Diploid hepatocytes drive physiological liver renewal in adult humans

Heinke, Paula; Rost, Fabian; Rode, Julian; Trus, Palina; Simonova, Irina; Lázár, Enikő; Feddema, Joshua; Welsch, Thilo; Alkass, Kanar; Salehpour, Mehran; Zimmermann, Andrea; Seehofer, Daniel; Possnert, Göran; Damm, Georg; Druid, Henrik; Brusch, Lutz; Bergmann, Olaf

doi:10.1016/j.cels.2022.05.001

Cited by 35 publications

(31 citation statements)

References 77 publications

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“…11 One interpretation of recent studies on the rate of physiological cell replacement in the mature liver could be that episomal gene augmentation might require re-administration to maintain efficacy. 12 As a more permanent alternative therapy, we are exploring genome editing. In a previous preclinical study, we generated G6pc-R83C mice carrying the G6PC-p.R83C variant in exon 2 of the G6pc gene and showed that the G6pc-R83C mice exhibit the symptoms of impaired glucose homeostasis mimicking human GSD-Ia.…”

Section: Discussionmentioning

confidence: 99%

“…8 Currently, there is insufficient clinical data to understand if multi-decade episomal transgene expression can be maintained in the human liver at a therapeutic level, 11 especially considering a recent report on the rate of physiological liver cell replacement. 12 Extended durability is important for pediatric disease. We therefore explored alternative genetic technologies for GSD-Ia therapy, such as CRISPR/Cas9-based gene editing.…”

Section: Introductionmentioning

confidence: 99%

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CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

Samanta

George

Arnaoutova

et al. 2023

J of Inher Metab Disea

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Glycogen storage disease type‐Ia (GSD‐Ia), characterized by impaired blood glucose homeostasis, is caused by a deficiency in glucose‐6‐phosphatase‐α (G6Pase‐α or G6PC). Using the G6pc‐R83C mouse model of GSD‐Ia, we explored a CRISPR/Cas9‐based double‐strand DNA oligonucleotide (dsODN) insertional strategy that uses the non‐homologous end joining repair mechanism to correct the pathogenic p.R83C variant in G6pc exon‐2. The strategy is based on the insertion of a short dsODN into G6pc exon‐2 to disrupt the native exon, and to introduce an additional splice acceptor site and the correcting sequence. When transcribed and spliced the edited gene would generate a wild‐type mRNA encoding the native G6Pase‐α protein. The editing reagents formulated in lipid nanoparticles (LNP) were delivered to the liver. Mice were treated either with one dose of LNP‐dsODN at age 4 weeks or with 2 doses of LNP‐dsODN at age 2 and 4 weeks. The G6pc‐R83C mice receiving successful editing expressed ~4% of normal hepatic G6Pase‐α activity, maintained glucose homeostasis, lacked hypoglycemic seizures, and displayed normalized blood metabolite profile. The outcomes are consistent with preclinical studies supporting previous gene augmentation therapy which is currently in clinical trials. This editing strategy may offer the basis for a therapeutic approach with an earlier clinical intervention than gene augmentation, with the additional benefit of a potentially permanent correction of the GSD‐Ia phenotype.This article is protected by copyright. All rights reserved.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

Samanta

George

Arnaoutova

et al. 2023

J of Inher Metab Disea

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“…36 Second, Heinke et al used retrospective radiocarbon ( 14 C) birth dating of cells to investigate physiological hepatocyte replacement in humans. 62 Hepatocyte nuclei from 29 human subjects aged 20 to 84 years were isolated by fluorescenceactivated cell sorting (FACS) into ploidy populations, and genomic 14 C concentrations were determined using accelerator mass spectrometry. Mathematical modeling was appliedincorporating historic atmosphere, memory effects, and cellcycle dynamics-to estimate hepatocyte age and renewal rates.…”

Section: Ploidy and Proliferationmentioning

confidence: 99%

The Ploidy State as a Determinant of Hepatocyte Proliferation

Wilson,

Duncan

2023

Semin Liver Dis

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The liver’s unique chromosomal variations, including polyploidy and aneuploidy, influence hepatocyte identity and function. Among the most well-studied mammalian polyploid cells, hepatocytes exhibit a dynamic interplay between diploid and polyploid states. The ploidy state is dynamic as hepatocytes move through the “ploidy conveyor,” undergoing ploidy reversal and re-polyploidization during proliferation. Both diploid and polyploid hepatocytes actively contribute to proliferation, with diploids demonstrating an enhanced proliferative capacity. This enhanced potential positions diploid hepatocytes as primary drivers of liver proliferation in multiple contexts, including homeostasis, regeneration and repopulation, compensatory proliferation following injury, and oncogenic proliferation. This review discusses the influence of ploidy variations on cellular activity. It presents a model for ploidy-associated hepatocyte proliferation, offering a deeper understanding of liver health and disease with the potential to uncover novel treatment approaches.

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“…The mammalian liver is an attractive model to study the regulation of non-canonical cell cycles, as mature hepatocytes have been reported to perform both canonical and endomitotic cell cycles throughout their lifetime. In humans, hepatocytes range in ploidy from 2N to 8N, and up to 20% are binucleated [30][31][32][33] . In a healthy liver, less than 0.01% of hepatocytes are actively cycling 34,35 .…”

Section: Introductionmentioning

confidence: 99%

Binucleated human hepatocytes arise through loss of membrane anchorage to the midbody during endomitosis

Darmasaputra

Lopes

Galli

2023

Preprint

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Many plant and animal cells transition from canonical to non-canonical cell cycles during development, resulting in the formation of polyploid cells. Two types of non-canonical cell cycles exist: endoreplication, where cells increase their DNA content without entering M phase, and endomitosis, where cells enter M phase but exit prematurely. Although endoreplication has been extensively studied in plants and insects, much less is known on the regulation of endomitosis, which is the most common mode of polyploidization in mammals. In this study, we use fetal-derived human hepatocyte organoids (Hep-Org), to investigate how human hepatocytes initiate and execute endomitosis. We find that cells in endomitosis M phase have normal mitotic timings, but lose membrane anchorage to the midbody during cytokinesis, resulting in regression of the cytokinetic furrow and formation of binucleate cells. Using immunofluorescence, we find that three cortical anchoring proteins, RacGAP1, anillin, and citron kinase (CIT-K), lose their association with the cell cortex during cytokinetic regression. Moreover, reduction of WNT activity by withdrawal of CHIR99021, a GSK3 inhibitor, from the culturing medium increases the percentage of binucleated cells in Hep-Orgs. This effect is lost in organoids with mutations in the atypical E2F proteins, E2F7 and E2F8, which have been implicated in binucleation of rodent hepatocytes. Together, our results identify how human hepatocytes inhibit cell division in endomitosis, and highlight an evolutionary recurrent mechanism to initiate non-canonical cell cycles in mammals.

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Diploid hepatocytes drive physiological liver renewal in adult humans

Cited by 35 publications

References 77 publications

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

The Ploidy State as a Determinant of Hepatocyte Proliferation

Binucleated human hepatocytes arise through loss of membrane anchorage to the midbody during endomitosis

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