The Utility of Human Kidney Organoids in Modeling Kidney Disease

2020 ◽  
Vol 40 (2) ◽  
pp. 188-198
Author(s):  
Aneta Przepiorski ◽  
Amanda E. Crunk ◽  
Eugenel B. Espiritu ◽  
Neil A. Hukriede ◽  
Alan J. Davidson
2018 ◽  
Author(s):  
Jennifer L. Harder ◽  
Rajasree Menon ◽  
Edgar A. Otto ◽  
Jian Zhou ◽  
Sean Eddy ◽  
...  

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.


2019 ◽  
Author(s):  
Esther Liu ◽  
Behram Radmanesh ◽  
Byungha H. Chung ◽  
Michael D. Donnan ◽  
Dan Yi ◽  
...  

ABSTRACTBackgroundDNA variants in APOL1 associate with kidney disease, but the pathophysiological mechanisms remain incompletely understood. Model organisms lack the APOL1 gene, limiting the degree to which disease states can be recapitulated. Here we present single-cell RNA sequencing (scRNA-seq) of genome-edited human kidney organoids as a platform for profiling effects of APOL1 risk variants in diverse nephron cell types.MethodsWe performed footprint-free CRISPR-Cas9 genome editing of human induced pluripotent stem cells (iPSCs) to knock in APOL1 high-risk G1 variants at the native genomic locus. iPSCs were differentiated into kidney organoids, treated with vehicle, IFN-γ, or the combination of IFN-γ and tunicamycin, and analyzed with scRNA-seq to profile cell-specific changes in differential gene expression patterns, compared to isogenic G0 controls.ResultsBoth G0 and G1 iPSCs differentiated into kidney organoids containing nephron-like structures with glomerular epithelial cells, proximal tubules, distal tubules, and endothelial cells. Organoids expressed detectable APOL1 only after exposure to IFN-γ. scRNA-seq revealed cell type-specific differences in G1 organoid response to APOL1 induction. Additional stress of tunicamycin exposure led to increased glomerular epithelial cell dedifferentiation in G1 organoids.ConclusionsSingle-cell transcriptomic profiling of human genome-edited kidney organoids expressing APOL1 risk variants provides a novel platform for studying the pathophysiology of APOL1-mediated kidney disease.SIGNIFICANCE STATEMENTGaps persist in our mechanistic understanding of APOL1-mediated kidney disease. The authors apply genome-edited human kidney organoids, combined with single-cell transcriptomics, to profile APOL1 risk variants at the native genomic locus in different cell types. This approach captures interferon-mediated induction of APOL1 gene expression and reveals cellular dedifferentiation after a secondary insult of endoplasmic reticulum stress. This system provides a human cellular platform to interrogate complex mechanisms and human-specific regulators underlying APOL1-mediated kidney disease.


Kidney360 ◽  
2020 ◽  
Vol 1 (3) ◽  
pp. 203-215
Author(s):  
Esther Liu ◽  
Behram Radmanesh ◽  
Byungha H. Chung ◽  
Michael D. Donnan ◽  
Dan Yi ◽  
...  

BackgroundDNA variants in APOL1 associate with kidney disease, but the pathophysiologic mechanisms remain incompletely understood. Model organisms lack the APOL1 gene, limiting the degree to which disease states can be recapitulated. Here we present single-cell RNA sequencing (scRNA-seq) of genome-edited human kidney organoids as a platform for profiling effects of APOL1 risk variants in diverse nephron cell types.MethodsWe performed footprint-free CRISPR-Cas9 genome editing of human induced pluripotent stem cells (iPSCs) to knock in APOL1 high-risk G1 variants at the native genomic locus. iPSCs were differentiated into kidney organoids, treated with vehicle, IFN-γ, or the combination of IFN-γ and tunicamycin, and analyzed with scRNA-seq to profile cell-specific changes in differential gene expression patterns, compared with isogenic G0 controls.ResultsBoth G0 and G1 iPSCs differentiated into kidney organoids containing nephron-like structures with glomerular epithelial cells, proximal tubules, distal tubules, and endothelial cells. Organoids expressed detectable APOL1 only after exposure to IFN-γ. scRNA-seq revealed cell type–specific differences in G1 organoid response to APOL1 induction. Additional stress of tunicamycin exposure led to increased glomerular epithelial cell dedifferentiation in G1 organoids.ConclusionsSingle-cell transcriptomic profiling of human genome-edited kidney organoids expressing APOL1 risk variants provides a novel platform for studying the pathophysiology of APOL1-mediated kidney disease.


