Human kidney organoids model the tacrolimus nephrotoxicity and elucidate the role of autophagy
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Background/Aims: Tacrolimus has been used as an immunosuppressive agent in organ transplantation. Despite the therapeutic benefits, tacrolimus’s use is limited due to its nephrotoxicity. To reduce tacrolimus nephrotoxicity, effective humanized experimental models may be helpful. Here, we modeled tacrolimus nephrotoxicity using kidney organoids derived from human inducible pluripotent stem cells (iPSCs) in vitro. Methods: Kidney organoids were differentiated from the CMC11 iPSC cell line, re-seeded in 96-well plates, and treated with tacrolimus at doses of 0, 30, or 60 μM for 24 hours. This in vitro model was compared to a mouse model of tacrolimus nephrotoxicity and the associated mechanisms were investigated. Results: The size of the kidney organoids and cell viability decreased in dose-dependent manners after treatment with tacrolimus. The number of tubular cells decreased with a loss of polarity, similar to the effects seen in mouse tacrolimus nephrotoxicity. Ultrastructural analysis showed numerous vacuoles in the proximal tubular cells of the kidney organoids treated with tacrolimus. Tacrolimus treatment induced oxidative stress and mitochondrial dysfunction, and autophagic activity was enhanced in the kidney organoids. Rapamycin, an autophagy inducer, accelerated cell death in the kidney organoid model of tacrolimus nephrotoxicity, which was attenuated by treatment with 3-methyladenine, an autophagy inhibitor. These findings indicate that the augmentation of autophagy by rapamycin treatment accelerated tacrolimus nephrotoxicity. Conclusions: Our data suggest that human kidney organoids are an effective in vitro model of tacrolimus nephrotoxicity and that autophagy plays a critical role in tacrolimus nephrotoxicity.
Abstract
Background/Aims: Tacrolimus has been used as an immunosuppressive agent in organ transplantation. Despite the therapeutic benefits, tacrolimus’s use is limited due to its nephrotoxicity. To reduce tacrolimus nephrotoxicity, effective humanized experimental models may be helpful. Here, we modeled tacrolimus nephrotoxicity using kidney organoids derived from human inducible pluripotent stem cells (iPSCs) in vitro. Methods: Kidney organoids were differentiated from the CMC11 iPSC cell line, re-seeded in 96-well plates, and treated with tacrolimus at doses of 0, 30, or 60 μM for 24 hours. This in vitro model was compared to a mouse model of tacrolimus nephrotoxicity and the associated mechanisms were investigated. Results: The size of the kidney organoids and cell viability decreased in dose-dependent manners after treatment with tacrolimus. The number of tubular cells decreased with a loss of polarity, similar to the effects seen in mouse tacrolimus nephrotoxicity. Ultrastructural analysis showed numerous vacuoles in the proximal tubular cells of the kidney organoids treated with tacrolimus. Tacrolimus treatment induced oxidative stress and mitochondrial dysfunction, and autophagic activity was enhanced in the kidney organoids. Rapamycin, an autophagy inducer, accelerated cell death in the kidney organoid model of tacrolimus nephrotoxicity, which was attenuated by treatment with 3-methyladenine, an autophagy inhibitor. These findings indicate that the augmentation of autophagy by rapamycin treatment accelerated tacrolimus nephrotoxicity. Conclusions: Our data suggest that human kidney organoids are an effective in vitro model of tacrolimus nephrotoxicity and that autophagy plays a critical role in tacrolimus nephrotoxicity.
