Tissue Transglutaminase (tTgase) in Native Pterygium and Primary Pterygium Cultures

Abstract

Abstract: : Introduction: Pterygium is characterized by inflammatory fibrovascular overgrowth of abnormal conjunctival tissue onto the clear cornea. A number of intrinisic and extrinsic factors have been identified in the pathogenesis of pterygium formation (i.e. UV radiation, IL–1, 6, 8, TNF–alpha, bFGF, TGF–beta, PDGF and others). Altered extracellular matrix (ECM) expression and composition appear to be relevant for pterygial stromal changes. In this context investigation of tTgase expression known to catalyse irreversible cross–linking of ECM proteins may allow further insite into pterygium pathogenesis and potential future therapies. Methods: Fresh pterygium specimen (n=3) and primary pterygium body fibroblast cultures were analyzed for co – expression of tTgase and fibronectin (Fn) as well as tTgase and epsilon–gama–glutamyl–lysine by immunhistochemistry using confocal laser scanning microscopy. Pterygium body fibroblast cultures and normal conjunctival fibroblast cultures (n=3 lines each) were treated with 10ng/mL IL–beta, PDGF–BB, EGF, bFGF, TNF–alpha and TGF–beta2 for 24h. mRNA and protein levels of tTgase were measued by RT–PCR and Western blot analysis. Results:Fresh pterygium samples showed tTgase expression in the stroma with a perivascular predominance. In confluent pterygium body fibroblast cultures a network like pattern of tTgase was identified. The enzyme showed co–expression with Fn and epsilon–gama–glutamyl–lysine. Expression of tTgase was increased by PDGF–BB and TGF–beta in pterygium fibroblast cultures but not in normal conjunctival fibroblast cultures as shown by RT–PCR (2 fold increase for PDGF–BB and 1.5 fold increase for TGF–beta) Western blot analysis showed similar results. All other substances did not alter tTgase expression. Conclusions: We were able to demonstrate that tTgase and its reaction product epsilon–gamma–glutamyl–lysine are expressed in the stroma of pterygium sections as well as in confluent pterygium fibroblast cultures. Furthermore an increase of tTgase was induced by PDGF–BB and TGF–beta, fibroangiogenic growth factors identified in different pterygium cell types. Intervention in this pathway may allow future approaches in the treatment of pterygium.

Keywords: Pterygium • cytokines/chemokines • extracellular matrix