Thinking out of the dish: what to learn about cortical development using pluripotent stem cells
2014; Elsevier BV; Volume: 37; Issue: 6 Linguagem: Inglês
10.1016/j.tins.2014.03.005
ISSN1878-108X
AutoresJelle van den Ameele, Luca Tiberi, Pierre Vanderhaeghen, Ira Espuny-Camacho,
Tópico(s)CRISPR and Genetic Engineering
Resumo•Pluripotent stem cell-derived corticogenesis recapitulates most features of in vivo temporal and regional patterning.•Pluripotent stem cell-derived corticogenesis displays species-specific features relevant to human brain evolution.•Pluripotent stem cell modelling reveals novel insights into neurodevelopmental diseases. The development of the cerebral cortex requires the tightly coordinated generation of dozens of neuronal subtypes that will populate specific layers and areas. Recent studies have revealed how pluripotent stem cells (PSC), whether of mouse or human origin, can differentiate into a wide range of cortical neurons in vitro, which can integrate appropriately into the brain following in vivo transplantation. These models are largely artificial but recapitulate a substantial fraction of the complex temporal and regional patterning events that occur during in vivo corticogenesis. Here, we review these findings with emphasis on the new perspectives that they have brought for understanding of cortical development, evolution, and diseases. The development of the cerebral cortex requires the tightly coordinated generation of dozens of neuronal subtypes that will populate specific layers and areas. Recent studies have revealed how pluripotent stem cells (PSC), whether of mouse or human origin, can differentiate into a wide range of cortical neurons in vitro, which can integrate appropriately into the brain following in vivo transplantation. These models are largely artificial but recapitulate a substantial fraction of the complex temporal and regional patterning events that occur during in vivo corticogenesis. Here, we review these findings with emphasis on the new perspectives that they have brought for understanding of cortical development, evolution, and diseases. The cerebral cortex is among the most complex of all biological structures, and the major site of higher cognitive functions specific to our species. The mechanisms underlying its development and evolution are at the core of what makes us humans, and could have major implications for a variety of human-specific diseases [1Lui J.H. et al.Development and evolution of the human neocortex.Cell. 2011; 146: 18-36Abstract Full Text Full Text PDF PubMed Scopus (898) Google Scholar]. In correlation with its elaborate functions, the cerebral cortex displays multiple levels of complexity. It contains dozens of different types of neurons populating specific cortical areas and layers, and cortical neuron number and diversity are thought to be at the core of its powerful computational capacities. A first subdivision among cortical neurons distinguishes two main cell classes. Pyramidal neurons constitute >85% of cortical neurons, they are glutamatergic, and send long-range projections to other cortical or subcortical targets. The remaining 15% of cortical neurons are GABA-ergic interneurons that display only local connectivity. Pyramidal neurons and interneurons can be further subdivided into dozens of subtypes, characterised by specific molecular and functional properties [2Molyneaux B.J. et al.Neuronal subtype specification in the cerebral cortex.Nat. Rev. Neurosci. 2007; 8: 427-437Crossref PubMed Scopus (1159) Google Scholar, 3DeFelipe J. et al.New insights into the classification and nomenclature of cortical GABAergic interneurons.Nat. Rev. Neurosci. 2013; 14: 202-216Crossref PubMed Scopus (546) Google Scholar]. Pluripotent embryonic stem cells (ESC; see Glossary) [4Smith A.G. Embryo-derived stem cells: of mice and men.Annu. Rev. Cell Dev. 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Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors.Cell. 2006; 126: 663-676Abstract Full Text Full Text PDF PubMed Scopus (19304) Google Scholar] has provided the opportunity to use ESC or iPSC-based neural differentiation to model human brain diseases [8Dolmetsch R. Geschwind D.H. The human brain in a dish: the promise of iPSC-Derived neurons.Cell. 2011; 145: 831-834Abstract Full Text Full Text PDF PubMed Scopus (228) Google Scholar, 9Gaspard N. Vanderhaeghen P. From stem cells to neural networks: recent advances and perspectives for neurodevelopmental disorders.Dev. Med. Child Neurol. 