Isolation and culture of rat and mouse oligodendrocyte precursor cells

Chen, Ying; Balasubramaniyan, Veerakumar; Peng, Jie; Hurlock, Edward C.; Tallquist, Michelle D.; Li, Jianrong; Lü, Qing

doi:10.1038/nprot.2007.149

Cited by 362 publications

(351 citation statements)

References 26 publications

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“…The ability of TNF-a to impair oligodendrocyte progenitor cell (OPC) differentiation in an in vitro system was examined following the protocol described by Chen et al 11 Immunostaining was performed using markers of the total oligodendrocyte population (oligodendrocyte-specific protein, or OSP) and for astrocytes (glial fibrillary acid protein, or GFAP) to verify the purity of the culture, which was found to contain 95.01 ± 1.0067% OSP þ cells and negligible numbers of GFAP þ cells. After 7 days of culture, the OPCs were treated for 24 h with a sublethal concentration of TNF-a, or 10 ng/ml, as suggested in a previous study.…”

Section: Resultsmentioning

confidence: 99%

Tumor necrosis factor-α impairs oligodendroglial differentiation through a mitochondria-dependent process

et al. 2014

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Mitochondrial defects, affecting parameters such as mitochondrial number and shape, levels of respiratory chain complex components and markers of oxidative stress, have been associated with the appearance and progression of multiple sclerosis. Nevertheless, mitochondrial physiology has never been monitored during oligodendrocyte progenitor cell (OPC) differentiation, especially in OPCs challenged with proinflammatory cytokines. Here, we show that tumor necrosis factor alpha (TNF-a) inhibits OPC differentiation, accompanied by altered mitochondrial calcium uptake, mitochondrial membrane potential, and respiratory complex I activity as well as increased reactive oxygen species production. Treatment with a mitochondrial uncoupler (FCCP) to mimic mitochondrial impairment also causes cells to accumulate at the progenitor stage. Interestingly, AMP-activated protein kinase (AMPK) levels increase during TNF-a exposure and inhibit OPC differentiation. Overall, our data indicate that TNF-a induces metabolic changes, driven by mitochondrial impairment and AMPK activation, leading to the inhibition of OPC differentiation.

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Section: Resultsmentioning

confidence: 99%

Tumor necrosis factor-α impairs oligodendroglial differentiation through a mitochondria-dependent process

et al. 2014

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“…Endothelial cells were placed on pre‐coated dishes with 3.2% bovine collagen type IV alpha 1 chain (COL4A1) (A1064401, Invitrogen) in PBS. Oligodendrocyte precursor cells (OPCs) were isolated from cerebral cortices from 1‐ to 2‐day‐old Sprague Dawley rats as previously described 8, 30.…”

Section: Methodsmentioning

confidence: 99%

Pericyte‐derived bone morphogenetic protein 4 underlies white matter damage after chronic hypoperfusion

et al. 2017

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Subcortical small vessel disease (SVD) is characterized by white matter damage resulting from arteriolosclerosis and chronic hypoperfusion. Transforming growth factor beta 1 (TGFB1) is dysregulated in the hereditary SVD, CARASIL (cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy). However, very little is known about the role of the largest group in the TGFB superfamily – the bone morphogenetic proteins (BMPs) – in SVD pathogenesis. The aim of this study was to characterize signaling abnormalities of BMPs in sporadic SVD. We examined immunostaining of TGFB1 and BMPs (BMP2/BMP4/BMP6/BMP7/BMP9) in a total of 19 post‐mortem human brain samples as follows: 7 SVD patients (4 males, 76–90 years old); 6 Alzheimer's disease (AD) patients (2 males, 67–93 years old) and 6 age‐matched disease controls (3 males, 68–78 years old). We subsequently investigated the effects of oxygen–glucose deprivation and BMP4 addition on cultured cells. Furthermore, adult mice were subjected to chronic cerebral hypoperfusion using bilateral common carotid artery stenosis, followed by continuous intracerebroventricular infusion of the BMP antagonist, noggin. In the SVD cases, BMP4 was highly expressed in white matter pericytes. Oxygen–glucose deprivation induced BMP4 expression in cultured pericytes in vitro. Recombinant BMP4 increased the number of cultured endothelial cells and pericytes and converted oligodendrocyte precursor cells into astrocytes. Chronic cerebral hypoperfusion in vivo also upregulated BMP4 with concomitant white matter astrogliogenesis and reduced oligodendrocyte lineage cells, both of which were suppressed by intracerebroventricular noggin infusion. Our findings suggest ischemic white matter damage evolves in parallel with BMP4 upregulation in pericytes. BMP4 promotes angiogenesis, but induces astrogliogenesis at the expense of oligodendrocyte precursor cell proliferation and maturation, thereby aggravating white matter damage. This may explain white matter vulnerability to chronic hypoperfusion. The regulation of BMP4 signaling is a potential therapeutic strategy for treating SVD.

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“…Multiple methods have been developed to isolate and expand NSCs, two of which have been recently published (Chen et al 2007;Brewer and Torricelli 2007). Each method has its own advantages.…”

Section: Discussionmentioning

confidence: 99%

Optimal time for passaging neurospheres based on primary neural stem cell cultures

et al. 2011

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Cultured neural stem cells (NSCs) provide a powerful means for investigating central nervous system disease, neuron development, differentiation, and regeneration. To obtain sufficient neurospheres, subculturing is essential following establishment of the primary NSC culture. Passaging the primary neurospheres is a key issue that is often ignored. We evaluated the influence of different passaging schedules on primary cultured NSCs. Passaging was performed on day 5, 7 or 9. We observed more neurospheres with diameters of 200-250 lm on day 7 than on day 5 or 9. Prolonging the time of primary culture reduced the cell metabolic activity by the MTT assay and cell proliferation by colony-forming assay and the differentiation to neurons from cells at P2 and later decreased. Additionally, more cells were in G0/G1 phase, and higher expression of p16INK4a and lower expression of cyclin D1 was found when the time of primary culture was prolonged to 9 days compared to 7-days cultures. Thus, in this study, we established that the optimal time for subculturing aggregated NSCs was on day 7 based on the primary culture.

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Isolation and culture of rat and mouse oligodendrocyte precursor cells

Cited by 362 publications

References 26 publications

Tumor necrosis factor-α impairs oligodendroglial differentiation through a mitochondria-dependent process

Tumor necrosis factor-α impairs oligodendroglial differentiation through a mitochondria-dependent process

Pericyte‐derived bone morphogenetic protein 4 underlies white matter damage after chronic hypoperfusion

Optimal time for passaging neurospheres based on primary neural stem cell cultures

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