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M. Ali Behzadian, Xi-Liang Wang, L. Jack Windsor, Nagla Ghaly, Ruth B. Caldwell; TGF-β Increases Retinal Endothelial Cell Permeability by Increasing MMP-9: Possible Role of Glial Cells in Endothelial Barrier Function. Invest. Ophthalmol. Vis. Sci. 2001;42(3):853-859.
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purpose. To determine transforming growth factor (TGF) β effects on matrix
metalloproteinases (MMPs) as a potential cause of the blood–retinal
barrier breakdown at the onset of angiogenesis. Previously, glial cells
were shown to play a role in the angiogenesis process and to express
the angiogenic regulating factor TGF-β, which becomes active under
hypoxia conditions. Here, the authors demonstrate that retinal
endothelial cells express MMP-9 when treated with TGF-β or cocultured
with glial cells and that both TGF-β and MMP-9 increase endothelial
methods. Primary cultures of bovine retinal endothelial (BRE) cells grown on
porous membranes were treated with TGF-β or purified MMP-9, and
permeability changes were assayed. The amount and distribution of the
tight junction protein occludin also was analyzed by
immunocytochemistry and Western blotting. Cell extracts or conditioned
media from TGF-β–treated BRE cells and from glial cell–BRE
cocultures were analyzed for MMP-9 content by substrate gel
electrophoresis (zymography) or Western blotting.
results. Both TGF-β and MMP-9 increased the permeability of BRE monolayers and
reduced the levels of the junction protein occludin. The effect of
MMP-9 on permeability was rapid, but the TGF-β–induced permeability
required longer incubation and was blocked by anti–TGF-β and
anti–MMP-9 antibodies as well as by TGF-β latency-associated
peptide. Zymography showed that MMP-9 activity, which was very low or
absent in untreated BRE cultures, was dramatically increased by TGF-β
as well as by coculturing with either astrocytes or Müller glial
cells. Anti–TGF-β antibody blocked the TGF-β effect, but not the
coculture effect on MMP-9 production.
conclusions. These data indicate a direct correlation between TGF-β–induced MMP-9
activity and increased endothelial cell permeability. Moreover,
endothelial cell production of MMP-9 is regulated by glial cells
through expression of TGF-β or by direct cell-to-cell contact. During
retinal disease, glial cell production of active TGF-β may contribute
to breakdown of the blood–retinal barrier by stimulating endothelial
cell MMP-9 production.
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