| 1. |
- Azevedo, Carla, et al.
(författare)
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Generation of an induced pluripotent stem cell line (CSC-32) from a patient with Parkinson's disease carrying a heterozygous variation p.A53T in the SNCA gene
- 2020
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Ingår i: Stem Cell Research. - : Elsevier BV. - 1873-5061. ; 43
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Tidskriftsartikel (refereegranskat)abstract
- Here, we describe the generation of an induced pluripotent stem cell (iPSC) line, from a male patient diagnosed with Parkinson's disease (PD). The patient carries a heterozygous variation p.A53T in the SNCA gene. Skin fibroblasts were reprogrammed using the non-integrating Sendai virus technology to deliver OCT3/4, SOX2, c-MYC and KLF4 factors. The generated iPSC line (CSC-32) preserved the mutation, displayed expression of common pluripotency markers, differentiated into derivatives of the three germ layers, and exhibited a normal karyotype. The clone CSC-32B is presented thereafter; it can be used to study the mechanisms underlying PD pathogenesis.
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| 2. |
- Azevedo, Carla, et al.
(författare)
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Parkinsons disease and multiple system atrophy patient iPSC-derived oligodendrocytes exhibit alpha-synuclein-induced changes in maturation and immune reactive properties
- 2022
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 119:12
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Tidskriftsartikel (refereegranskat)abstract
- Limited evidence has shed light on how aSYN proteins affect the oligodendrocyte phenotype and pathogenesis in synucleinopathies that include Parkinsons disease (PD) and multiple system atrophy (MSA). Here, we investigated early transcriptomic changes within PD and MSA O4(+) oligodendrocyte lineage cells (OLCs) generated from patient-induced pluripotent stem cells (iPSCs). We found impaired maturation of PD and MSA O4(+) OLCs compared to controls. This phenotype was associated with changes in the human leukocyte antigen (HLA) genes, the immunoproteasome subunit PSMB9, and the complement component C4b for aSYN p.A53T and MSA O4(+) OLCs, but not in SNCA(trip) O4(+) OLCs despite high levels of aSYN assembly formation. Moreover, SNCA overexpression resulted in the development of O4(+) OLCs, whereas exogenous treatment with aSYN species led to significant toxicity. Notably, transcriptome profiling of genes encoding proteins forming Lewy bodies and glial cytoplasmic inclusions revealed clustering of PD aSYN p.A53T O4(+) OLCs with MSA O4(+) OLCs. Our work identifies early phenotypic and pathogenic changes within human PD and MSA O4(+) OLCs.
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| 3. |
- Chumarina, Margarita, et al.
(författare)
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Cellular alterations identified in pluripotent stem cell-derived midbrain spheroids generated from a female patient with progressive external ophthalmoplegia and parkinsonism who carries a novel variation (p.Q811R) in the POLG1 gene
- 2019
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Ingår i: Acta Neuropathologica Communications. - : Springer Science and Business Media LLC. - 2051-5960. ; 7:1
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Tidskriftsartikel (refereegranskat)abstract
- Variations in the POLG1 gene encoding the catalytic subunit of the mitochondrial DNA polymerase gamma, have recently been associated with Parkinson's disease (PD), especially in patients diagnosed with progressive external ophthalmoplegia (PEO). However, the majority of the studies reporting this association mainly focused on the genetic identification of the variation in POLG1 in PD patient primary cells, and determination of mitochondrial DNA copy number, providing little information about the cellular alterations existing in patient brain cells, in particular dopaminergic neurons. Therefore, through the use of induced pluripotent stem cells (iPSCs), we assessed cellular alterations in novel p.Q811R POLG1 (POLG1Q811R) variant midbrain dopaminergic neuron-containing spheroids (MDNS) from a female patient who developed early-onset PD, and compared them to cultures derived from a healthy control of the same gender. Both POLG1 variant and control MDNS contained functional midbrain regionalized TH/FOXA2-positive dopaminergic neurons, capable of releasing dopamine. Western blot analysis identified the presence of high molecular weight oligomeric alpha-synuclein in POLG1Q811R MDNS compared to control cultures. In order to assess POLG1Q811R-related cellular alterations within the MDNS, we applied mass-spectrometry based quantitative proteomic analysis. In total, 6749 proteins were identified, with 61 significantly differentially expressed between POLG1Q811R and control samples. Pro-And anti-inflammatory signaling and pathways involved in energy metabolism were altered. Notably, increased glycolysis in POLG1Q811R MDNS was suggested by the increase in PFKM and LDHA levels and confirmed using functional analysis of glycolytic rate and oxygen consumption levels. Our results validate the use of iPSCs to assess cellular alterations in relation to PD pathogenesis, in a unique PD patient carrying a novel p.Q811R variation in POLG1, and identify several altered pathways that may be relevant to PD pathogenesis.
