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Performance Performance. Performance cookies are used to understand and analyze the key performance indexes of the website which helps in delivering a better user experience for the visitors. Analytics Analytics. Analytical cookies are used to understand how visitors interact with the website. These cookies help provide information on metrics the number of visitors, bounce rate, traffic source, etc. And researchers have, indeed, been comparing the development of hand gestures in chimpanzees, bonobos, and humans.
The evidence, they say, points toward the use of these gestures as the early onset of language. This evolution of language was present in a common ancestor of humans, chimpanzees, and bonobos. All three species demonstrated an increase in their use of gestures while developing. Just as we go to supermarkets, chimpanzees collect food to assemble a feast.
Research shows that like humans, chimps prefer cooked food over raw food; they also understand the transformation process of cooking. Chimpanzee Trekking. Photo Credit: Tad Bradley. Humans are not the only ones with a moral conscious, research has proven chimpanzees can distinguish behavior that is right from wrong. A University of Zurich study reveals that chimps angrily react to scenes of a baby chimp being harmed or killed by its own kind.
This suggests chimps have a strong moral conscious like humans. The common DNA, we share with chimpanzees, accounts for many of our similarities. Networks of brain cells in the cerebral cortex also behave differently in the two species.
How these species differences arise is not clear, but it likely occurs in the earliest phases of development when brain stem and progenitor cells divide and give rise to cerebral cortex cells in the growing brain.
To study the earliest stages of brain development, researchers often use human brain cells grown in the laboratory. Under the right conditions, cells collected from adult humans and other animals can be reprogrammed to behave like brain stem cells. Recently, researchers have been able to use these reprogrammed cells to make tissue that resembles the brain in petri dishes, known as brain organoids.
The experiments showed that the human and chimpanzee brain organoids were remarkably similar in many ways including in the mix of cell types and in how these cells were arranged. This suggests that a longer metaphase may be a feature of brain stem cells. Further studies are now needed to find out how the length of time these progenitor cells spend in metaphase affects how chimpanzee and human brains develop; and whether this can help explain why the human brain is so much larger.
The expansion of the neocortex during primate evolution is thought to contribute to the higher cognitive capacity of humans compared to our closest living relatives, the great apes, and notably the chimpanzees Geschwind and Rakic, ; Rakic, ; Striedter, Neocortex expansion in humans relative to chimpanzees involves an increase in the number of cortical neurons generated during fetal development Borrell and Reillo, ; Florio and Huttner, ; Herculano-Houzel, ; Lui et al. This reflects primarily a greater and prolonged proliferative capacity of human neural stem and progenitor cells NSPCs within the germinal zones of the developing neocortex Lewitus et al.
Unravelling differences between human and chimpanzee NSPC behaviour is therefore a key issue, yet very little is known about such differences. The neocortex develops from two principal germinal zones, the ventricular zone VZ and the subventricular zone SVZ Angevine et al.
APs neuroepithelial cells, apical radial glia, and apical intermediate progenitors divide at the ventricular surface, keep ventricular contact and exhibit apical cell polarity, whereas BPs basal or outer radial glia and basal intermediate progenitors lack this contact and type of cell polarity Taverna et al.
Studies dissecting the switch between NSPC proliferation and differentiation have demonstrated that a central aspect of the cell division process, the orientation of the mitotic spindle, has a pivotal role, particularly in the case of APs Lancaster and Knoblich, ; Mora-Bermudez and Huttner, ; Mora-Bermudez et al.
Comparing spindle orientation in mitotic APs may therefore provide insight into the cell biological basis underlying the differences between humans and chimpanzees in NSPC proliferation versus differentiation during neocortex development. Protocols to generate structured cerebral tissue cerebral organoids from pluripotent stem cells in vitro constitute a major advance for studying neocortex development, in particular with regard to humans and non-human primates where fetal brain tissue is hard or impossible to obtain and manipulate Kadoshima et al.
Human cerebral organoids form a variety of tissues that resemble specific brain regions, including the cerebral cortex, ventral forebrain, midbrain-hindbrain boundary, hippocampus, and retina. Moreover, their cerebral cortex-like regions exhibit distinct germinal zones, that is, a VZ containing APs and an SVZ containing BPs, as well as basal-most neuronal layers.
Cerebral organoid APs include apical radial glia-like NSPCs that contact a ventricle-like lumen, express radial glia marker genes, undergo interkinetic nuclear migration, and divide at the apical surface, similar to their in vivo counterparts, and cerebral organoid BPs comprise both basal radial glia-like and basal intermediate progenitor-like NSPCs Lancaster et al.
Finally, we have previously shown by single-cell RNA sequencing that the gene expression programs controlling neocortex development in human cerebral organoids are remarkably similar to those in the developing fetal tissue Camp et al.
