Objective To investigate the activation and role of signal transduction pathway of epidermal growth factor (EGF)-epidermal growth factor receptor (EGFR)-mitogen activated protein kinase (MAPK) in proliferation of human retinal pigment epithelial (RPE) cells. Methods Human RPE cells were stimulated with 0.1%，10% foetal calfserum (FCS) and EGF(0.1, 1, 10, 50 and 100 ng/ml)in 0.1% FCS Dulbeco′s modified Eagle′s medium (DMEM) and in 10% FCS DMEM for 3 days, respectively. Immunohistochemical staining and in situ hybridization were used to observe the expressions of EGFR protein and EGFR mRNA，respectively. Activation of MAPK was detected by immunohistochemical method with specific anti-phosphorylated ERK 1/2 antibody. Results The optimal concentrations of EGF were 10 ng/ml in 0.1% FCS DMEM and 1 ng/ml in 10% FCS DMEM. After 3 days of stimulation with EGF, phosphorylated ERK 1/2 staining was detectable in nucleus of RPE cells, whereas cells presented immunostaining for phosphorylated ERK 1/2 in the cytoplasm before stimulation. Conclusions EGF may improve the expression of EGFR protein and EGFR mRNA of RPE cells, and induced MAPK nuclear translocation in a concentration-dependent manner. EGF-EGFR-MAPK signal transduction pathway may play a key role in RPE cells proliferation, and serum exerts an important acceclerating function in the process. (Chin J Ocul Fundus Dis，2004，20：67-132)
ObjectiveTo investigate the expression and mechanism of miR-1470 in plasma of diabetic retinopathy (DR) patients.MethodsThirty patients with DR (DR group), 30 patients with diabetes (DM group) and 30 normal healthy subjects (normal group) were enrolled in the study. Three groups of subjects were taken 5 ml of venous blood, and total plasma RNA was extracted and purified. The differentially expressed miRNAs in the plasma of DR patients were screened by gene chip, and the results of gene chip detection were verified by reverse transcription polymerase chain reaction (RT-PCR). Bioinformatics was used to predict potential target genes for miRNA regulation, and miR-1470 and its target gene epidermal growth factor receptor (EGFR) were screened. Human retinal microvascular endothelial cells (hREC) were divided into normal group (sugar concentration 5.5 mmol/L) and high glucose group (sugar concentration 25.0 mmol/L). hREC was transfected into miR-1470 mimics to establish a miR-1470 high expression cell model, which was divided into blank control group, high expression group and negative control group. The expression of miR-1470 was detected by RT-PCR. The expression of EGFR protein was detected by Western blot. The measurement data of the two groups were compared using the independent sample t test. The comparison of the measurement data between the two groups was analyzed by ANOVA. The comparison between the measurement data of the groups was compared by multiple comparisons.ResultsThe results of RT-PCR were consistent with those of the gene chip. The expression of miR-1470 in the plasma of the DR group, the DM group and the normal group was statistically significant (F=63.486, P=0.049). Compared with the DM group and the normal group, the expression of miR-1470 in the DR group was significantly decreased, and the difference was statistically significant (q=111.2, 73.9; P＜0.05). The expression of miR-1470 in hREC in the high glucose group was significantly lower than that in the normal group (t=42.082, P=0.015). The expression of EGFR protein in hREC of high glucose group was significantly higher than that of normal group (t=−39.939, P=0.016). The expression of miR-1470 (F=637.069, P=0.000) and EGFR (F=122.908, P=0.000) protein expression in hREC of blank control group, negative control group and high expression group were statistically significant . Compared with the blank control group and the negative control group, the expression of miR-1470 in hREC of high expression group was significantly increased (q=329.7, 328.8; P＜0.05), and the expression of EGFR protein was significantly decreased (q=242.5, 234.6; P＜0.05). There was no significant difference in the expression of miR-1470 and EGFR protein in hREC between the negative control group and the blank control group (q=1.5, 7.9; P＞0.05).ConclusionThe expression of miR-1470 in the plasma of patients with DR is significantly down-regulated, and the increase of EGFR expression may be related to it.