Protective effect of human umbilical cord mesenchymal stem cells on light-damaged retinal pigment epithelial cells

Objective

To investigate the protective effect of human umbilical cord mesenchymal stem cells (UCMSCs) on light-damaged retinal pigment epithelial (RPE) cells in vitro.

Methods

Human UCMSCs were cultured, and then identified by flow cytometry.Human RPE cells were isolated and cultured, and then the model of light-damaged RPE cells was prepared.The noncontact co-culture system of light-damaged RPE cells and UCMSCs was established by Transwell chamber.RPE cells were divided into normal control group, model control group and UCMSCs co-culture group.RPE cells in the normal control group were not treated.RPE cells in the model control group were treated with blue light to induce RPE cell damage.Co-culture system of light-damaged RPE cells and UCMSCs was established in the UCMSCs co-culture group.The proliferative ability of RPE cells was measured by methyl thiazolyl tetrazolium (MTT) assay at 24 hours and 48 hours after co-culture.ELISA kits were used to quantitatively measure the levels of pigment epithelium-derived factor (PEDF) and basic fibroblast growth factor (bFGF) in the culture supernatant at 48 hours after co-culture.And the photoreceptor outer segments (POS) phagocytosis assay of RPE cells was also conducted.

Results

UCMSCs displayed spindle-shaped morphology, positively expressed CD29, CD44, CD90 and CD105, while negatively expressed CD34 and CD45.RPE cells were polygonal in morphology and positive for the specific marker RPE65 protein.The proliferative ability(A value) of RPE cells in the three groups at different timepoints were significantly different (F_group=132.388, P=0.000; F_time=231.440, P=0.000), the A values of RPE cells in model control group and UCMSCs co-culture group were significantly lower than that in the normal control group, the Avalue of RPE cells in UCMSCs co-culture group was significantly higher than that in the model control group, and the differences were statistically significant both at 24 hours and 48 hours after co-culture (all at P<0.01). POS phagocytosis test showed that there was a significant difference in the average number of POS particles phagocytized by RPE cells among the three groups(F=28.087, P=0.000). The average number of POS particles phagocytized by RPE cells in model control group was significantly lower than that in normal control group, and the average number of POS particles phagocytized by RPE cells in UCMSCs co-culture group was significantly more than that in model control group (all at P<0.01). ELISA showed that the concentrations of PEDF in RPE cell supernatant of normal control group, model control group and UCMSCs co-culture group were (18.8±1.9), (10.0±1.7) and (20.2±6.0)ng/ml, respectively.The concentrations of bFGF in RPE cell supernatant were (25.2±1.5), (26.3±3.6) and (61.9±14.3)pg/ml, respectively.There were significant differences in PEDF and bFGF concentrations among the three groups (F=8.654, P=0.008; F=23.698, P=0.000). The concentration of PEDF in the model control group was significantly lower than that in the normal control group, and the concentration of PEDF in the UCMSCs co-culture group was significantly higher than that in the model control group (all at P<0.01). The concentration of bFGF in UCMSCs co-culture group was significantly higher than that in the normal control group and model control group (all at P<0.01).

Conclusions

Cocultivation with UCMSCs can improve the proliferation ability and phagocytosis function of light-damaged RPE cells, and promote the secretion of PEDF by RPE cells.UCMSCs may protect RPE cells from light damage through paracrine secretion of bFGF.