scholarly journals NODAL/TGFβ signalling mediates the self-sustained stemness induced by PIK3CAH1047R homozygosity in pluripotent stem cells

2019 ◽  
Author(s):  
Ralitsa R. Madsen ◽  
James Longden ◽  
Rachel G. Knox ◽  
Xavier Robin ◽  
Franziska Völlmy ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Human Cancer ◽  
Human Pluripotent Stem Cells ◽  
Depth Analysis ◽  
Pi3k Activation ◽  
Tgfβ Signalling ◽  
Pi3k Signalling ◽  
Pathway Inhibition ◽  
Dose Dependent

AbstractActivating PIK3CA mutations are known “drivers” of human cancer and developmental overgrowth syndromes. We recently demonstrated that the “hotspot” PIK3CAH1047R variant exerts unexpected allele dose-dependent effects on stemness in human pluripotent stem cells (hPSCs). In the present study, we combine high-depth transcriptomics, total proteomics and reverse-phase protein arrays to reveal potentially disease-related alterations in heterozygous cells, and to assess the contribution of activated TGFβ signalling to the stemness phenotype of PIK3CAH1047R homozygous cells. We demonstrate signalling rewiring as a function of oncogenic PI3K signalling dose, and provide experimental evidence that self-sustained stemness is causally related to enhanced autocrine NODAL/TGFβ signalling. A significant transcriptomic signature of TGFβ pathway activation in PIK3CAH1047R heterozygous was observed but was modest and was not associated with the stemness phenotype seen in homozygous mutants. Notably, the stemness gene expression in PIK3CAH1047R homozygous iPSCs was reversed by pharmacological inhibition of TGFβ signalling, but not by pharmacological PI3Kα pathway inhibition. Altogether, this provides the first in-depth analysis of PI3K signalling in human pluripotent stem cells and directly links dose-dependent PI3K activation to developmental NODAL/TGFβ signalling.

scholarly journals NODAL/TGFβ signalling mediates the self-sustained stemness induced by PIK3CAH1047R homozygosity in pluripotent stem cells

2021 ◽  
pp. dmm.048298
Author(s):  
Ralitsa R. Madsen ◽  
James Longden ◽  
Rachel G. Knox ◽  
Xavier Robin ◽  
Franziska Völlmy ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Human Cancer ◽  
Human Pluripotent Stem Cells ◽  
Human Genetic Disease ◽  
Allele Dosage ◽  
Depth Analysis ◽  
Tgfβ Signalling ◽  
Pi3k Signalling ◽  
Pathway Inhibition

Activating PIK3CA mutations are known “drivers” of human cancer and developmental overgrowth syndromes. We recently demonstrated that the "hotspot" PIK3CAH1047R variant exerts unexpected allele dose-dependent effects on stemness in human pluripotent stem cells (hPSCs). In the present study, we combine high-depth transcriptomics, total proteomics and reverse-phase protein arrays to reveal potentially disease-related alterations in heterozygous cells, and to assess the contribution of activated TGFβ signalling to the stemness phenotype of homozygous PIK3CAH1047R cells. We demonstrate signalling rewiring as a function of oncogenic PI3K signalling strength, and provide experimental evidence that self-sustained stemness is causally related to enhanced autocrine NODAL/TGFβ signalling. A significant transcriptomic signature of TGFβ pathway activation in heterozygous PIK3CAH1047R was observed but was modest and was not associated with the stemness phenotype seen in homozygous mutants. Notably, the stemness gene expression in homozygous PIK3CAH1047R iPSCs was reversed by pharmacological inhibition of NODAL/TGFβ signalling, but not by pharmacological PI3Kα pathway inhibition. Altogether, this provides the first in-depth analysis of PI3K signalling in human pluripotent stem cells and directly links strong PI3K activation to developmental NODAL/TGFβ signalling. This work illustrates the importance of allele dosage and expression when artificial systems are used to model human genetic disease caused by activating PIK3CA mutations.

scholarly journals Identification of TGFβ signalling as a regulator of interneuron neurogenesis in a human pluripotent stem cell model

2021 ◽  
Author(s):  
Maria Santos Cruz ◽  
Meng Li
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Cell Model ◽  
Neurological Diseases ◽  
Human Pluripotent Stem Cells ◽  
Tgfβ Signaling ◽  
Neuropsychiatric Diseases ◽  
Cortical Interneurons ◽  
Complex Development ◽  
Tgfβ Signalling

Cortical interneurons are GABAergic inhibitory cells that connect locally in the neocortex and play a  pivotal role in shaping cortical network activities. Dysfunction of these cells is believed to lead to runaway excitation underlying seizure-based diseases, such as epilepsy, autism, and schizophrenia. There is a growing interest in using cortical interneurons derived from human pluripotent stem cells for understanding their complex development and for modeling neuropsychiatric diseases. Here, we report the identification of a novel role of TGFβ signaling in modulating interneuron progenitor maintenance and neuronal differentiation. TGFβ signaling inhibition suppresses terminal differentiation of interneuron progenitors while exogenous TGFβ3 accelerates the transition of progenitors into postmitotic neurons. We provide evidence that TGFb signaling exerts this function via regulating cell cycle length of the NKX2.1+ neural progenitors. Together, this study represents a useful platform for studying human interneuron development and interneuron associated neurological diseases with human pluripotent stem cells.

