基于HIF-1α/VEGF信号通路探讨丹参提取物对糖尿病视网膜病变大鼠模型的影响

Effect of Danshen(Salvia miltiorrhiza)Extracts on Prevention and Treatment of Diabetic Retinopathy in Rat Model Based on HIF-1α/VEGF Signaling Pathway

目的 探讨丹参提取物通过低氧诱导因子-1α (Hypoxic inducible factor-1α,HIF-1α)/血管内皮生长因子(Vascular endothelial growth factor, VEGF)信号通路对糖尿病视网膜病变(Diabetic retinopathy, DR)大鼠模型的影响。方法 高脂高糖饲养+腹腔注射链脲佐菌素(Streptozotocin, STZ)建立DR大鼠模型,将其分为模型对照组、阳性对照组(芪明颗粒1.42 g/kg)、丹参水提物组(0.16 g/kg)和丹参酮提取物组(0.12 g/kg),连续灌胃4周,然后检测大鼠的空腹血糖(Fasting blood glucose, FBG)、甘油三酯(Triglyceride, TG)、总胆固醇(Total cholesterol, TC)、低密度脂蛋白胆固醇(Low density lipoprotein cholesterol, LDL-C)和高密度脂蛋白胆固醇(High density lipoprotein cholesterol, HDL-C)的水平;HE染色观察视网膜组织形态变化,测量视网膜外核层(Outer nuclear layer, ONL)、内核层(Inner nuclear layer, INL)和神经节细胞层(Ganglion cell layer, GCL)的厚度;酶联免疫吸附测定法(Enzyme linked immunosorbent assay, ELISA)检测血清中糖化血红蛋白(Glycosylated hemoglobin, HbA1c)、糖基化终末产物(Advanced glycation end products, AGEs)和视网膜中白细胞介素-1β (Interleukin-1β,IL-1β)、白细胞介素-10 (Interleukin-10,IL-10)、肿瘤坏死因子-α (Tumor necrosis factor-α,TNF-α)与转化生长因子-β1 (Transforming growth factor-β1,TGF-β1)的水平,免疫组化法检测视网膜中HIF-1α和VEGF的表达。结果 较正常对照组,模型对照组中视网膜组织结构排列紊乱,各结构层厚度减少,FBG、TG、TC和LDL-C水平显著上升(P<0.01),HDL-C水平显著下降(P<0.05),血清中HbA1c和AGEs水平显著升高(P<0.01),视网膜中IL-1β、TNF-α、TGF-β1、HIF-1α和VEGF表达水平显著升高(P<0.01),IL-10表达水平显著降低(P<0.01);与模型对照组相比,丹参水提物和丹参酮提取物减轻了视网膜组织病变程度,降低了血糖和血脂的水平(P<0.05或P<0.01),减少了血清中HbA1c和AGEs的含量(P<0.01),IL-10和TGF-β1改善无统计学意义,IL-1β、TNF-α、HIF-1α和VEGF表达降低(P<0.05或P<0.01)。结论 丹参提取物能改善DR大鼠视网膜组织的病变,其机制可能是改善血糖血脂、炎性反应以及抑制HIF-1α/VEGF通路。

Objective To investigate the effect of Danshen(Salvia miltiorrhiza)extracts on diabetic retinopathy(DR)rat model via hypoxic inducible factor-1α(HIF-1α)/vascular endothelial growth factor(VEGF)signaling pathway.s Methods High fat and high sugar feeding+intraperitoneal injection of streptozotocin(STZ)were used to establish the DR model rat.DR rats were divided into model control group,positive control group[Qiming Granules(芪明颗粒),1.42 g/kg],water extract of Danshen(Salvia miltiorrhiza)group(0.16 g/kg)and tanshinones group(0.12 g/kg),and the drugs were given by gavage continuously for 4 weeks.Then,the levels of fasting blood glucose(FBG),triglyceride(TG),total cholesterol(TC),low density lipoprotein choles-terol(LDL-C)and high density lipoprotein cholesterol(HDL-C)in rats were determined.The morphological changes of reti-nal tissue were observed by HE staining,and the thickness of the outer nuclear layer(ONL),inner nuclear layer(INL)and gan-glion cell layer(GCL)of the retina were measured.The levels of glycosylated hemoglobin(HbA1c),advanced glycation end products(AGEs)in serum and levels of interleukin-1β(IL-1β),interleukin-10(IL-10),tumor necrosis factor-α(TNF-α)and transforming growth factor-β1(TGF-β1)in the retina were detected by enzyme linked immunosorbent assay(ELISA),and the expressions of HIF-1αand VEGF in the retina were detected by immunohistochemistry.s Results Compared with those of the normal control group,the retinal tissue structure was disorganized,and the thickness of each structural layer was reduced in the model control group.The levels of FBG,TG,TC and LDL-C were significantly increased(P<0.01),and the levels of HDL-C were significantly decreased(P<0.05),and the levels of HbA1c and AGEs in serum were significantly increased(P<0.01).The levels of IL-1β,TNF-α,TGF-β1,HIF-1αand VEGF in the retina were significantly increased(P<0.01),and the levels of IL-10 were significantly decreased(P<0.01).Compared with those of the model control group,water extract of Danshen(Salvia miltiorrhiza)and tanshinones reduced the degree of retinal histopathy,decreased the blood glucose and lipids(P<0.05 or P<0.01),reduced the serum levels of HbA1c and AGEs(P<0.01),improved the levels of IL-10 and TGF-β1,and the levels of IL-1β,TNF-α,HIF-1αand VEGF decreased(P<0.05 or P<0.01).s Conclusions Danshen(Salvia miltiorrhiza)extracts improved retinal tissue lesions in DR rats,and the mechanism may be related to its improvement of glucose lipids,inflammatory response and inhibition of HIF-1α/VEGF pathway.