R-3 Repository :: Browsing by Author "Chhabra, Sapna"

Browsing by Author "Chhabra, Sapna"

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BMP-treated human embryonic stem cells transcriptionally resemble amnion cells in the monkey embryo
(The Company of Biologists, 2021) Chhabra, Sapna; Warmflash, Aryeh
Human embryonic stem cells (hESCs) possess an immense potential to generate clinically relevant cell types and unveil mechanisms underlying early human development. However, using hESCs for discovery or translation requires accurately identifying differentiated cell types through comparison with their in vivo counterparts. Here, we set out to determine the identity of much debated BMP-treated hESCs by comparing their transcriptome to recently published single cell transcriptomic data from early human embryos ( Xiang et al., 2020). Our analyses reveal several discrepancies in the published human embryo dataset, including misclassification of putative amnion, intermediate and inner cell mass cells. These misclassifications primarily resulted from similarities in pseudogene expression, highlighting the need to carefully consider gene lists when making comparisons between cell types. In the absence of a relevant human dataset, we utilized the recently published single cell transcriptome of the early post implantation monkey embryo to discern the identity of BMP-treated hESCs. Our results suggest that BMP-treated hESCs are transcriptionally more similar to amnion cells than trophectoderm cells in the monkey embryo. Together with prior studies, this result indicates that hESCs possess a unique ability to form mature trophectoderm subtypes via an amnion-like transcriptional state.This article has an associated First Person interview with the first author of the paper.
Dissecting signaling dynamics underlying self-organized fate patterning of 2D human gastruloids
(2020-04-13) Chhabra, Sapna; Warmflash, Aryeh
Gastrulation is a crucial stage in embryonic development when the epiblast self- organizes to form three germ layers – the ectoderm, mesoderm and endoderm, which eventually form the entire organism. Decades of genetic and biochemical studies in the mouse embryo have revealed that a signaling cascade involving the BMP, WNT and NODAL pathways is necessary for gastrulation. In the developing mouse embryo, WNT and NODAL ligands are expressed near the site of gastrulation, and knockout of these ligands leads to a failure to gastrulate. These data have led to the prevailing view that a signaling gradient in WNT and NODAL underlies germ layer fate patterning during gastrulation. However, the activities of these pathways in space and time have never been directly observed, as gastrulation occurs after the mouse embryo implants in the mother’s uterus, and thus, precludes, direct observation and analyses. Thus, whether a stable signaling gradient of WNT and NODAL activities underlies formation of the three germ layers remains unknown. In this study, we utilize our previously developed model of human gastrulation (2D human gastruloids) where circular colonies of human embryonic stem cells (hESCs), treated with BMP4 ligands, self-organize to form a radial pattern of the three germ layers surrounded by extra-embryonic cells at the colony edge; to measure signaling activities of WNT and NODAL. Our data shows that BMP signaling initiates a wave of WNT signaling that initiates a wave of NODAL signaling. Both WNT and NODAL signaling activities spread towards the colony center at a constant rate. Using a simple mathematical model, we show that this spreading behavior is inconsistent with a reaction- diffusion–based Turing system, indicating that there is no stable signaling gradient of WNT/NODAL signaling activity. Instead, the final signaling state is homogeneous, and spatial differences arise only from boundary effects. We further show that although both WNT and NODAL signaling synergize to enable mesodermal differentiation, neither of them forms a spatial pattern that maps directly to the mesodermal region, suggesting that mesoderm differentiation is controlled by the dynamics of multiple signals. In contrast to this, extra-embryonic differentiation at the colony edge is controlled by the duration of BMP signaling, which is consistently high in extra-embryonic cells. The identity of these extra-embryonic cells has been controversial, with some studies suggesting a trophoblast cell fate and others suggesting an extra-embryonic mesodermal fate. Using RNA- sequencing, we show that these cells are transcriptionally similar to trophoblast cells in the day 7 human embryo. Taken together, our results show that that the dynamics of signaling events in the BMP, WNT, and NODAL cascade in the absence of a stable signaling gradient, instruct germ layer fate patterning in human gastruloids.
Dissecting the dynamics of signaling events in the BMP, WNT, and NODAL cascade during self-organized fate patterning in human gastruloids
(Public Library of Science, 2019) Chhabra, Sapna; Liu, Lizhong; Goh, Ryan; Kong, Xiangyu; Warmflash, Aryeh
microfiber mesh function to maintain scaffold cellularity under serum-free conditions as well as aid the deposition of GAGs. This supports the hypothesis that scaffolds with constituents more closely resembling native ECM components may be beneficial for cartilage regeneration.
Nodal is a short-range morphogen with activity that spreads through a relay mechanism in human gastruloids
(Springer Nature, 2022) Liu, Lizhong; Nemashkalo, Anastasiia; Rezende, Luisa; Jung, Ji Yoon; Chhabra, Sapna; Guerra, M. Cecilia; Heemskerk, Idse; Warmflash, Aryeh
Morphogens are signaling molecules that convey positional information and dictate cell fates during development. Although ectopic expression in model organisms suggests that morphogen gradients form through diffusion, little is known about how morphogen gradients are created and interpreted during mammalian embryogenesis due to the combined difficulties of measuring endogenous morphogen levels and observing development in utero. Here we take advantage of a human gastruloid model to visualize endogenous Nodal protein in living cells, during specification of germ layers. We show that Nodal is extremely short range so that Nodal protein is limited to the immediate neighborhood of source cells. Nodal activity spreads through a relay mechanism in which Nodal production induces neighboring cells to transcribe Nodal. We further show that the Nodal inhibitor Lefty, while biochemically capable of long-range diffusion, also acts locally to control the timing of Nodal spread and therefore of mesoderm differentiation during patterning. Our study establishes a paradigm for tissue patterning by an activator-inhibitor pair.
Rapid changes in morphogen concentration control self-organized patterning in human embryonic stem cells
(eLife Sciences Publications, Ltd, 2019) Heemskerk, Idse; Burt, Kari; Miller, Matthew; Chhabra, Sapna; Guerra, M.Cecilia; Liu, Lizhong; Warmflash, Aryeh
During embryonic development, diffusible signaling molecules called morphogens are thought to determine cell fates in a concentration-dependent way. Yet, in mammalian embryos, concentrations change rapidly compared to the time for making cell fate decisions. Here, we use human embryonic stem cells (hESCs) to address how changing morphogen levels influence differentiation, focusing on how BMP4 and Nodal signaling govern the cell-fate decisions associated with gastrulation. We show that BMP4 response is concentration dependent, but that expression of many Nodal targets depends on rate of concentration change. Moreover, in a self-organized stem cell model for human gastrulation, expression of these genes follows rapid changes in endogenous Nodal signaling. Our study shows a striking contrast between the specific ways ligand dynamics are interpreted by two closely related signaling pathways, highlighting both the subtlety and importance of morphogen dynamics for understanding mammalian embryogenesis and designing optimized protocols for directed stem cell differentiation.

Browsing by Author "Chhabra, Sapna"

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