Protection of cultured brain endothelial cells from cytokine-induced damage by α-melanocyte stimulating hormone

The blood-brain barrier (BBB), an interface between the systemic circulation and the nervous system, can be a target of cytokines in inflammatory conditions. Pro-inflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) induce damage in brain endothelial cells and BBB dysfunction which contribute to neuronal injury. The neuroprotective effects of a-melanocyte stimulating hormone (alpha-MSFI) were investigated in experimental models, but there are no data related to the BBB. Based on our recent study, in which a-MSH reduced barrier dysfunction in human intestinal epithelial cells induced by TNF-alpha and IL-1 beta, we hypothesized a protective effect of alpha-MSH on brain endothelial cells. We examined the effect of these two pro-inflammatory cytokines, and the neuropeptide alpha-MSH on a culture model of the BBB, primary rat brain endothelial cells cocultured with rat brain pericytes and glial cells. We demonstrated the expression of melanocortin-1 receptor in isolated rat brain microvessels and cultured brain endothelial cells by RT-PCR and immunohistochemistry. TNF-alpha and IL-1 beta induced cell damage, measured by impedance and MTT assay, which was attenuated by alpha-MSH (1 and 10 pM). The peptide inhibited the cytokine-induced increase in brain endothelial permeability, and restored the morphological changes in cellular junctions visualized by immunostaining for elaudin-5 and beta-catenin. Elevated production of reactive oxygen species and the nuclear translocation of NF-kappa B were also reduced by alpha-MSH in brain endothelial cells stimulated by cytokines. We demonstrated for the first time the direct beneficial effect of alpha-MSH on cultured brain endothelial cells, indicating that this neurohormone may be protective at the BBB.