Allostimulatory capacity of conditionally immortalized proximal tubule cell lines for bioartificial kidney application

Milos Mihajlovic,Rosalinde Masereeuw,Joost G Hoenderop,Lambertus P Van Den Heuvel,Dimitrios Stamatialis,Wil A Allebes,Luuk B Hilbrands,Martijn J Wilmer,Jitske Jansen,Annemarie J F Westheim

doi:10.1038/s41598-017-07582-1

Abstract

Novel renal replacement therapies, such as a bioartificial kidney (BAK), are needed to improve current hemodialysis treatment of end-stage renal disease (ESRD) patients. As BAK applications may reveal safety concerns, we assessed the alloimmunization and related safety aspects of readily available conditionally immortalized human proximal tubule epithelial cell (ciPTEC) lines to be used in BAK. Two ciPTEC lines, originally derived from urine and kidney tissue, were characterized for the expression and secretion of relevant molecules involved in alloimmunization and inflammatory responses, such as HLA class-I, HLA-DR, CD40, CD80, CD86, as wells as soluble HLA class I and proinflammatory cytokines (IL-6, IL-8 and TNF-α). A lack of direct immunogenic effect of ciPTEC was shown in co-culture experiments with peripheral blood mononuclear cells (PBMC), after appropriate stimulation of ciPTEC. Tight epithelial cell monolayer formation on polyethersulfone flat membranes was confirmed by zonula occludens-1 (ZO-1) expression in the ciPTEC tight junctions, and by restricted inulin-FITC diffusion. Co-culture with (activated) PBMC did not jeopardize the transepithelial barrier function of ciPTEC. In conclusion, the absence of allostimulatory effects and the stability of ciPTEC monolayers show that these unique cells could represent a safe option for BAK engineering application.

Highlights

Novel renal replacement therapies, such as a bioartificial kidney (BAK), are needed to improve current hemodialysis treatment of end-stage renal disease (ESRD) patients
The surface expression of Human Leukocyte Antigens (HLA) class I molecules was evaluated in both conditionally immortalized human proximal tubule epithelial cell (ciPTEC)-U and –T1 lines in basal, as well as in several stimulatory conditions, using W6/32 monoclonal antibody directed against the core of HLA-A, B and C antigens
We demonstrated the lack of immunogenic effects of ciPTEC studying several aspects of the immune system as a part of safety assessment required during the BAK device development

Summary

Introduction

Novel renal replacement therapies, such as a bioartificial kidney (BAK), are needed to improve current hemodialysis treatment of end-stage renal disease (ESRD) patients. Novel therapies for CKD are needed, and one of the most promising options is the bioartificial kidney (BAK) device, composed of proximal tubule epithelial cells (PTEC) cultured on hollow fiber membranes www.nature.com/scientificreports/. We developed conditionally immortalized proximal tubule epithelial cell lines (ciPTEC), derived from human urine or kidney tissue, as an unlimited and invariable cell source for BAK application . CiPTEC were immortalized with the temperature-sensitive mutant U19tsA58 of SV40 large T antigen (SV40T) and the essential catalytic subunit of human telomerase (hTERT), as described[9,10,11] This allows the cells to proliferate at the permissive temperature of 33 °C and to fully differentiate to mature PTEC at non-permissive temperature of 37 °C. This allows the cells to proliferate at the permissive temperature of 33 °C and to fully differentiate to mature PTEC at non-permissive temperature of 37 °C. ciPTEC were extensively characterized for most proximal tubule functions such as reabsorption and excretory transport activities and successfully cultured on biofunctionalized HFM6–8, 12

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Conclusion