Together, these findings suggest that cerebral organoids constitute a valid system to explore potential differences in NSPC proliferation versus differentiation between humans and chimpanzees Otani et al. Here, we have generated cerebral organoids from chimpanzee-derived induced pluripotent stem cells iPSCs , and used single-cell transcriptomics, immunohistofluorescence and live imaging to compare relevant features of chimpanzee NSPCs to human NSPCs in cerebral organoids and fetal neocortex.
While most NSPC characteristics are found to be similar, we show that the prometaphase-metaphase in mitotic APs is longer in humans than in chimpanzees, indicating that a fundamental difference exists in the regulation of mitosis during neocortex development between the two species. Our data also provide a resource for further studies on human and chimpanzee differences in cortical development, and demonstrate the usability of cerebral organoids as a means to be able to perform such studies.
We generated cerebral organoids from iPSCs derived from chimpanzee fibroblasts and lymphocytes Figure 1A left, Figure 1—figure supplement 1. These chimpanzee cerebral organoids formed complex tissue structures that resembled the developing primate brain Figure 1A right , as reported previously for human cerebral organoids Lancaster et al.
Similar to human iPSC-derived cerebral organoids [ Camp et al. Consistent with this, cells immunoreactive for the deep-layer neuron marker CTIP2 were observed in the basal region of the developing cortical wall Figure 1B left , corresponding to an early cortical plate. In the context of the time-lapse live imaging of apical mitoses described below, we observed apically directed nuclear migration prior to, and basally directed nuclear migration after, mitosis, consistent with the existence of interkinetic nuclear migration.
Our results suggest that chimpanzee cerebral organoids recapitulate important aspects of fetal chimpanzee brain development and allow comparisons with cerebral cortex development in human cerebral organoids and fetal neocortex. CP, cortical plate; N, neuron. E Heatmap showing normalized correlation Z-score of single-cell transcriptomes from chimpanzee cerebral organoid cortex with bulk RNA-seq data from laser-microdissected zones Fietz et al.
CP, cortical plate. F Scatterplot showing NSPC and neuronal signature scores derived from analysis of fetal cerebral cortex single-cell transcriptomes Figure 1—figure supplement 1 calculated for each chimpanzee cerebral organoid cortical cell. Each column represents a single cell, each row a gene. Cell type and maximum correlation to bulk RNA-seq data from cortical zones are shown in the top sidebar. Cells are coloured based on cortical zone top left or cell type assignment bottom left.
APs, BPs, and neurons were classified based on maximum correlation with single-cell transcriptomes from the human fetal neocortex. List of genes identified by PCA on all chimpanzee organoid single-cell transcriptomes as being most informative for defining cell populations. In line with what would be expected with regard to neuron production, the proportion of PAX6—TBR2— cells, located in the basal-most zones of the developing cortical wall, was very low at D28 but increased by DD54 to about a third of the total cells for both, human and chimpanzee cerebral organoids Figure 2B.
Immunostaining for CTIP2 corroborated the neuronal identity of these cells data not shown. Error bars, SEM. Consistent with the observation that the total proportion of NSPCs relative to neurons was virtually identical in human and chimpanzee organoids Figure 2B , the abundance of cycling cells, as revealed by KI67 immunostaining, was essentially similar Figure 2C,D.
To survey the cellular composition and cell type-specific transcriptomes of the chimpanzee organoids, we analysed single cell transcriptomes from 7 organoids ranging in age from 45 to 80 days Figure 1D , Figure 1—source data 1.
We combined all transcriptomes and identified the genes most informative for defining cell populations by principal component analysis PCA Figure 1—source data 2. Using these genes, we used tSNE analysis to cluster cells into transcriptionally distinct groups representing cerebral cortex, hindbrain, ventral midbrain and peripheral mesenchyme Figure 1—figure supplement 2.
These groups are similar to those identified in human cerebral organoids Camp et al. We identified cortex-like cells based on strong expression of canonical NSPC and neuron marker genes i. We sub-classified the cerebral cortex-like cells based on the correlation between their transcriptomes and the bulk transcriptomes of laser-capture microdissected VZ, iSVZ, oSVZ, and cortical plate of fetal human neocortex GSE, [ Fietz et al. We found that groups of cells correlated best with one of the four zones, suggesting that the range of cell types present in the human fetal and organoid cerebral cortex are represented in our chimpanzee data Figure 1E.
Consistent with this, each chimpanzee cell represents a cell state on a continuum from NPSCs to neurons based on gene expression signatures extracted from fetal human cerebral cortex transcriptomes Figure 1F , Figure 1—figure supplement 3 Camp et al. We next classified the chimpanzee cerebral cortex cells by determining the fetal cell type with which each cell most strongly correlates, resulting in 73 APs, 25 BPs, and 80 neurons.
