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Modeling the ischemic neurovascular unit in a dish using patient‐specific induced pluripotent stem cells
Author(s) -
Page Shyanne
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.659.20
Subject(s) - stroke (engine) , neuroprotection , induced pluripotent stem cell , medicine , neurovascular bundle , neuroscience , neural stem cell , blood–brain barrier , programmed cell death , bioinformatics , stem cell , biology , pathology , microbiology and biotechnology , central nervous system , apoptosis , embryonic stem cell , mechanical engineering , biochemistry , engineering , gene
The blood‐brain barrier (BBB) is a component of the neurovascular unit formed by specialized brain microvascular endothelial cells (BMECs), astrocytes and neurons. It plays an important function in the maintenance of brain homeostasis by tightly regulating the entrance of water and solutes into the brain and by blocking the entrance of potentially neurotoxic molecules. Dysfunction of the BBB is a key element observed in various neurological disorders and, in particular, following stroke injury. Stroke is the fifth cause of death in the US and the third leading cause of death in women. It is also a major cause of permanent disability. A key element of ischemic stroke (the most common type of stroke) pathophysiology is the opening of the BBB leading to cerebral edema onset and neural cell death by excitotoxicity. Notably, there is a growing literature highlighting the presence of a sexual dimorphism in stroke. Pre‐menopausal women have lesser stroke events than age‐matched men, however stroke outcome is worsened in post‐menopausal women. Interestingly, the nature such neuroprotection in pre‐menopausal women remains yet to be determined, as current in vitro models of the BBB cannot differentiate the effect of sex from the effects of sexual hormones. In this project, we designed a sex‐specific in vitro model of the BBB using patient‐derived induced pluripotent stem cells (iPSCs), using one male (CTR66M) and one female cell line (IMR90). We were capable of generating three different cell types from these iPSCs: astrocytes, BMECs and neurons. We did not observe a sexual dimorphism in the differentiation of these cell types in normal conditions, suggesting that biological sex may have not an impact on the differentiation and maturation of the BBB. Next, we challenged our cells to oxygen‐glucose deprivation (OGD) stress to model ischemic stroke injury in vitro . Cells were given DMEM –glucose and placed in a normobaric, hypoxic chamber for six hours. Following OGD stress, cells were given DMEM +glucose and re‐oxygenated for 24 hours to determine the effect of re‐oxygenation on barrier recovery. In order to assess barrier integrity, transendothelial electrical resistance (TEER) and permeability to sodium fluorescein was used. To determine any changes in cellular metabolism, an MTS or LDH assay was used. Preliminary data suggests the ability of the different cell types to respond to OGD stress in similar fashion as in vivo studies, as we noted a reversible BBB opening, astrocyte activation and pruning of neurites. In conclusion, our study demonstrates the possibility to obtain a sex‐specific in vitro model of the neurovascular unit, as well as its ability to respond to OGD stress. We are currently investigating the role of the HIF1‐alpha pathway in the response to ischemic stroke in astrocytes, neurons and BMECs.