scholarly journals Selective linkage of mitochondrial enzymes to intracellular calcium stores differs between human‐induced pluripotent stem cells, neural stem cells, and neurons

2020 ◽  
Author(s):  
Huanlian Chen ◽  
Abigail C. Cross ◽  
Ankita Thakkar ◽  
Hui Xu ◽  
Aiqun Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Intracellular Calcium ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Calcium Stores ◽  
Mitochondrial Enzymes ◽  
Intracellular Calcium Stores ◽  
Induced Pluripotent

scholarly journals Selective linkage of mitochondrial enzymes to intracellular calcium stores differs between human induced pluripotent stem cells, neural stem cells and neurons

2020 ◽  
Author(s):  
Huanlian Chen ◽  
Ankita Thakkar ◽  
Abigail C. Cross ◽  
Hui Xu ◽  
Aiqun Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Neuronal Development ◽  
Calcium Stores ◽  
Mitochondrial Enzymes ◽  
Induced Pluripotent ◽  
Er Calcium ◽  
Dehydrogenase Complex

AbstractThe coupling of the endoplasmic reticulum (ER) with mitochondria modulates neuronal calcium signaling. Whether this link changes with neuronal development is unknown. The current study first determined whether ER calcium stores are similar during development of human neurons, and then tested if the ER/mitochondrial coupling varied with development. The release of ER calcium to the cytosol by the IP3 agonist bradykinin was determined in human induced-pluripotent stem cells (iPSC), neural stem cells (NSC) and neurons. The concentration dependence for the release of ER calcium was similar at different stages of development. Metabolism changes dramatically with development. Glycolysis is the main energy source in iPSC and NSC whereas mitochondrial metabolism is more prominent in neurons. To test whether the coupling of mitochondria and ER changed with development, bombesin or bradykinin releasable calcium stores (BRCS) were monitored after inhibiting either of two key mitochondrial enzyme complexes: the alpha-ketoglutarate dehydrogenase complex (KGDHC) or the pyruvate dehydrogenase complex (PDHC). Inhibition of KGDHC did not alter BRCS in either iPSC or NSC. Inhibition of PDHC in neurons diminished BRCS whereas decreased KGDHC activity exaggerated BRCS. The latter finding may help understand the pathology of Alzheimer’s disease (AD). BRCS is exaggerated in cells from AD patients and KGDHC is reduced in brains of patients with AD. In summary, a prominent ER/mitochondrial link in neurons is associated with selective mitochondrial enzymes. The ER/mitochondrial link changes with human neuronal development and plausibly links ER calcium changes to AD.

scholarly journals Modeling CNS Involvement in Pompe Disease Using Neural Stem Cells Generated from Patient-Derived Induced Pluripotent Stem Cells

Cells ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 8
Author(s):  
Yu-Shan Cheng ◽  
Shu Yang ◽  
Junjie Hong ◽  
Rong Li ◽  
Jeanette Beers ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pompe Disease ◽  
Drug Efficacy ◽  
Cns Involvement ◽  
Glycogen Accumulation ◽  
Induced Pluripotent ◽  
Alpha Glucosidase

Pompe disease is a lysosomal storage disorder caused by autosomal recessive mutations in the acid alpha-glucosidase (GAA) gene. Acid alpha-glucosidase deficiency leads to abnormal glycogen accumulation in patient cells. Given the increasing evidence of central nervous system (CNS) involvement in classic infantile Pompe disease, we used neural stem cells, differentiated from patient induced pluripotent stem cells, to model the neuronal phenotype of Pompe disease. These Pompe neural stem cells exhibited disease-related phenotypes including glycogen accumulation, increased lysosomal staining, and secondary lipid buildup. These morphological phenotypes in patient neural stem cells provided a tool for drug efficacy evaluation. Two potential therapeutic agents, hydroxypropyl-β-cyclodextrin and δ-tocopherol, were tested along with recombinant human acid alpha-glucosidase (rhGAA) in this cell-based Pompe model. Treatment with rhGAA reduced LysoTracker staining in Pompe neural stem cells, indicating reduced lysosome size. Additionally, treatment of diseased neural stem cells with the combination of hydroxypropyl-β-cyclodextrin and δ-tocopherol significantly reduced the disease phenotypes. These results demonstrated patient-derived Pompe neural stem cells could be used as a model to study disease pathogenesis, to evaluate drug efficacy, and to screen compounds for drug discovery in the context of correcting CNS defects.

scholarly journals In vivo differentiation of induced pluripotent stem cells into neural stem cells by chimera formation

PLoS ONE ◽  
2017 ◽  
Vol 12 (1) ◽  
pp. e0170735 ◽  
Author(s):  
Hyun Woo Choi ◽  
Yean Ju Hong ◽  
Jong Soo Kim ◽  
Hyuk Song ◽  
Ssang Gu Cho ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
In Vivo Differentiation ◽  
Induced Pluripotent

scholarly journals Generation of Porcine Induced Neural Stem Cells Using the Sendai Virus

2022 ◽  
Vol 8 ◽  
Author(s):  
Warunya Chakritbudsabong ◽  
Ladawan Sariya ◽  
Phakhin Jantahiran ◽  
Nattarun Chaisilp ◽  
Somjit Chaiwattanarungruengpaisan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Sendai Virus ◽  
Genetic Alterations ◽  
Human Medicine ◽  
Large Animal ◽  
Induced Pluripotent ◽  
Induced Neural Stem Cells

