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U.C. Welge–Lussen, A.L. Yu, H. Bloemendal, A. Kampik; Hypoxia/Reoxygenation and TGF–Beta2 Increase AlphaB–Crystallin Expression in Human Optic Nerve Head Astrocytes . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1257.
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© ARVO (1962-2015); The Authors (2016-present)
Optic nerve changes in glaucomatous disease are characterized by an increased expression of the small heat shock protein αB–crystallin and the growth factor transforming growth factor beta 2 (TGF–ß2) in reactive astrocytes. In the pathogenesis of glaucomatous optic nerve damage, ischemia/reperfusion injury may play an important role. The goal of the present study was to determine the influence of hypoxia/reoxygenation and TGF–ß2 on the expression of αB–crystallin in cultured human astrocytes of the optic nerve head (ONH).
Astrocytes were isolated from eyes of 5 human donors and cultured monolayers were incubated under hypoxic conditions (1% O2 for 4 to 12 hours) with following reoxygenation (12 to 24 hours). Additionally, cells were treated with 1.0 ng/ml TGF–ß2 for 12 and 24 hours. Expression of αB–crystallin was examined by northern and western blots. Levels of TGF–ß2 in response to hypoxia/reoxygenation were analyzed by RT–PCR analysis and ELISA. To determine the effect of the secreted TGF–ß2, untreated astrocytes were incubated with the supernatant of reoxygenated astrocytes combined with a neutralizing TGF–ß2 antibody.
Hypoxia/reoxygenation markedly increased the expression of αB–crystallin on the mRNA as well as on the protein level. Fold inductions ranged between factor 2.8 to 3.1 on the mRNA and 1.8 to 2.1 on the protein level. Treatment with 1.0 ng/ml TGF–ß2 for 12 to 24 hours elevated αB–crystallin mRNA up to 3.8 fold and αB–crystallin protein to 2.6 fold. TGF–ß2 showed an increase after 12 and 24 hours of reoxygenation on the mRNA level as well on the protein level about 2 fold. Supernatant of reoxygenated astrocytes induced a 2 fold mRNA expression of αB–crystallin. Simultaneous incubation of this supernatant with a neutralizing antibody against TGF–ß prevented the upregulation of αB–crystallin.
The process of hypoxia/reoxygenation is capable of inducing the expression of αB–crystallin in cultured ONH astrocytes. Therefore optimization of conditions leading to hypoxia/reoxygenation in the optic nerve head of glaucomatous patients could help to lower the incidence of characteristic changes in the optic nerve.
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