Embryonic Stem Cell-Derived Pitx3-Enhanced Green Fluorescent Protein Midbrain Dopamine Neurons Survive Enrichment by Fluorescence-Activated Cell Sorting and Function in an Animal Model of Parkinson's Disease

Hedlund, Eva; Pruszak, Jan; Lardaro, Thomas; Ludwig, Wesley W.; Viñuela, Angel; Kim, Kwangsoo; Isacson, Ole

doi:10.1634/stemcells.2007-0996

Cited by 131 publications

(111 citation statements)

References 58 publications

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“…This finding supports and extends previous work in this area, in which fluorescent transgenes have been used to isolate mDA progenitors that can form functional mDA neurons after transplantation (2,3,5,6). Although the concept of targeting surface antigens is not in itself new, its practical development has been limited by a lack of knowledge of transmembrane proteins with some degree of selectivity for mDA neurons.…”

Section: Discussionsupporting

confidence: 86%

“…Recent studies have used genetic labeling approaches to demonstrate that such an approach is feasible using fetal ventral mesencephalon (VM) tissue from transgenic mice (2,3) as well as differentiated embryonic stem cell reporter lines (4)(5)(6). Fluorescent protein expression driven by regulatory elements for genes involved in the embryonic development of mDA neurons, including neurogenin2 (Ngn2), Nr4a2 (Nurr1), and Pitx3, allows for isolation of cells within these gene linages through fluorescent activated cell sorting (FACS) and a significant enrichment of mDA neurons in the resulting grafts.…”

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confidence: 99%

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Transcriptome analysis reveals transmembrane targets on transplantable midbrain dopamine progenitors

Bye

Jönsson

Björklund

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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An important challenge for the continued development of cell therapy for Parkinson's disease (PD) is the establishment of procedures that better standardize cell preparations for use in transplantation. Although cell sorting has been an anticipated strategy, its application has been limited by lack of knowledge regarding transmembrane proteins that can be used to target and isolate progenitors for midbrain dopamine (mDA) neurons. We used a "FACS-array" approach to identify 18 genes for transmembrane proteins with high expression in mDA progenitors and describe the utility of four of these targets (Alcam, Chl1, Gfra1, and Igsf8) for isolating mDA progenitors from rat primary ventral mesencephalon through flow cytometry. Alcam and Chl1 facilitated a significant enrichment of mDA neurons following transplantation, while targeting of Gfra1 allowed for robust separation of dopamine and serotonin neurons. Importantly, we also show that mDA progenitors isolated on the basis of transmembrane proteins are capable of extensive, functional innervation of the host striatum and correction of motor impairment in a unilateral model of PD. These results are highly relevant for current efforts to establish safe and effective stem cell-based procedures for PD, where clinical translation will almost certainly require safety and standardization measures in order to deliver well-characterized cell preparations.Parkinson's disease | cell sorting | Alcam | microarray | transplantation

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Section: Discussionsupporting

confidence: 86%

mentioning

confidence: 99%

Transcriptome analysis reveals transmembrane targets on transplantable midbrain dopamine progenitors

Bye

Jönsson

Björklund

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Toward this goal, a straightforward approach is to direct in vitro differentiation to a specific cell lineage using physiological signaling molecules and/or key transcription factors. This strategy has been successfully used for the efficient generation of several neuronal cells, such as dopamine and motor neurons (22)(23)(24)(25)(26)(27). However, this strategy has limitations such as nonexpandability, high vulnerability, and/or heterogeneity of the differentiated neuronal cells.…”

Section: Discussionmentioning

confidence: 99%

ES cell-derived renewable and functional midbrain dopaminergic progenitors

Chung

Moon

Leung

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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During early development, midbrain dopaminergic (mDA) neuronal progenitors (NPs) arise from the ventral mesencephalic area by the combined actions of secreted factors and their downstream transcription factors. These mDA NPs proliferate, migrate to their final destinations, and develop into mature mDA neurons in the substantia nigra and the ventral tegmental area. Here, we show that such authentic mDA NPs can be efficiently isolated from differentiated ES cells (ESCs) using a FACS method combining two markers, Otx2 and Corin. Purified Otx2 + Corin + cells coexpressed other mDA NP markers, including FoxA2, Lmx1b, and Glast. Using optimized culture conditions, these mDA NPs continuously proliferated up to 4 wk with almost 1,000-fold expansion without significant changes in their phenotype. Furthermore, upon differentiation, Otx2 + Corin + cells efficiently generated mDA neurons, as evidenced by coexpression of mDA neuronal markers (e.g., TH, Pitx3, Nurr1, and Lmx1b) and physiological functions (e.g., efficient DA secretion and uptake). Notably, these mDA NPs differentiated into a relatively homogenous DA population with few serotonergic neurons. When transplanted into PD model animals, aphakia mice, and 6-OHDA-lesioned rats, mDA NPs differentiated into mDA neurons in vivo and generated well-integrated DA grafts, resulting in significant improvement in motor dysfunctions without tumor formation. Furthermore, grafted Otx2 + Corin + cells exhibited significant migratory function in the host striatum, reaching >3.3 mm length in the entire striatum. We propose that functional and expandable mDA NPs can be efficiently isolated by this unique strategy and will serve as useful tools in regenerative medicine, bioassay, and drug screening.neural precursors | dopaminergic neurons | transplantation

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“…A reasonable future focus is testing the functional outcome of various cell transplantation innovations, including the use of PD patient individualized induced pluripotent stem (iPS) cells as a source for donor cells. Such new technology could possibly improve both transplant quality and content, by providing cell sorted pure DA populations [13,47] compared to elective abortion dissected, mixed fetal tissue including tissue with bloodvessel donor antigens and inflammatory reactions [21]. In particular, breakthroughs in the use of human stem cells are necessary to provide a reliable cell source for a large-scale clinical application.…”

Section: Cell Preparation Techniques and Patient Recipient Status Maymentioning

confidence: 99%

Lack of functional relevance of isolated cell damage in transplants of Parkinson’s disease patients

et al. 2009

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Postmortem analyses from clinical neural transplantation trials of several subjects with Parkinson's disease revealed surviving grafted dopaminergic neurons after more than a decade. A subset of these subjects displayed isolated dopaminergic neurons within the grafts that contained Lewy body-like structures. In this review, we discuss why this isolated cell damage is unlikely to affect the overall graft function and how we can use these observations to help us to understand age-related neurodegeneration and refine our future cell replacement therapies.

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Embryonic Stem Cell-Derived Pitx3-Enhanced Green Fluorescent Protein Midbrain Dopamine Neurons Survive Enrichment by Fluorescence-Activated Cell Sorting and Function in an Animal Model of Parkinson's Disease

Cited by 131 publications

References 58 publications

Transcriptome analysis reveals transmembrane targets on transplantable midbrain dopamine progenitors

Transcriptome analysis reveals transmembrane targets on transplantable midbrain dopamine progenitors

ES cell-derived renewable and functional midbrain dopaminergic progenitors

Lack of functional relevance of isolated cell damage in transplants of Parkinson’s disease patients

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