Suppression of inflammatory and fibrotic responses in allergic inflammation by the amniotic membrane stromal matrix

Background: The amniotic membrane (AM), which is the innermost layer of the placenta, was shown to possess anti-inflammatory and anti-fibrotic properties in various in vitro and clinical studies. Purpose: To evaluate the anti-fibrotic and anti-inflammatory effects of the AM matrix (AMM) on human conjunctival and lung fibroblasts in an in vitro system that tests fibrotic and inflammatory responses at the effector stages of allergic inflammation. Methods: Human conjunctival or lung fibroblasts were seeded on plastic or on the stromal aspect of the AM, which was mounted on plastic inserts. Sonicates of human peripheral blood eosinophils activated with lipopolysacharide (LPS), or human mast cell (HMC-1) leukaemia cell sonicates, were added to sub-confluent fibroblast monolayers. Proliferation of the sub-confluent fibroblasts was assessed using the [³H]-thymidine incorporation assay. The production of transforming growth factor (TGF)-β₁, granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-8 in conjunctival or lung fibroblasts was measured in conditioned media from these cultures by ELISA. Results: After 4 days in culture, the [³H]-thymidine incorporation assay indicated a reduced proliferation of activated conjunctival and lung fibroblasts when cultured directly on the AMM. The production of both TGF-β₁ and IL-8 was significantly suppressed in activated conjunctival fibroblasts cultured on the AMM compared with those cultured on plastic, while the production of both TGF-β₁ and GM-CSF was decreased in human lung fibroblast cultured on the AMM. Conclusions: The AMM is capable of suppressing fibrotic responses in an in vitro system of effector stages of ocular allergic inflammation. These data may provide a basis for exploring matrix components in the AM for the treatment of allergic eye disease.