A Tissue-Bioengineering Strategy for Modeling Rare Human Kidney Diseases In Vivo

The lack of animal models for certain human diseases precludes our understanding of disease mechanisms and our ability to test new therapies in vivo. Here we generated kidney organoids from Tuberous Sclerosis Complex (TSC) patient-derived-hiPSCs to recapitulate a rare kidney tumor called angiomylipoma (AML). Organoids derived from TSC2-/- hiPSCs but not from isogenic TSC2+/- or TSC2+/+ hiPSCs shared a common transcriptional signature and a myomelanocytic cell phenotype with kidney AMLs, and developed epithelial cysts, replicating two major TSC-associated kidney lesions driven by genetic mechanisms that cannot be robustly and consistently recapitulated with transgenic mice. Transplantation of multiple TSC2-/- kidney organoids into the kidneys of immunodeficient rats allowed us to recapitulate AML and cystic kidney disease in vivo, in a scalable fashion and with fidelity, and to test the efficiency of rapamycin-loaded nanoparticles as a novel approach to ablate AMLs by inducing apoptosis triggered by mTOR-inhibition. Collectively, these methods represent a novel tissue-bioengineering strategy for rare disease modeling in vivo.

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A tissue-bioengineering strategy for modeling rare human kidney diseases in vivo

Nature Communications ◽

10.1038/s41467-021-26596-y ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

J. O. R. Hernandez ◽

X. Wang ◽

M. Vazquez-Segoviano ◽

M. Lopez-Marfil ◽

M. F. Sobral-Reyes ◽

...

Keyword(s):

Kidney Diseases ◽

Human Kidney ◽

Cell Phenotype ◽

Transcriptional Signature ◽

Disease Mechanisms ◽

Genetic Mechanisms ◽

Experimental Approaches ◽

Kidney Lesions ◽

Kidney Organoids

AbstractThe lack of animal models for some human diseases precludes our understanding of disease mechanisms and our ability to test prospective therapies in vivo. Generation of kidney organoids from Tuberous Sclerosis Complex (TSC) patient-derived-hiPSCs allows us to recapitulate a rare kidney tumor called angiomyolipoma (AML). Organoids derived from TSC2−/− hiPSCs but not from isogenic TSC2+/− or TSC2+/+ hiPSCs share a common transcriptional signature and a myomelanocytic cell phenotype with kidney AMLs, and develop epithelial cysts, replicating two major TSC-associated kidney lesions driven by genetic mechanisms that cannot be consistently recapitulated with transgenic mice. Transplantation of multiple TSC2−/− renal organoids into the kidneys of immunodeficient rats allows us to model AML in vivo for the study of tumor mechanisms, and to test the efficacy of rapamycin-loaded nanoparticles as an approach to rapidly ablate AMLs. Collectively, our experimental approaches represent an innovative and scalable tissue-bioengineering strategy for modeling rare kidney disease in vivo.

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Maturation of Nephrons by Implanting hPSC-Derived Kidney Progenitors Under Kidney Capsules of Unilaterally Nephrectomized Mice

10.21203/rs.3.rs-847264/v1 ◽

2021 ◽

Author(s):

Xin Yu ◽

Shan Jiang ◽

Kailin Li ◽

Xianzhen Yang ◽

Zhihe Xu ◽

...

Keyword(s):

