Podocyte Expression of Membrane Transporters Involved in Puromycin Aminonucleoside-Mediated Injury

Cristina Zennaro,Mary Artero,Marco Stebel,Claudio Tiribelli,Michele Carraro,Elisa Trevisan,Lorella Pascolo,Vittorio Di Maso,Maria Pia Rastaldi,Hendrik W Van Veen

doi:10.1371/journal.pone.0066159

Abstract

Several complex mechanisms contribute to the maintenance of the intricate ramified morphology of glomerular podocytes and to interactions with neighboring cells and the underlying basement membrane. Recently, components of small molecule transporter families have been found in the podocyte membrane, but expression and function of membrane transporters in podocytes is largely unexplored. To investigate this complex field of investigation, we used two molecules which are known substrates of membrane transporters, namely Penicillin G and Puromycin Aminonucleoside (PA).We observed that Penicillin G pre-administration prevented both in vitro and in vivo podocyte damage caused by PA, suggesting the engagement of the same membrane transporters by the two molecules. Indeed, we found that podocytes express a series of transporters which are known to be used by Penicillin G, such as members of the Organic Anion Transporter Polypeptides (OATP/Oatp) family of influx transporters, and P-glycoprotein, a member of the MultiDrug Resistance (MDR) efflux transporter family.Expression of OATP/Oatp transporters was modified by PA treatment. Similarly, in vitro PA treatment increased mRNA and protein expression of P-glycoprotein, as well as its activity, confirming the engagement of the molecule upon PA administration.In summary, we have characterized some of the small molecule transporters present at the podocyte membrane, focusing on those used by PA to enter and exit the cell. Further investigation will be needed to understand precisely the role of these transporter families in maintaining podocyte homeostasis and in the pathogenesis of podocyte injury.

Highlights

Pathology of the podocyte underlies several glomerular diseases, the most severe being focal segmental glomerulosclerosis, which is clinically characterized by heavy proteinuria or nephrotic syndrome and frequently progresses to renal insufficiency
The effect was abolished in a dose-dependent manner by 15 min pre-incubation with Penicillin G and complete inhibition appeared at a concentration of 700 IU/ml, but a significant effect was already present at a concentration of 500 IU/ml (Fig. 1b)
When isolated glomeruli were exposed to Puromycin Aminonucleoside (PA) (10 ug/ ml) before Penicillin (700 Unit/ml), Palb remained elevated (0.7560.05 vs. 0.8960.20) (Fig. 1d), supporting the hypothesis that the Penicillin effect is exerted by blocking PA entrance into the cell

Summary

Introduction

Pathology of the podocyte underlies several glomerular diseases, the most severe being focal segmental glomerulosclerosis, which is clinically characterized by heavy proteinuria or nephrotic syndrome and frequently progresses to renal insufficiency. The membrane transport system, known as a major transmembrane component involved in the movement of ions, small molecules, and drugs, has been extensively studied in some organs and cellular systems, such as the intestine, the liver, and the bloodbrain barrier. Several members of the main membrane transporter families have been described in the renal tubule in the context of ion and drug transport [1]. Two transporters with broad specificity for small molecules have been identified in mouse podocytes in relation to drug clearance [2,3]. Small molecule transporters constitutively expressed in the podocyte contribute to maintain the cellular structure and barrier function, possibly by interacting with endogenous substrates and metabolites; and they could be involved in the uptake and efflux of drugs in these cells

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Conclusion