Isogenic blood-brain barrier models based on patient-derived stem cells display inter-individual differences in cell maturation and functionality.

The blood-brain barrier (BBB) constitutes an important component of the neurovascular unit formed by specialized brain microvascular endothelial cells (BMECs) surrounded by astrocytes, pericytes, and neurons. Recently, isogenic in vitro models of the BBB based on human pluripotent stem cells have been documented, yet the impact of inter-individual variability on the yield and phenotype of such models remains to be documented. In this study, we investigated the impact of inter-individual variability on the yield and phenotype of isogenic models of the BBB, using patient-derived induced pluripotent stem cells (iPSCs). Astrocytes, BMECs, and neurons were differentiated from four asymptomatic patient-derived iPSCs (two males, two females). We differentiated such cells using existing differentiation protocols and quantified expression of cell lineage markers, as well as BBB phenotype, barrier induction, and formation of neurite processes. iPSC-derived BMECs showed barrier properties better than hCMEC/D3 monolayers; however, we noted differences in the expression and activity among iPSC lines. In addition, we noted differences in the differentiation efficiency of these cells into neural stem cells and progenitor cells (as noted by differences in expression of cell lineage markers). Such differences were reflected later in the terminal differentiation, as seen as ability to induce barrier function and to form neurite processes. Although we demonstrated our ability to obtain an isogenic model of the BBB with different patients' iPSCs, we also noted subtle differences in the expression of cell lineage markers and cell maturation processes, suggesting the presence of inter-individual polymorphisms.

Neurological diseases at the blood-brain barrier: Stemming new scientific paradigms using patient-derived induced pluripotent cells.

The blood-brain barrier (BBB) is a component of the neurovascular unit formed by specialized brain microvascular endothelial cells (BMECs) surrounded by a specific basement membrane interacting with astrocytes, neurons, and pericytes. The BBB plays an essential function in the maintenance of brain homeostasis, by providing a physical and chemical barrier against pathogens and xenobiotics. Although the disruption of the BBB occurs with several neurological disorders, the scarcity of patient material source and lack of reliability of current in vitro models hindered our ability to model the BBB during such neurological conditions. The development of novel in vitro models based on patient-derived stem cells opened new venues in modeling the human BBB in vitro, by being more accurate than existing in vitro models, but also bringing such models closer to the in vivo setting. In addition, patient-derived models of the BBB opens the avenue to address the contribution of genetic factors commonly associated with certain neurological diseases on the BBB pathophysiology. This review provides a comprehensive understanding of the BBB, the current development of stem cell-based models in the field, the current challenges and limitations of such models.

Oxygen-Glucose Deprivation/Reoxygenation-Induced Barrier Disruption at the Human Blood-Brain Barrier is Partially Mediated Through the HIF-1 Pathway.

The blood-brain barrier (BBB) plays an important role in brain homeostasis. Hypoxia/ischemia constitutes an important stress factor involved in several neurological disorders by inducing the disruption of the BBB, ultimately leading to cerebral edema formation. Yet, our current understanding of the cellular and molecular mechanisms underlying the BBB disruption following cerebral hypoxia/ischemia remains limited. Stem cell-based models of the human BBB present some potentials to address such issues. Yet, such models have not been validated in regard of its ability to respond to hypoxia/ischemia as existing models. In this study, we investigated the cellular response of two iPSC-derived brain microvascular endothelial cell (BMEC) monolayers to respond to oxygen-glucose deprivation (OGD) stress, using two induced pluripotent stem cells (iPSC) lines. iPSC-derived BMECs responded to prolonged (24 h) and acute (6 h) OGD by showing a decrease in the barrier function and a decrease in tight junction complexes. Such iPSC-derived BMECs responded to OGD stress via a partial activation of the HIF-1 pathway, whereas treatment with anti-angiogenic pharmacological inhibitors (sorafenib, sunitinib) during reoxygenation worsened the barrier function. Taken together, our results suggest such models can respond to hypoxia/ischemia similarly to existing in vitro models and support the possible use of this model as a screening platform for identifying novel drug candidates capable to restore the barrier function following hypoxic/ischemic injury.

Cerebral hypoxia/ischemia selectively disrupts tight junctions complexes in stem cell-derived human brain microvascular endothelial cells.

BACKGROUND: Cerebral hypoxia/ischemia (H/I) is an important stress factor involved in the disruption of the blood-brain barrier (BBB) following stroke injury, yet the cellular and molecular mechanisms on how the human BBB responds to such injury remains unclear. In this study, we investigated the cellular response of the human BBB to chemical and environmental H/I in vitro. METHODS: In this study, we used immortalized hCMEC/D3 and IMR90 stem-cell derived human brain microvascular endothelial cell lines (IMR90-derived BMECs). Hypoxic stress was achieved by exposure to cobalt chloride (CoCl2) or by exposure to 1 % hypoxia and oxygen/glucose deprivation (OGD) was used to model ischemic injury. We assessed barrier function using both transendothelial electrical resistance (TEER) and sodium fluorescein permeability. Changes in cell junction integrity were assessed by immunocytochemistry and cell viability was assessed by trypan-blue exclusion and by MTS assays. Statistical analysis was performed using one-way analysis of variance (ANOVA). RESULTS: CoCl2 selectively disrupted the barrier function in IMR90-derived BMECs but not in hCMEC/D3 monolayers and cytotoxic effects did not drive such disruption. In addition, hypoxia/OGD stress significantly disrupted the barrier function by selectively disrupting tight junctions (TJs) complexes. In addition, we noted an uncoupling between cell metabolic activity and barrier integrity. CONCLUSIONS: In this study, we demonstrated the ability of IMR90-derived BMECs to respond to hypoxic/ischemic injury triggered by both chemical and environmental stress by showing a disruption of the barrier function. Such disruption was selectively targeting TJ complexes and was not driven by cellular apoptosis. In conclusion, this study suggests the suitability of stem cell-derived human BMECs monolayers as a model of cerebral hypoxia/ischemia in vitro.