The reprogramming of cells into induced neural stem cells (iNSCs), which are faster and safer to generate than induced pluripotent stem cells, holds tremendous promise for fundamental and frontier research, as well as personalized cell-based therapies for neurological diseases. However, reprogramming cells with viral vectors increases the risk of tumor development due to vector and transgene integration in the host cell genome. To circumvent this issue, the Sendai virus (SeV) provides an alternative integration-free reprogramming method that removes the danger of genetic alterations and enhances the prospects of iNSCs from bench to bedside. Since pigs are among the most successful large animal models in biomedical research, porcine iNSCs (piNSCs) may serve as a disease model for both veterinary and human medicine. Here, we report the successful generation of piNSC lines from pig fibroblasts by employing the SeV. These piNSCs can be expanded for up to 40 passages in a monolayer culture and produce neurospheres in a suspension culture. These piNSCs express high levels of NSC markers (PAX6, SOX2, NESTIN, and VIMENTIN) and proliferation markers (KI67) using quantitative immunostaining and western blot analysis. Furthermore, piNSCs are multipotent, as they are capable of producing neurons and glia, as demonstrated by their expressions of TUJ1, MAP2, TH, MBP, and GFAP proteins. During the reprogramming of piNSCs with the SeV, no induced pluripotent stem cells developed, and the established piNSCs did not express OCT4, NANOG, and SSEA1. Hence, the use of the SeV can reprogram porcine somatic cells without first going through an intermediate pluripotent state. Our research produced piNSCs using SeV methods in novel, easily accessible large animal cell culture models for evaluating the efficacy of iNSC-based clinical translation in human medicine. Additionally, our piNSCs are potentially applicable in disease modeling in pigs and regenerative therapies in veterinary medicine.

191 ROBUST GENERATION OF NEURAL STEM CELLS FROM PIG INDUCED PLURIPOTENT STEM CELLS FOR TRANSLATIONAL NEURAL REGENERATIVE MEDICINE

2014 ◽  
Vol 26 (1) ◽  
pp. 210
Author(s):  
A. Gallegos-Cardenas ◽  
K. Wang ◽  
E. T. Jordan ◽  
R. West ◽  
F. D. West ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Neural Differentiation ◽  
Neural Induction ◽  
Differentiation Potential ◽  
Induced Pluripotent ◽  
Human Ipsc

The generation of pig induced pluripotent stem cells (iPSC) opened the possibility to evaluate autologous neural cell therapy as a viable option for human patients. However, it is necessary to demonstrate whether pig iPSC are capable of in vitro neural differentiation similar to human iPSC in order to perform in vitro and in vivo comparative studies. Multiple laboratories have generated pig iPSC that have been characterised using pluripotent markers such as SSEA4 and POU5F1. However, correlations of pluripotent marker expression profiles among iPSC lines and their neural differentiation potential has not been fully explored. Because neural rosettes (NR) are composed of neural stem cells, our goal was to demonstrate that NR from pig iPSC can be generated, isolated, and expanded in vitro from multiple porcine iPSC lines similar to human iPSC and that the level of pluripotency in the starting porcine iPSC population (POUF51 and SSEA4 expression) could influence NRs development. Three lines of pig iPSC L1, L2, and L3 were cultured on matrigel-coated plates in mTeSR1 medium (Stemcell Technologies Inc., Vancouver, BC, Canada) and passaged every 3 to 4 days. For neural induction (NI), pig iPSC were disaggregated using dispase and plated. After 24 h, cells were maintained in N2 media [77% DMEM/F12, 10 ng mL–1 bovine fibroblast growth factor (bFGF), and 1X N2] for 15 days. To evaluate the differentiation potential to neuron and glial cells, NR were isolated, expanded in vitro and cultured for three weeks in AB2 medium (AB2, 1X ANS, and 2 mM L-Glutamine). Immunostaining assays were performed to determine pluripotent (POU5F1 and SSEA4), tight junction (ZO1), neural epithelial (Pax6 and Sox1), neuron (Tuj1), astrocyte (GFAP), and oligodendrocyte (O4) marker expression. Line L2 (POU5F1high and SSEA4low) showed a high potential to form NR (6.3.5%, P < 0.05) in comparison to the other 2 lines L1 (POU5F1low and SSEA4low) and L3 (POU5F1low and SSEA4high) upon NI. The NR immunocytochemistry results from Line L2 showed the presence of Pax6+ and Sox1– NRs cells at day 9 post-neural induction and that ZO1 started to localise at the apical border of NRs. At day 13, NRs cells were Pax6+ and Sox1+, and ZO1 was localised to the lumen of NR. After isolation and culture in vitro, NR cells expressed transcription factors PLAGL1, DACH1, and OTX2 through 2 passages, but were not detected in later passages. However, rosette cytoarchitecture was present up until passage 7 and were still Pax6+/Sox1+. NRs at passage 2 were cryopreserved and upon thaw showed normal NR morphology and were Pax6+/Sox1+. To characterise the plasticity of NRs, cells were differentiated. Tuj1 expression was predominant after differentiation indicating a bias towards a neuron phenotype. These results demonstrate that L2 pig iPSC (POUF51high and SSEA4low) have a high potential to form NR and neural differentiation parallels human iPSC neurulation events. Porcine iPSC should be considered as a large animal model for determining the safety and efficacy of human iPSC neural cell therapies.

Sign in / Sign up

Export Citation Format

Share Document