2011; 53: 13-17Crossref PubMed Scopus (45) Google Scholar]. Here, we review recent progress on the generation of cortical neurons from PSC, illustrating that much, but not all of the complexity of cortical development can be recapitulated with surprisingly simple in vitro conditions, providing novel insights into corticogenesis. The cerebral cortex is formed within the telencephalon, the anterior-most part of the forebrain. Forebrain or telencephalon identity is thought to constitute a primitive pattern of neural identity, which is acquired and retained through local inhibition of caudalising morphogen signals [10Wilson S.W. Houart C. Early steps in the development of the forebrain.Dev. Cell. 2004; 6: 167-181Abstract Full Text Full Text PDF PubMed Scopus (359) Google Scholar]. In vitro studies using ESC have confirmed and extended this model in both mice and humans (Figure 1A) . 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The telencephalon then undergoes patterning along the dorsoventral axis, primarily through induction of ventral identities by the morphogen Sonic Hedgehog (SHH) [21Hebert J.M. Fishell G. The genetics of early telencephalon patterning: some assembly required.Nat. Rev. Neurosci. 2008; 9: 678-685Crossref PubMed Scopus (271) Google Scholar]. This regionalisation process is intimately linked to the specification of the two main populations of cortical neurons: pyramidal neurons and interneurons are generated from distinct populations of progenitors located in the dorsal and the ventral part of the telencephalon, respectively [21Hebert J.M. Fishell G. The genetics of early telencephalon patterning: some assembly required.Nat. Rev. Neurosci. 2008; 9: 678-685Crossref PubMed Scopus (271) Google Scholar, 22Sur M. Rubenstein J.L. Patterning and plasticity of the cerebral cortex.Science. 2005; 310: 805-810Crossref PubMed Scopus (512) Google Scholar, 23Wonders C.P. Anderson S.A. The origin and specification of cortical interneurons.Nat. Rev. Neurosci. 2006; 7: 687-696Crossref PubMed Scopus (707) Google Scholar]. The same binary logic is observed during ESC-derived telencephalic induction (Figure 1A). During mouse ESC differentiation, SHH inhibition leads to the generation of dorsal telencephalic progenitors, which subsequently generate mostly pyramidal neurons [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar, 17Watanabe K. et al.Directed differentiation of telencephalic precursors from embryonic stem cells.Nat. 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Intriguingly, SHH inhibition is not strictly required during human ESC corticogenesis, which is likely to be due to lower endogenous SHH signalling levels [18Espuny-Camacho I. et al.Pyramidal neurons derived from human pluripotent stem cells integrate efficiently into mouse brain circuits in vivo.Neuron. 2013; 77: 440-456Abstract Full Text Full Text PDF PubMed Scopus (382) Google Scholar, 26Li X.J. et al.Coordination of sonic hedgehog and Wnt signaling determines ventral and dorsal telencephalic neuron types from human embryonic stem cells.Development. 2009; 136: 4055-4063Crossref PubMed Scopus (249) Google Scholar, 27Shi Y. et al.Human cerebral cortex development from pluripotent stem cells to functional excitatory synapses.Nat. Neurosci. 2012; 15: 477-486Crossref PubMed Scopus (96) Google Scholar]. Conversely, specification of ventral telencephalic cells from both human and mouse ESC does require SHH stimulation, alone or together with Wnt inhibition [17Watanabe K. et al.Directed differentiation of telencephalic precursors from embryonic stem cells.Nat. Neurosci. 2005; 8: 288-296Crossref PubMed Scopus (615) Google Scholar, 25Danjo T. et al.Subregional specification of embryonic stem cell-derived ventral telencephalic tissues by timed and combinatory treatment with extrinsic signals.J. 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The concentration and timing (onset and duration) of exposure to SHH will lead to different types of ventral progenitors and, hence, to distinct subtypes of neurons, from hypothalamic and striatal projection neurons to cortical and striatal interneurons [25Danjo T. et al.Subregional specification of embryonic stem cell-derived ventral telencephalic tissues by timed and combinatory treatment with extrinsic signals.J. Neurosci. 