丹蒌片通过Keap1-Nrf2/HO-1信号通路缓解视网膜缺血-再灌注损伤

Danlou tablet alleviates retinal ischemia-reperfusion injury through Keap1-Nrf2/HO-1 pathway

摘要:

目的：探讨丹蒌片对小鼠视网膜缺血-再灌注损伤(RIRI)的保护作用及其机制。

方法：将40只ApoE^-/-小鼠给予高脂饲料喂养6 wk，通过前房灌注加压法建立RIRI模型，分为对照组(给予生理盐水灌胃8 wk)、RIRI模型组(给予生理盐水灌胃8 wk)及丹蒌片低、中、高剂量组\〖分别给予丹蒌片溶液1、2、4 g/(kg·d)灌胃8 wk\〗。采用苏木精-伊红(HE)染色观察各组小鼠视网膜组织形态学变化情况，TUNEL染色检测各组小鼠视网膜细胞凋亡情况，Western-blot法检测各组小鼠视网膜组织中Kelch样环氧氯丙烷相关蛋白-1(Keap1)、转录因子核因子E2相关因子2(Nrf2)、血红素加氧酶-1(HO-1)、超氧化物歧化酶(Sod2)蛋白表达情况。

结果：与对照组比较，RIRI模型组小鼠视网膜萎缩，厚度变薄，细胞凋亡增加，视网膜组织中Sod2蛋白表达下调，Keap1蛋白表达上调(均P<0.01)； 与RIRI模型组比较，丹蒌片中、高剂量组小鼠视网膜厚度增加(均P<0.01)，丹蒌片低、中、高剂量组小鼠视网膜细胞凋亡降低(均P<0.05)，丹蒌片低剂量组小鼠视网膜组织中Keap1和HO-1蛋白表达水平无明显差异(P>0.05)，丹蒌片中、高剂量组小鼠视网膜组织中Sod2、Nrf2、HO-1蛋白表达上调，Keap1蛋白表达下调(均P<0.05)。

结论：丹蒌片能缓解RIRI小鼠模型视网膜萎缩变薄，减少视网膜细胞凋亡，其作用是通过调控Keap1-Nrf2/HO-1信号通路降低RIRI氧化应激反应来实现。

Abstract:

AIM: To investigate the protective effect and mechanism of Danlou tablet on retinal ischemia-reperfusion injury(RIRI)in mice.

METHODS: A total of 40 ApoE^-/- mice were fed with high fat diet for 6 wk, and the RIRI model was established by anterior chamber infusion of pressurized saline. The mice were divided into control group(normal saline for 8 wk), RIRI model group(normal saline for 8 wk), and low-, medium-, and high-dose Danlou tablets groups \〖1, 2, and 4 g/(kg·d), respectively, for 8 wk\〗. The morphological changes of retina were observed by hematoxylin-eosin(HE)staining, retinal cell apoptosis was detected by terminal-deoxynucleoitidyl transferase mediated Nick-End Labeling(TUNEL)staining. The Western-blot assay was used to detect the expression of retinal tissue sample Kelch-like ech-associated protein 1(Keap1), nuclear factor E2 related factor 2(Nrf2), heme oxygenase 1(HO-1), and superoxide dismutase(Sod2)proteins.

RESULTS: Compared with the control group, the mouse retina was atrophic with thinning thickness and increasing cell apoptosis, down-regulation of Sod2 protein expression, and up-regulation of Keap1 protein expression in the RIRI model group(all P<0.01). Compared with the RIRI model group, the retinal thickness increased in the medium- and high-dose of Danlou tablets groups(all P<0.01), and the cell apoptosis of retina decreased in the low-, medium- and high-dose of Danlou tablets groups(all P<0.05). There were no significant differences in the expression of Keap1 and HO-1 proteins of mouse retina tissue in the low-dose of Danlou tablets group(P>0.05). The expression of Sod2, Nrf2 and HO-1 proteins up regulated, and the expression of Keap1 protein down regulated in the medium- and high-dose of Danlou tablets groups(all P<0.05).

CONCLUSION: Danlou tablet can alleviate RIRI-induced atrophy and thinning of retina and retinal cell apoptosis by regulating Keap1-Nrf2/HO-1 signal pathway and reducing oxidative stress.