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2.
J Comp Neurol ; 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34525221

RESUMO

Increasing interest in studies of prenatal human brain development, particularly using new single-cell genomics and anatomical technologies to create cell atlases, creates a strong need for accurate and detailed anatomical reference atlases. In this study, we present two cellular-resolution digital anatomical atlases for prenatal human brain at post-conceptional weeks (PCW) 15 and 21. Both atlases were annotated on sequential Nissl-stained sections covering brain-wide structures on the basis of combined analysis of cytoarchitecture, acetylcholinesterase staining and an extensive marker gene expression dataset. This high information content dataset allowed reliable and accurate demarcation of developing cortical and subcortical structures and their subdivisions. Furthermore, using the anatomical atlases as a guide, spatial expression of 37 and 5 genes from the brains respectively at PCW 15 and 21 was annotated, illustrating reliable marker genes for many developing brain structures. Finally, the present study uncovered several novel developmental features, such as the lack of an outer subventricular zone in the hippocampal formation and entorhinal cortex, and the apparent extension of both cortical (excitatory) and subcortical (inhibitory) progenitors into the prenatal olfactory bulb. These comprehensive atlases provide useful tools for visualization, segmentation, targeting, imaging and interpretation of brain structures of prenatal human brain, and for guiding and interpreting the next generation of cell census and connectome studies. This article is protected by copyright. All rights reserved.

3.
Acta Neuropathol ; 142(4): 761-776, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34347142

RESUMO

Dandy-Walker malformation (DWM) and Cerebellar vermis hypoplasia (CVH) are commonly recognized human cerebellar malformations diagnosed following ultrasound and antenatal or postnatal MRI. Specific radiological criteria are used to distinguish them, yet little is known about their differential developmental disease mechanisms. We acquired prenatal cases diagnosed as DWM and CVH and studied cerebellar morphobiometry followed by histological and immunohistochemical analyses. This was supplemented by laser capture microdissection and RNA-sequencing of the cerebellar rhombic lip, a transient progenitor zone, to assess the altered transcriptome of DWM vs control samples. Our radiological findings confirm that the cases studied fall within the accepted biometric range of DWM. Our histopathological analysis points to reduced foliation and inferior vermian hypoplasia as common features in all examined DWM cases. We also find that the rhombic lip, a dorsal stem cell zone that drives the growth and maintenance of the posterior vermis is specifically disrupted in DWM, with reduced proliferation and self-renewal of the progenitor pool, and altered vasculature, all confirmed by transcriptomics analysis. We propose a unified model for the developmental pathogenesis of DWM. We hypothesize that rhombic lip development is disrupted through either aberrant vascularization and/or direct insult which causes reduced proliferation and failed expansion of the rhombic lip progenitor pool leading to disproportionate hypoplasia and dysplasia of the inferior vermis. Timing of insult to the developing rhombic lip (before or after 14 PCW) dictates the extent of hypoplasia and distinguishes DWM from CVH.

4.
JAMA Neurol ; 78(10): 1236-1248, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34459874

RESUMO

Importance: Juvenile amyotrophic lateral sclerosis (ALS) is a rare form of ALS characterized by age of symptom onset less than 25 years and a variable presentation. Objective: To identify the genetic variants associated with juvenile ALS. Design, Setting, and Participants: In this multicenter family-based genetic study, trio whole-exome sequencing was performed to identify the disease-associated gene in a case series of unrelated patients diagnosed with juvenile ALS and severe growth retardation. The patients and their family members were enrolled at academic hospitals and a government research facility between March 1, 2016, and March 13, 2020, and were observed until October 1, 2020. Whole-exome sequencing was also performed in a series of patients with juvenile ALS. A total of 66 patients with juvenile ALS and 6258 adult patients with ALS participated in the study. Patients were selected for the study based on their diagnosis, and all eligible participants were enrolled in the study. None of the participants had a family history of neurological disorders, suggesting de novo variants as the underlying genetic mechanism. Main Outcomes and Measures: De novo variants present only in the index case and not in unaffected family members. Results: Trio whole-exome sequencing was performed in 3 patients diagnosed with juvenile ALS and their parents. An additional 63 patients with juvenile ALS and 6258 adult patients with ALS were subsequently screened for variants in the SPTLC1 gene. De novo variants in SPTLC1 (p.Ala20Ser in 2 patients and p.Ser331Tyr in 1 patient) were identified in 3 unrelated patients diagnosed with juvenile ALS and failure to thrive. A fourth variant (p.Leu39del) was identified in a patient with juvenile ALS where parental DNA was unavailable. Variants in this gene have been previously shown to be associated with autosomal-dominant hereditary sensory autonomic neuropathy, type 1A, by disrupting an essential enzyme complex in the sphingolipid synthesis pathway. Conclusions and Relevance: These data broaden the phenotype associated with SPTLC1 and suggest that patients presenting with juvenile ALS should be screened for variants in this gene.