2011; 31: 1919-1933Crossref PubMed Scopus (138) Google Scholar, 32Germain N.D. et al.Derivation and isolation of NKX2.1-positive basal forebrain progenitors from human embryonic stem cells.Stem Cells Dev. 2013; 22: 1477-1489Crossref PubMed Scopus (46) Google Scholar, 33Ma L. et al.Human embryonic stem cell-derived GABA neurons correct locomotion deficits in quinolinic acid-lesioned mice.Cell Stem Cell. 2012; 10: 455-464Abstract Full Text Full Text PDF PubMed Scopus (234) Google Scholar], reminiscent of the time dependence of SHH signalling in vivo [34Briscoe J. 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Collectively, these data demonstrate that ESC and iPSC, whether of human or mouse origin, can generate either ventral or dorsal telencephalic progenitors, depending on the levels of SHH signalling. Eventually, these progenitors produce specific types of neurons, whereby dorsal cells generate essentially glutamatergic pyramidal neurons, whereas ventral cells generate mostly GABA-ergic neurons. Interestingly, human and mouse PSC do not differ in this respect, in line with results obtained in human ex vivo cultures where, as in the rodent, the ventral telencephalon appears to be the site of origin of most, if not all, cortical interneurons [36Hansen D.V. et al.Non-epithelial stem cells and cortical interneuron production in the human ganglionic eminences.Nat. Neurosci. 2013; 16: 1576-1587Crossref PubMed Scopus (199) Google Scholar, 37Ma T. et al.Subcortical origins of human and monkey neocortical interneurons.Nat. Neurosci. 2013; 16: 1588-1597Crossref PubMed Scopus (201) Google Scholar]. The mammalian neocortex is organised into six different layers, each of which comprises a collection of neurons displaying specific patterns of gene expression and connectivity [2Molyneaux B.J. et al.Neuronal subtype specification in the cerebral cortex.Nat. Rev. Neurosci. 2007; 8: 427-437Crossref PubMed Scopus (1159) Google Scholar, 38Gaspard N. Vanderhaeghen P. Laminar fate specification in the cerebral cortex.F1000 Biol. Rep. 2011; 3: 6Crossref PubMed Scopus (23) Google Scholar, 39Leone D.P. et al.The determination of projection neuron identity in the developing cerebral cortex.Curr. Opin. Neurobiol. 2008; 18: 28-35Crossref PubMed Scopus (288) Google Scholar, 40Greig L.C. et al.Molecular logic of neocortical projection neuron specification, development and diversity.Nat. Rev. Neurosci. 2013; 14: 755-769Crossref PubMed Scopus (518) Google Scholar] (Figure 1B). Which layer a neuron settles in, is tightly linked to its birthdate, with deeper layer neurons being generated earlier than upper layer neurons. This process of temporal patterning is central to the generation of layer-specific types of cortical neurons. In vivo and in vitro studies have shown that it involves the progressive restriction of progenitor competence, which is controlled by both intrinsic, cell-autonomous programmes and by environmental influences [2Molyneaux B.J. et al.Neuronal subtype specification in the cerebral cortex.Nat. Rev. Neurosci. 2007; 8: 427-437Crossref PubMed Scopus (1159) Google Scholar, 38Gaspard N. Vanderhaeghen P. Laminar fate specification in the cerebral cortex.F1000 Biol. Rep. 2011; 3: 6Crossref PubMed Scopus (23) Google Scholar, 39Leone D.P. et al.The determination of projection neuron identity in the developing cerebral cortex.Curr. Opin. 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Surprisingly, both mouse and human PSC-derived corticogenesis recapitulate robustly temporal patterning in vitro: neurons that express molecular markers and project to targets specific of upper layer neurons are generated consistently later than neurons with a deep layer identity [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar, 18Espuny-Camacho I. et al.Pyramidal neurons derived from human pluripotent stem cells integrate efficiently into mouse brain circuits in vivo.Neuron. 2013; 77: 440-456Abstract Full Text Full Text PDF PubMed Scopus (382) Google Scholar, 24Eiraku M. et al.Self-organized formation of polarized cortical tissues from ESCs and its active manipulation by extrinsic signals.Cell Stem Cell. 2008; 3: 519-532Abstract Full Text Full Text PDF PubMed Scopus (990) Google Scholar, 41Tiberi L. et al.BCL6 controls neurogenesis through Sirt1-dependent epigenetic repression of selective Notch targets.Nat. 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However, whereas in vitro systems of corticogenesis display remarkable similarities with in vivo developmental processes, it still differs in significant ways from in vivo corticogenesis, depending on culture conditions. Indeed, whereas in vivo deep and upper layer neurons each represent approximately half of the cortex, ESC-derived pyramidal neurons are strongly skewed towards a deep layer identity following monoadherent culture in minimal differentiation conditions in the absence of added morphogens [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar, 18Espuny-Camacho I. et al.Pyramidal neurons derived from human pluripotent stem cells integrate efficiently into mouse brain circuits in vivo.Neuron. 