Abstract
Simple SummaryThe potential of breast cancer to spread to the brain increases the clinical complications of the disease; breast cancer is considered to have the second-highest capacity to spread to the brain after lung cancer. The brain is protected by highly specialized endothelial cells, forming a barrier against the entry of circulating molecules and cells. The ability of breast cancer cells to penetrate the protective endothelial barrier is still not completely understood. Here, we aimed to investigate the effect of breast cancer cells on the brain’s endothelial cells. We showed that breast cancer cells induce changes in endothelial cells by releasing factors that target the mitochondria, affecting the endothelial cells and their attachment to each other and, therefore, their function as a protective barrier of the brain. Understanding the mechanism that breast cancer cells utilize to affect endothelial cells under normoxic and hypoxic conditions contributes to the development of treatments to prevent the metastasis of cancer cells to the brain.Background: The blood-brain barrier (BBB) is a central nervous system protective barrier formed primarily of endothelial cells that regulate the entry of substances and cells from entering the brain. However, the BBB integrity is disrupted in disease, including cancer, allowing toxic substances, molecules, and circulating cells to enter the brain. This study aimed to determine the mitochondrial changes in brain endothelial cells co-cultured with cancer cells. Method: Brain endothelial cells (bEnd.3) were co-cultivated with various concentrations of breast cancer (MCF7) conditioned media (CM) generated under normoxic (21% O2) and hypoxic conditions (5% O2). The mitochondrial activities (including; dehydrogenases activity, mitochondrial membrane potential (ΔΨm), and ATP generation) were measured using Polarstar Omega B.M.G-Plate reader. Trans-endothelial electrical resistance (TEER) was evaluated using the EVOM system, followed by quantifying gene expression of the endothelial tight junction (ETJs) using qPCR. Results: bEnd.3 cells had reduced cell viability after 72 h and 96 h exposure to MCF7CM under hypoxic and normoxic conditions. The ΔΨm in bEnd.3 cells were hyperpolarized after exposure to the hypoxic MCF7CM (p < 0.0001). However, the normoxic MCF7CM did not significantly affect the state of ΔΨm in bEnd.3 cells. ATP levels in bEnd.3 co-cultured with hypoxic and normoxic MCF7CM was significantly reduced (p < 0.05). The changes in brain endothelial mitochondrial activity were associated with a decrease in TEER of bEnd.3 monolayer co-cultured with MCF7CM under hypoxia (p = 0.001) and normoxia (p < 0.05). The bEnd.3 cells exposed to MCF7CM significantly increased the gene expression level of ETJs (p < 0.05). Conclusions: MCF7CM modulate mitochondrial activity in brain endothelial cells, affecting the brain endothelial barrier function.
Highlights
The Blood-brain barrier (BBB) is a multicellular barrier located between the blood and the brain tissues
Similar results were obtained after bEnd.3 cells were exposed to selected concentrations of MCF7 conditioned media (MCF7CM)
The viability of bEnd.3 cells were significantly reduced after 24 h exposure to the media composed of 75% of hypoxic MCF7CM (* p = 0.0450)
Summary
The Blood-brain barrier (BBB) is a multicellular barrier located between the blood and the brain tissues. At the point that the cancer cells are arrested at the BEC’s apical surface, and before penetrating the endothelial barrier [8], potential interactions occur between cancer cells and endothelial cells These interactions are characterized by either physical adhesion to the BECs or via secretory paracrine factors [10,11], which induces disruption of the brain endothelial cells and facilitates the migration of cancer cells into the brain. The mechanism proposed for this interaction, in general, suggest that physiological changes occur in endothelial cells under the stimulation of cancer cells, resulting in the activation of adhesion molecules such as E-selectin in endothelial cells Once these BEC plasma membrane proteins interact with their ligand on the metastatic cancer cell, a series of signaling pathways affect the opening of the cell-cell junctions allowing cancer cells to cross the endothelial barrier [12,13]. Conclusions: MCF7CM modulate mitochondrial activity in brain endothelial cells, affecting the brain endothelial barrier function
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