Inhibitory effects of jiajianzhujing decoction containing serum on proliferation of rat choroidal vascular endothelial cells

Background

Choroidal neovascularization (CNV) of macular zone is one of blinding eye diseases.Conventional treatment methods include photodynamic therapy and intravitreal injection of anti-vascular endothelial growth factor (VEGF) drugs.However, there are some adverse effects following these treatments.Researches showed that jiajianzhujing decoction, a Chinese herbal medicine, can inhibit CNV.

Objective

This study was to investigate the impact of drug serum of jiajianzhujing decocation (drug serum) on the proliferation of rat choroidal vascular endothelial cells (RCVECs).

Methods

Jiajianzhujing decoction (0.525 g/ml) was used by intragastric administration with the dosage 20 ml/kg for 3 days, and the abdominal aortic blood was collected 2 hours after last dosage to prepare the drug serum.Cultured RCVECs were divided into 4 groups.CoCl₂ with the concentration of 100 μmol/L (100 μl) was added in the medium to establish the anoxic models in the CoCl₂ group, and 20% drug serum was added in the anoxic culture medium in the drug serum+ CoCl₂ group.Regular culture medium was used in the blank control group, and 20% pure serum was added in the anoxic culture medium in the pure serum+ CoCl₂ group.The cells were incubated for consecutive 24 hours, and the growth of the cells was detected by monotetrazolium (MTT) assay as absorbance (A). The expressions of gene and protein of hypoxia-inducible factor-1α (HIF-1α) and VEGF in the cells were detected by reverse transcription PCR and Western blot, respectively.This study was approved by the Ethics Committee of Eye Hospital of China Academy of Chinese Medical Sciences.

Results

Cultured cells grew well with fusiform shape.Hypoxic cell models were created by adding CoCl₂.The mean Avalues were 0.659±0.051, 0.757±0.553, 0.683±0.037 and 0.731±0.038 in the blank control group, CoCl₂ group, drug serum+ CoCl₂ group and pure serum+ CoCl₂ group, respectively, and the A value in the CoCl₂ group was significantly elevated in comparison with the blank control group and drug serum+ CoCl₂ group (both at P<0.01). The significant differences were found in the relative expressions of HIF-1α and VEGF among the blank control group, CoCl₂ group, drug serum+ CoCl₂ group and pure serum+ CoCl₂ group (HIF-1α: F=3.100, P<0.05; VEGF mRNA: F=3.420, P<0.05; HIF-1α protein: F=470.600, P=0.000; VEGF protein: F=146.700, P=0.000), and the relative expressions of HIF-1α mRNA, VEGF mRNA, HIF-1α protein and VEGF protein in the CoCl₂ group were significantly higher than those in the blank control group and drug serum+ CoCl₂ group (all at P<0.05).

Conclusions

Drug serum of jiajianzhujing decoction can inhibit the growth and proliferation of hypoxic RCVECs by down-regulating the secretion of HIF-1α and VEGF.

Hypoxia/chemistry-induced; Choroidal vascular endothelial cells; Chinese herbal medicine; Rats, Sprague-Dawley; Drug serum; Jiajianzhujing decoction; Cultured, cells