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V. N. Simirskii, Y. Wang, M. K. Duncan; Gene Expression Changes in the Mouse Lens After Conditional Knockout of the β1-Integrin Gene. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4351.
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Integrins are ECM and cell-cell adhesion receptors that are at the cross-roads of many signaling pathways. β1 integrin is a key integrin subunit of the mouse lens that forms a diversity of functional integrins. We previously demonstrated that conditional deletion of β1-integrin in both epithelial and fiber cells of the lens results in expression of -smooth muscle actin (SMA) in, and apoptosis of, lens epithelial cells. This work seeks to investigate the possible role of TGFβ/BMP-mediated signal transduction in this phenotype.
Mice homozygous for the floxed β1 integrin gene harboring MLR10-CRE were used. Semi-quantitative rt-PCR and confocal immunofluorescence were performed to characterize integrin expression and localization in embryonic and newborn lenses. Real-time rt-PCR arrays were used to profile the expression of 168 genes related to TGFβ/BMP-mediated signal transduction and/or cell-cell and cell-matrix interactions.
Three distinct steps were recognized in the progression of the β1-integrin null lens phenotype: Step 1 (11.5-15.5 dpc). Lens is morphologically normal and few changes in gene expression are detected. Particularly surprising was the maintained expression of integrins (2, 3, 4, 5) that are reported to be dependent on β1-integrin for stability. However, the down-regulation of 6 integrin and some laminins (Lama2, Lamc1) as well as the up-regulation of laminin β3, E-cadherin and Tgfβ2 was detected. Step 2 (16.5 dpc up to birth). The appearance and gradual intensification of cell abnormalities and the up-regulation of expression of epithelial-mesenchymal transition (EMT) markers (SMA, collagen I, Tgfβi, Igfbp3), coincident with the total disappearance of 2, 3 and 6 integrins. In contrast, V, β5 and β8 integrins are down-regulated in most lens fiber cells, but are up-regulated in lens epithelial cells. Step 3 (first days after birth). The epithelial cells undergo apoptosis coincident with the up-regulation of Tgfβ1 and Tgfβ3 expression and further up-regulation of collagen I, Tgfβi and Igfbp3. All steps are also characterized by up-regulation of JunB, inhibins (Inhba, Inhbb) and inhibitor of Cdk4 (Cdkn2b).
Conditional knockout of β1-integrin in the lens results in gradual loss of other integrins accompanying by remodeling of ECM, cell cycle changes and stepwise activation of EMT-specific genes beginning from 16.5 dpc. This is accompanied by increases in the expression of known EMT mediators including TGFβs.
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