2021 ◽  
Vol 22 (14) ◽  
pp. 7642
Author(s):  
Zoran V. Popovic ◽  
Felix Bestvater ◽  
Damir Krunic ◽  
Bernhard K. Krämer ◽  
Raoul Bergner ◽  
...  

The CD73 pathway is an important anti-inflammatory mechanism in various disease settings. Observations in mouse models suggested that CD73 might have a protective role in kidney damage; however, no direct evidence of its role in human kidney disease has been described to date. Here, we hypothesized that podocyte injury in human kidney diseases alters CD73 expression that may facilitate the diagnosis of podocytopathies. We assessed the expression of CD73 and one of its functionally important targets, the C-C chemokine receptor type 2 (CCR2), in podocytes from kidney biopsies of 39 patients with podocytopathy (including focal segmental glomerulosclerosis (FSGS), minimal change disease (MCD), membranous glomerulonephritis (MGN) and amyloidosis) and a control group. Podocyte CD73 expression in each of the disease groups was significantly increased in comparison to controls (p < 0.001–p < 0.0001). Moreover, there was a marked negative correlation between CD73 and CCR2 expression, as confirmed by immunohistochemistry and immunofluorescence (Pearson r = −0.5068, p = 0.0031; Pearson r = −0.4705, p = 0.0313, respectively), thus suggesting a protective role of CD73 in kidney injury. Finally, we identify CD73 as a novel potential diagnostic marker of human podocytopathies, particularly of MCD that has been notorious for the lack of pathological features recognizable by light microscopy and immunohistochemistry.


2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yuting Guan ◽  
Xiujie Liang ◽  
Ziyuan Ma ◽  
Hailong Hu ◽  
Hongbo Liu ◽  
...  

AbstractGenome-wide association studies (GWAS) have identified loci for kidney disease, but the causal variants, genes, and pathways remain unknown. Here we identify two kidney disease genes Dipeptidase 1 (DPEP1) and Charged Multivesicular Body Protein 1 A (CHMP1A) via the triangulation of kidney function GWAS, human kidney expression, and methylation quantitative trait loci. Using single-cell chromatin accessibility and genome editing, we fine map the region that controls the expression of both genes. Mouse genetic models demonstrate the causal roles of both genes in kidney disease. Cellular studies indicate that both Dpep1 and Chmp1a are important regulators of a single pathway, ferroptosis and lead to kidney disease development via altering cellular iron trafficking.


2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
Sandrine Lemoine ◽  
Alireza Akbari ◽  
Taylor Marcus ◽  
Christopher McIntyre

