Megalin expression in human term and preterm placental villous tissues: Effect of gestational age and sample processing and storage time

Introduction: The aims of this study were to characterize megalin expression in human term and preterm placental villous tissues and to assess the impact of gestational age and sample storage on receptor expression. Methods: Placental tissue samples were collected from pregnant women undergoing term and preterm Cesarean deliveries. Placental villous tissues were used to quantify megalin protein and mRNA expression by western blotting and quantitative polymerase chain reaction (q-PCR), respectively. Stability of megalin expression was also evaluated under various processing and storage conditions. Results: Megalin mRNA was detected in term and preterm placental villous tissues. Expression in early preterm samples was 6-fold higher than in late preterm and term samples. Refrigeration of processed term samples at 4 degrees C for up to 18 h had a slight impact on megalin mRNA expression with stored samples exhibiting mRNA levels approximately 1.5-fold lower than those frozen immediately after processing. A greater decrease in mRNA expression (up to 33-fold) was observed when processed samples were snap-frozen immediately and thawed at 4 degrees C. Processing of samples prior to refrigeration also appeared to improve mRNA stability with significantly higher expression levels noted in processed vs. unprocessed samples at all points for up to 48 h. Discussion: These data suggest that expression of megalin mRNA in term placental villous tissue is relatively stable for up to 18 h when samples are processed immediately and refrigerated at 4 degrees C prior to freezing. Processing prior to storage also appears to improve mRNA stability. This paper demonstrates the practical feasibility of analyzing stored tissue samples, thus, it will help with placental mRNA analysis. Additionally, megalin expression appears to vary inversely with gestational age with the greatest expression noted in the most premature samples. Age-dependent differences in placental megalin may therefore influence fetal exposure. (C) 2014 Elsevier Inc. All rights reserved.