槲皮素脂质体对视网膜组织血管生成素样蛋白2及其受体表达的影响

Effect of quercetin liposome on angiopoietin-like protein 2 and its receptor Tie2 expression in the retina

背景糖尿病视网膜病变（DR）的特征性病理改变为微血管病变，血管生成素样蛋白2（Ang-2）是与血管发生密切相关的新蛋白，槲皮素具有多种药理作用，可改善微循环、降低毛细血管通透性，但其对DR的作用机制尚不清楚。目的探讨槲皮素脂质体对视网膜组织Ang-2及其受体（Tie2）表达的影响。方法选取60只清洁级雄性成年Wistar大鼠以随机数字表法分为6个组，其中10只大鼠作为空白对照组，其他60只大鼠采用链脲佐菌素（STZ）35mg／kg一次性腹腔内注射制作糖尿病模型。槲皮素分为50mg／（kg·d）（低剂量）、150mg／（kg·d）（中剂量）和250mg／（kg·d）（高剂量），并用脂质体包裹，分别对模型鼠进行灌胃3—5ml共12周，其他模型鼠分别用生理盐水和羟苯磺酸钙混悬液灌胃作为阴性对照和阳性对照。给药12周后处死各组大鼠并分离视网膜，用ELISA法检测视网膜组织中Ang-2的吸光度（A450）值，并用逆转录聚合酶链反应（RT—PCR）检测视网膜中Tie2mRNA的A值（Tie2mRNA／GAPDHmRNA）。结果ELISA检测表明，槲皮素中剂量组、高剂量组Ang-2的A450值分别为0．796±0．057和0．8424±0．043，较生理盐水组的1．012±0．046明显下降，差异均有统计学意义（q=2．95、2．698，P〈0．05）。RT—PCR检测表明，槲皮素中剂量组、高剂量组的Tie2mRNA在视网膜中的表达量分别为0．712±0．092和0．821±0．087，均明显低于阴性对照组的1．182±0．098，差异均有统计学意义（q=3．497、2．852，P〈0．05）。Ang-2蛋白及Tie2mRNA表达量的下降均以中剂量组最为明显。结论槲皮素脂质体对视网膜中Ang-2及其受体Tie2的表达有抑制作用，可对糖尿病大鼠的视网膜微循环异常有改善作用。

Background The special pathological change of diabetic retinopathy （DR）is microvascular disorder. Angiopoietin-like protein 2（Ang-2） is a new protein associated with genesis of blood vessels. Quercetin has multiple pharmacological action, including improving the microcircularion and the permeability of blood capillary. However,the action mechanism of Ang-2 on DR was unclear. Objective The present study was to investigate the effects of quercetin on Ang-2 and its receptor Tie2 expression in retina with diabetes mellitus. Methods Sixty clean male Wistar rats were randomized into 7 groups and 10 rats for each group,and 10 rats served as blank control group. Streptozotocin of 35 mg/kg was intraperitoneally injected in 60 rats to establish the diabetic models. Quercetins encapsulated by liposome with the doses of 50,150 and 250 mg/（ kg·d） （3-5 ml）were used to garage in different groups of models for 12 weeks,and normal saline solution and calcium dobesilate were used at the same fashion as the negative control group and positive control group, respectively. Twelve weeks later, the animals were sacrificed and retinas were isolated. Expressions of Ang-2 protein and Tie2 mRNA in retinas were detected by ELISA and RT-PCR, respectively. The usage and rearing of the animals complied with the Regulations for the Administration of Affair Concerning Experimental Animals by State Sciences and Technology Commission. Results ELISA showed that the A450 of Ang-2 in 150 and 250 mg/（ kg·d） quercetin groups was 0. 796±0. 057 and 0. 842±0. 043 respectively and was lower than that negative group （ 1. 012 ± 0. 046 ） , showing statistically significant differences （ q = 2.95,2. 698,P〈0.05 ）. RT-PCR assay showed that expression of Tie2 mRNA （Tie2 mRNA/GAPDH mRNA）in retinas was 0. 712±0. 092 and 0. 821 ±0. 087, presenting statistically significant differences in comparison with negative group（ 1. 182± 0. 098） （ q = 3. 497,2. 852, P〈0.05 ）. The expression levels of Ang-2 and Tie2 mRNA in retina were lowest in 150 mg/（kg·d） quereetin group. Conclusions Quercetin can improve the retinal microcireulation by downregulating the expressions of Ang-2 and its receptor in early period of diabetic rats.