Disease Modeling ◽

Vascular System ◽

Human Kidney ◽

Target Cells ◽

Mice Model ◽

Immunodeficient Mice ◽

Filtration Barrier ◽

Kidney Organoids

Abstract Background Human pluripotent stem cell (hPSCs)-derived kidney organoids may contribute to disease modeling and generation of kidney replacement tissues. However, realization of such applications requires the induction of hPSCs into functional mature organoids. One of the key questions for this process is whether a specific vascular system exists for nephrogenesis. Our previous study showed that implantation of hPSC-derived organoids below the kidney capsules of unilaterally nephrectomized immunodeficient mice for a short-term (2 weeks) resulted in the enlargement of organoids and production of vascular cells, although signs of maturation were lacking. Methods In this study, organoids are induced in vitro during 15 days and then sub-capsularly grafted into kidneys, we used the same unilaterally nephrectomized immunodeficient mice model to examine whether a medium -term (4 weeks) implantation could improve organoid maturation and vascularization, as evaluated by immunofluorescence and transmission electron microscopy(TEM). Results We demonstrate that after 2–4 weeks implantation, implanted renal organoids can form host-derived vascularization and mature in the absence of any exogenous vascular endothelial growth factor. Glomerular filtration barrier maturation was evidenced by glomerular basement membrane deposition, perforated glomerular endothelial cell development, as well as apical to basal podocyte polarization. A polarized monolayer epithelium and extensive brush border were also observed for tubular epithelial cells. Conclusions Our results indicate that the in vivo microenvironment is important for the maturation of human kidney organoids. Stromal expansion and a reduction of nephron structures were observed following longer-term (12 weeks) implantation,suggesting effects on off-target cells during the induction process. Accordingly, induction efficiency and transplantation models should be improved in the future.

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Towards Modelling Genetic Kidney Diseases with Human Pluripotent Stem Cells

Nephron ◽

10.1159/000514018 ◽

2021 ◽

pp. 1-12

Author(s):

Kirsty M. Rooney ◽

Adrian S. Woolf ◽

Susan J. Kimber

Keyword(s):

Stem Cell ◽

Kidney Disease ◽

Cell Biology ◽

Kidney Diseases ◽

Premature Mortality ◽

Cystic Kidney Disease ◽

Cystic Kidney ◽

Metanephric Mesenchyme ◽

Potential Therapeutic Application ◽

Made In

Background: Kidney disease causes major suffering and premature mortality worldwide. With no cure for kidney failure currently available, and with limited options for treatment, there is an urgent need to develop effective pharmaceutical interventions to slow or prevent kidney disease progression. Summary: In this review, we consider the feasibility of using human pluripotent stem cell-derived kidney tissues, or organoids, to model genetic kidney disease. Notable successes have been made in modelling genetic tubular diseases (e.g., cystinosis), polycystic kidney disease, and medullary cystic kidney disease. Organoid models have also been used to test novel therapies that ameliorate aberrant cell biology. Some progress has been made in modelling congenital glomerular disease, even though glomeruli within organoids are developmentally immature. Less progress has been made in modelling structural kidney malformations, perhaps because sufficiently mature metanephric mesenchyme-derived nephrons, ureteric bud-derived branching collecting ducts, and a prominent stromal cell population are not generated together within a single protocol. Key Messages: We predict that the field will advance significantly if organoids can be generated with a full complement of cell lineages and with kidney components displaying key physiological functions, such as glomerular filtration. The future economic upscaling of reproducible organoid generation will facilitate more widespread research applications, including the potential therapeutic application of these stem cell-based technologies.

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Stem cells and fluid flow drive cyst formation in an invertebrate excretory organ

eLife ◽

10.7554/elife.07405 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 37

Author(s):

Hanh Thi-Kim Vu ◽

Jochen C Rink ◽

Sean A McKinney ◽

Melainia McClain ◽

Naharajan Lakshmanaperumal ◽

...

Keyword(s):

Molecular Mechanisms ◽

Kidney Diseases ◽

Human Kidney ◽

Cyst Formation ◽

Cystic Kidney ◽

Flow Sensing ◽

Cell Proliferation And Differentiation ◽

Proliferation And Differentiation ◽

Genetic Interference ◽

Cystic Kidney Diseases

Cystic kidney diseases (CKDs) affect millions of people worldwide. The defining pathological features are fluid-filled cysts developing from nephric tubules due to defective flow sensing, cell proliferation and differentiation. The underlying molecular mechanisms, however, remain poorly understood, and the derived excretory systems of established invertebrate models (Caenorhabditis elegans and Drosophila melanogaster) are unsuitable to model CKDs. Systematic structure/function comparisons revealed that the combination of ultrafiltration and flow-associated filtrate modification that is central to CKD etiology is remarkably conserved between the planarian excretory system and the vertebrate nephron. Consistently, both RNA-mediated genetic interference (RNAi) of planarian orthologues of human CKD genes and inhibition of tubule flow led to tubular cystogenesis that share many features with vertebrate CKDs, suggesting deep mechanistic conservation. Our results demonstrate a common evolutionary origin of animal excretory systems and establish planarians as a novel and experimentally accessible invertebrate model for the study of human kidney pathologies.

