ABSTRACT
The blood-brain barrier (BBB) is a biological unit composed of capillary endothelial cells and astrocytes. Here we examined the effects of various types of advanced glycation end-products (AGEs) on astrocytes and BBB-forming endothelial cells. While no type of AGE we examined changed the permeability of endothelial sheets, glyceraldehyde-derived AGE induced VEGF expression most significantly in astrocytes. The expression of glial cell line-derived neurotrophic factor (GDNF), which reduces the vascular permeability, was decreased in the astrocytes by treatment with glyceraldehyde-derived AGE. These results indicate that glyceraldehyde-derived AGE is the biologically active substance for astrocytes by regulating the VEGF and GDNF expression, which is causally contributing to an increase in the permeability of the BBB.
Subject(s)
Astrocytes/metabolism , Gene Expression Regulation/physiology , Glycation End Products, Advanced/physiology , Glyceraldehyde/metabolism , Nerve Growth Factors/genetics , Vascular Endothelial Growth Factor A/genetics , Astrocytes/cytology , Base Sequence , Blood-Brain Barrier , Blotting, Western , DNA Primers , Glial Cell Line-Derived Neurotrophic Factor , HumansABSTRACT
Cyclic AMP (cAMP) promotes functions of tight junctions in endothelial cells, although its target remains unknown. We showed here that cAMP increased gene expression of claudin-5 and decreased that of claudin-1 in porcine blood-brain-barrier endothelial cells via protein kinase A (PKA)-independent and -dependent pathways, respectively. cAMP also enhanced immunoreactivity of claudin-5 along cell borders and in the cytoplasm, reorganized actin filaments, and altered signals of claudin-5, occludin, ZO-1, and ZO-2 along cell boundaries from zipperlike to linear patterns. In contrast, claudin-1 was detected only in the cytoplasm in a dotlike pattern, and its immunolabeling was reduced by cAMP. Interestingly, 31- and 62-kDa claudin-5 immunoprecipitates in the NP-40-soluble and -insoluble fractions, respectively, were highly phosphorylated on threonine residue(s) upon cAMP treatment. All these changes induced by cAMP, except for claudin-5 expression and its signals in the cytoplasm, were reversed by an inhibitor of PKA, H-89. We also demonstrated that cAMP elevated the barrier function of tight junctions in porcine blood-brain-barrier endothelial cells in PKA-dependent and -independent manners. These findings indicate that both PKA-induced phosphorylation of claudin-5 immunoprecipitates and cAMP-dependent but PKA-independent induction of claudin-5 expression could be involved in promotion of tight-junction function in endothelial cells.
