L Zhong1, M Li. 1. Department of Ophthalmology, First School of Clinical Medicine, Beijing Medical University, Beijing 100034.
Abstract
OBJECTIVE: To understand transforming growth factor-beta(1) (TGF-beta(1)) inducing cultured human trabecular cells (HTCs) to produce elastin and to approach the effect of resistance of aqueous outflow in the pathogenesis and mechanism of primary open angle glaucoma (POAG). METHODS: Trabecular meshwork's specimens were collected from twelve normal eyeballs of six human donors under six years of age within 24 hours after death. HTCs were primarily cultured and subcultured. Cultured cells were observed under light and electron microscopes. Laminin (LN) and IV collagen (IV C) in extracellular matrix (ECM) were immunohistochemically stained by S-P method. The third passage confluent cells after two weeks were divided into three groups: control group, TGF-beta(1) group and neutralizing antibody of TGF-beta(1) group. Concentration of neutralizing antibody of TGF-beta(1) was 30 microg/ml. After culturing for two weeks, elastin in the media of three groups was respectively determined by western blotting from sodium dodecylsulfate-polyacrylamide gel electrophoresis using anti-human aortic alpha-elastin rabbit serum, biotin conjugated goat anti-rabbit IgG and anti-biotin conjugated horse radish peroxidase. RESULTS: Cultured HTCs were identified from cultured cell's growing characteristics, morphological features under light and electron microscopes, and stained peculiarities of LN and IV C in ECM. With western blotting, elastin in the control group was not detected, elastin in TGF-beta(1) group was detected, and elastin in neutralizing antibody of TGF-beta(1) group was not detected. CONCLUSION: TGF-beta(1) induces cultured HTCs to produce elastin, which the neutralizing antibody of elastin can antagonize; the increase of the amount of elastin may aggravate its stacking in ECM and increase the resistance of aqueous outflow in trabecular meshwork, leading to the occurrence of glaucoma.
OBJECTIVE: To understand transforming growth factor-beta(1) (TGF-beta(1)) inducing cultured human trabecular cells (HTCs) to produce elastin and to approach the effect of resistance of aqueous outflow in the pathogenesis and mechanism of primary open angle glaucoma (POAG). METHODS: Trabecular meshwork's specimens were collected from twelve normal eyeballs of six human donors under six years of age within 24 hours after death. HTCs were primarily cultured and subcultured. Cultured cells were observed under light and electron microscopes. Laminin (LN) and IV collagen (IV C) in extracellular matrix (ECM) were immunohistochemically stained by S-P method. The third passage confluent cells after two weeks were divided into three groups: control group, TGF-beta(1) group and neutralizing antibody of TGF-beta(1) group. Concentration of neutralizing antibody of TGF-beta(1) was 30 microg/ml. After culturing for two weeks, elastin in the media of three groups was respectively determined by western blotting from sodium dodecylsulfate-polyacrylamide gel electrophoresis using anti-human aortic alpha-elastinrabbit serum, biotin conjugated goat anti-rabbit IgG and anti-biotin conjugated horseradish peroxidase. RESULTS: Cultured HTCs were identified from cultured cell's growing characteristics, morphological features under light and electron microscopes, and stained peculiarities of LN and IV C in ECM. With western blotting, elastin in the control group was not detected, elastin in TGF-beta(1) group was detected, and elastin in neutralizing antibody of TGF-beta(1) group was not detected. CONCLUSION:TGF-beta(1) induces cultured HTCs to produce elastin, which the neutralizing antibody of elastin can antagonize; the increase of the amount of elastin may aggravate its stacking in ECM and increase the resistance of aqueous outflow in trabecular meshwork, leading to the occurrence of glaucoma.