Dataset Information

Profiling APOL1 Nephropathy Risk Variants in Genome-Edited Kidney Organoids with Single-Cell Transcriptomics.

ABSTRACT:

Background

DNA 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.

Methods

We 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.

Results

Both 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.

Conclusions

Single-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.

SUBMITTER: Liu E

PROVIDER: S-EPMC7351353 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Publications

Profiling <i>APOL1</i> Nephropathy Risk Variants in Genome-Edited Kidney Organoids with Single-Cell Transcriptomics.

Liu Esther E Radmanesh Behram B Chung Byungha H BH Donnan Michael D MD Yi Dan D Dadi Amal A Smith Kelly D KD Himmelfarb Jonathan J Li Mingyao M Freedman Benjamin S BS Lin Jennie J

Kidney360 20200301 3

<h4>Background</h4>DNA variants in <i>APOL1</i> associate with kidney disease, but the pathophysiologic mechanisms remain incompletely understood. Model organisms lack the <i>APOL1</i> 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 <i>APOL1</i> risk variants in diverse nephron cell types.<h4>Methods</h4>We performed footprint-free CRISPR-C ...[more]

PMID: 32656538

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Project description:Background and objectivesThe APOL1 risk variants (G1 and G2) are associated with kidney disease among Black adults, but the clinical presentation is heterogeneous. In mouse models and cell systems, increased gene expression of G1 and G2 confers cytotoxicity. How APOL1 risk variants relate to the circulating proteome warrants further investigation.Design, setting, participants, & measurementsAmong 461 African American Study of Kidney Disease and Hypertension (AASK) participants (mean age: 54 years; 41% women; mean GFR: 46 ml/min per 1.73 m2), we evaluated associations of APOL1 risk variants with 6790 serum proteins (measured via SOMAscan) using linear regression models. Covariates included age, sex, percentage of European ancestry, and protein principal components 1-5. Associated proteins were then evaluated as mediators of APOL1-associated risk for kidney failure. Findings were replicated among 875 Atherosclerosis Risk in Communities (ARIC) study Black participants (mean age: 75 years; 66% women; mean eGFR: 67 ml/min per 1.73 m2).ResultsIn the AASK study, having two (versus zero or one) APOL1 risk alleles was associated with lower serum levels of APOL1 (P=3.11E-13; P=3.12E-06 [two aptamers]), APOL2 (P=1.45E-10), CLSTN2 (P=2.66E-06), MMP-2 (P=2.96E-06), SPOCK2 (P=2.57E-05), and TIMP-2 (P=2.98E-05) proteins. In the ARIC study, APOL1 risk alleles were associated with APOL1 (P=1.28E-11); MMP-2 (P=0.004) and TIMP-2 (P=0.007) were associated only in an additive model, and APOL2 was not available. APOL1 high-risk status was associated with a 1.6-fold greater risk of kidney failure in the AASK study; none of the identified proteins mediated this association. APOL1 protein levels were not associated with kidney failure in either cohort.ConclusionsAPOL1 risk variants were strongly associated with lower circulating levels of APOL1 and other proteins, but none mediated the APOL1-associated risk for kidney failure. APOL1 protein level was also not associated with kidney failure.