5.
Am J Hum Genet ; 108(8): 1436-1449, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34216551

RESUMO

Despite widespread clinical genetic testing, many individuals with suspected genetic conditions lack a precise diagnosis, limiting their opportunity to take advantage of state-of-the-art treatments. In some cases, testing reveals difficult-to-evaluate structural differences, candidate variants that do not fully explain the phenotype, single pathogenic variants in recessive disorders, or no variants in genes of interest. Thus, there is a need for better tools to identify a precise genetic diagnosis in individuals when conventional testing approaches have been exhausted. We performed targeted long-read sequencing (T-LRS) using adaptive sampling on the Oxford Nanopore platform on 40 individuals, 10 of whom lacked a complete molecular diagnosis. We computationally targeted up to 151 Mbp of sequence per individual and searched for pathogenic substitutions, structural variants, and methylation differences using a single data source. We detected all genomic aberrations-including single-nucleotide variants, copy number changes, repeat expansions, and methylation differences-identified by prior clinical testing. In 8/8 individuals with complex structural rearrangements, T-LRS enabled more precise resolution of the mutation, leading to changes in clinical management in one case. In ten individuals with suspected Mendelian conditions lacking a precise genetic diagnosis, T-LRS identified pathogenic or likely pathogenic variants in six and variants of uncertain significance in two others. T-LRS accurately identifies pathogenic structural variants, resolves complex rearrangements, and identifies Mendelian variants not detected by other technologies. T-LRS represents an efficient and cost-effective strategy to evaluate high-priority genes and regions or complex clinical testing results.


Assuntos
Aberrações Cromossômicas , Análise Citogenética/métodos , Doenças Genéticas Inatas/diagnóstico , Doenças Genéticas Inatas/genética , Predisposição Genética para Doença , Genoma Humano , Mutação , Variações do Número de Cópias de DNA , Feminino , Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Cariotipagem , Masculino , Análise de Sequência de DNA
6.
Nat Neurosci ; 24(8): 1163-1175, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34140698

RESUMO

The human neonatal cerebellum is one-fourth of its adult size yet contains the blueprint required to integrate environmental cues with developing motor, cognitive and emotional skills into adulthood. Although mature cerebellar neuroanatomy is well studied, understanding of its developmental origins is limited. In this study, we systematically mapped the molecular, cellular and spatial composition of human fetal cerebellum by combining laser capture microscopy and SPLiT-seq single-nucleus transcriptomics. We profiled functionally distinct regions and gene expression dynamics within cell types and across development. The resulting cell atlas demonstrates that the molecular organization of the cerebellar anlage recapitulates cytoarchitecturally distinct regions and developmentally transient cell types that are distinct from the mouse cerebellum. By mapping genes dominant for pediatric and adult neurological disorders onto our dataset, we identify relevant cell types underlying disease mechanisms. These data provide a resource for probing the cellular basis of human cerebellar development and disease.


