Objective To investigate the feasibility of Y27632 to induce transdifferentiation from human retinal pigment epithelial (hRPE) cells into neuron-like cells in vitro. Methods The third to sixth generation of primary hRPE cells were cultured with 2% fetal bovine serum + Dulbecco's modified eagle medium/F12 culture solution, with (experimental group) or without (control group) 10 mu;mol/L Y27632. At 3, 6 hours and 1, 3, 5, 7 days after induction, the morphologic changes of RPE cells were observed by inverted microscope. The expression rate of CK18, Map2, NF200 and Pax6 at 3 days after induction in the experimental and control group were detected by immunofluorescent staining. chi;2 test was employed for comparison between the two groups. Results 50.1% cells of the experimental group formed axon-like processes and interconnected each other with typical neuron-like appearance. The expression rates of CK18, Map2, NF200 and Pax6 in the experimental group were 43.88%, 31.90%, 57.45% and 65.79%, while the above indexes in the control group were 93.97%, 4.49%, 22.37% and 8.33% respectively. Compared the expression rate of CK18 (chi;2=64.763), Map2 (chi;2=23.634), NF200 (chi;2=21.261) and Pax6 (chi;2=25.946) between the two groups, the differences were significant (P＜0.01). Conclusion The hRPE cells can be trans-differentiated into neuron-like cells in vitro by Y27632.
ObjectiveTo study the role of Rac1 in the epithelial-mesenchymal transition (EMT) process of retinal pigment epithelial cells (RPE) induced by transforming growth factorβ(TGF-β). MethodsHuman ARPE-19 cells were divided into 4 groups including control group, TGF-βgroup, TGF-β+NSC23766 group, NSC23766 group. NSC23766 was added to medium 2 hours before TGF-βtreatment to block the Rac1 receptors.α-smooth muscle actin (α-SMA) expression was measured by immunofluorescence and Western blot. Cell scratch assay, invasion assay and gel contraction experiments were used to measure cell migration, invasion, cell contraction. ResultsThe expression ofα-SM A was higher in TGF-βgroup, compared with the control group, TGF-β+NSC23766 group (F=825.314, P < 0.05). Cell scratch assay showed that the cellular gap was less in GF-βgroup, compared with the control group, TGF-β+NSC23766 group, NSC23766 group (F=177.351, P < 0.05). Cell invasion assay showed that, the number of cells pass through the fiber membrane was the same in TGF-βgroup and other 3 groups (F=0.371, P=0.055). Gel contraction assay showed that TGF-βcan promote the cellular contraction, compare to the control group, TGF-β+NSC23766 group, NSC23766 group, the difference was statistically significant (F=40.473, P < 0.05). ConclusionRac1 play a role in TGF-β-induced behavioral changes of RPE cells; NSC23766 inhibit RPE cellular behavior change by regulating Rac1 activation.