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The bulk cell density as decisive parameter and simple method for mesendodermal patterning of human pluripotent stem cells

ABSTRACT: We showed that the relation of pluripotent stem cell count to culture volume influences cardiomyogenic differentiation by affecting the concentration of secreted proteins that function as paracrine factors

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stem Cell

SUBMITTER: Falk Büttner

LAB HEAD: Rita Gerardy-Schahn

PROVIDER: PXD004874 | Pride | 2018-10-17

REPOSITORIES: Pride

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			Action	DRS
	HK01_1ml_06h_A.raw	Raw
	HK01_1ml_06h_C.raw	Raw
	HK01_1ml_06h_D.raw	Raw
	HK02_1ml_24h_A.raw	Raw
	HK02_1ml_24h_C.raw	Raw

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Publications

Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells.

Kempf Henning H Olmer Ruth R Haase Alexandra A Franke Annika A Bolesani Emiliano E Schwanke Kristin K Robles-Diaz Diana D Coffee Michelle M Göhring Gudrun G Dräger Gerald G Pötz Oliver O Joos Thomas T Martinez-Hackert Erik E Haverich Axel A Buettner Falk F R FFR Martin Ulrich U Zweigerdt Robert R

Nature communications 20161209

In vitro differentiation of human pluripotent stem cells (hPSCs) recapitulates early aspects of human embryogenesis, but the underlying processes are poorly understood and controlled. Here we show that modulating the bulk cell density (BCD: cell number per culture volume) deterministically alters anteroposterior patterning of primitive streak (PS)-like priming. The BCD in conjunction with the chemical WNT pathway activator CHIR99021 results in distinct paracrine microenvironments codifying hPSCs ...[more]

PMID: 27934856

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Modulation of the bulk cell density as decisive parameter and simple method for mesendodermal patterning of human pluripotent stem cells

Project description:1 million human pluripotent cells cultured as suspension aggregates (hES3-NKX2.5 cell line) were treated with 0, 7.5 or 15µM CHIR99021 in 1 or 3 ml RPMI/B27 (w/o insulin) for 24h to induce the differentiation of human pluripotent stem cells (hPSCs). CHIR99021 is a potent GSK3 inhibitor that results in prominent WNT pathway activation and thereby induces mesendodermal differentiation.

2016-12-30 | E-MTAB-5051 | biostudies-arrayexpress

LINE-1 DNA methylation levels in DPPA3 KO overexpressing DPPA3 mutants

Project description:DPPA3 mutants were overexpressed using a doxycycline system to identify regions in DPPA3 necessary for DNA methylation maintenance suppression

2020-10-22 | E-MTAB-9653 | biostudies-arrayexpress

Amplicon bisulfite sequencing of LINE-1 elements in TET catalytic mutants and DPPA3 KO ESCs rescued by doxycylin induced overexpression

Project description:Tet1, Tet2 and Tet1/Tet2 catalytic mutants as well as DPPA3 KO ESCs harboring a doxycyclin inducible Dppa3 transgene were induced for several days to monitor LINE-1 methylation levels.

2020-10-23 | E-MTAB-9654 | biostudies-arrayexpress

Flutamide treated rat hepatocytes in microfluidic biochips

Project description:We investigated the effects of the flutamide (FLU) -induced liver injury in primary rat hepatocytes using our liver microfluidic biochips. Flutamide is used a non-steroidal anti-androgenic drug. Two flutamide concentrations, 10M-BM-5M and 100M-BM-5M, were used to expose the hepatocytes for 24h under perfusion. Primary rat hepatocytes were cultivated in microfluidic biochips and treated with 10 and 100M-BM-5M of flutamide for 24h

2013-12-01 | E-GEOD-44317 | biostudies-arrayexpress

Bulk mRNA Seq CAR.CCR7 vs. CAR

Project description:The study aimed to elucidate the transcriptional changes introduced by genetic engineering of CAR T cells to stably express the homeostatic chemokine receptor CCR7. In vitro killing assays hinted to a higher cytolytic capacity that could be tied to an altered cytoskeletal rearrangement. Human CAR and CAR.CCR7 T cells were generated by retroviral transduction and enriched by FACS at the end of the expansion phase (day 10), either without or with a previous over-night co-culture with target Jeko1 tumor cells that express the CAR target antigen CXCR5. Enriched cells were immediately processed for bulk mRNA Seq