Assuntos
Cerebelo/embriologia , Neurogênese , Feto , Humanos , Microdissecção e Captura a Laser , Análise de Célula Única , Transcriptoma
7.
Cell ; 184(12): 3281-3298.e22, 2021 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-34019796

RESUMO

Organs are composed of diverse cell types that traverse transient states during organogenesis. To interrogate this diversity during human development, we generate a single-cell transcriptome atlas from multiple developing endodermal organs of the respiratory and gastrointestinal tract. We illuminate cell states, transcription factors, and organ-specific epithelial stem cell and mesenchyme interactions across lineages. We implement the atlas as a high-dimensional search space to benchmark human pluripotent stem cell (hPSC)-derived intestinal organoids (HIOs) under multiple culture conditions. We show that HIOs recapitulate reference cell states and use HIOs to reconstruct the molecular dynamics of intestinal epithelium and mesenchyme emergence. We show that the mesenchyme-derived niche cue NRG1 enhances intestinal stem cell maturation in vitro and that the homeobox transcription factor CDX2 is required for regionalization of intestinal epithelium and mesenchyme in humans. This work combines cell atlases and organoid technologies to understand how human organ development is orchestrated.

8.
Am J Hum Genet ; 108(1): 8-15, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33417889

RESUMO

The delineation of disease entities is complex, yet recent advances in the molecular characterization of diseases provide opportunities to designate diseases in a biologically valid manner. Here, we have formalized an approach to the delineation of Mendelian genetic disorders that encompasses two distinct but inter-related concepts: (1) the gene that is mutated and (2) the phenotypic descriptor, preferably a recognizably distinct phenotype. We assert that only by a combinatorial or dyadic approach taking both of these attributes into account can a unitary, distinct genetic disorder be designated. We propose that all Mendelian disorders should be designated as "GENE-related phenotype descriptor" (e.g., "CFTR-related cystic fibrosis"). This approach to delineating and naming disorders reconciles the complexity of gene-to-phenotype relationships in a simple and clear manner yet communicates the complexity and nuance of these relationships.


Assuntos
Doenças Genéticas Inatas/diagnóstico , Doenças Genéticas Inatas/genética , Genômica/métodos , Fibrose Cística/diagnóstico , Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Genótipo , Humanos , Mutação/genética , Fenótipo
9.
Neuropediatrics ; 52(3): 186-191, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33445191

RESUMO

We describe two novel missense variants in CACNA1A segregating in a family with variable severity of ataxia/oculomotor dysfunction, neurobehavioral impairments, and epilepsy. The most severe outcome occurred in a compound heterozygous proband, which could represent variable expression of the paternal allele or biallelic modulation of calcium channel function. Acetazolamide and lamotrigine were effective for seizure control.

10.
Cell Stem Cell ; 28(3): 568-580.e4, 2021 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-33278341

RESUMO

The human intestinal stem cell niche supports self-renewal and epithelial function, but little is known about its development. We used single-cell mRNA sequencing with in situ validation approaches to interrogate human intestinal development from 7-21 weeks post conception, assigning molecular identities and spatial locations to cells and factors that comprise the niche. Smooth muscle cells of the muscularis mucosa, in close proximity to proliferative crypts, are a source of WNT and RSPONDIN ligands, whereas EGF is expressed far from crypts in the villus epithelium. Instead, an PDGFRAHI/F3HI/DLL1HI mesenchymal population lines the crypt-villus axis and is the source of the epidermal growth factor (EGF) family member NEUREGULIN1 (NRG1). In developing intestine enteroid cultures, NRG1, but not EGF, permitted increased cellular diversity via differentiation of secretory lineages. This work highlights the complexities of intestinal EGF/ERBB signaling and delineates key niche cells and signals of the developing intestine.


Assuntos
Intestinos , Nicho de Células-Tronco , Diferenciação Celular , Humanos , Mucosa Intestinal , Células-Tronco
11.
Am J Med Genet A ; 2020 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-33205886