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3D kidney organoids for bench-to-bedside translation

Journal of Molecular Medicine ◽

10.1007/s00109-020-01983-y ◽

2020 ◽

Author(s):

Navin Gupta✉ ◽

Emre Dilmen ◽

Ryuji Morizane

Keyword(s):

Collecting Duct ◽

Developmental Toxicity ◽

Human Kidney ◽

Great Promise ◽

Duct Systems ◽

Recent Developments ◽

Induced Pluripotent ◽

Kidney Organoids

Abstract The kidneys are essential organs that filter the blood, removing urinary waste while maintaining fluid and electrolyte homeostasis. Current conventional research models such as static cell cultures and animal models are insufficient to grasp the complex human in vivo situation or lack translational value. To accelerate kidney research, novel research tools are required. Recent developments have allowed the directed differentiation of induced pluripotent stem cells to generate kidney organoids. Kidney organoids resemble the human kidney in vitro and can be applied in regenerative medicine and as developmental, toxicity, and disease models. Although current studies have shown great promise, challenges remain including the immaturity, limited reproducibility, and lack of perfusable vascular and collecting duct systems. This review gives an overview of our current understanding of nephrogenesis that enabled the generation of kidney organoids. Next, the potential applications of kidney organoids are discussed followed by future perspectives. This review proposes that advancement in kidney organoid research will be facilitated through our increasing knowledge on nephrogenesis and combining promising techniques such as organ-on-a-chip models.

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A Sphingosine-1-Phosphate Modulator Ameliorates Polycystic Kidney Disease in Han:SPRD Rats

American Journal of Nephrology ◽

10.1159/000502855 ◽

2019 ◽

Vol 51 (1) ◽

pp. 1-10 ◽

Cited By ~ 1

Author(s):

Xin Li ◽

Ming Wu ◽

Limin Chen ◽

Junyan Lu ◽

Guo Li ◽

...

Keyword(s):

Kidney Disease ◽

Polycystic Kidney Disease ◽

Inflammatory Cytokines ◽

Inflammatory Responses ◽

Kidney Diseases ◽

Cordyceps Sinensis ◽

Cystic Kidney Disease ◽

Cystic Kidney ◽

Polycystic Kidney ◽

Sphingosine 1 Phosphate

Background: Inflammation plays an important role in polycystic kidney disease (PKD). Cordyceps sinensis, a prized Chinese medicinal herb, exerts anti-tumor, anti-inflammatory and anti-metastatic effects and benefits patients with kidney diseases. The aim of this study was to test the efficacy of FTY720, an immunosuppressant derived from C. sinensis, in a rat cystic kidney disease model, and explore its underlining mechanism. Methods: Male wild type and Cy/+ Han:SPRD rats were treated with FTY720 at 3 and 10 mg/kg/day for 5 weeks and 12 weeks by gavage. Blood and kidney were collected for functional, morphological, RNA, and protein analysis. Results: Inflammation is activated in Cy/+ Han:SPRD rats. Inflammatory cytokines including interleukin 6 and tumor necrosis factor alpha were upregulated and inflammation-related pathways were activated, such as nuclear factor κB and signal transducer and activator of transcription 3 (STAT3) pathways. Furthermore, the bioactive sphingolipid mediator sphingosine-1-phosphate (S1P), a regulator of inflammation, was accumulated in the Cy/+ Han:SPRD rats. FTY720 significantly reduced cyst growth and delayed disease progression by reducing the accumulation of S1P, thereby inhibiting inflammatory responses. Conclusion: FTY720 treatment reduced the expression of inflammatory cytokines and attenuated the activation of NK-κB and STAT3 pathways in Cy/+ Han:SPRD rats. It suggests that FTY720 may serve as a therapeutic agent for clinical autosomal dominant PKD treatment.

