Triptolide Of Ifn-¦Ã Stimulate The Descendants Of The Eye Into Fiber Cells

Objective To study the effects of triptolide, the principal active diterpenoid from the Chinese Medicinal Herb Tripterygium Wilfordii Hook. F which have immunosuppressive and anti-inflammatory properties, on cell proliferation , the synthesis of hyaluronic acid and collagen type I, and the expression of human leukocyte antigen (HLA)-DR, intercellular adhesion molecule 1(ICAM-1),CD40 expression on the cell surface of cultured normal human retroocular fibroblasts (RFs) in vitro induced by recombinant human interferon-γ (IFN-γ) .Methods1. Culture and identifiction of RFs. The retrobulbar connective tissue were obtained from the normal human, and the tissues were transported to the laboratory soon after removal and were cultured in condition of sterility in our laboratory. After two to eight passages, cultured normal human RFs were identificted by inverted microscope and immunocytochemitry methodes.2. After incubated with medium alone or with various concentrations of triptolide (0- 1OOug/L) for 48 hours, the proliferation of cultured RFs in vitro was assessed by MTT colorimetry assay.3. After incubated with medium alone or with ectogenesis cytokine--recombinant human interferon γ (IFN-γ, 100U/mL) in the presenceof various concentrations of triptolide (0 - 10ug/L) for 48 hours, the synthesis content of hyaluronic acid and collagen type I secreted by RFs were measured by radioimmunoassay and immunocytochemitry methodes.4. Cultured cells incubated with medium alone or in the presence of IFN-γ (100U/mL) and various concentrations of triptolide (0-10ug/L) for 48 hours. Then we used flow cytometry to investigate the effects of triptolide on the cell-surface expression of ICAM-1, HLA-DR and CD40 on cultured RFs induced by IFN-γ.Results1. These incubated cells were fibrous type cells observed by inverted microscope, and express Vimentin and fail to express Cytokeratin, and thus the cultures are not contaminated with endothelial or smooth muscle cells.2. Triptolide inhibited the proliferation of RFs significantly in a dose-dependent manner, and the proliferation was inhibited entirely with the concentration of lOOug/L.3. After incubated with ectogenesis cytokine--IFN-γ ( 100U/mL ) for48 hours, the synthesis of hyaluronic acid and collagen type I increasedsignificantly. Triptolide could inhibit the synthesis of hyaluronic acid and collagen type I significantly induced by IFN-γ significant ly in a dose-dependent manner.4. Cultured normal human RFs spontaneously and partly expressed ICAM-1, but did not express HLA-DR and CD40. IFN-γ ( 100U/mL ) significantly enhanced the expression of ICAM-1, HLA-DR and CD40 (P<0.01 ) on the surface of cultured normal human RFs and triptolide could antagonize the inflammatory effect of IFN-γ, down-regulation the expression of ICAM-1, HLA-DR and CD40 on RFs significantly in a dose-dependent manners.Conclusions Triptolide inhibits the activation of cultured normal human RFs induced by IFN-y and thus may decrease the autoimmune injury to the orbit in Graves' ophthalmopathy.