RESUMO

Twins have an increased risk for congenital malformations and disruptions, including defects in brain morphogenesis. We analyzed data on brain imaging, zygosity, sex, and fetal demise in 56 proband twins and 7 less affected co-twins with abnormal brain imaging and compared them to population-based data and to a literature series. We separated our series into malformations of cortical development (MCD, N = 39), cerebellar malformations without MCD (N = 13), and brain disruptions (N = 11). The MCD group included 37/39 (95%) with polymicrogyria (PMG), 8/39 (21%) with pia-ependymal clefts (schizencephaly), and 15/39 (38%) with periventricular nodular heterotopia (PNH) including 2 with PNH but not PMG. Cerebellar malformations were found in 19 individuals including 13 with a cerebellar malformation only and another 6 with cerebellar malformation and MCD. The pattern varied from diffuse cerebellar hypoplasia to classic Dandy-Walker malformation. Brain disruptions were seen in 11 individuals with hydranencephaly, porencephaly, or white matter loss without cysts. Our series included an expected statistically significant excess of monozygotic (MZ) twin pairs (22/41 MZ, 54%) compared to population data (482/1448 MZ, 33.3%; p = .0110), and an unexpected statistically significant excess of dizygotic (DZ) twins (19/41, 46%) compared to the literature cohort (1/46 DZ, 2%; p < .0001. Recurrent association with twin-twin transfusion syndrome, intrauterine growth retardation, and other prenatal factors support disruption of vascular perfusion as the most likely unifying cause.

12.
Science ; 370(6518)2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-33184180

RESUMO

The chromatin landscape underlying the specification of human cell types is of fundamental interest. We generated human cell atlases of chromatin accessibility and gene expression in fetal tissues. For chromatin accessibility, we devised a three-level combinatorial indexing assay and applied it to 53 samples representing 15 organs, profiling ~800,000 single cells. We leveraged cell types defined by gene expression to annotate these data and cataloged hundreds of thousands of candidate regulatory elements that exhibit cell type-specific chromatin accessibility. We investigated the properties of lineage-specific transcription factors (such as POU2F1 in neurons), organ-specific specializations of broadly distributed cell types (such as blood and endothelial), and cell type-specific enrichments of complex trait heritability. These data represent a rich resource for the exploration of in vivo human gene regulation in diverse tissues and cell types.


Assuntos
Cromatina/metabolismo , Feto/citologia , Feto/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Análise de Célula Única , Atlas como Assunto , Humanos , Neurônios/metabolismo , Fatores de Transcrição/metabolismo
13.
Science ; 370(6518)2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-33184181

RESUMO

The gene expression program underlying the specification of human cell types is of fundamental interest. We generated human cell atlases of gene expression and chromatin accessibility in fetal tissues. For gene expression, we applied three-level combinatorial indexing to >110 samples representing 15 organs, ultimately profiling ~4 million single cells. We leveraged the literature and other atlases to identify and annotate hundreds of cell types and subtypes, both within and across tissues. Our analyses focused on organ-specific specializations of broadly distributed cell types (such as blood, endothelial, and epithelial), sites of fetal erythropoiesis (which notably included the adrenal gland), and integration with mouse developmental atlases (such as conserved specification of blood cells). These data represent a rich resource for the exploration of in vivo human gene expression in diverse tissues and cell types.


Assuntos
Cromatina/metabolismo , Feto/citologia , Feto/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Análise de Célula Única , Atlas como Assunto , Humanos , Neurônios/metabolismo , Fatores de Transcrição/metabolismo
15.
Dev Cell ; 54(4): 516-528.e7, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32841595

RESUMO

Human pluripotent stem cell (hPSC)-derived intestinal organoids (HIOs) lack some cellular populations found in the native organ, including vasculature. Using single-cell RNA sequencing (scRNA-seq), we have identified a population of endothelial cells (ECs) present early in HIO differentiation that declines over time in culture. Here, we developed a method to expand and maintain this endogenous population of ECs within HIOs (vHIOs). Given that ECs possess organ-specific gene expression, morphology, and function, we used bulk RNA-seq and scRNA-seq to interrogate the developing human intestine, lung, and kidney in order to identify organ-enriched EC gene signatures. By comparing these gene signatures and validated markers to HIO ECs, we find that HIO ECs grown in vitro share the highest similarity with native intestinal ECs relative to kidney and lung. Together, these data demonstrate that HIOs can co-differentiate a native EC population that is properly patterned with an intestine-specific EC transcriptional signature in vitro.


