Gene expression analysis of midbrain dopamine progenitors in embryonic mouse
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE65094 | GEO | 2015/02/28
SECONDARY ACCESSION(S): PRJNA272970
REPOSITORIES: GEO
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Project description:To improve the standardization of cell therapies for Parkinson’s disease, methods for the selection and isolation of midbrain dopaminergic progenitors for transplantation are required. To facilitate this we established an expression profile for genes selectively expressed on transplantable midbrain dopaminergic progenitors using microarray analysis. Expression of GFP in the ventral mesencephalon of embryonic E12.5 Ngn2-GFP mice identifies a distinct sub-population of cells containing virtually all of the midbrain dopaminergic progenitors. Gene expression profiles from 3 biological replicates of FACS isolated GFP-positive cells from mouse Ngn2-GFP ventral mesencephalon were generated using microarrays. To reduce the likelihood of identifying transcripts from non-dopaminergic progenitors, 3 biological replicates of FACS isolated GFP-negative cells from mouse Lmx1a-GFP ventral mesencephalon (definitively non-dopaminergic) were used as a reference population.
2015-02-28 | E-GEOD-65094 | biostudies-arrayexpress
Project description:Despite the progress in safety and efficacy of cell therapy with pluripotent stem cells (PSCs), the presence of residual undifferentiated stem cells or proliferating neural progenitor cells (NPCs) with rostral identity has remained a major challenge. Here we reported the generation of an LMX1A knock-in GFP reporter human embryonic stem cell (hESC) line that marks the early dopaminergic progenitors during neural differentiation. Purified GFP positive cells in vitro exhibited expression of mRNA and proteins that characterized and matched the midbrain dopaminergic identity. Further proteomic analysis of enriched LMX1A+ cells identified several membrane associated proteins including CNTN2, enabling prospective isolation of LMX1A+ progenitor cells. Transplantation of hPSC-derived purified CNTN2+ progenitors enhanced dopamine release from transplanted cells in the host brain and alleviated Parkinson’s disease symptoms in animal models. Our study establishes an efficient approach for purification of large numbers of hPSC-derived dopaminergic progenitors for therapeutic applications.
2018-06-06 | PXD007837 | Pride
Project description:Embryonic stem (ES) cells were differentiated in culture to midbrain dopaminergic (mDA) progenitors and subjected to ChIP-seq analysis to resolve genome-wide binding sites of forkhead box protein A2 (Foxa2). Foxa2 was found to directly regulate multiple lineage pathways to specify midbrain dopaminergic and floor plate progenitor identity.
2012-07-01 | E-MTAB-1137 | biostudies-arrayexpress
Project description:Current ventral midbrain dopaminergic progenitor differentiation protocols utilize small molecule inhibitors targeting Glycogen synthase kinase-3 (GSK3) to activate Wnt signaling, a step required for the anterior-posterior patterning of the nervous system and acquisition of the midbrain fate. However, GSK3a and GSK3β are pleiotropic kinases involved in multiple signaling pathways and their inhibition is a known trigger of neurogenesis. We predicted that direct activation of specific Wnt receptors naturally involved during neural patterning would allow for a more precise spatiotemporal control of Wnt/β-catenin signaling and mimic endogenous cell-cell communication mechanisms to improve hPSC differentiation. Here, we characterized the expression of FZD receptors at the surface of neural progenitor cells with different regional identity. Our data shows that FZD5 expression is uniquely upregulated in anterior neural progenitors and is rapidly downregulated as cells adopt a posterior fate. This spatial regulation of Frizzled cell surface expression constitutes a novel regulatory mechanism adjusting the levels of β-catenin signaling along the anterior-posterior axis and possibly contribute to midbrain-hindbrain boundary formation. Using a tetravalent antibody that selectively triggers FZD5 and LRP6 clustering to activate Wnt/β-catenin signaling, we show that the resulting midbrain-patterned neural progenitors exhibit a gene expression program more closely aligned with the anatomical origin of dopaminergic neurons and could hence represent a more efficient source for cell transplant therapies.
2024-02-21 | GSE241042 | GEO
Project description:Current ventral midbrain dopaminergic progenitor differentiation protocols utilize small molecule inhibitors targeting Glycogen synthase kinase-3 (GSK3) to activate Wnt signaling, a step required for the anterior-posterior patterning of the nervous system and acquisition of the midbrain fate. However, GSK3a and GSK3β are pleiotropic kinases involved in multiple signaling pathways and their inhibition is a known trigger of neurogenesis. We predicted that direct activation of specific Wnt receptors naturally involved during neural patterning would allow for a more precise spatiotemporal control of Wnt/β-catenin signaling and mimic endogenous cell-cell communication mechanisms to improve hPSC differentiation. Here, we characterized the expression of FZD receptors at the surface of neural progenitor cells with different regional identity. Our data shows that FZD5 expression is uniquely upregulated in anterior neural progenitors and is rapidly downregulated as cells adopt a posterior fate. This spatial regulation of Frizzled cell surface expression constitutes a novel regulatory mechanism adjusting the levels of β-catenin signaling along the anterior-posterior axis and possibly contribute to midbrain-hindbrain boundary formation. Using a tetravalent antibody that selectively triggers FZD5 and LRP6 clustering to activate Wnt/β-catenin signaling, we show that the resulting midbrain-patterned neural progenitors exhibit a gene expression program more closely aligned with the anatomical origin of dopaminergic neurons and could hence represent a more efficient source for cell transplant therapies.
