Podocytes in culture: past, present, and future

Abstract

Human genetic and in vivo animal studies have helped to define the critical importance of podocytes for kidney function in health and disease. However, as in any other research area, by default these approaches do not allow for mechanistic studies. Such mechanistic studies require the availability of cells grown ex vivo (i.e., in culture) with the ability to directly study mechanistic events and control the environment such that specific hypotheses can be tested. A seminal breakthrough came about a decade ago with the documentation of differentiation in culture of primary rat and human podocytes and the subsequent development of conditionally immortalized differentiated podocyte cell lines that allow deciphering the decisive steps of differentiation and function of 'in vivo' podocytes. Although this paper is not intended to provide a comprehensive review of podocyte biology, nor their role in proteinuric renal diseases or progressive glomerulosclerosis, it will focus specifically on several aspects of podocytes in culture. In particular, we will discuss the scientific and research rationale and need for cultured podocytes, how podocyte cell-culture evolved, and how cultured podocytes are currently being used to uncover novel functions of podocytes that can then be validated in vivo in animal or human studies. In addition, we provide a detailed description of how to properly culture and characterize podocytes to avoid potential pitfalls.

Establishment of Conditionally Immortalized Mouse Glomerular Parietal Epithelial Cells in Culture

“…These cells were larger in size, more irregular in shape, and flatter; had multiple cellular processes; and grew at a slower rate. On the basis of our experience, 8 the morphology of these cells was consistent with podocytes ( Figure 2, F and G), similar to those that we 8 and others previously characterized. 9,10 Decapsulated glomeruli were easily distinguished from capsulated glomeruli under the microscope by their characteristic shape ( Figure 2, H and I).…”

“…17 Kidney cortex from ImmortoMice was removed, minced, and subjected to standard differential sieving under sterile conditions to isolate glomeruli as previously reported. 8,18,19 All animal procedures were conducted in accordance with the Institutional Animal Care and Use Committee. Isolated capsulated (contains the glomerular tuft, Bowman's membrane, and PEC) and decapsulated (contains the glomerular tuft only) glomeruli were plated onto 10-cm primaria plates (BD Biosciences, San Jose, CA) coated with type I collagen (BD Biosciences).…”

“…Because it is critical to ensure that the targeted cell population generated was indeed mPEC and not podocytes, separate studies were performed in parallel on a well-characterized conditionally immortalized mouse podocyte cell line in culture also derived from ImmortoMice 8,19 for comparisons with mPEC.…”

Wnt/β-Catenin Pathway in Podocytes Integrates Cell Adhesion, Differentiation, and Survival

“…FloxE3/FloxE3 mice (30,37). Podocyte clones (n ϭ 7) were established by serial dilution and selected based on their expression of podocyte-specific markers (Nphs1, Nphs2, Synpo, and Wt1).…”

Curcumin attenuates high glucose-induced podocyte apoptosis by regulating functional connections between caveolin-1 phosphorylation and ROS

“…Podocytes are terminally differentiated and highly specialized glomerular visceral epithelial cells that form part of the glomerular filtration barrier, which prevents urinary protein loss [2] . Several recent studies suggested that podocyte damage may play a key role in the pathogenesis of DN [3][4][5] . Thus, finding an effective therapy that protects podocytes from injury may provide a promising therapeutic modality for DN treatment.…”