Analysis of known cell type markers revealed expression patterns consistent with what has been observed in human organoid and fetal cerebral cortex Figure 1G Camp et al. Though this classification is convenient to describe the cell types present in the chimpanzee organoid, we note that many of the cells can be described as intermediates between APs, BPs, and different stages of neuron maturation. We inferred lineage relationships among the chimpanzee cerebral cortex in an adjacency network based on pairwise correlations between cells Figure 1H , revealing a structured topology where VZ-APs connect to cortical plate neurons through SVZ-BPs.
These lineage relationships were corroborated using a minimal spanning tree algorithm Figure 1—figure supplement 3G Trapnell et al. Together, these data allowed reconstruction of the chimpanzee organoid cerebral cortex from single-cell transcriptomes. To further explore transcriptome similarities and differences between chimpanzee and human cerebral cortex cells, we compared them to the single-cell transcriptomes of fetal human cortex cells 12—13 weeks post-conception wpc , published in Camp et al.
Hierarchical clustering of organoid and fetal cells showed that human and chimpanzee organoid and human fetal cells were distributed together within the two main sub-clusters representing NSPCs and neurons not shown , and showed highly correlated expression of marker gene patterns Figure 3B.
PCA was performed on all single-cell transcriptomes using genes expressed in more than two cells and with a non-zero variance. B Quasibinomial fit line of representative marker gene expression across cells ordered by correlation with PC1.
D Lineage network see C coloured by scaled expression level of marker genes. E Scatterplots showing z-scored significance estimates from single-cell differential expression SCDE analysis based on Bayesian probabilistic models. Reads from human and chimpanzee were mapped to a consensus genome, and human gene annotations were used for expression counting.
Genes coloured as white triangles represent marker genes from Figure 1 and are generally not differentially expressed between human and chimpanzee, but do vary between APs and neurons, validating the SCDE analysis.
Yellow and purple circles represent genes upregulated specifically in human APs and neurons, respectively. Circles are sized based on differential expression between human APs and neurons. Figure 3—figure supplement 1 shows a similar plot from the chimpanzee perspective. F Gene ontology enrichments -log 10 P-value for differentially expressed gene groups shown in panel E.
Left, human APs yellow and neurons N, purple that are not differential between human and chimpanzee. Center, upregulated in human APs top or neurons N, bottom compared to chimpanzee. Right, upregulated in chimpanzee APs top or neurons N, bottom from Figure 3—figure supplement 1. H The same bulk RNA-seq data was used to confirm and estimate the origin of differential gene expression in APs versus neurons from single-cell organoid data.
Pie chart shows the proportion of AP-enriched yellow or neuron-enriched N, purple genes that are observed in human, chimpanzee, and mouse. Pie charts also show the proportion of genes differential between APs and neurons that are observed only in human and chimpanzee, but not mouse human-chimp ancestor , or genes specific to human or chimpanzee. Generally, APs, BPs, and neurons from human and chimpanzee intermixed, confirming that cells in the chimpanzee organoid cortices have a zonal organization consistent with what is observed histologically Figure 3C,D.
In conclusion, the major proportion of the variation in these data is not between in vitro and in vivo tissues or between species, but among cell states during neurogenesis, confirming that the major features of the genetic programs regulating the NSPC-to-neuron lineage are conserved between human and chimpanzees, and are recapitulated in cerebral organoids. To identify genes differentially expressed between chimpanzee and human cortex-like cells, we remapped all single-cell transcriptome reads to a consensus human-chimpanzee genome and used human annotations to identify 1-to-1 orthologous genes.
We then used a Bayesian approach to identify differentially expressed genes by comparing cerebral organoid APs and neurons between species ignoring BPs due to the low number of BPs identified. We identified and genes that were more highly expressed in human APs and neurons, respectively, and and genes that were more highly expressed in chimpanzee APs and neurons, respectively Figure 3E , Figure 3—source data 2.
In addition to the between-species comparisons, we identified genes differentially expressed between human or chimpanzee APs and neurons to identify cell-type specific genes for human: AP-specific, neuron-specific; for chimpanzee: AP-specific, neuron-specific. Of the differentially expressed genes between species, we identified 93 genes that are strongly upregulated in human organoid APs and 72 genes upregulated in human organoid neurons. Gene ontology enrichments suggest that the proteins encoded by some of these genes are integral to cell membranes and involved in intercellular signalling Figure 3F , Figure 3—source data 2 , for example integrin beta 8 ITGB8 in APs and insulin receptor INSR in human neurons.
Notably, a similar proportion of AP- and neuron-specific genes were gained on the chimpanzee and human branch subsequent to their separation, suggesting that our analysis did not have a strong human bias. We used an established live imaging method Mora-Bermudez et al. We did not observe signs of strong perturbation during live image acquisition in either system, such as mitotic arrest Figure 4A,C,E ; see also Figure 5A—C and Video 1 or lack of nuclear movements and cell death.