2013; 77: 440-456Abstract Full Text Full Text PDF PubMed Scopus (382) Google Scholar]. Importantly, this was also the case when native mouse cortical progenitors were grown ex vivo at clonal densities [43Shen Q. et al.The timing of cortical neurogenesis is encoded within lineages of individual progenitor cells.Nat. Neurosci. 2006; 9: 743-751Crossref PubMed Scopus (471) Google Scholar]. Conversely, a higher proportion of upper layer neurons appears to be generated when ESC are first differentiated at high density and/or supplemented with extrinsic cues, such as retinoic acid [27Shi Y. et al.Human cerebral cortex development from pluripotent stem cells to functional excitatory synapses.Nat. Neurosci. 2012; 15: 477-486Crossref PubMed Scopus (96) Google Scholar] or as cell aggregates [24Eiraku M. et al.Self-organized formation of polarized cortical tissues from ESCs and its active manipulation by extrinsic signals.Cell Stem Cell. 2008; 3: 519-532Abstract Full Text Full Text PDF PubMed Scopus (990) Google Scholar, 44Mariani J. et al.Modeling human cortical development in vitro using induced pluripotent stem cells.Proc. Natl. Acad. Sci. U.S.A. 2012; 109: 12770-12775Crossref PubMed Scopus (369) Google Scholar]. Although direct comparison between various studies is not always straightforward because of different markers being analysed, these findings suggest that extrinsic cues that may be missing in minimal culture systems, are required for the proper generation of the upper layer neurons [45Tiberi L. et al.Cortical neurogenesis and morphogens: diversity of cues, sources and functions.Curr. Opin. Cell Biol. 2012; 24: 269-276Crossref PubMed Scopus (76) Google Scholar]. Consistent with this hypothesis, whereas human PSC-derived cortical progenitors cultured in minimal conditions generate only a few upper layer neurons even after prolonged periods in vitro, they generate many upper layer neurons following transplantation into the mouse newborn cortex [18Espuny-Camacho I. et al.Pyramidal neurons derived from human pluripotent stem cells integrate efficiently into mouse brain circuits in vivo.Neuron. 2013; 77: 440-456Abstract Full Text Full Text PDF PubMed Scopus (382) Google Scholar]. These data suggest that cortical progenitors competent to generate neurons of all six layers can be generated following minimal in vitro conditions from ESC, but that cues present in the mouse newborn brain are necessary to enhance upper layer neuron production from these cells. Such cues could be either produced by cortical progenitors and neurons themselves, or derived from extrinsic sources, such as meninges, cerebrospinal fluid, or the vascular niche [45Tiberi L. et al.Cortical neurogenesis and morphogens: diversity of cues, sources and functions.Curr. Opin. Cell Biol. 2012; 24: 269-276Crossref PubMed Scopus (76) Google Scholar]. PSC-derived corticogenesis may provide a useful platform to identify these cues and their mechanisms of action. Specifically, it will be interesting to determine whether and how these cues act to change the competence of cortical progenitors to generate upper layer neurons, and if they act on the specification or amplification of specific types of progenitors, such as intermediate progenitors or outer radial glial cells, as explained further below. In parallel with temporal patterning, a key transition for the generation of cortical neurons is neurogenesis itself, whereby cortical progenitors differentiate into cortical neurons, either directly or indirectly following brief amplification through intermediate progenitors [46Kriegstein A. Alvarez-Buylla A. The glial nature of embryonic and adult neural stem cells.Annu. Rev. Neurosci. 2009; 32: 149-184Crossref PubMed Scopus (1668) Google Scholar, 47Fietz S.A. Huttner W.B. Cortical progenitor expansion, self-renewal and neurogenesis: a polarized perspective.Curr. Opin. Neurobiol. 2011; 21: 23-35Crossref PubMed Scopus (91) Google Scholar]. The rate and timing of neuronal differentiation from cortical progenitors is dynamically regulated by various intrinsic and extrinsic cues, including Notch and proneural factors, such as neurogenins, which collectively control the number and fate of cortical neurons [45Tiberi L. et al.Cortical neurogenesis and morphogens: diversity of cues, sources and functions.Curr. Opin. Cell Biol. 2012; 24: 269-276Crossref PubMed Scopus (76) Google Scholar, 48Gotz M. Huttner W.B. The cell biology of neurogenesis.Nat. Rev. Mol. Cell Biol. 2005; 6: 777-788Crossref PubMed Scopus (1570) Google Scholar, 49Okano H. Temple S. Cell types to order: temporal specification of CNS stem cells.Curr. Opin. Neurobiol. 