Involvement of Muller Glial Autoinduction of TGF-β in Diabetic Fibrovascular Proliferation Via Glial-Mesenchymal Transition

PURPOSE. Muller glial-mesenchymal transition (GMT) is reported as the fibrogenic mechanism promoted by TGF-beta-SNAIL axis in Muller cells transdifferentiated into myofibroblasts. Here we show the multifaceted involvement of TGF-beta in diabetic fibrovascular proliferation via Muller GMT and VEGF-A production. METHODS. Surgically excised fibrovascular tissues from the eyes of patients with proliferative diabetic retinopathy were processed for immunofluorescence analyses of TGF beta downstream molecules. Human Muller glial cells were used to evaluate changes in gene and protein expression with real-time quantitative PCR and ELISA, respectively. Immunoblot analyses were performed to detect TGF-beta signal activation. RESULTS. Muller glial cells in patient fibrovascular tissues were immunopositive for GMT-related molecular markers, including SNAIL and smooth muscle protein 22, together with colocalization of VEGF-A and TGF-beta receptors. In vitro administration of TGF-beta 1/2 upregulated TGFB1 and TGFB2, both of which were suppressed by inhibitors for nuclear factor-kappa B, glycogen synthase kinase-3, and p38 mitogen-activated protein kinase. Of the various profibrotic cytokines, TGF-beta 1/2 application exclusively induced Muller glial VEGFA mRNA expression, which was decreased by pretreatment with small interfering RNA for SMAD2 and inhibitors for p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase. Supporting these findings, TGF-beta 1/2 stimulation to Muller cells increased the phosphorylation of these intracellular signaling molecules, all of which were also activated in Muller glial cells in patient fibrovascular tissues. CONCLUSIONS. This study underscored the significance of Muller glial autoinduction of TGF-beta as a pathogenic cue to facilitate diabetic fibrovascular proliferation via TGF-beta-driven GMT and VEGF-A-driven angiogenesis.