PurposeTransforming growth factor (TGF)-β is a key mediator of proliferative vitreoretinopathy, but the cellular mechanisms by which TGF-β induces extracellular matrix protein (ECM) synthesis are not fully understood. This study examined whether the PI3K/Akt pathway is involved in TGF-β2-induced collagen expression in human retinal pigment epithelial cells.MethodsHuman retinal pigment epithelial cells ARPE-19 were cultured and stimulated with TGF-β2. The role of the PI3K/Akt pathway was evaluated using the biochemical inhibitor, wortmannin. The effect of wortmannin on the expression of type I collagen mRNA (COL1A1, COL1A2) induced by TGF-β2 was evaluated by real-time RT-PCR. The effect of wortmannin on the synthesis of type I collagen induced by TGF-β2 was assessed by an immunocytochemical analysis with anti-type I collagen antibody. Luciferase reporter assays were performed to examine the effect of wortmannin on the transcriptional activities of COL1A2. A luciferase assay using a mutation construct of the Smad binding site in COL1A2 promoter (Smad-mut/Luc) was also performed to examine the crosstalk between the Smad pathway and the PI3K/Akt pathway. The effects of wortmannin on the transcriptional activity of Smad3 were also examined using CAGA12-Luc. Moreover, the effect of wortmannin on TGF-β2-induced Smad7 mRNA expression was evaluated.ResultsThe biochemical blockade of PI3K/Akt activation inhibited TGF-β2-induced type I collagen mRNA expression and type I collagen synthesis. The blockade of PI3K/Akt pathway inhibited the increase in COL1A2 promoter activities when induced by TGF-β2 and reduced TGF-β2 induction of Smad-mut/Luc promoter activity and CAGA12-Luc activity. Moreover, wortmannin increased the TGF-β2-induced Smad7 mRNA expression levels.ConclusionsThe PI3K/Akt pathway plays a role in relaying the TGF-β2 signal to induce type I collagen synthesis in the retinal pigment epithelium through Smad-dependent and Smad-independent pathways.

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In bilaterally enucleated White-crowned Sparrows, Zonotrichia leucophrys gambelii, testicular growth and development at rates similar to those of intact birds subjected to long days, can be induced by illumination of sites within and near the ventromedial hypothalamus from the tips of single, chronically implanted, light-conducting fibers. Essentially identical results are obtained with the same mode of encephalic illumination of birds with intact eyes held on short days that are otherwise nonstimulatory. Examination of the distribution of the sites of the fibers from which weak illumination induced testicular growth, increase in plasma level of luteinizing hormone or increase in plasma level of testosterone suggests that most of the photoreceptors must lie either within the ventromedial hypothalamus or in sites ventral thereto, such as the tuberal complex. For the time being, at least, we find it necessary to assume that some photosensitive elements of the system occur outside of the ventral hypothalamus.

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