Polarization of primary human renal proximal tubular epithelial cells - a pilot study

Abstract

Introduction: Epithelial cells are specialized cells with an apicobasal polarity that allows them to exert their functions, such as acting as a barrier and controlling flow and transport of molecules across the epithelial layer. Polarized epithelial cell models have been developed for several organs and yield a more in vivo-like organization than cell monolayers. BK Polyomavirus (BKPyV) is a ubiquitous virus that infects polarized epithelial cells in the reno-urinary tract. High-level replication in these cells may cause severe disease. Much research has been done on BKPyV, but the majority of studies on BKPyV have been done in non-polarized epithelial cell culture models. We therefore chose to develop a polarized renal cell culture model for assessing the replication cycle of BKPyV. Materials and methods: Adult and fetal human primary renal proximal tubule epithelial cells (RPTECs), immortalized RPTECs and urinary cells were seeded on collagen-coated permeable supports and allowed to differentiate. After differentiation, cells were examined for polarity markers by immunofluorescence and/or electron microscopy. For infection studies, RPTECs were infected with BKPyV, and after three days the cells were fixed and immunofluorescence staining with primary antibodies directed against BKPyV VP1 and agnoprotein was performed. Results: Immunofluorescence microscopy demonstrated that adult RPTECs, immortalized RPTECs and urinary cells developed a polarized morphology with microvilli, primary cilium and apicobasal polarity. Electron microscopy of adult RPTECs confirmed the presence of microvilli. In contrast, fetal RPTECs did not polarize in culture. After polarization, adult RPTECs were still permissive for BKPyV-infection as shown by agnoprotein- and VP1-staining three days after infection. Conclusions: Adult human RPTECs, immortalized RPTECs and urinary cells develop a polarized morphology on permeable supports. Polarized adult RPTECs are permissive for BKPyV infection. These cell culture models will be useful for research on BKPyV.