Outgrowth Endothelial Cell Conditioned Medium Negates TNF-α-Evoked Cerebral Barrier Damage: A Reverse Translational Research to Explore Mechanisms

Abstract

Improved understanding of the key mechanisms underlying cerebral ischemic injury is essential for the discovery of efficacious novel therapeutics for stroke. Through detailed analysis of plasma samples obtained from a large number of healthy volunteers (n = 90) and ischemic stroke patients (n = 81), the current study found significant elevations in the levels of TNF-α at baseline (within the first 48 h of stroke) and on days 7, 30, 90 after ischaemic stroke. It then assessed the impact of this inflammatory cytokine on an invitro model of human blood–brain barrier (BBB) and revealed dramatic impairments in both barrier integrity and function, the main cause of early death after an ischemic stroke. Co-treatment of BBB models in similar experiments with outgrowth endothelial cell-derived conditioned media (OEC-CM) negated the deleterious effects of TNF-α on BBB. Effective suppression of anti-angiogenic factor endostatin, stress fiber formation, oxidative stress, and apoptosis along with concomitant improvements in extracellular matrix adhesive and tubulogenic properties of brain microvascular endothelial cells and OECs played an important role in OEC-CM-mediated benefits. Significant increases in pro-angiogenic endothelin-1 and monocyte chemoattractant protein-1 in OEC-CM compared to the secretomes of OEC and HBMEC, detected by proteome profiling assay, accentuate the beneficial effects of OEC-CM. In conclusion, this reverse translational study identifies TNF-α as an important mediator of post-ischemic cerebral barrier damage and proposes OEC-CM as a potential vasculoprotective therapeutic strategy by demonstrating its ability to regulate a wide range of mechanisms associated with BBB function. Clinical trial registration NCT02980354. Graphical Abstract

Evaluation of Endothelial Progenitor Cell Characteristics as Clinical Biomarkers for Elderly Patients with Ischaemic Stroke

“…Biochemical analyses of HV and IS patient plasma samples revealed consistent increases in pro-inflammatory cytokine, TNF-α in IS patients. This confirmed a shift to pro-inflammatory state after stroke [ 27 , 28 ]. However, increased availability of pro-angiogenic cytokine, VEGF and the diminished availability of anti-angiogenic factors that normally inhibit the activity of VEGF i.e.…”

The Role of Stem Cells as Therapeutics for Ischaemic Stroke

“…The exogenous addition of EPCs repairs the integrity of an in vitro BBB model under OGD conditions and attenuates ischaemia-evoked oxidative stress and the apoptosis of endothelial cells [77,85,86]. Besides administering the stem cells directly, the condition media of outgrowth endothelial cells were found to negate the deleterious effects of TNF-α on BBB function in an in vitro triculture model of the BBB [87]. EPCs are able to secret a wide range of substances to regulate angiogenesis, migration, proliferation, and antipoptosis, including angiogenin, HGF, PDGF, VEGF, and PBEF [88].…”

“…EPCs are able to secret a wide range of substances to regulate angiogenesis, migration, proliferation, and antipoptosis, including angiogenin, HGF, PDGF, VEGF, and PBEF [88]. A clinical study indicated that post-stroke patients have elevated levels of TNF-α, and an in vitro study determined that TNF-α significantly impairs the function and integrity of the BBB [87,89]. Aside from their therapeutic potential, EPCs can also be used as diagnostic and prognostic biomarkers.…”

Endothelial Progenitor Cells in Neurovascular Disorders—A Comprehensive Overview of the Current State of Knowledge

“…An exogenous addition of OECs to an in vitro model of the BBB (established with astrocytes, pericytes and BMECs) repaired the wound scratch-induced on a layer of BMECs in serum-free conditions [ 63 ]. Additionally, a very recent study demonstrated that an outgrowth endothelial cell-derived conditioned media (OEC-CM) prevents the damaging effects of TNF-α on the BBB since the levels of TNF-α were found to be significantly elevated on days 2, 7, 30 and 90 after ischemic stroke and TNF-α impairs function and integrity of the BBB, which is the main early cause of death after IS [ 64 ].…”