Identification of housekeeping genes for microRNA expression analysis in kidney tissues of Pkd1 deficient mouse models

• Published in: Scientific reports Vol. 10; no. 1; p. 231
• Main Authors: Muñoz, J. J., Anauate, A. C., Amaral, A. G., Ferreira, F. M., Meca, R., Ormanji, M. S., Boim, M. A., Onuchic, L. F., Heilberg, I. P.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 14.01.2020; Nature Publishing Group
• Subjects: 38; 631/208/199; 631/337/384/331; 64; 64/60; 692/4022/1585/1589; Animal models; Animals; Gene Expression Profiling; Genes, Essential - genetics; Haploinsufficiency; Humanities and Social Sciences; Kidney - metabolism; Kidney diseases; Kidneys; Mice; MicroRNAs; MicroRNAs - genetics; miRNA; multidisciplinary; Polycystic kidney; Protein Kinase C - deficiency; Protein Kinase C - genetics; Rodents; Science; Science (multidisciplinary)
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-019-57112-4

Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1 -deficient mouse models from a panel of five candidates genes ( miR-20a , miR- 25, miR-26a , miR-191 and U6 ) and 3 target genes ( miR-17 , miR-21 and let-7a ) using samples from kidneys of cystic mice ( Pkd1 flox/flox : Nestin cre , CY), non-cystic controls ( Pkd1 flox/flox , NC), Pkd1 -haploinsufficient ( Pkd1 +/− , HT), wild-type controls ( Pkd1 +/+ , WT), severely cystic mice ( Pkd1 V/V , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR- 25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a , miR- 25 and miR-26a . Our findings established miR-20a , miR- 25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1 -deficient mouse models.