Three-dimensional spheroid culture induces apical-basal polarity and the original characteristics of immortalized human renal proximal tubular epithelial cells

概要

The kidney plays an essential role in maintaining the homeostasis of the human body by regulating blood components and pressure, and producing urine. Proximal tubules, which are a sub-region of the tubules, are an epithelial tissue located between the Bowman's sac and Henle’s loop. The proximal tubules play an essential role in regulating blood homeostasis by absorbing amino acids, glucose, water, and ions such as sodium, potassium, and bicarbonate. Impaired function of the proximal tubules is known to cause various diseases such as diabetic nephropathy, proximal tubular acidosis, and Renal Fanconi syndrome. Further, proximal tubules are known to be particularly sensitive to ischemia and xenobiotic nephrotoxicity. The kidney is one of the most complex structures among all organs, along with the brain, and it is difficult to assess the mechanism of proximal tubular damage and the function of proximal tubules on a cell-by-cell basis in in vivo experiments using animal models. Therefore, in vitro experiments are necessary to elucidate the characteristics, functions, disease mechanisms, and drug toxicity of the proximal tubules in detail. Renal proximal tubular epithelial cells (RPTECs) are responsible for the main function of the proximal tubules; thus, studies focusing on RPTECs will greatly enhance our understanding of nephrotoxicity, proximal tubule function, and the diseases associated with their dysfunction. RPTECs, which retain the properties of in vivo cells, are necessary to study the physiology and pathobiology of the proximal tubules accurately. However, human primary cells, which can provide the most ideal models, are limited by donor availability and variability. Moreover, RPTECs derived from non-human animal species cannot be an accurate assessment model for human research, because of the species difference. Therefore, it is preferable to use immortalized RPTECs, such as HK-2 cells, which are derived from humans and maintain indefinite cell growth. However, they are not suitable for use as a model cell in conventional two-dimensional (2D) culture systems because of the loss of their original properties as RPTECs. To overcome this limitation, we developed a three-dimensional (3D) spheroid culture method for HK-2 cells using an extracellular matrix. HK-2 spheroids in 3D culture formed a tubule-like architecture with cellular polarity and showed markedly restored Na transport function. 3D culture of HK-2 cells also increased kidney development-related genes, including WNT9B. Models of human renal tubules using HK-2 spheroids will greatly improve our understanding of the physiology and pathobiology of the kidney.