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H. Yang, H. E. Grossniklaus; Pigment Epithelium-Derived Factor RNA Interference Promotes Ocular Melanoma Growth in a Mouse Model. Invest. Ophthalmol. Vis. Sci. 2010;51(13):864.
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Our previous studies demonstrated that overexpression of pigment epithelium-derived factor (PEDF) inhibited ocular melanoma growth and metastasis. The purpose of this study is to determine if down-regulation of endogenous PEDF affects the growth of ocular melanoma, and to know the functional consequences of PEDF down-regulation during malignant progression of ocular melanoma.
To establish a melanoma cell line which expresses the down-regulated PEDF gene, PEDF shRNA or control shRNA lentiviral particles were transduced into mouse melanoma cell line B16LS9. After puromycin selection for 2 weeks, PEDF expression was detected by RT-PCR and western blot. CyQUANT® cell proliferation assay was performed in B16LS9 with PEDF down-regulation (B16LS9-PEDFshRNA) and B16LS9 with control shRNA transduction (B16LS9-shRNA). 2.5x105/2µl of B16LS9-PEDFshRNA and B16LS9-shRNA cells were inoculated into posterior chamber in C57/BL6 mice. At the 7th day after inoculation, the eye was checked with histological methods, and the largest area of ocular tumor was measured under microscope with Image J.
PEDF expression was found in B16LS9-shRNA. The loss of PEDF expression in B16LS9-PEDFshRNA cells was confirmed using RT-PCR and western blot. Tumor cell proliferation in vitro was increased by PEDF knockout. In vivo experiments showed the area of ocular tumor with B16LS9-PEDFshRNA was significantly larger than one with B16LS9-shRNA (P<0.01).
Endogenous PEDF plays an important role in the proliferation of ocular melanoma. Loss of PEDF promotes the growth in ocular melanoma.
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