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| 4. |
- Chumarina, Margarita, et al.
(författare)
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Derivation of mouse embryonic stem cell lines from tyrosine hydroxylase reporter mice crossed with a human SNCA transgenic mouse model of Parkinson's disease
- 2017
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Ingår i: Stem Cell Research. - : Elsevier BV. - 1876-7753 .- 1873-5061. ; 19, s. 17-20
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Tidskriftsartikel (refereegranskat)abstract
- Mouse embryonic stem cell (mESC) lines were derived by crossing heterozygous transgenic (tg) mice expressing green fluorescent protein (GFP) under the control of the rat tyrosine hydroxylase (TH) promoter, with homozygous alpha-synuclein (aSYN) mice expressing human mutant SNCAA53T under the control of the mouse Prion promoter (MoPrP), or wildtype (WT) mice. The expression of GFP and human aSYN was validated by immunocytochemistry in midbrain neuron cultures upon differentiation of mESC lines using stromal cell-derived inducing activity. These mESC lines can help to study the impact of human aSYN expression in neurons and oligodendrocytes, and also trace GFP-expressing midbrain neurons.
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| 5. |
- Chumarina, Margarita
(författare)
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Stem cell-based models to study cellular alterations in Parkinson’s Disease
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Parkinson’s Disease (PD) is the most prevalent neurodegenerative movement disorder. Despite investigative efforts into the pathogenesis of the disease, the aetiology and mechanism of PD remains unknown, with multiple factors and cellular pathways known to play a part. But the genetic component of the disease, as well as enhanced vulnerability of the midbrain dopaminergic neurons of the Substantia nigra pars compacta make PD a suitable candidate for stem cell-based studies of the disease initiation and progression.The work presented in this thesis aims to generate and utilize PD patient-derived induced pluripotent stem cells (iPSCs), making it possible to assess cellular alterations among patients with different genetic disease backgrounds and disease phenotypes. We also highlight the potential role of other brain cell types, particularly glial cells, in the PD pathogenesis, and employ the same iPSC-based strategies to derive oligodendrocytes, a cell type not well-studied in relation to PD.Our work demonstrated that as oligodendrocytes express synuclein-alpha (SNCA), the expression of PD-associated genetic variant p.A53T impairs oligodendrocyte maturation. The same effect was observed using mouse embryonic stem cell (mESC)-derived oligodendrocytes from a transgenic mouse model overexpressing human SNCA variant p.A53T. These findings validate the use of stem cells to further investigate oligodendrocyte pathology and its possible implications in the pathogenesis of PD.We also obtained iPSCs from a patient with a novel, not previously characterised variant p.Q811R in the POLG1, a risk factor for PD. Differentiation of these iPSCs into midbrain dopaminergic neuron containing spheroids (MDNS) enabled for the first time to assess the neuronal phenotype of POLG1-related PD, and identified several disease phenotypes, associated with PD, including SNCA pathology, disturbance of dopamine homeostasis and impaired energy metabolism.Using omics approaches for investigation of key altered networks in MDNS in POLG1Q811R and SNCAA53T lines revealed distinct altered pathways, highlighting the heterogeneity and multifactorial nature of PD.
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| 6. |
- Gustavsson, Nadja, et al.
(författare)
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Generation of an induced pluripotent stem cell line (CSC-46) from a patient with Parkinson's disease carrying a novel p.R301C mutation in the GBA gene
- 2019
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Ingår i: Stem Cell Research. - : Elsevier BV. - 1873-5061. ; 34
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Tidskriftsartikel (refereegranskat)abstract
- Mutations in the glucocerebrosidase (GBA) gene have been associated with the development of Parkinson's disease (PD). An induced pluripotent stem cell (iPSC) line was generated from a 60-year old patient diagnosed with PD and carrying a new mutation variant p.R301C in GBA. Using non-integrating Sendai virus-based technology, we utilized OCT3/4, SOX2, c-MYC and KLF4 transcription factors to reprogram skin fibroblasts into iPSCs. The generated iPSC line retained the mutation, displayed expression of common pluripotency markers, differentiated into the three germ layers, and exhibited normal karyotype. The iPSC line can be further used for studying PD pathogenesis.
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