EXTRACELLULAR-MATRIX PRODUCTION REGULATION BY TGF-BETA IN CORNEAL ENDOTHELIAL-CELLS

PURPOSE. Production of extracellular matrix (ECM) by corneal endotheli al cells is related to physiologic functions and pathologic conditions and is regulated by many cytokines, including transforming growth fac tor-beta (TGF-beta). In this study, the molecular mechanism of ECM pro duction regulation by TGF-beta was investigated in cultured corneal en dothelial cells. METHODs. The production of ECM components (laminin an d fibronectin) was detected in cultured corneal endothelial cells by w estern blot analysis. To determine the signal transduction pathways, m utant TGF-beta type I receptor (T beta R-I) and/or Smad protein family members (intracellular signal transducers in TGF-beta signaling) were overexpressed by transfecting their cDNA into the cultured cells, and the effects on ECM production were observed. RESULTS. The production of laminin and fibronectin was stimulated by treatment with TGF-beta(1 ) or TGF-beta(2). After transient transfection of cDNA of the constitu tively active (CA) mutant of T beta R-I, the production of laminin and fibronectin was stimulated even in the absence of TGF-beta. The trans fection of the dominant negative mutant of T beta R-I counteracted the effects of TGF-beta. These results confirm that TGF-beta directly sti mulates ECM production from corneal, endothelial cells through T beta R-I. The ECM production stimulation by TGF-beta or CA T beta R-I was a ccelerated by the overexpression of Smad2, Smad3, and/or Smad4 and inh ibited by that of Smad7. These results show that TGF-beta signals conn ected to ECM production are regulated by Smad family members, located downstream of T beta R-I. CONCLUSIONS. The results of this study show that TGF-beta stimulates ECM production from corneal endothelial cells through T beta R-I and Smad family transducers.