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

Author(s)

Munoz, J. J. ^[1] ; Anauate, A. C. ^[1] ; Amaral, A. G. ^{[2, 3]} ; Ferreira, F. M. ^[4] ; Meca, R. ^[1] ; Ormanji, M. S. ^[1] ; Boim, M. A. ^[1] ; Onuchic, L. F. ^{[2, 3]} ; Heilberg, I. P. ^[1]

Affiliation

^[1] Univ Fed Sao Paulo, Dept Med, Nephrol Div, Sao Paulo - Brazil

^[2] Univ Sao Paulo, Div Mol Med, Sch Med, Sao Paulo - Brazil

^[3] Univ Sao Paulo, Div Nephrol, Sch Med, Sao Paulo - Brazil

^[4] Univ Sao Paulo, Heart Inst, Lab Immunol, Sch Med, Sao Paulo - Brazil

• Document type: Journal article
• Source: SCIENTIFIC REPORTS; v. 10, n. 1 JAN 14 2020.
• Web of Science Citations: 0

Abstract

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 Delta Ct 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. (AU)

• FAPESP's process: 15/17152-3 - Smoking Effects on the Renal and Cardiac Phenotypes in Cystic Mice due to Pkd1 Inactivation
• Grantee: Luiz Fernando Onuchic
• Support Opportunities: Regular Research Grants
• FAPESP's process: 11/21593-4 - Caffeine effects on polycystic kidney disease development and progression, using a murine model Pkd1 cond/cond: Balcre
• Grantee: Ita Pfeferman Heilberg
• Support Opportunities: Regular Research Grants
• FAPESP's process: 18/09135-0 - Evaluation of fibrosis factors and microRNAs in the progression of the polycystic renal disease in a PKD1 deficient murine model
• Grantee: Juan José Augusto Moyano Muñoz
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 15/23345-9 - MicroRNAs, extracellular vesicles and stem cells: physiology, pathophysiological role and therapeutic potential in renal diseases
• Grantee: Mirian Aparecida Boim
• Support Opportunities: Research Projects - Thematic Grants