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Organoid single-cell profiling identifies a transcriptional signature of glomerular disease

10.1101/468850 ◽

2018 ◽

Author(s):

Jennifer L. Harder ◽

Rajasree Menon ◽

Edgar A. Otto ◽

Jian Zhou ◽

Sean Eddy ◽

...

Keyword(s):

Gene Expression ◽

Kidney Disease ◽

Single Cells ◽

Gene Expression Signature ◽

Human Kidney ◽

Glomerular Disease ◽

Podocyte Injury ◽

Compensation Mechanisms ◽

Kidney Organoids

ABSTRACTPodocyte injury is central to many forms of kidney disease, but transcriptional signatures reflecting podocyte injury and compensation mechanisms are challenging to analyze in vivo. Human kidney organoids derived from pluripotent stem cells (PSCs), a new model for disease and regeneration, present an opportunity to explore the transcriptional plasticity of podocytes. Here, transcriptional profiling of over 12,000 single cells from human PSC-derived kidney organoid cultures was used to identify robust and reproducible cell-lineage gene expression signatures shared with developing human kidneys based on trajectory analysis. Surprisingly, the gene expression signature characteristic of developing glomerular epithelial cells was also observed in glomerular tissue from a kidney disease cohort. This signature correlated with proteinuria and inverse eGFR, and was confirmed in an independent podocytopathy cohort. Three genes in particular were further identified as critical components of the glomerular disease signature. We conclude that cells in human PSC-derived kidney organoids reliably recapitulate the developmental transcriptional program of podocytes and other cell lineages in the human kidney, and that the early transcriptional profile seen in developing podocytes is reactivated in glomerular disease. Our findings demonstrate an innovative approach to identifying novel molecular programs involved in the pathogenesis of glomerulopathies.

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Interleukin-1β Activates a MYC-Dependent Metabolic Switch in Kidney Stromal Cells Necessary for Progressive Tubulointerstitial Fibrosis

Journal of the American Society of Nephrology ◽

10.1681/asn.2017121283 ◽

2018 ◽

Vol 29 (6) ◽

pp. 1690-1705 ◽

Cited By ~ 45

Author(s):

Dario R. Lemos ◽

Michael McMurdo ◽

Gamze Karaca ◽

Julia Wilflingseder ◽

Irina A. Leaf ◽

...

Keyword(s):

Stromal Cells ◽

Target Genes ◽

Kidney Injury ◽

Human Kidney ◽

Metabolic Switch ◽

Metabolic Derangement ◽

Sequestosome 1 ◽

Kidney Organoids

Background Kidney injury is characterized by persisting inflammation and fibrosis, yet mechanisms by which inflammatory signals drive fibrogenesis remain poorly defined.Methods RNA sequencing of fibrotic kidneys from patients with CKD identified a metabolic gene signature comprising loss of mitochondrial and oxidative phosphorylation gene expression with a concomitant increase in regulators and enzymes of glycolysis under the control of PGC1α and MYC transcription factors, respectively. We modeled this metabolic switch in vivo, in experimental murine models of kidney injury, and in vitro in human kidney stromal cells (SCs) and human kidney organoids.Results In mice, MYC and the target genes thereof became activated in resident SCs early after kidney injury, suggesting that acute innate immune signals regulate this transcriptional switch. In vitro, stimulation of purified human kidney SCs and human kidney organoids with IL-1β recapitulated the molecular events observed in vivo, inducing functional metabolic derangement characterized by increased MYC-dependent glycolysis, the latter proving necessary to drive proliferation and matrix production. MYC interacted directly with sequestosome 1/p62, which is involved in proteasomal degradation, and modulation of p62 expression caused inverse effects on MYC expression. IL-1β stimulated autophagy flux, causing degradation of p62 and accumulation of MYC. Inhibition of the IL-1R signal transducer kinase IRAK4 in vivo or inhibition of MYC in vivo as well as in human kidney organoids in vitro abrogated fibrosis and reduced tubular injury.Conclusions Our findings define a connection between IL-1β and metabolic switch in fibrosis initiation and progression and highlight IL-1β and MYC as potential therapeutic targets in tubulointerstitial diseases.

