Placental expression of RNU44, RNU48 and miR-16-5p: stability and relations with fetoplacental growth

Background/Objectives The current study aimed to identify suitable reference miRNA for placental miRNA expression analysis in a set of well-characterized and fetal-sex balanced small- (SGA) and appropriate- (AGA) for gestational age full-term singleton pregnancies. Subjects/Methods In this retrospective study, placental samples ( n = 106) from 35 SGA (19 male and 16 female) and 71 AGA (30 male and 41 female) full-term singleton pregnancies were utilized. Placental transcript abundance of three widely used reference miRNAs [miR-16-5p and Small nucleolar RNAs (snoRNAs) RNU44 and RNU48] were assessed by real-time quantitative PCR. Raw cycle threshold (C t ) analysis and RefFinder tool analysis were conducted for evaluating stability of expression of these miRNAs. Results Raw C t values of miR-16-5p were similar between SGA and AGA births ( P = 0.140) and between male and female births within SGA ( P = 0.159) and AGA ( P = 0.060) births while that of RNU44 and RNU48 were higher in SGA births ( P = 0.008 and 0.006 respectively) and in male births within the SGA group ( P = 0.005) for RNU44 and in female births within the AGA group ( P = 0.048) for RNU48. Across all 106 samples tested using the RefFinder tool, miR-16-5p and RNU44 were equally stable reference miRNAs. Conclusion We recommend miR-16-5p and RNU44 as suitable reference miRNAs for placental samples from settings similar to our study.