Pro-Inflammatory Tnf Alpha And Il-1 Beta Differentially Regulate The Inflammatory Phenotype Of Brain Microvascular Endothelial Cells

Background: The vasculature of the brain is composed of endothelial cells, pericytes and astrocytic processes. The endothelial cells are the critical interface between the blood and the CNS parenchyma and are a critical component of the blood-brain barrier (BBB). These cells are innately programmed to respond to a myriad of inflammatory cytokines or other danger signals. IL-1 beta and TNF alpha are well recognised pro-inflammatory mediators, and here, we provide compelling evidence that they regulate the function and immune response profile of human cerebral microvascular endothelial cells (hCMVECs) differentially.Methods: We used xCELLigence biosensor technology, which revealed global differences in the endothelial response between IL-1 beta and TNF alpha. xCELLigence is a label-free impedance-based biosensor, which is ideal for acute or long-term comparison of drug effects on cell behaviour. In addition, flow cytometry and multiplex cytokine arrays were used to show differences in the inflammatory responses from the endothelial cells.Results: Extensive cytokine-secretion profiling and cell-surface immune phenotyping confirmed that the immune response of the hCMVEC to IL-1 beta was different to that of TNF alpha. Interestingly, of the 38 cytokines, chemokines and growth factors measured by cytometric bead array, the endothelial cells secreted only 13. Of importance was the observation that the majority of these cytokines were differentially regulated by either IL-1 beta or TNF alpha. Cell-surface expression of ICAM-1 and VCAM-1 were also differentially regulated by IL-1 beta or TNF alpha, where TNF alpha induced a substantially higher level of expression of both key leukocyte-adhesion molecules. A range of other cell-surface cellular and junctional adhesion molecules were basally expressed by the hCMVEC but were unaffected by IL-1 beta or TNF alpha.Conclusions: To our knowledge, this is the most comprehensive analysis of the immunological profile of brain endothelial cells and the first direct evidence that human brain endothelial cells are differentially regulated by these two key pro-inflammatory mediators.