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Targeted exon skipping of a CEP290 mutation rescues Joubert syndrome phenotypes in vitro and in a murine model

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1809432115 ◽

2018 ◽

Vol 115 (49) ◽

pp. 12489-12494 ◽

Cited By ~ 16

Author(s):

Simon A. Ramsbottom ◽

Elisa Molinari ◽

Shalabh Srivastava ◽

Flora Silberman ◽

Charline Henry ◽

...

Keyword(s):

Kidney Disease ◽

Mouse Model ◽

Exon Skipping ◽

Joubert Syndrome ◽

Cystic Kidney Disease ◽

Cystic Kidney ◽

Kidney Cells ◽

Promising Therapeutic Approach

Genetic treatments of renal ciliopathies leading to cystic kidney disease would provide a real advance in current therapies. Mutations in CEP290 underlie a ciliopathy called Joubert syndrome (JBTS). Human disease phenotypes include cerebral, retinal, and renal disease, which typically progresses to end stage renal failure (ESRF) within the first two decades of life. While currently incurable, there is often a period of years between diagnosis and ESRF that provides a potential window for therapeutic intervention. By studying patient biopsies, patient-derived kidney cells, and a mouse model, we identify abnormal elongation of primary cilia as a key pathophysiological feature of CEP290-associated JBTS and show that antisense oligonucleotide (ASO)-induced splicing of the mutated exon (41, G1890*) restores protein expression in patient cells. We demonstrate that ASO-induced splicing leading to exon skipping is tolerated, resulting in correct localization of CEP290 protein to the ciliary transition zone, and restoration of normal cilia length in patient kidney cells. Using a gene trap Cep290 mouse model of JBTS, we show that systemic ASO treatment can reduce the cystic burden of diseased kidneys in vivo. These findings indicate that ASO treatment may represent a promising therapeutic approach for kidney disease in CEP290-associated ciliopathy syndromes.

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CaMKII as a pathological mediator of ER stress, oxidative stress, and mitochondrial dysfunction in a murine model of nephronophthisis

AJP Renal Physiology ◽

10.1152/ajprenal.00426.2015 ◽

2016 ◽

Vol 310 (11) ◽

pp. F1414-F1422 ◽

Cited By ~ 17

Author(s):

Christina Bracken ◽

Philippe Beauverger ◽

Olivier Duclos ◽

Ryan J. Russo ◽

Kelly A. Rogers ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Endoplasmic Reticulum Stress ◽

Oxidative Damage ◽

Molecular Mechanisms ◽

Kidney Diseases ◽

Cystic Kidney ◽

Effective Inhibition ◽

Stress Oxidative

Polycystic kidney diseases (PKDs) are genetic diseases characterized by renal cyst formation with increased cell proliferation, apoptosis, and transition to a secretory phenotype at the expense of terminal differentiation. Despite recent progress in understanding PKD pathogenesis and the emergence of potential therapies, the key molecular mechanisms promoting cystogenesis are not well understood. Here, we demonstrate that mechanisms including endoplasmic reticulum stress, oxidative damage, and compromised mitochondrial function all contribute to nephronophthisis-associated PKD. Ca2+/calmodulin-dependent protein kinase II (CaMKII) is emerging as a critical mediator of these cellular processes. Therefore, we reasoned that pharmacological targeting of CaMKII may translate into effective inhibition of PKD in jck mice. Our data demonstrate that CaMKII is activated within cystic kidney epithelia in jck mice. Blockade of CaMKII with a selective inhibitor results in effective inhibition of PKD in jck mice. Mechanistic experiments in vitro and in vivo demonstrated that CaMKII inhibition relieves endoplasmic reticulum stress and oxidative damage and improves mitochondrial integrity and membrane potential. Taken together, our data support CaMKII inhibition as a new and effective therapeutic avenue for the treatment of cystic diseases.

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