Early neonatal adaptation to extrauterine life is i.a. dependent on effective mitochondrial biogenesis during foetal development. Understanding of mitochondrial biogenesis is limited, because only scarce data are available from prenatal studies including RNA analyses in human foetal tissues. Aims of the study were focused on the factors affecting RNA quality in human placental tissue (HPT) including temperature, time period before HPT freezing and the Apgar score. In addition, optimal reference genes for mRNA quantification by real-time PCR in HPT were studied. Samples of HPT were obtained after the birth of 20 term neonates. Seven HPT were used for the time-course study of RNA degradation in two different temperatures (0 °C and 24 °C). Various instruments NanoDrop (NanoDrop Technologies), Experion (Bio-Rad Laboratories), Agilent 2100 Bioanalyzer (Agilent Technologies) were used for analysis of RNA integrity, purity and yield. Identification of suitable reference genes was achieved by analysing six candidate genes (ATP5O, SDHA, TBP, HPRT, PMBS, ATP6) for their expression stability (GeNorm application). The results showed that the HPT samples for RNA analyses must be frozen immediately after birth in –80 °C or stored at 0 °C maximally for 1 hour. The reference genes ATP50 and SDHA were the most stable for mRNA quantification in HPT. Human placenta represents easily obtainable source of foetal tissue for studies concerning mitochondrial biogenesis. We demonstrated that the critical limit for optimal storage and handling of HPT are the temperature and the time period before freezing of the samples.

Department of Pediatrics and Adolescent Medicine 1st Faculty of Medicine Charles University Prague and General University Hospital Prague Prague Czech Republic

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