Effect of Zhixue Mingmu Formula on Retinal Vasopermeability of Streptozotocin-Induced Diabetic Retinopathy in Rats
Authors: Jia Xin , Zhang Yue , Zhang Minglian , Yang Zanzhang , Liu Yinglin , Li Chengquan , Li Mingran 
Published: March 2022

Aims: To investigate the efficacy of Zhixue Mingmu Formula (ZXMM), a traditional Chinese herbal compound, for retinal vasopermeability in streptozotocin (STZ)-induced diabetic retinopathy (DR) in rats.
Materials and Methods: This experimental study was performed at the Hebei Provincial Key Laboratory of Ophthalmology, Hebei Provincial Eye Hospital within 6 months from January 2021 to June 2021. A total of 70 Sprague-Dawley rats aged 7-8 weeks old and weighing 200-240 g were selected. Animals were categorized into 2 groups:

A control group (n = 10): normal rats - receive vehicle.
A model group (n = 60): diabetic rats - receive 1% STZ in 0.1 mol/L sodium citrate buffer (45 mg/kg, i.p.).

Diabetes was induced via a single intraperitoneal injection of 45 mg/kg STZ. After 72 hours, fasting blood glucose (FBG) levels were measured and FBG ≥16.7 mmol/L was taken as diabetes. After 4 weeks of rearing, 10 normal rats and 50 diabetic rats which were diagnosed with retinopathy by fundus fluorescence angiography were further divided into 6 groups:

A normal control group (NG, n=10): normal rats - receive 0.9% normal saline.
A model control group (MG, n=10): DR rats - receive 0.9% normal saline.
A calcium dobesilate group (CD, n=10): DR rats + calcium dobesilate (135 mg/kg/day, p.o.) daily.
A low-dose ZXMM group (LZXMM, n=10): DR rats + low-dose ZXMM (15.3 g/kg/day, p.o.) daily.
A moderate-dose ZXMM group (MZXMM, n=10): DR rats + moderate-dose ZXMM (30.6 g/kg/day, p.o.) daily.
A high-dose ZXMM group (HZXMM, n=10): DR rats + high-dose ZXMM (45.9 g/kg/day, p.o.) daily.

Drug treatments were administered daily for 4 weeks. FBG levels were monitored every 2 weeks. Retinal vasopermeability was measured using an Evans blue assay. Then, serum samples and retinal tissues were harvested to assess the levels of inflammatory factors and the activity of antioxidative enzymes by enzyme-linked immunosorbent assay (ELISA). Retinal hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor-A (VEGF-A) protein levels were evaluated by ELISA.
Results: In the MG, FBG levels were significantly (P < 0.001) increased, and retinal vasopermeability was markedly (P < 0.001) elevated compared with those in the NG. After oral administration of ZXMM or calcium dobesilate, the elevated retinal vasopermeability was significantly (P < 0.001) decreased in DR rats. Concurrently, ZXMM treatment significantly (P < 0.05) attenuated the levels of the inflammatory factors interleukin-6 (IL-6) and C-reactive protein (CRP) in serum and ameliorated the activity of the antioxidative enzymes superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in the retina of DR rats. ZXMM treatment significantly (P < 0.05) decreased HIF-1α and VEGF-A expression in retinal tissues compared with MG treatment. Furthermore, the effects of ZXMM were dose dependent.
Conclusion: ZXMM was shown to have a protective effect against retinal vasopermeability in DR comparable to that of calcium dobesilate, and its mechanism may be attributed to inhibition of the downstream pathway of glycaemia-induced hypoxia, inflammation and oxidative stress of retinal tissue.