Background Retinal pigment epithelial (RPE) cell transplantation is a novel approach to the treatment of hereditary retinal diseases, however, human-derived RPE cell line occurs de-differentiation during in vitro cell culture.Studies showed that early abnormal activation of Akt/mammalian target of rapamycin (mTOR) signal pathway is the primary cause of RPE cell line de-differentiation, therefore, the inhibition of mTOR pathway will be helpful for the retard of de-differentiation of RPE cells.Objective This study aimed to investigate whether rapamycin can suppress the activation of mTOR pathway and promote differentiation of ARPE19 cells.Methods ARPE-19 cells were incubated in 12-well plate and divided into control group and rapamycin-treated group.DMSO or rapamycin with the final concentration of 400 nmol/L was added in the medium of the control group and the rapamycin-treated group, respectively.The cells of each group were collected 24 hours and 48 hours after cultured.The expression of zonula occludens-1 (ZO-1) in the cells was examined by immunofluorescence.The relative expression levels of RPE cell specific genes and proteins were assayed by using real-time quantitative PCR and Western blot, respectively.The detected results were compared between the two groups.Results ZO-1 was expressed in both group,but the fluorescence intensity was evidently enhanced in the rapamycin-treated group.The relative expression levels of RPE65, MERKT and LRAT mRNA in the cells increased by 25.97％ , 29.71％ and 13.00％ in the rapamycin treated group compared with the control group 24 hours after cultured (P=0.04,0.04,0.04) , and the expression levels of RPE65, LRAT, rLBP1, BEST1 , keratin18 and MERKT mRNA elevated by 174.00％ , 88.00％ , 56.18％ ,193.81％ ,10.83％ and 35.02％ in the rapamycin-treated group in comparison with the control group 48 hours after cultured (P =0.00,0.04,0.01,0.04,0.04,0.03).In addition, the expressions of p-mTOR, p-P70S6 and p-S6 protein were weaker in the rapamyein-treated group than those in the control group both 24 hours and 48 hours after cultured.Twenty four hours after cultured,the expression level of ZO-1 protein raised by 40％ in the rapamycin-treated group compared with the control group (P =0.01);while 48 hours after cultured,the expression levels of ZO-1 ,MERKT, catenin and LRAT proteins elevated by 36.00％ ,57.37％, 13.68％ and 41.07％ in the rapamycintreated group in comparison with the control group (P=0.01,0.00,0.04,0.04).Conclusions Rapamycin can suppress the activation of mTOR signaling pathway and up-regulate the expressions of RPE specific genes in ARPE19 cells.Inhibition of mTOR pathway might be an effective way for culturing RPE cells in vitro.

Objective To observe the correlation between long-term chronic pharyngitis and rheumatic heart vavular disease (RHD) caused by to long-term latent chronic rheumatic activity and to understand the progressive course of rheumatic heart vavular disease. Methods In 1126 cases with chronic pharyngitis, 319 cases with serum antistreptolysin O (ASO) level between 400-500 U/ml were followed-up. ASO, creatine kinase enzyme MB (CK-MB) and echocardiography were measured for follow-up since 1986. Of the 319 cases, 158 were male and 161 were female with the average age of 29.4 years old. By the end of 2009, 6 cases were lost during follow up, data of 313 cases including 155 male patients and 158 female patients whose average age was 49.6 were analyzed. As the number of every kind of rheumatic heart vavular lesion was so fewer for statistical analysis that the data were only listed in tables. The student's t test was performed to compare of the ASO, CK-MB between the group with vavular lesionss and the group without vavular lesion. Results ①Of the 313 cases, 9 cases suffered from rheumatic fever at the fourth year since 1986, and 29 cases had rheumatic fever 1, 2 or 3 years after the ASOs decreased to lower than 400 U/ml and no one developed heart valvular lesion.②Two hundred and seventy-five cases whose ASO in the range of 400-500 U/ml but with normal CK-MB were found by the end of 5, 10, 15, 20 years, 9, 42, 65, 78 cases had developed heart vavular diseases respectively. ③ The levels of CK-MB in the heart valvular disease groups were significantly higher than those in the non-vavular disease group, while the levels of ASO were not. Conclusion Some of the cases suffering from long-term chronic pharyngitis can have high levels of ASO, but with normal CK-MB. These patients may have latent long-term chronic rheumatic activity and develop rheumatic heart valvular disease years later.