Effect of Jiajian-Zhujing Decoction on the AKT/mTOR signaling pathway in ARPE-19 cells after AKT transfection

Abstract

Objective To study the mechanism of the effect of Jiajian-Zhujing Decoction on the expression of VEGF on ARPE-19 cells after AKT transfection. Methods To prepare the serum and blank serum of Jiajian-Zhujing Decoction and divide ARPE-19 cells into the normal group, model group, blank serum group, medicated serum group, Conbercept group and combined group. Except normal group, this research established AKT transfected cell model. Then cultured the normal group and model group with conventional method, and the blank serum group was cultured with 10% blank serum, the medicated serum group was cultured with 10% medicated serum, the Conbercept group was cultured with 20 μg/ml Conbercept, the combined group was cultured with 10% medicated serum and 20 μg/m Conbercept. The proliferation of ARPE-19 cells in each group was detect by the CCK-8 method. The levels of AKT, mTOR and VEGF mRNA were detected by real-time quantitative PCR. Western blot was used to detect the expression of AKT, mTOR and VEGF. Results After being cultured for 24, 48 and 72 hours, compared with the model group, the cell proliferation rate in blank serum group, medicated serum group, Conbercept group and combined group significantly decreased (P<0.05). Compared with the model group, the expression of AKT mRNA (24 h: 3.10 ± 0.48, 1.97 ± 0.14, 1.26 ± 0.24 vs. 4.77 ± 0.68; 48 h: 3.52 ± 0.82, 2.62 ± 0.77, 1.10 ± 0.19 vs. 6.12 ± 1.21), mTOR mRNA (24 h: 3.02 ± 0.26, 2.45 ± 0.75, 1.13 ± 0.15 vs. 4.48 ± 0.80; 48 h: 1.29 ± 0.30, 1.30 ± 0.57, 0.65 ± 0.19 vs. 2.54 ± 0.62), VEGF mRNA (24 h: 3.33 ± 0.62, 2.18 ± 0.20, 1.55 ± 0.28 vs. 5.53 ± 1.02; 48 h: 2.35 ± 0.54, 1.23 ± 0.28, 0.93 ± 0.25 vs. 3.59 ± 0.40), AKT protion (24 h: 0.45 ± 0.09, 0.25 ± 0.05, 0.14 ± 0.04 vs. 0.62 ± 0.04; 48 h: 0.36 ± 0.06, 0.23 ± 0.04, 0.14 ± 0.03 vs. 0.54 ± 0.08), mTOR protion (24 h: 0.35 ± 0.05, 0.24 ± 0.02, 0.18 ± 0.02 vs. 0.52 ± 0.09; 48 h: 0.23 ± 0.04, 0.29 ± 0.04, 0.14 ± 0.03 vs. 0.40 ± 0.10), VEGF protion (24 h: 0.14 ± 0.03, 0.33 ± 0.04, 0.24 ± 0.03 vs. 0.54 ± 0.10; 48 h: 0.24 ± 0.03, 0.17 ± 0.02, 0.11 ± 0.02 vs. 0.42 ± 0.10) significantly decreased (P<0.05), and the combined group was significantly lower than that of the Conbercept group (P<0.05). Conclusions AKT transfection can promote the proliferation of ARPE-19 cells, and Jiajian-Zhujing Decoction can significantly inhibit this proliferation. Jiajian-Zhujing Decoction may inhibit the activity of AKT/mTOR signaling pathway to reduce the expression of VEGF. Key words: Serum pharmacology (TCD); Retinal pigment epithelium; Transfection; Vascular endothelial growth factors; Jiajian-Zhujing Decoction