scholarly journals A role for TGFβ signalling in medium spiny neuron differentiation of human pluripotent stem cells

2020 ◽  
Vol 4 (2) ◽  
Author(s):  
Marija Fjodorova ◽  
Zoe Noakes ◽  
Meng Li
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Activin A ◽  
Human Pluripotent Stem Cells ◽  
Medium Spiny Neuron ◽  
Fine Tuning ◽  
Fate Specification ◽  
Differentiation Protocol ◽  
Tgfβ Signalling ◽  
Bmp Signalling

Abstract Activin A and other TGFβ family members have been shown to exhibit a certain degree of promiscuity between their family of receptors. We previously developed an efficient differentiation protocol using Activin A to obtain medium spiny neurons (MSNs) from human pluripotent stem cells (hPSCs). However, the mechanism underlying Activin A-induced MSN fate specification remains largely unknown. Here we begin to tease apart the different components of TGFβ pathways involved in MSN differentiation and demonstrate that Activin A acts exclusively via ALK4/5 receptors to induce MSN progenitor fate during differentiation. Moreover, we show that Alantolactone, an indirect activator of SMAD2/3 signalling, offers an alternative approach to differentiate hPSC-derived forebrain progenitors into MSNs. Further fine tuning of TGFβ pathway by inhibiting BMP signalling with LDN193189 achieves accelerated MSN fate specification. The present study therefore establishes an essential role for TGFβ signalling in human MSN differentiation and provides a fully defined and highly adaptable small molecule-based protocol to obtain MSNs from hPSCs.

scholarly journals Interspecies Chimeric Conditions Affect the Developmental Rate of Human Pluripotent Stem Cells

2020 ◽  
Author(s):  
Jared Brown ◽  
Christopher Barry ◽  
Matthew T. Schmitz ◽  
Cara Argus ◽  
Jennifer M. Bolin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryonic Stem ◽  
Cell Types ◽  
In Utero ◽  
Human Pluripotent Stem Cells ◽  
Human Organs ◽  
Species Specific ◽  
Dose Dependent

ABSTRACTHuman pluripotent stem cells hold significant promise for regenerative medicine. However, long differentiation protocols and immature characteristics of stem cell-derived cell types remain challenges to the development of many therapeutic applications. In contrast to the slow differentiation of human stem cells in vitro that mirrors a nine-month gestation period, mouse stem cells develop according to a much faster three-week gestation timeline. Here, we tested if co-differentiation with mouse pluripotent stem cells could accelerate the differentiation speed of human embryonic stem cells. Following a six-week RNA-sequencing time course of neural differentiation, we identified 929 human genes that were upregulated earlier and 535 genes that exhibited earlier peaked expression profiles in chimeric cell cultures than in human cell cultures alone. Genes with accelerated upregulation were significantly enriched in Gene Ontology terms associated with neurogenesis, neuron differentiation and maturation, and synapse signaling. Moreover, chimeric mixed samples correlated with in utero human embryonic samples earlier than human cells alone, and acceleration was dose-dependent on human-mouse co-culture ratios. Differences in the timing and expression levels of genes corresponding to neuron cell types and brain region identity under chimeric conditions were also observed. The altered developmental rates and lineage outcomes described in this report have implications for accelerating human stem cell differentiation and the use of interspecies chimeric embryos in developing human organs for transplantation.Author SummaryHuman pluripotent stem cells often require long in vitro protocols to form mature cell types of clinical relevance for potential regenerative therapies, a ramification of a nine-month developmental clock in utero that also runs ex utero. What controls species-specific developmental time and whether the timer is amenable to acceleration is unknown. Further, interspecies chimeric embryos are increasingly being created to study early human development or explore the potential growth of human organs for transplantation. How the conflicting developmental speeds of cells from different species co-differentiating together affect each other is not understood. Here, using genome-wide transcriptional analysis of RNA-sequencing time courses, we show that 1) co-differentiating human embryonic stem cells intermixed with mouse stem cells accelerated elements of human developmental programs, 2) the acceleration was dose-dependent on the proportion of mouse cells, and 3) human cells in chimeric samples correlated to in utero samples earlier than human only samples. Our results provide evidence that some components of species-specific developmental clocks may be susceptible to acceleration.

Differentiation of human pluripotent stem cells into airway epithelial cells

Pneumologie ◽  
2015 ◽  
Vol 69 (07) ◽  
Author(s):  
S Ulrich ◽  
S Weinreich ◽  
R Haller ◽  
S Menke ◽  
R Olmer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Epithelial Cells ◽  
Pluripotent Stem Cells ◽  
Airway Epithelial Cells ◽  
Human Pluripotent Stem Cells ◽  
Airway Epithelial

326-LB: BCL-xL/BCL2L1 Is a Critical Anti-Apoptotic Protein that Suppresses BAK to Promote Pancreatic Specification from Human Pluripotent Stem Cells

Diabetes ◽  
2019 ◽  
Vol 68 (Supplement 1) ◽  
pp. 326-LB
Author(s):  
LARRY SAI WENG LOO ◽  
ADRIAN TEO ◽  
SOUMITA GHOSH ◽  
ANDREAS ALVIN PURNOMO SOETEDJO ◽  
LINH NGUYEN ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Human Pluripotent Stem Cells ◽  
Apoptotic Protein
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