Chromosome dynamics and spindle orientation of APs, as revealed by the orientation of the metaphase plate, were similar in human developing neocortex and human organoids, both before anaphase Figure 4A—D,G and during anaphase Figure 4A—D,H,I , when cell cleavage initiates.
This strongly suggests that cerebral organoids are a suitable model to study live NSPC division and spindle orientation dynamics. Live tissue imaging of spindle orientation, as reported by chromosome plate orientation, in organotypic slice culture of developing neocortex and cerebral organoids. Measurements were started after all chromosomes had formed a tight metaphase plate. The time indicated on each image is when that image was taken, relative to anaphase onset 0 min.
White dashed lines, ventricular surface. Yellow dashed lines indicate the two metaphase plate orientations with the greatest difference to each other. B , D , F Quantification of all orientations of the chromosome plates from the beginning of the metaphase plate stage to anaphase, for APs in the three respective tissues described in A , C , E.
G Maximal range of chromosome plate orientations for APs, from the beginning of the metaphase plate stage to anaphase onset, as determined in the measurements shown in B , D , F. H , I , J Orientation of chromosome plates at 2.
Live tissue imaging of mitotic phases, as reported by chromosomes, in organotypic slice culture of developing neocortex and cerebral organoids. This video cannot be played in place because your browser does support HTML5 video.
You may still download the video for offline viewing. Related to Figure 5B and C Live tissue imaging of mitotic phases, as reported by chromosomes, in organotypic slice culture of cerebral organoids. Datasets are the same as in Figure 5B and C. Growing colour bars at the bottom indicate time progression of the respective dividing AP and are synchronized to the beginning of prometaphase in green.
Metaphase plate time is in yellow and anaphase time is in red. Note the slower progression of the dividing human AP on the left. Spindle orientation can determine symmetric vs. We compared spindle orientation dynamics between human and chimpanzee APs in cerebral organoids. However, our data revealed no clear differences in spindle orientation, either during metaphase Figure 4C—G or shortly after anaphase onset Figure 4C—F, I—J.
Oblique and near-horizontal orientations were also observed, but at a much lower abundance and at similar frequencies in chimpanzee and human organoids Figure 4H—J.
This shows that the frequency of asymmetric cell division caused by oblique spindle orientation is most likely not a major difference between human and chimpanzee APs. We noticed, however, unexpected differences between human and chimpanzee APs in their progression through mitosis. By comparison, prometaphase-metaphase of APs in slice culture of mouse neocortex, a well-characterized model system for neurogenesis, lasted for only approximately half the amount of time than human APs Figure 5D,E ; Figure 5—source data 1.
To trace the specific phase of mitosis when this difference arises, we used chromosome morphology and dynamics to determine the time chromosomes spent congressing toward the equatorial plane of the cell defined here as 'prometaphase' and the time they spent tightly aligned as a metaphase plate defined here as 'metaphase'.
By contrast, in mouse APs, both prometaphase and metaphase were found to be significantly shorter than the respective mitotic phases in human and chimpanzee APs Figure 5D,F,G ; Figure 5—source data 1. None of the other mitotic phases prophase, anaphase, telophase differed in length between APs in human fetal neocortex and human cerebral organoids vs. However, anaphase of mouse APs was found to be significantly shorter than that of human and chimpanzee APs Figure 5—figure supplement 1A ; Figure 5—source data 1.
These differences between species in the individual mitotic phases were reflected in the cumulative length of total mitosis, which was significantly shorter in mouse APs than human and chimpanzee APs Figure 5—figure supplement 1B. To search for potential functional implications of these observations, we next quantified and compared the length of prometaphase-metaphase in human and chimpanzee APs of day 52 D52 cerebral organoids, and compared the results with those of D30 organoids.
Prometaphase-metaphase Figure 5—figure supplement 2A and metaphase alone Figure 5—figure supplement 2C ; Figure 5—source data 1 were shorter in D52 than in D30 human APs, and not anymore statistically significantly different in length from D52 chimpanzee APs. The longer metaphase of human than chimpanzee organoid APs may therefore characterise early phases of cortical development, when proliferative AP divisions are predominant.
We also generated cerebral organoids from an orangutan iPSC line and determined the length of AP prometaphase-metaphase. This revealed that the length of prometaphase-metaphase in orangutan D30 organoid APs was similar to that of chimpanzee APs and significantly shorter than that of human organoid APs Figure 5—figure supplement 3A,B.
As was the case for the human-chimpanzee AP comparison, the shorter prometaphase-metaphase of orangutan than human APs was due to a shorter metaphase Figure 5—figure supplement 3A,D rather than prometaphase Figure 5—figure supplement 3A,C ; Figure 5—source data 1.
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