2009; 19: 112-119Crossref PubMed Scopus (186) Google Scholar, 50Guillemot F. Cell fate specification in the mammalian telencephalon.Prog. Neurobiol. 2007; 83: 37-52Crossref PubMed Scopus (194) Google Scholar]. ESC-derived cortical neurogenesis closely recapitulates these processes [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar, 24Eiraku M. et al.Self-organized formation of polarized cortical tissues from ESCs and its active manipulation by extrinsic signals.Cell Stem Cell. 2008; 3: 519-532Abstract Full Text Full Text PDF PubMed Scopus (990) Google Scholar], and has been used to gain mechanistic insights into the factors involved in this process (Figure 1A). The mouse ESC cell-based model of corticogenesis was used to screen for novel transcription factors involved in neurogenesis, leading to the identification of a novel potent pro-neurogenic gene, B cell CLL/lymphoma 6 (BCL6) [41Tiberi L. et al.BCL6 controls neurogenesis through Sirt1-dependent epigenetic repression of selective Notch targets.Nat. Neurosci. 2012; 15: 1627-1635Crossref PubMed Scopus (92) Google Scholar]. Whereas BCL6 was uncovered in an ESC-based screen, it was subsequently found to be expressed in cortical progenitors in vivo, during the transition from cortical progenitors to pyramidal neurons. Furthermore, the analysis of BCL6 knockout mice led to the conclusion that BCL6 is required for proper cortical neurogenesis in vivo [41Tiberi L. et al.BCL6 controls neurogenesis through Sirt1-dependent epigenetic repression of selective Notch targets.Nat. Neurosci. 2012; 15: 1627-1635Crossref PubMed Scopus (92) Google Scholar]. Combined studies on the ESC system and in vivo cortex converged to demonstrate that BCL6 acts through direct and stable epigenetic repression of the Notch target Hes5 promoter, thereby enabling the neurogenic transition to proceed irreversibly. This study illustrates how the use of ESC-based models of corticogenesis can lead to novel insights into the in vivo mechanisms of cortical neuron generation. In addition to layer-specific identity, neurons from different cortical areas also develop selective patterns of gene expression and connectivity. The patterning of cortical areas is a complex process resulting from the interplay between factors intrinsic to the cortex, as well as extrinsic factors from outside the brain [22Sur M. Rubenstein J.L. Patterning and plasticity of the cerebral cortex.Science. 2005; 310: 805-810Crossref PubMed Scopus (512) Google Scholar, 51O'Leary D.D. Sahara S. Genetic regulation of arealization of the neocortex.Curr. Opin. Neurobiol. 2008; 18: 90-100Crossref PubMed Scopus (179) Google Scholar]. Surprisingly, in vivo transplantation experiments revealed that mouse ESC-derived cortical neurons seem to acquire mainly limbic and visual (occipital) identities [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar]. Following transplantation, ESC-derived cortical neurons send axons to specific visual and limbic targets, with a pattern of projection that is strikingly similar to grafted embryonic visual cortical tissue [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar, 52Gaillard A. Roger M. Early commitment of embryonic neocortical cells to develop area-specific thalamic connections.Cereb. Cortex. 2000; 10: 443-453Crossref PubMed Scopus (33) Google Scholar]. Importantly, these results were all obtained with grafts within the frontal cortex, suggesting that the highly selective pattern of projections was not due to in vivo respecification of the grafted neurons. Confirming this hypothesis, examination of the molecular identity of ESC-derived cortical progenitors and neurons before grafting revealed that most of them expressed typical markers of the occipital cortex, in particular chicken ovalbumin upstream promoter transcription factors (Coup-TF) I and II [11Gaspard N. et al.An intrinsic mechanism of corticogenesis from embryonic stem cells.Nature. 2008; 455: 351-357Crossref PubMed Scopus (490) Google Scholar]. By contrast, the areal fate of ESC-derived cortical progenitors in vitro could be modified by the addition of extrinsic cues known
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