Abstract Background and Aims Maintenance of a cortico-medullary concentration gradient (CMG) required for urine concentration, is one of most important tubular function. However, we are lacking of functional tubular parameters to explore this function. The only tool available to assess it currently, is urinary osmolarity that is an indirect and nonspecific maker of CMG. In this study, we explore the ability of 23NaMRI in measuring 1) the dynamics of CMG for the first time compared to urinary osmolarity after a water load 2) the CMG in kidney disease. Method We conducted an exploratory pilot study for 10 healthy controls with water load then 5 cardiorenal patients with kidney disease. 1) Healthy controls were asked to be fasting since midnight. Urines sample were collected to measure fasting osmolarity and a first MRIscan were performed to acquire baseline anatomical and sodium images. Once the baseline was completed, healthy participants were asked to ingest water (15 mL/kg) within 15 minutes. Four subsequent sodium pictures were acquired an hour after water ingestion. Urine samples were obtained after each sodium acquisition every 15 min during one hours. 2) Cardiorenal patients underwent an MRI scan, provided a spot urine sample and have blood work collected. All MR experiments were carried out on a GE MR750 3T (GE Healthcare, WI). A custom-built two-loop (18cm in diameter) butterfly radiofrequency surface coil tuned for 23Na frequency (33.786 MHz) was used to acquire renal 23Na images. Results Mean age of the 10 healthy controls was 41.8 ± 15.3 years, mean body mass index (BMI) was 24.3 ± 3.8 kg/m2. Mean water intake was 1092 ± 233 mL, total water excreted was 1250 ± 301 mL . Mean age of the 5 cardiorenal patients was 76.6 ± 12.2 years, mean BMI was 28.1 ± 6.9 kg/m2. eGFR was 54 ± 37 mL/min/1.73m2. Urinary osmolarity was 498 ± 145 mosm/L and medulla/cortex ratio was 1.35 ± 0.11. Sodium imaging was successfully acquired in all volunteers. In the morning fasting, medulla/cortex ratio was 1.55 ± 0.11 regarding to a urinary osmolarity to 814 ± 121 mosm/L. Mean ± SD fasting urinary osmolarity dropped significantly to 73 ± 14 mosm/L for maximal dilution, p=0.001. Mean medulla/cortex ratio dropped significantly to 1.31 ± 0.09 mosm/L for maximal dilution, p=0.002. Figure 1 displays changes of 23NaMRI pictures before (A) then 1h (B), 1H15 (C), 1h30 (D) and 1h45 (E) after a water load. Urinary osmolarity and medulla/cortex ratio are significantly correlated, r=0.54, p=0.0001. We measured corticomedullary gradient in cardiorenal patient with different level of eGFR to show the ability and feasibility to measure this gradient in pathological settings. We were able to measure medulla/cortex ratio in patients with CKD with a mean SNR of 20.45 ± 9.45. Conclusion We explored CMG dynamically every 15 min and we were able to discriminate significant changes after a water load. We were also able to provide efficient 23NaMRI pictures in cardiorenal patients with kidney disease. CMG exploration would provide a relevant assessment of tubular dysfunction independently of glomerular alteration and thus could be of prognostic value.


2021 ◽  
Author(s):  
Jin Wook Hwang ◽  
Christophe Desterke ◽  
Julien Loisel-Duwattez ◽  
Frank Griscelli ◽  
Annelise Bennaceur-Griscelli ◽  
...  

AbstractBackgroundIn mammalians, hematopoietic stem cells (HSC) arise in the dorsal aorta from the hemogenic endothelium, followed by their migration to fetal liver and to bone marrow. In zebrafish, kidney is the site of primary hematopoiesis. In humans, the presence of HSC in the fetal or adult kidney has not been established.MethodsWe analyzed the presence of HSC markers in human fetal kidneys by analysis of single-cell datasets. We then analyzed in kidney organoids derived from iPSC, the presence of hematopoietic markers using transcriptome analyses.Results12 clusters were identified of stromal, endothelial, and nephron cell type-specific markers in the two fetal stage (17 weeks) kidney datasets. Among these, expression of hematopoietic cells in Cluster 9 showed expression of primitive markers. Moreover, whole transcriptome analysis of our iPSC-derived kidney organoids revealed induction of the primitive hematopoietic transcription factor RUNX1 as found in the human fetal kidney cortex.ConclusionsThese finding support the presence of cells expressing HSC transcriptome in human kidney. The mechanisms of the appearance of the cells with the same transcriptional features during iPSC-derived kidney organoid generation requires further investigation.


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