Effect Of TNF-α On MMPs Expression In Cultured Human Trabecular Meshwork Cells

Purpose:To study the culture of human trabecular meshwork cells(HTCs) in vitro and observe their biological character. To investigate the effect of tumor necrosis factor-α(TNF-α) on the expression of matrix metalloproteinases(MMPs) in cultured human trabecular cells (HTCs) , and to study the effect of NF-κB inhibitor PDTC on MMPs in cultured HTCs.Methods:(1) In the firt part, the trabecular meshwork tissues from human eyes were primarily cultured. The morphologic features and growing characteristics of the cultured cells were observed by inverted microscope, electron microscope and immunocytochemical medthod.The proliferative curve of the cultured cells was obtained by colorimetric assay with MTT.(2) MMP-3 and MMP-9 expressed in cultured HTCs were measured by semi-quantitative RT-PCR and casein/gelatin zymography after treated with 0.0ng/ml(control),1ng/ml,10ng/ml,25ng/ml TNF-αfor 24h.(3)PDTC,a inhibitor of nuclear fator–kappa B(NF-κB),was used before HTCs were treated with 10ng/ml,25ng/ml TNF-αfor 24h.Rusults:(1)The primary cells in cultured were observed from the edge of the cultured tissues after 10-15 days.The cultured cells were round ,oval ,fusiform and multiangular. The trabecular cells showed apical villons projections and had a high density of secondary lysosome. The junctions between the trabecular cells observed most frequently were gap junction. Laminin (LN),fibronectin(FN),and neuron specific enolase (NSE) of the cultured HTCs expressed were identified with immunocytochemical medthod.(2) The relative gray value of MMP-3/β-actin of cells treated with 0ng/ml(control),1ng/ml,10ng/ml,25ng/ml TNF-αfor 24h was 0.65±0.01,0.88±0.02,1.02±0.01,1.20±0.02, respectively. The difference between these 1ng/ml,10ng/ml,25ng/ml TNF-αtreated group and that of the control group was statistically significant. The relative gray value of MMP-9/β-actin of cells treated with 0ng/ml(control),1ng/ml,10ng/ml,25ng/ml TNF-αfor 24h was0.75±0.02, 0.94±0.01,0.97±0.05,1.13±0.02, respectively. The difference between these 1ng/ml,10ng/ml,25ng/ml TNF-αtreated...