Effects of Circulating Exosomes in Pediatric Obstructive Sleep Apnea with or Without Neurocognitive Deficits in an In Vitro 3D-Blood-Brain Barrier Spheroid Model
Year: 2021
Type: conference-abstract
Abstract: Rational: Pediatric obstructive sleep apnea (P-OSA) has emerged as a prevalent condition that imposes a large array of morbidities including neurocognitive deficits. Altered blood-brain barrier (BBB) integrity is associated many neurological disorders. The BBB consists of specialized endothelial cells (ECs) interconnected by junctional complexes including tight junctions (TJs) and adherent junctions, surrounded by astrocytes and pericytes. Here, we developed a 3D in vitro spheroid model of the human BBB that incorporates the 3 cell types to study the effects of exosomes on the 3D-spheroids. Methods: Thirty children were included and divided into three groups: (a) children normal polysomnographic studies and cognitive function (CO, n=6), (b) children with OSA and no evidence of cognitive deficits (OSA-NC(-); n=12), and (c) children with OSA with evidence of neurocognitive deficits (OSA-NC(+), n=12). All children were age-, gender-, ethnicity-, BMI z score-matched, and for those with OSA they were also apnea -hypopnea index (AHI)-matched. Plasma exosomes were characterized, quantified, and applied on a previously validated human 3D spheroid BBB model for 24 hr. We measured trans-endothelial electrical resistance (TEER) using trans-well cultures to evaluate the barrier function for each individual cell type and the 3D spheroids. Tight junction integrity was evaluated using ZO1 and VE cadherin immunostaining. In addition, exosomes were applied into the 3D spheroids cells to assess angiogenesis (tube formation) using a Matrigel assay. A time course of vessel tube formation was assessed in ECs alone, and in 3D spheroids for up to 48 hrs using confocal microscopy. Results: Using the same number of exosomes, those derived from OSA-NC(+) children disrupted 3D spheroids compared to OSA-(NC-) and CO using TEER, permeability to fluorescent dye, and ZO1 and VE cadherin staining. Also, OSA-(NC+) and OSA-(NC-) exosomes increased permeability and dysfunction of each of cell types plated as a monolayer when compared to CO. Finally, tube formation was clearly accelerated following exposure to CO-derived exosomes compared to OSA-(NC-) and OSA-(NC+). Conclusion: 3D spheroids generated using the prototypic cell types of the BBB display a robust typical barrier function. Exosomes derived from children with OSA disrupt BBB function, particularly among OSA-NC(+). Thus, this human 3D BBB model has unique biological features that should provide a promising high-throughput platform for personalized functional reporter studies.
Authors: A. Khalyfa, Z. Qiao,, G. David, K.-G. Leila
Institution: University of Missouri
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W4253377535