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Functional characterization of ion channels expressed in kidney organoids derived from human induced pluripotent stem cells.

ABSTRACT: Kidney organoids derived from human or rodent pluripotent stem cells have glomerular structures and differentiated/polarized nephron segments. Although there is an increasing understanding of the patterns of expression of transcripts and proteins within kidney organoids, there is a paucity of data regarding functional protein expression, in particular on transporters that mediate the vectorial transport of solutes. Using cells derived from kidney organoids, we examined the functional expression of key ion channels that are expressed in distal nephron segments: the large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel, the renal outer medullary K⁺ (ROMK, Kir1.1) channel, and the epithelial Na⁺ channel (ENaC). RNA-sequencing analyses showed that genes encoding the pore-forming subunits of these transporters, and for BK_Ca channels, key accessory subunits, are expressed in kidney organoids. Expression and localization of selected ion channels was confirmed by immunofluorescence microscopy and immunoblot analysis. Electrophysiological analysis showed that BK_Ca and ROMK channels are expressed in different cell populations. These two cell populations also expressed other unidentified Ba²⁺-sensitive K⁺ channels. BK_Ca expression was confirmed at a single channel level, based on its high conductance and voltage dependence of activation. We also found a population of cells expressing amiloride-sensitive ENaC currents. In summary, our results show that human kidney organoids functionally produce key distal nephron K⁺ and Na⁺ channels.NEW & NOTEWORTHY Our results show that human kidney organoids express key K⁺ and Na⁺ channels that are expressed on the apical membranes of cells in the aldosterone-sensitive distal nephron, including the large-conductance Ca²⁺-activated K⁺ channel, renal outer medullary K⁺ channel, and epithelial Na⁺ channel.

SUBMITTER: Montalbetti N

PROVIDER: S-EPMC9529267 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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Functional characterization of ion channels expressed in kidney organoids derived from human induced pluripotent stem cells.

Montalbetti Nicolas N Przepiorski Aneta J AJ Shi Shujie S Sheng Shaohu S Baty Catherine J CJ Maggiore Joseph C JC Carattino Marcelo D MD Vanichapol Thitinee T Davidson Alan J AJ Hukriede Neil A NA Kleyman Thomas R TR

American journal of physiology. Renal physiology 20220818 4

Kidney organoids derived from human or rodent pluripotent stem cells have glomerular structures and differentiated/polarized nephron segments. Although there is an increasing understanding of the patterns of expression of transcripts and proteins within kidney organoids, there is a paucity of data regarding functional protein expression, in particular on transporters that mediate the vectorial transport of solutes. Using cells derived from kidney organoids, we examined the functional expression ...[more]

PMID: 35979965

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Project description:ImportanceThree-dimensional cerebral organoids generated from patient-derived induced pluripotent stem cells (iPSCs) may be used to interrogate cellular-molecular underpinnings of schizophrenia.ObjectiveTo determine transcriptomic profiles and functional characteristics of cerebral organoids from patients with schizophrenia using gene expression studies, complemented with investigations of mitochondrial function through measurement of real-time oxygen consumption rate, and functional studies of neuronal firing with microelectrode arrays.Design, setting, and participantsThis case-control study was conducted at Massachusetts General Hospital between 2017 and 2019. Transcriptomic profiling of iPSC-derived cerebral organoids from 8 patients with schizophrenia and 8 healthy control individuals was undertaken to identify cellular pathways that are aberrant in schizophrenia. Induced pluripotent stem cells and cerebral organoids were generated from patients who had been diagnosed as having schizophrenia and from heathy control individuals.Main outcomes and measuresTranscriptomic analysis of iPSC-derived cerebral organoids from patients with schizophrenia show differences in expression of genes involved in synaptic biology and neurodevelopment and are enriched for genes implicated in schizophrenia genome-wide association studies (GWAS).ResultsThe study included iPSC lines generated from 11 male and 5 female white participants, with a mean age of 38.8 years. RNA sequencing data from iPSC-derived cerebral organoids in schizophrenia showed differential expression of genes involved in synapses, in nervous system development, and in antigen processing. The differentially expressed genes were enriched for genes implicated in schizophrenia, with 23% of GWAS genes showing differential expression in schizophrenia and control organoids: 10 GWAS genes were upregulated in schizophrenia organoids while 15 GWAS genes were downregulated. Analysis of the gene expression profiles suggested dysregulation of genes involved in mitochondrial function and those involved in modulation of excitatory and inhibitory pathways. Studies of mitochondrial respiration showed lower basal consumption rate, adenosine triphosphate production, proton leak, and nonmitochondrial oxygen consumption in schizophrenia cerebral organoids, without any differences in the extracellular acidification rate. Microelectrode array studies of cerebral organoids showed no differences in baseline electrical activity in schizophrenia but revealed a diminished response to stimulation and depolarization.Conclusions and relevanceInvestigations of patient-derived cerebral organoids in schizophrenia revealed gene expression patterns suggesting dysregulation of a number of pathways in schizophrenia, delineated differences in mitochondrial function, and showed deficits in response to stimulation and depolarization in schizophrenia.

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Functional characterization of ion channels expressed in kidney organoids derived from human induced pluripotent stem cells.

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Functional characterization of ion channels expressed in kidney organoids derived from human induced pluripotent stem cells.

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