目的探讨白藜芦醇对糖尿病大鼠晶状体混浊的防治作用及其生物学机制。
方法选取8周龄SPF级健康雄性SD大鼠50只，采用随机体重排序法将大鼠分为空白对照组、模型组、格列齐特组、低剂量白藜芦醇组和高剂量白藜芦醇组，其中空白对照组10只，其他各组制备糖尿病性白内障模型后各剩余9只。模型组、格列齐特组、低剂量白藜芦醇组、高剂量白藜芦醇组均采用链脲佐菌素腹腔内注射法建立糖尿病模型，建模后第3天格列齐特组以2 mg/(kg·d)格列齐特混悬液灌胃，低、高剂量白藜芦醇组分别以20 mg/(kg·d)和40 mg/(kg·d)白藜芦醇灌胃，空白对照组和模型组大鼠采用等容量生理盐水灌胃，1次/d，连续4周。采用血糖仪检测大鼠空腹血糖浓度；采用酶联免疫吸附法检测大鼠空腹胰岛素水平及过氧化物歧化酶(SOD)1、SOD2、SOD3、谷胱甘肽过氧化物酶(GPX1)含量；采用裂隙灯显微镜检查治疗后晶状体混浊程度；采用苏木精－伊红染色观察晶状体细胞形态学变化；采用TUNEL法检测晶状体上皮细胞(LECs)凋亡情况；采用Western blot法检测晶状体组织中核因子E2相关因子2(Nrf2)、血红素氧合酶1(HO-1)蛋白相对表达量。
结果与空白对照组比较，模型组空腹血糖浓度、空腹胰岛素水平、LECs凋亡率均升高，SOD1、SOD2、SOD3、GPX1水平均降低，差异均有统计学意义(均P<0.05)；与模型组比较，格列齐特组、低剂量白藜芦醇组、高剂量白藜芦醇组大鼠空腹血糖浓度、空腹胰岛素水平、LECs凋亡率均降低，SOD1、SOD2、SOD3、GPX1均升高，差异均有统计学意义(均P<0.05)；与低剂量白藜芦醇组比较，格列齐特组和高剂量白藜芦醇组空腹血糖浓度、空腹胰岛素水平、LECs凋亡率均降低，SOD1、SOD2、SOD3、GPX1水平均升高，差异均有统计学意义(均P<0.05)。空白对照组治疗后晶状体混浊分期均为0期，模型组0期、Ⅰ期、Ⅱ期各占0.00%、66.67%、33.33%，格列齐特组各占77.78%、22.22%、0.00%，低剂量白藜芦醇组各占22.22%、44.44%、33.33%，高剂量白藜芦醇组各占66.67%、33.33%、0.00%，总体比较差异有统计学意义(H＝7.514，P<0.001)，其中模型组晶状体混浊程度较空白对照组严重；格列齐特组、低剂量白藜芦醇组、高剂量白藜芦醇组晶状体混浊程度较模型组减轻，且格列齐特组和高剂量白藜芦醇组晶状体混浊程度均较低剂量白藜芦醇组更轻，差异均有统计学意义(均P<0.05)。与模型组比较，格列齐特组、低剂量白藜芦醇组、高剂量白藜芦醇组晶状体细胞及亚细胞器异常改变减轻，其中以格列齐特组、高剂量白藜芦醇组异常改变更轻微。与空白对照组比较，模型组Nrf2、HO-1蛋白相对表达量降低，差异均有统计学意义(均P<0.05)；格列齐特组、低剂量白藜芦醇组、高剂量白藜芦醇组Nrf2、HO-1蛋白相对表达量较模型组升高，格列齐特组、高剂量白藜芦醇组较低剂量白藜芦醇组升高，差异均有统计学意义(均P<0.05)。
结论白藜芦醇可减轻糖尿病大鼠晶状体混浊程度，其作用机制可能与调节Nrf2/HO-1信号通路，发挥抗氧化应激作用有关。
ObjectiveTo investigate the preventive and therapeutic effects of resveratrol on lens opacification in diabetic rats and its biological mechanism.
