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- Elabi, Osama F, et al.
(författare)
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Human Embryonic Stem Cell-Derived Dopaminergic Grafts Alleviate L-DOPA Induced Dyskinesia
- 2022
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Ingår i: Journal of Parkinson's Disease. - 1877-718X. ; 12:6, s. 1881-1896
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: First-in-human studies to test the efficacy and safety of human embryonic stem cells (hESC)-derived dopaminergic cells in the treatment of Parkinson's disease (PD) are imminent. Pre-clinical studies using hESC-derived dopamine neuron transplants in rat models have indicated that the benefits parallel those shown with fetal tissue but have thus far failed to consider how ongoing L-DOPA administration might impact on the graft.OBJECTIVE: To determine whether L-DOPA impacts on survival and functional recovery following grafting of hESC-derived dopaminergic neurons.METHODS: Unilateral 6-OHDA lesioned rats were administered with either saline or L-DOPA prior to, and for 18 weeks following surgical implantation of dopaminergic neural progenitors derived from RC17 hESCs according to two distinct protocols in independent laboratories.RESULTS: Grafts from both protocols elicited reduction in amphetamine-induced rotations. Reduced L-DOPA-induced dyskinesia preceded the improvement in amphetamine-induced rotations. Furthermore, L-DOPA had no effect on overall survival (HuNu) or dopaminergic neuron content of the graft (TH positive cells) but did lead to an increase in the number of GIRK2 positive neurons.CONCLUSION: Critically, we found that L-DOPA was not detrimental to graft function, potentially enhancing graft maturation and promoting an A9 phenotype. Early improvement of L-DOPA-induced dyskinesia suggests that grafts may support the handling of exogenously supplied dopamine earlier than improvements in amphetamine-induced behaviours indicate. Given that one of the protocols will be employed in the production of cells for the European STEM-PD clinical trial, this is vital information for the management of patients and achieving optimal outcomes following transplantation of hESC-derived grafts for PD.
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| 2. |
- Holmqvist, Staffan, et al.
(författare)
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Generation of human pluripotent stem cell reporter lines for the isolation of and reporting on astrocytes generated from ventral midbrain and ventral spinal cord neural progenitors.
- 2015
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Ingår i: Stem Cell Research. - : Elsevier BV. - 1876-7753 .- 1873-5061. ; 15:1, s. 203-220
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Tidskriftsartikel (refereegranskat)abstract
- Astrocytes play a critical role during the development and the maintenance of the CNS in health and disease. Yet, their lack of accessibility from fetuses and from the brain of diseased patients has hindered our understanding of their full implication in developmental and pathogenic processes. Human pluripotent stem cells (PSCs) are an alternative source to obtain large quantities of astrocytes in vitro, for mechanistic studies of development and disease. However, these studies often require highly pure populations of astrocytes, which are not always achieved, depending on the PSC lines and protocols used. Here, we describe the generation and characterization of human PSC reporter lines expressing TagRFP driven by the ABC1D region of the human GFAP promoter, as new cellular model for generating homogenous population of astrocytes generated from CNS regionally defined PSC-derived neural progenitors. GFAABC1D::TagRFP-expressing astrocytes can be purified by fluorescent-activated cell sorting and maintain a bright expression for several additional weeks. These express canonical astrocyte markers NF1A, S100β, CX43, GLAST, GS and CD44. These new cellular models, from which highly pure populations of fluorescence-expressing astrocytes can be obtained, provide a new platform for studies where pure or fluorescently labeled astrocyte populations are necessary, for example to assess pro-inflammatory cytokine and chemokine release in response to specific treatment, and uptake and degradation of fluorescently labeled pathogenic proteins, as reported in this study.
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| 3. |
- Reyes, Juan F, et al.
(författare)
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A cell culture model for monitoring α-synuclein cell-to-cell transfer.
- 2015
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Ingår i: Neurobiology of Disease. - : Elsevier BV. - 0969-9961. ; 77:Jul 16, s. 266-275
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Tidskriftsartikel (refereegranskat)abstract
- The transfer of α-synuclein (α-syn) between cells has been proposed to be the primary mechanism of disease spreading in Parkinson's disease. Several cellular models exist that monitor the uptake of recombinant α-syn from the culture medium. Here we established a more physiologically relevant model system in which α-syn is produced and transferred between mammalian neurons. We generated cell lines expressing either α-syn tagged with fluorescent proteins or fluorescent tags alone then we co-cultured these cell lines to measure protein uptake. We used live-cell imaging to demonstrate intercellular α-syn transfer and used flow cytometry and high content analysis to quantify the transfer. We then successfully inhibited intercellular protein transfer genetically by down-regulating dynamin or pharmacologically using dynasore or heparin. In addition, we differentiated human induced pluripotent stem cells carrying a triplication of the α-syn gene into dopaminergic neurons. These cells secreted high levels of α-syn, which was taken up by neighboring neurons. Collectively, our co-culture systems provide simple but physiologically relevant tools for the identification of genetic modifiers or small molecules that inhibit α-syn cell-to-cell transfer.
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