Expression of Robo4 in the fibrovascular membranes from patients with proliferative diabetic retinopathy and its role in RF/6A and RPE cells

[摘要] Purpose: Robo4, a member of theroundabout (Robo) family, acts as a neuronal guidance receptor andplays some role in vasculogenesis and angiogenesis. This studyinvestigated the effect of Robo4 on the formation of fibrovascularmembranes (FVMs) from patients with proliferative diabetic retinopathyand its roles in choroid–retina endothelial (RF/6A) and human retinalpigment epithelial (RPE) cells. Methods: RT–PCR and immunohistochemistrywere used to determine the levels of mRNA and the presence anddistribution of Robo4 in FVMs. Small interfering RNA (siRNA) technologywas used to knock down Robo4 expression and to study its effects onRF/6A and RPE cells in vitro. Cell proliferation, migration, spreading,cycling, and apoptosis were assessed with MTT assay, Boyden chamberassay, immunocytochemistry, and flow cytometry. Tube formation by RF/6Aon Matrigel was also analyzed. Results: The level of Robo4 mRNAwas high in FVMs. Robo4 was expressed in the vessels andfibrous-like tissue co-immunostained for CD31 and GFAP, respectively. Robo4siRNA knockdown inhibited cell proliferation and migration. Tubeformation by RF/6A cells was also disturbed. Under hypoxic conditions,more apoptotic cells were evident among the knockdown cells than amongthe control cells (p<0.01). Conclusions: Robo4 may play a role inthe formation of FVMs. Silencing the expression of Robo4 inRF/6A and RPE cells inhibited their proliferation and reduced theirtolerance of hypoxic conditions, suggesting physiologic functions ofRobo4 in the cells of the retina.

[学科分类] 生物化学/生物物理