2024-02-21 | GSE241041 | GEO
Project description:Ventral midbrain (VM) dopaminergic progenitor cells derived from human pluripotent stem cells have the potential to replace endogenously lost dopamine neurons and are currently in preclinical and clinical development for treatment of Parkinson’s Disease (PD). However, one main challenge in the quality control of the cells is that rostral and caudal VM progenitors are extremely similar transcriptionally though only the caudal VM cells give rise to dopaminergic neurons with functionality in PD. Therefore, it is critical to develop assays which can rapidly and reliably discriminate rostral from caudal VM cells during clinical manufacturing. Here, we applied shotgun proteomics to search for novel secreted biomarkers specific for caudal VM progenitors compared to rostral VM progenitors and validated key hits by ELISA. From this, we identified novel secreted markers (CPE, LGI1 and PDGFC) significantly enriched in caudal versus rostral VM progenitor cultures, whereas the markers CNTN2 and CORIN were significantly enriched in rostral VM cultures. With this data, we suggest and test in clinical grade samples a panel of coupled ELISA assays that can be applied as a quality control tool for assessing the correct patterning of cells during clinical manufacturing.
2023-12-13 | PXD039510 | Pride
Project description:Human induced pluripotent stem cells (iPSCs) can provide a promising source of midbrain dopaminergic (DA) neurons for cell replacement therapy for Parkinson’s disease. However, iPSC-derived donor cells may inevitably contain tumorigenic or inappropriate cells. Purification of neural progenitor cells or DA neurons as suitable donor cells has been attempted, but the isolation of DA progenitor cells derived from human pluripotent stem cells has so far been unsuccessful. Here we show human iPSC-derived DA progenitor cells can be efficiently isolated by cell sorting using a floor plate marker, Corin. we were able to develop a method for 1) scalable DA neuron induction on human laminin fragment and 2) sorting DA progenitor cells using an anti-Corin antibody. Furthermore, we determined the optimal timing for the cell sorting and transplantation. The grafted cells survived well and functioned as midbrain DA neurons in the 6-OHDA-lesioned rats, and showed minimal risk of tumor formation. The sorting of Corin-positive cells is favorable in terms of both safety and efficiency, and our protocol will contribute to the clinical application of human iPSCs for Parkinson’s disease. Differentiated human iPSC-derived neural progenitors just after sorting (day12 unsorted, day12 Corin+) and dopaminergic progenitors after an aggregation culture (day28 and day42, unsorted and day12-sorted, respectively), and human fetal ventral mesencephalon and dorsal mesencephalon (gestational age of 7.5 weeks) were subjected to RNA extraction and hybrdization on Affymetrix microarrays. Each sample except for human mesencephalon, undifferentiated iPSC, and day12-unsorted, day42-sample has 3 or 4 repeats.
2014-03-07 | E-GEOD-51214 | biostudies-arrayexpress
Project description:Parkinson’s disease, an aging-associated neurodegenerative disorder, is characterised by nigrostriatal pathway dysfunction caused by the gradual loss of dopaminergic neurons in the substantia nigra pars compacta of the midbrain. Human in vitro models are enabling the study of the dopaminergic neurons’ loss, but not the dysregulation within the dopaminergic network in the nigrostriatal pathway. Additionally, these models do not incorporate aging characteristics which potentially contribute to the development of Parkinson’s disease. Here we present a nigrostriatal pathway model based on midbrain-striatum assembloids with inducible aging. We show that these assembloids can develop characteristics of the nigrostriatal connectivity, with catecholamine release from the midbrain to the striatum and synapse formation between midbrain and striatal neurons. Moreover, Progerin-overexpressing assembloids acquire aging traits that lead to early neurodegenerative phenotypes. This model shall help to reveal the contribution of aging as well as nigrostriatal connectivity to the onset and progression of Parkinson’s disease.
2023-10-28 | GSE236458 | GEO
Project description:Parkinson’s disease, an aging-associated neurodegenerative disorder, is characterised by nigrostriatal pathway dysfunction caused by the gradual loss of dopaminergic neurons in the substantia nigra pars compacta of the midbrain. Human in vitro models are enabling the study of the dopaminergic neurons’ loss, but not the dysregulation within the dopaminergic network in the nigrostriatal pathway. Additionally, these models do not incorporate aging characteristics which potentially contribute to the development of Parkinson’s disease. Here we present a nigrostriatal pathway model based on midbrain-striatum assembloids with inducible aging. We show that these assembloids can develop characteristics of the nigrostriatal connectivity, with catecholamine release from the midbrain to the striatum and synapse formation between midbrain and striatal neurons. Moreover, Progerin-overexpressing assembloids acquire aging traits that lead to early neurodegenerative phenotypes. This model shall help to reveal the contribution of aging as well as nigrostriatal connectivity to the onset and progression of Parkinson’s disease.
2023-10-28 | GSE241631 | GEO
Project description:Advances in stem cell technologies open up new avenues for modelling development and diseases. The success of these pursuits however rely on the use of cells most relevant to those targeted by the disease of interest, for example, midbrain dopaminergic neurons for Parkinson’s disease. In the present study, we report the generation of a human induced pluripotent stem cell (iPSC) line capable of purifying and tracing nascent midbrain dopaminergic progenitors and their differentiated progeny via the expression of a Blue Fluorescent Protein (BFP). This was achieved by CRISPR/Cas9 assisted knock-in of BFP and Cre into the safe harbour locus AAVS1 and an early midbrain dopaminergic lineage marker gene LMX1A, respectively. Immunocytochemical analysis and single cell RNA sequencing of iPSC-derived neural cultures confirms developmental recapitulation of the human fetal midbrain and high quality midbrain cells. By modelling Parkinson’s disease-related drug toxicity using 1-Methyl-4-phenylpyridinium (MPP+), we showed preferential reduction of BFP+ cells, a finding demonstrated independently by cell death assays and single cell transcriptomic analysis of MPP+ treated neural cultures. Together, these results highlight the importance of disease relevant cell type in stem cell modelling.
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