Role of Caspases in Cytokine-Induced Barrier Breakdown in Human Brain Endothelial Cells

Abstract

During neuroinflammation, cytokines such as TNF-α and IFN-γ secreted by activated leukocytes and/or CNS resident cells have been shown to alter the phenotype and function of brain endothelial cells (BECs) leading to blood–brain barrier breakdown. In this study, we show that the human BEC line hCMEC/D3 expresses the receptors for TNF-α, TNF receptor 1 and TNF receptor 2, and for IFN-γ. BEC activation with TNF-α alone or in combination with IFN-γ induced endothelial leakage of paracellular tracers. At high cytokine concentrations (10 and 100 ng/ml), this effect was associated with caspase-3/7 activation and apoptotic cell death as evidenced by annexin V staining and DNA fragmentation (TUNEL) assays. In addition, inhibition of JNK and protein kinase C activation at these doses partially prevented activation of caspase-3/7, although only JNK inhibition was partially able to prevent the increase in BEC paracellular permeability induced by cytokines. By contrast, lower cytokine concentrations (1 ng/ml) also led to effector caspase activation, increased paracellular flux, and redistribution of zonula occludens-1 and VE-cadherin but failed to induce apoptosis. Under these conditions, specific caspase-3 and caspase-9, but not caspase-8, inhibitors partially blocked cytokine-induced disruption of tight and adherens junctions and BEC paracellular permeability. Our results suggest that the concentration of cytokines in the CNS endothelial microenvironment determines the extent of caspase-mediated barrier permeability changes, which may be generalized as a result of apoptosis or more subtle as a result of alterations in the organization of junctional complex molecules.

Glial Cell–Elicited Activation of Brain Microvasculature in Response to Brucella abortus Infection Requires ASC Inflammasome–Dependent IL-1β Production

“…Although both IL-1b and TNF-a have been involved in the activation of the BBB (53,55), it seems that TNF-a, at least in our model, is dispensable for the activation of the brain endothelial cells, as shown in the activation of brain endothelial cells in murine Toxoplasma encephalitis (58), whereas IL-1b is responsible for this phenomenon. The reasons for these discrepancies are not known, but they could be related to the concentrations used in the experiments in which TNF-a has been indicated as the main activator of the brain microvascular endothelium (30,59,60), which are at least two orders of magnitude higher than the range induced by B. abortus-infected glial cell in this article and our previous observations (10,24). Thus, at least in our model, IL-1b signaling seems to have a major role in BBB activation.…”

Junctional proteins of the blood-brain barrier: New insights into function and dysfunction

“…Redistribution of transmembrane junction proteins, particularly claudin-5 and occludin as well Ve-cadherin, is closely associated with loosening of adhesion at the TJ complex, which ultimately leads to paracellular gap formation. 64,103,106,131 Caveolae-mediated internalization is the underlying mechanism of occludin and claudin-5 redistribution during the BBB dysfunction found in inflammation, infection (HIV-1), ischemic injury and with toxins (methamphetamine). 55,64,106 The destiny of the redistributed/internalized claudin-5 and occludin is still largely unknown.…”

Pathogens Penetrating the Central Nervous System: Infection Pathways and the Cellular and Molecular Mechanisms of Invasion

“…In vivo and in vitro studies have shown that group B streptococci, pneumococci, and S. suis induce the production of a range of cytokines and chemokines, including IL-1␤, IL-6, IL-8, IL-10, TNF-␣, macrophage chemoattractant protein 1 (MCP-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), Gro-␣, and Gro-␤, in brain microvascular endothelial cells (109,(256)(257)(258)) and brain tissue (94,259,260). The stimulation of HBMECs with cytokines (including TNF-␣, IL-1␤, IL-6, and IL-17F) leads to cytoskeletal rearrangements and a redistribution of tight junction and adherens junction proteins, resulting in a decrease in barrier integrity (261)(262)(263)(264)(265)(266)(267). Therefore, the host inflammatory response to group B streptococcus, S. pneumoniae, and S. suis meningitis may contribute to the disruption of tight junctions.…”