The Effects Of Rapamycin On Proliferation Of Human Retinal Pigment Epithelial Cell Cultured In Vitro

A large number of studies have shown that retinal pigment epithelial (RPE) cells are the major source of all PVR cell membranes. How to control the abnormal behavior of RPE cells and inhibit the proliferation of other related cells play a important role in researchs preventions and cures of PVR. Rapamycin (RAPA)is a new macrolide immunosuppressant, which can inhibit the proliferation of many kinds of cell. In Ophthalmology,it has been used in drugs test in animal and cell cultured in vitro.Objective:This study aimed to observe the bioactivity of human retinal pigment epithelial (hRPE) cell cultured in vitro,and investigate the inhibitory reaction of different concentrations rapamycin on the morphology, proliferation and other biological activity of hRPE cells.Methods:To culture hRPE cells in vitro and divide into six groups with concentrations of rapamycin (Ong/ml,5ng/ml,10ng/ml,20ng/ml,40ng/ml,80ng/ml). To observe hPRE cells under the microscope, and use the ways of doubling time, cell viability, growth curve and the tetrazolium salt (MTT) assay for measuring the impact of RAPA on the proliferation of hRPE cells.Results:Recovered hRPE cell lines and obtained well vitality cells with visible.At a certain concentration range,RAPA inhibited the proliferation of hRPE cells. It could be seen that the number of cells gradually decreased with the concentration of RAPA increased under light microscope.Cells sticked to culture plates extened with the shape of shuttle or polygon. After HE stained, cells in control group were fusiform or irregular polygon with abundant cytoplasm showing Eosin Red; nuclear was slightly smaller, center and lightly stained. Cells that treated with80ng/ml rapamycin after72h stretched or collapsed, occasionally being floating. Cells showed obvious decrease. The cell division time of control group was1.83days, cell division time of all groups extended with the concentration of RAPA increased. Growth curve showed that hRPE cells significantly inhibited with increasing concentrations and extending time of rapamycin. It proved time-dose dependent manner of inhibition. Comparing cell growth state of control group and treatment groups, showed that the growth index of control group was higher than that of the treatment groups. Trypan blue cell viability determination showed each group cell viability was92.3%,89.3%,88.7%,87.7%,86.0%,82.6%.MTT assay showed the basic law:the results indicated that the inhibition rate of RAPA were8.70%,22.06%,40.69%,42.31%,43.12%for24h and12.57%,26.35%,42.23%44.15%,47.12%for72h.Conclusions:1Rapamycin can inhibit the proliferation of hRPE cells cultured in vitro It showed time-dose dependent manner of inhibition. During the10-80ng/ml concentration range, RAPA inhibited hRPE cell proliferation significantly (P<0.01); but during the20-80ng/ml concentration range, inhibition among the three groups was not different significantly (P>0.05).2rapamycin extended doubling time of hRPE cell,and reduced the activity of hRPE cells cultured in vitro.3The prohibition rate was about50%at the concentration of80ng/ml.4This work provided some experimental basis in further animal and clinical experiments.The inhibition effects of rapamycin on hRPE cell might provide a new clinical prevention and treatment option of PVR to further expand its application in the field of Ophthalmology.