Within the glomerulus, podocytes are highly specialized visceral epithelial cells, that are part of the glomerular filtration barrier. Human podocyte cell culture is rather challenging for primary or immortalized cells, due to the non-proliferative state of the cells. In addition, de-differentiation is often observed rapidly. Hence, iPSC-derived podocytes offer an exciting alternative to culture podocyte-like cells from different donors over a prolonged time. Here we report an improved protocol of a simple and rapid one-step differentiation that drives iPSC into podocyte-like cells in 10 days and offers the possibility to transfer the podocyte-like cells into another plate format once.

Good Cell and Tissue Culture Practice (GCCP) 2.0 is an updated guidance document from GCCP 1.0 (published by ECVAM in 2005), which was developed for practical use in the laboratory to assure the reproducibility of in vitro (cellbased) work. The update in the guidance was essential as cell models have advanced dramatically to more complex culture systems and need more comprehensive quality management to ensure reproducibility and high-quality scientific data. This document describes six main principles to consider when performing cell culture including characterization and maintenance of essential characteristics, quality management, documentation and reporting, safety, education and training, and ethics. The document does not intend to impose detailed procedures but to describe potential quality issues. It is foreseen that the document will require further updates as the science and technologies evolve over time.

The renal proximal tubule is responsible for re-absorption of the majority of the glomerular filtrate and its proper function is necessary for whole-body homeostasis. Aging, certain diseases and chemical-induced toxicity are factors that contribute to proximal tubule injury and chronic kidney disease progression. To better understand these processes, it would be advantageous to generate renal tissues from human induced pluripotent stem cells (iPSC). Here, we report the differentiation and characterization of iPSC lines into proximal tubular-like cells (PTL). The protocol is a step wise exposure of small molecules and growth factors, including the GSK3 inhibitor (CHIR99021), the retinoic acid receptor activator (TTNPB), FGF9 and EGF, to drive iPSC to PTL via cell stages representing characteristics of early stages of renal development. Genome-wide RNA sequencing showed that PTL clustered within a kidney phenotype. PTL expressed proximal tubular-specific markers, including megalin (LRP2), showed a polarized phenotype, and were responsive to parathyroid hormone. PTL could take up albumin and exhibited ABCB1 transport activity. The phenotype was stable for up to 7 days and was maintained after passaging. This protocol will form the basis of an optimized strategy for molecular investigations using iPSC derived PTL.

Women are not small men. Sex-specific differences do not only affect the classical target organs of sexual differentiation and reproduction, but have been found to involve most, if not all the organs and tissues in the body. One of the consequences of this dimorphism is that diseases manifest in a sex- and gender-specific way. Key to maintenance of a healthy state is functioning tissue able to cope with insults. Regulated death of damaged cells and replacement with new cells by proliferation is a prerequisite for maintaining tissue function taking place at different pace in the different organs. The intent of this chapter is to review current evidence for sex-specific differences in tissue homeostasis focusing on the variability of hormone exposure characteristic for the female reproductive life stages.