Assuntos
Células Endoteliais/metabolismo , Mucosa Intestinal/crescimento & desenvolvimento , Intestinos/crescimento & desenvolvimento , Especificidade de Órgãos/genética , Diferenciação Celular/genética , Linhagem Celular , Regulação da Expressão Gênica/genética , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Mucosa Intestinal/metabolismo , Rim/crescimento & desenvolvimento , Rim/metabolismo , Pulmão/crescimento & desenvolvimento , Pulmão/metabolismo , Organoides/crescimento & desenvolvimento , Organoides/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , RNA-Seq
16.
Am J Med Genet A ; 182(9): 2037-2048, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32710489

RESUMO

The SET domain containing 2, histone lysine methyltransferase encoded by SETD2 is a dual-function methyltransferase for histones and microtubules and plays an important role for transcriptional regulation, genomic stability, and cytoskeletal functions. Specifically, SETD2 is associated with trimethylation of histone H3 at lysine 36 (H3K36me3) and methylation of α-tubulin at lysine 40. Heterozygous loss of function and missense variants have previously been described with Luscan-Lumish syndrome (LLS), which is characterized by overgrowth, neurodevelopmental features, and absence of overt congenital anomalies. We have identified 15 individuals with de novo variants in codon 1740 of SETD2 whose features differ from those with LLS. Group 1 consists of 12 individuals with heterozygous variant c.5218C>T p.(Arg1740Trp) and Group 2 consists of 3 individuals with heterozygous variant c.5219G>A p.(Arg1740Gln). The phenotype of Group 1 includes microcephaly, profound intellectual disability, congenital anomalies affecting several organ systems, and similar facial features. Individuals in Group 2 had moderate to severe intellectual disability, low normal head circumference, and absence of additional major congenital anomalies. While LLS is likely due to loss of function of SETD2, the clinical features seen in individuals with variants affecting codon 1740 are more severe suggesting an alternative mechanism, such as gain of function, effects on epigenetic regulation, or posttranslational modification of the cytoskeleton. Our report is a prime example of different mutations in the same gene causing diverging phenotypes and the features observed in Group 1 suggest a new clinically recognizable syndrome uniquely associated with the heterozygous variant c.5218C>T p.(Arg1740Trp) in SETD2.


Assuntos
Predisposição Genética para Doença , Histona-Lisina N-Metiltransferase/genética , Deficiência Intelectual/genética , Transtornos do Neurodesenvolvimento/genética , Tubulina (Proteína)/genética , Criança , Pré-Escolar , Códon/genética , Epigênese Genética/genética , Feminino , Estudos de Associação Genética , Humanos , Lactente , Deficiência Intelectual/patologia , Mutação com Perda de Função/genética , Masculino , Mutação de Sentido Incorreto , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/patologia , Transtornos do Neurodesenvolvimento/fisiopatologia
17.
Cell Rep ; 31(1): 107489, 2020 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-32268104

RESUMO

Gene expression levels vary across developmental stage, cell type, and region in the brain. Genomic variants also contribute to the variation in expression, and some neuropsychiatric disorder loci may exert their effects through this mechanism. To investigate these relationships, we present BrainVar, a unique resource of paired whole-genome and bulk tissue RNA sequencing from the dorsolateral prefrontal cortex of 176 individuals across prenatal and postnatal development. Here we identify common variants that alter gene expression (expression quantitative trait loci [eQTLs]) constantly across development or predominantly during prenatal or postnatal stages. Both "constant" and "temporal-predominant" eQTLs are enriched for loci associated with neuropsychiatric traits and disorders and colocalize with specific variants. Expression levels of more than 12,000 genes rise or fall in a concerted late-fetal transition, with the transitional genes enriched for cell-type-specific genes and neuropsychiatric risk loci, underscoring the importance of cataloging developmental trajectories in understanding cortical physiology and pathology.


