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D. Gupta, D. Kenchegowda, J. Piatigorsky, S. K. Swamynathan; Role of Krüppel-Like Factor 4 in Mouse Lens Gene Expression, Development and Function. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3457.
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© ARVO (1962-2015); The Authors (2016-present)
We have previously reported that the Klf4-conditional null (Klf4CN) lens is defective. Here, we have compared the wild type (WT) and Klf4CN lens gene expression patterns to understand the role of Klf4 in the lens gene expression, function and development.
Developmental expression of Klf4 was studied by Q-RT-PCR using standard curve method. Lens morphology was studied by light and electron microscopy. WT and Klf4CN lens gene expression patterns were compared by microarray, validated by RT-PCR and analyzed using BRB-ArrayTools and Ingenuity Pathway Analysis tools. Influence of KLF4 on selected promoter activities was measured by cotransfection assays. Fluorimetric method was used to measure glutathione levels in WT and Klf4CN lens.
Expression of Klf4, first detected in the embryonic day-12 (E12) mouse lens, peaked at E16, and steadily declined with age. Mean diameter of the 8 week-old Klf4CN lens was 58% of the age matched WT lens. Klf4CN lens developed structural deformity and central opacity, unlike the normal WT lens. Gene expression comparison by microarray identified 226 and 276 genes up- and down-regulated by more than 2-fold, respectively, in the Klf4CN lens. Alox12 and Alox15 were up-regulated while several crystallins were down-regulated in the Klf4CN lens. Co-transfection with pCI-Klf4 stimulated the Alox12, Alox12e, Alox15 and Shsp/αB-crystallin promoter activities by 4- to 20-fold. Pathway analysis identified the lipoxygenase pathway as one of the significantly affected pathways in the Klf4CN lens. Reduced glutathione (GSH) levels were lower while oxidized glutathione (GSSG) levels were higher in the Klf4CN lens than the WT, indicating that the Klf4CN lens is oxidatively stressed.
The expression of Klf4 is developmentally regulated in the mouse lens, where it contributes to different pathways including arachidonic acid metabolism, aryl hydrocarbon receptor signaling, and glutathione metabolism.
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