Abstract

Background: Finding the right drug-dosage for neonates is still a challenge. Until now, neonatal doses are extrapolated from adults and children doses. However, there are differences between neonatal and adult kidney physiology that should be considered, especially when it comes to drug metabolism and/or transport. Studying renal drug disposition in neonates is limited by the lack of reliable human cell models.

Objective: To illustrate the feasibility of developing an in vitro model for neonatal proximal tubule epithelial cells (nPTECs) to study renal drug disposition at this age.

Method: nPTECs were isolated from urine samples of neonates of different gestational ages and were conditionally immortalized using a temperature sensitive SV40T antigen and human telomerase hTERT. Cell clones were characterized on gene expression level for PTEC markers such as P-glycoprotein (ABCB1), aquaporin1 (AQP1), and organic cation transport protein 2 (SLC22A2), and for kidney progenitor cell and podocyte markers. In addition, protein expression and functional assessment were performed for P-gp and OCT2.

Results: We established 101 clonal cell lines of conditionally immortalized nPTECs derived from neonatal urines. Characterization of primary cells lines showed expression of genes from different cell types such as progenitors, PTECs and podocytes, however the developed conditionally immortalized nPTECs only expressed proximal tubule markers. Quantitative PCR analysis confirmed the expression of proximal tubule markers in nPTECs similar to the adult control PTECs. P-gp was expressed in nPTECs derived from the different gestational ages with a similar functionality compared with adult derived PTECs. In contrast, OCT2 functionality was significantly lower in nPTEC cell lines compared with adult PTECs.

Conclusion: We demonstrate the feasibility of culturing proximal tubule epithelial cells with high efficiency from urine of neonates. These cells expressed PTEC-specific genes and functional drug transporters. The cell model presented is a valuable tool to study proximal tubule physiology and pharmacology in newborns. In addition, we demonstrate the physiological differences between the neonatal and adult kidney, which emphasizes the importance of studying drug disposition in neonatal models instead of extrapolating from adult data.

Keywords: Drug transporters, neonates, pharmacology, PTECs, AQP1, OCT2, P-gp.