Assuntos
Encéfalo/embriologia , Biologia Computacional/métodos , Córtex Pré-Frontal/metabolismo , Sequência de Bases/genética , Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Bases de Dados Genéticas , Predisposição Genética para Doença/genética , Variação Genética/genética , Estudo de Associação Genômica Ampla/métodos , Genômica/métodos , Humanos , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Análise de Sequência de RNA/métodos , Transcriptoma/genética , Sequenciamento Completo do Exoma/métodos , Sequenciamento Completo do Genoma/métodos
18.
Bio Protoc ; 10(1)2020 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-32190713

RESUMO

Precise and reproducible isolation of desired cell types or layers from heterogeneous tissues is crucial to analyze specific gene profiles and molecular interactions in vivo. Forebrain is the core site of higher functions, like cognition and memory consolidation. It is composed of heterogeneous and distinct cell types, interconnected to form functional neural circuits. Any alteration in the development or function often leads to brain disorders with profound consequences. Thus, precise molecular understanding of forebrain development in normal and diseased scenarios is important. For quantitative studies, most traditional analytical methods require pooling of large cell populations, that results in loss of in vivo tissue integrity and of spatial, molecular and cellular resolution. Laser capture microdissection (LCM) is a fast and extremely precise method of obtaining uncontaminated, homogeneous sets of specific cell types and layers. Our current procedure involves cryo-sectioning and laser microdissection of fresh-frozen mouse forebrains, that are genetically modified and treated with small-molecule therapeutics. Using LCM, specific regions of interest, such as neural layers, cells from adjacent yet distinct subregions within a tissue layer, are obtained under RNase-free conditions. These small cellular cohorts are further used for downstream, high-throughput genomic or transcriptomic assays. Here, we have introduced break-points at multiple stages throughout our protocol. This makes our method simpler and more user-friendly to follow, without compromising on the quality. The current protocol can easily be adapted for different brain regions, as well as for other model organisms/human tissue.

19.
Cell Rep ; 30(5): 1644-1659.e4, 2020 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-32023475

RESUMO

To study the development of the human retina, we use single-cell RNA sequencing (RNA-seq) at key fetal stages and follow the development of the major cell types as well as populations of transitional cells. We also analyze stem cell (hPSC)-derived retinal organoids; although organoids have a very similar cellular composition at equivalent ages as the fetal retina, there are some differences in gene expression of particular cell types. Moreover, the inner retinal lamination is disrupted at more advanced stages of organoids compared with fetal retina. To determine whether the disorganization in the inner retina is due to the culture conditions, we analyze retinal development in fetal retina maintained under similar conditions. These retinospheres develop for at least 6 months, displaying better inner retinal lamination than retinal organoids. Our single-cell RNA sequencing (scRNA-seq) comparisons of fetal retina, retinal organoids, and retinospheres provide a resource for developing better in vitro models for retinal disease.


Assuntos
Feto/citologia , Organoides/citologia , Células-Tronco Pluripotentes/citologia , Retina/citologia , Análise de Célula Única , Transcriptoma/genética , Células Cultivadas , Humanos , Mitose/genética , Esferoides Celulares/citologia , Fatores de Tempo
20.
Dev Cell ; 53(1): 117-128.e6, 2020 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-32109386

RESUMO

Bud tip progenitor cells give rise to all murine lung epithelial lineages and have been described in the developing human lung; however, the mechanisms controlling human bud tip differentiation into specific lineages are unclear. Here, we used homogeneous human bud tip organoid cultures and identified SMAD signaling as a key regulator of the bud tip-to-airway transition. SMAD induction led to the differentiation of airway-like organoids possessing functional basal cells capable of clonal expansion and multilineage differentiation. To benchmark in vitro-derived organoids, we developed a single-cell mRNA sequencing atlas of the human lung from 11.5 to 21 weeks of development, which revealed high degrees of similarity between the in vitro-derived and in vivo airway. Together, this work sheds light on human airway differentiation in vitro and provides a single-cell atlas of the developing human lung.


Assuntos
Diferenciação Celular/fisiologia , Células Epiteliais/citologia , Organoides/citologia , Células-Tronco Pluripotentes/citologia , Humanos , Pulmão/citologia , Engenharia Tecidual/métodos
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