2023-06-30 | E-MTAB-12778 | biostudies-arrayexpress

A Modular Bioinstructive Platform For Additive-Free Control of Stem Cell Differentiation and Patterning

Project description:Recreating 3D bone formation in vitro without biochemical inducers remains a longstanding challenge in preclinical testing. We present a modular, bioinstructive platform based on polylactic acid microparticles with controlled dimpled surface features that direct mesenchymal stem cell differentiation through endogenous mechanotransduction, establishing a mechanistically validated, additive-free platform. These dimpled 3D topographical cues drive cytoskeletal reorganisation and induce osteogenesis via canonical Hedgehog signalling. RNA-Seq revealed early significant upregulation of cytoskeletal components and osteochondral transcription factors, including runt-related transcription factor 2 (RUNX2) and SRY-box transcription factor 9 (SOX9), followed by activation of the insulin growth factor-II pathway and osteogenic commitment. To demonstrate functional utility, two-photon polymerisation lithography was employed to engineer precisely-patterned 3D topographies, inducing graded GLI1 expression without added soluble cues. This establishes a scalable, versatile platform for stem cell engineering, offering a non-genetic, topography-driven analogue to mechanogenetics with transformative potential for mechanobiology research and development of human-relevant bone models.

2026-07-01 | E-MTAB-15393 | biostudies-arrayexpress

Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells.

Project description: Not available

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BIGenomics_Agilent_15k_templates

Project description:Hybridization of ligation probes to a zipcode (tag) microarray. The purpose of the experiment was to establish functionality and target specificity of the ligation probes by using pools of synthetic templates with all of the probes. The templates are 80-mer synthetic dsDNA molecules, each specific for a single probe. Each probe is a ssDNA molecule containing target recognition sequences, PCR primer binding sequences and a unique tag sequence. If a matching target sequence is present in the mixture, the probes is ligated into a circular molecule, which can be PCR amplified and hybrized on a microarray harbouring complementary tag sequences.

2012-02-24 | E-MEXP-3538 | biostudies-arrayexpress

Pre-patterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B

Project description:Dynamic interactions of nuclear lamins with chromatin through so-called lamin-associated domains (LADs) contribute to spatial arrangements of the genome. Here, we provide evidence for pre-patterning of differentiation-driven formation of lamin A/C LADs by domains of histone H2B modified by the nutrient sensor O-linked N-acetylglucosamine (H2BGlcNAc), which we term GADs. We demonstrate a two-step process of lamin A/C LAD formation during in vitro adipogenesis, involving (i) a spreading of lamin A/C-chromatin interactions during the transition from progenitor cell proliferation to cell cycle arrest, and (ii) a genome-scale redistribution these interactions through a process of LAD ‘exchange’ within hours of adipogenic induction. Chromatin state modeling reveals that lamin A/C LADs can be found both in active and repressive chromatin contexts which can be influenced by cell differentiation status. De novo formation of adipogenic lamin A/C LADs occurs non-randomly on GADs, consisting of megabase-size intergenic and repressive chromatin domains. Accordingly, while pre-differentiation lamin A/C LADs are gene-rich, post-differentiation LADs harbor repressive features reminiscent of lamin B1 LADs. Moreover, release of lamin A/C from genes directly involved in glycolysis concurs with their transcriptional upregulation after adipogenic induction, and with concordant downstream elevations in H2BGlcNAc levels and O-GlcNAc cycling. Our results unveil an epigenetic pre-patterning of adipogenic LADs by GADs, suggesting a coupling of developmentally regulated lamin A/C-genome interactions to a metabolically sensitive chromatin modification. Examination of LMNA and H2BGlcNAc binding in ASCs across differentiation

2015-09-04 | E-GEOD-63346 | biostudies-arrayexpress

Multiscale Anisotropic Tissue Biofabrication via Bulk Acoustic Patterning of Cells and Functional Additives in Hybrid Bioinks.

Project description: Not available

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