Hypoxia‑Induced Inflammation in In Vitro Model of Human Blood–Brain Barrier: Modulatory Effects of the Olfactory Ensheathing Cell‑Conditioned Medium

Hypoxia compromises the integrity of the blood–brain barrier (BBB) and increases its permeability, thereby inducing inflam-mation. Olfactory ensheathing cells (OECs) garnered considerable interest due to their neuroregenerative and anti-inflammatoryproperties. Here, we aimed to investigate the potential modulatory effects of OEC-conditioned medium (OEC-CM) on theresponse of human brain microvascular endothelial cells (HBMECs), constituting the BBB, when exposed to hypoxia. HBMECswere utilized to establish the in vitro BBB model. OECs were isolated from mouse olfactory bulbs, and OEC-CM was collectedafter 48 h of culture. The effect of OEC-CM treatment on the HBMEC viability was evaluated under both normoxic and hypoxicconditions at 6 h, 24 h, and 30 h. Western blot and immunostaining techniques were employed to assess NF-κB/phospho-NF-κBexpression. HIF-1α, VEGF-A, and cPLA 2 mRNA expression levels were quantified using digital PCR. ELISA assays wereperformed to measure PGE2, VEGF-A, IL-8 secretion, and cPLA2 specific activity. The in vitro formation of HBMEC capillary-like structures was examined using a three-dimensional matrix system. OEC-CM attenuated pro-inflammatory responses andmitigated the HIF-1α/VEGFA signaling pathway activation in HBMECs under hypoxic condition. Hypoxia-induced damageof the BBB can be mitigated by novel therapeutic strategies harnessing OEC potential.