capability as endogenous reference genes in zebrafish experiments. Expression levels of potential candidates were measured in 3 different zebrafish strains, different developmental stages, and in different kidney disease models by RT-qPCR. Expression values were analyzed with NormFinder, BestKeeper, GeNorm, and DeltaCt and were tested for inter-group differences. All candidates show an abundant expression throughout all samples and relatively high stability. The most stable candidate without significant inter-group differences was dre-miR-92b-3p making it a suitable endogenous reference gene for RT-qPCR-based miR expression zebrafish studies.
Highlights
MicroRNAs regulate protein expression by translational suppression via RNA interference
While the proportion of embryos showing edema of any severity was similar in both groups, the phenotype of the nphs[1] knockdown embryos was generally more severe compared to the wt1a knockdown (Fig. 1B)
There is no consensus about endogenous reference controls for reverse transcription (RT)-qPCR derived miR expression data in zebrafish larvae
Summary
MicroRNAs (miRs) regulate protein expression by translational suppression via RNA interference. The fact that zebrafish develop a fully filtrating glomerulus attached to a pair of tubules with high structural and molecular homology two days past fertilization (dpf) makes this model highly relevant for the investigation of human glomerular kidney diseases[6]. Most of these diseases arise from highly specialized glomerular cells, the podocytes. There is no consensus about a normalization strategy in zebrafish miR expression data This is even more obvious when it comes to kidney research: Most studies make use of endogenous reference genes typically used in human and rodent studies. To the best of our knowledge there is no study dealing with suitable endogenous normalizers focusing on zebrafish development and kidney disease models
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