참고문헌 (36)
- Liptak P, "Primer : histopathology of calcineurin-inhibitor toxicity in renal allografts", Nat Clin Pract Nephrol, 2, pp.398~404 (2006)
- 윤혜은, "Established and Newly Proposed Mechanisms of Chronic Cyclosporine Nephropathy", The Korean Journal of Internal Medicine, 24(2), pp.81~92 (2009), 대한내과학회
- Wu Q, "Mechanism of cyclosporine A nephrotoxicity : oxidative stress, autophagy, and signalings", Food Chem Toxicol, 118, pp.889~907 (2018)
- Kumar A, "Oxidative stress in kidney transplant biopsies", Exp Clin Transplant, 13(Suppl 1), pp.207~213 (2015)
- Freedman BS, "Modeling kidney disease with iPS cells", Biomark Insights, 10(Suppl 1), pp.153~169 (2015)
- Wilmer MJ, "Kidney-on-a-chip technology for drug-induced nephrotoxicity screening", Trends Biotechnol, 34, pp.156~170 (2016)
- Cukierman E, "Taking cell-matrix adhesions to the third dimension", Science, 294, pp.1708~1712 (2001)
- Cukierman E, "Cell interactions with three-dimensional matrices", Curr Opin Cell Biol, 14, pp.633~639 (2002)
- Astashkina AI, "A 3-D organoid kidney culture model engineered for high-throughput nephrotoxicity assays", Biomaterials, 33, pp.4700~4711 (2012)
- Freedman BS, "Modelling kidney disease with CRISPR-mutant kidney organoids derived from human pluripotent epiblast spheroids", Nat Commun, 6, pp.8715~ (2015)
- Kim YK, "Gene-edited human kidney organoids reveal mechanisms of disease in podocyte development", Stem Cells, 35, pp.2366~2378 (2017)
- Morizane R, "Nephron organoids derived from human pluripotent stem cells model kidney development and injury", Nat Biotechnol, 33, pp.1193~1200 (2015)
- Taguchi A, "Redefining the in vivo origin of metanephric nephron progenitors enables generation of complex kidney structures from pluripotent stem cells", Cell Stem Cell, 14, pp.53~67 (2014)
- Takasato M, "Generation of kidney organoids from human pluripotent stem cells", Nat Protoc, 11, pp.1681~1692 (2016)
- Wu H, "Comparative analysis and refinement of human PSC-derived kidney organoid differentiation with single-cell transcriptomics", Cell Stem Cell, 23, pp.869~881 (2018)
- Sun Ah Nam, "Graft immaturity and safety concerns in transplanted human kidney organoids", Experimental and Molecular Medicine, 51, pp.1~13 (2019), 생화학분자생물학회[DOI]
- Czerniecki SM, "High-throughput screening enhances kidney organoid differentiation from human pluripotent stem cells and enables automated multidimensional phenotyping", Cell Stem Cell, 22, pp.929~940 (2018)
- Takasu C, "Treatment with dimethyl fumarate attenuates calcineurin inhibitor-induced nephrotoxicity", Transplantation, 99, pp.1144~1150 (2015)
- Yu JH, "Coenzyme Q10 alleviates tacrolimus-induced mitochondrial dysfunction in kidney", FASEB J, 33, pp.12288~12298 (2019)
- Lim SW, "Effect of Klotho on autophagy clearance in tacrolimus-induced renal injury", FASEB J, 33, pp.2694~2706 (2019)
- Ott M, "Mitochondria, oxidative stress and cell death", Apoptosis, 12, pp.913~922 (2007)
- Hajnoczky G, "Mitochondrial calcium signalling and cell death : approaches for assessing the role of mitochondrial Ca2+ uptake in apoptosis", Cell Calcium, 40, pp.553~560 (2006)
- Lin DS, "Oxidative insults and mitochondrial DNA mutation promote enhanced autophagy and mitophagy compromising cell viability in pluripotent cell model of mitochondrial disease", Cells, 8, pp.65~ (2019)
- Klionsky DJ, "Autophagy : from phenomenology to molecular understanding in less than a decade", Nat Rev Mol Cell Biol, 8, pp.931~937 (2007)
- Levine B, "Autophagy in the pathogenesis of disease", Cell, 132, pp.27~42 (2008)
- Komatsu M, "p62", SQSTM1, Pharmacol Res, pp.2012, 66[DOI]
- Chan LL, "A novel image-based cytometry method for autophagy detection in living cells", Autophagy, 8, pp.1371~1382 (2012)
- Yu L, "Termination of autophagy and reformation of lysosomes regulated by mTOR", Nature, 465, pp.942~946 (2010)
- DeVorkin L, "LysoTracker staining to aid in monitoring autophagy in Drosophila", Cold Spring Harb Protoc, 2014, pp.951~958 (2014)
- Wu L, "Rapamycin upregulates autophagy by inhibiting the mTOR-ULK1 pathway, resulting in reduced podocyte injury", PLoS One, 8, pp.e63799~ (2013)
- Faria J, "Kidney-based in vitro models for drug-induced toxicity testing", Arch Toxicol, 93, pp.3397~3418 (2019)
- Troth SP, "Kidney safety assessment : current practices in drug development", Semin Nephrol, 39, pp.120~131 (2019)
- Singh NK, "Three-dimensional cell-printing of advanced renal tubular tissue analogue", Biomaterials, 232, pp.119734~ (2020)
- Pallet N, "Autophagy protects renal tubular cells against cyclosporine toxicity", Autophagy, 4, pp.783~791 (2008)
- Lim SW, "Chronic cyclosporine nephropathy is characterized by excessive autophagosome formation and decreased autophagic clearance", Transplantation, 94, pp.218~225 (2012)
- Gallon L, "Long-term renal allograft function on a tacrolimus-based, pred-free maintenance immunosuppression comparing sirolimus vs. MMF", Am J Transplant, 6, pp.1617~1623 (2006)