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J. Zhang, K.-Y. Xing, M. F. Lou; The Biochemical Alterations in Lenses of Thioltransferase (TTase) Knockout Mice. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4582.
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Thioltransferase knockout (TTase KO) model has been established in this laboratory and showed cataract developed sooner than the wild type (WT). The purpose of this study is to examine the relationship of morphological changes with the biochemical alterations in the lenses of TTase KO and WT mice as a function of age.
Mice of matching age of WT (1.1, 4.5, 9.2 m) and KO (1.3, 4.6, 8.9 m) were examined for lens opacity with portable slit lamp and Fundus camera. Each lens was homogenized in lysis buffer and processed for measurement of glutathione (GSH) level by DTNB colorimetric method and examination of protein-GSH mixed disulfides (PSSG) formation by Western blot analysis using an anti-GSH specific antibody. Dethiolation of lens proteins was carried out using either DTT or purified human recombinant TTase.
The slit lamp examination and Fundus camera showed a gradual cataract development in both eyes of WT and KO mice as a function of age. Nuclear cataract began to appear sooner in KO mice (4 m) compared with WT mice (9 m). The opacity in both lenses of the 9 months-old KO mouse also covered cortical and subcapsular regions. Western blot analysis showed that PSSG gradually increased with age and corroborated with the severity of opacity. PSSG was more abundant in the KO group than the WT group. These GSH-conjugated proteins could be eliminated when the lens homogenate was either treated with DTT or TTase. Interestingly GSH levels in these lenses remained relatively unchanged.
The present results showed that deletion of TTase gene in the mouse could lead to an early age-dependent cataract formation. The spontaneously accumulated PSSG in these lenses appeared to link directly to lens opacity.
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