MethodsFifty 8-week-old healthy male SPF grade SD rats were selected and randomly divided into blank control group, model group, gliclazide group, low-dose resveratrol group and high-dose resveratrol group according to their body weight, with 10 rats in each group.The diabetes model was established by intraperitoneal injection of streptozotocin in model group, gliclazide group, low-dose resveratrol group and high-dose resveratrol group.On the third day after modeling, rats in gliclazide group was gavaged with 2 mg/(kg·d) gliclazide suspension, and rats in low-dose and high-dose resveratrol groups were gavaged with 20 and 40 mg/(kg·d) resveratrol, respectively, for four weeks.Rats in blank control group and model group were gavaged with the same volume of normal saline once a day, also for four weeks.After the diabetes model was established, there were 10 rats in blank control group and 9 rats in the other four groups.The fasting blood glucose concentration of the rats was measured with a blood glucose meter.The concentrations of fasting insulin, superoxide dismutase (SOD) 1, SOD2, SOD3, and glutathione peroxidase (GPX1) were determined by enzyme-linked immunosorbent assay.Lens opacification after treatment was observed by slit lamp microscopy.Morphologic changes in lens cells were examined by hematoxylin-eosin staining.Apoptosis of lens epithelial cells (LECs) was detected using TUNEL.The relative expressions of nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) proteins in lens tissues were determined by Western blot.The study protocol was approved by the Welfare Ethics Committee of Experimental Animal of Zhengzhou University (No.IACYC2019-02).
ResultsFasting blood glucose concentration, fasting insulin level, and apoptosis rate of LECs were increased and the concentrations of SOD1, SOD2, SOD3, and GPX1 were decreased in model group in comparison with blank control group, and the differences were statistically significant (all atP<0.05). Fasting blood glucose concentration, fasting insulin level, and apoptosis rate of LECs were decreased and the concentrations of SOD1, SOD2, SOD3, and GPX1 were increased in gliclazide group, low-dose resveratrol group, and high-dose resveratrol group compared with model group, and the differences were statistically significant (all atP<0.05). Fasting blood glucose concentration, fasting insulin level, and apoptosis rate of LECs were decreased and the concentrations of SOD1, SOD2, SOD3, and GPX1 were increased in gliclazide group and high-dose resveratrol group compared with low-dose resveratrol group, and the differences were statistically significant (all atP<0.05). The proportions of grade 0, 1 and 2 lens opacities after treatment were 100.00%, 0.00% and 0.00% in blank control group, 0.00%, 66.67% and 33.33% in model group, 77.78%, 22.22% and 0.00% in gliclazide group, 22.22%, 44.44% and 33.33% in low-dose resveratrol group, and 66.67%, 33.33% and 0.00% in high-dose resveratrol group, respectively, with a statistically significant difference (H=7.514, P<0.001). Compared with model group, lens opacification was less severe in blank control group, gliclazide group, low-dose resveratrol group, and high-dose resveratrol group, with statistically significant differences (all atP<0.05). Lens opacification was less severe in gliclazide group and high-dose resveratrol group compared with low-dose resveratrol group, showing statistically significant differences (both atP<0.05). Compared with model group, there were fewer abnormal changes of lens cells and sub-organelles in gliclazide group, low-dose resveratrol group and high-dose resveratrol group, and the abnormalities in gliclazide group and high-dose resveratrol group were slighter.Compared with model group, the relative expression levels of Nrf2 and HO-1 were higher in blank control group, gliclazide group, low-dose resveratrol group, and high-dose resveratrol group, with statistically significant differences (all atP<0.05). The relative expression levels of Nrf2 and HO-1 were higher in gliclazide group and high-dose resveratrol group compared with low-dose resveratrol group, showing statistically significant differences (both atP<0.05).
ConclusionsResveratrol can reduce lens opacification in diabetic rats and its mechanism may be related to the regulation of the Nrf2/HO-1 signaling pathway by exerting antioxidative stress effects.
葛晓芳,朱大强,刘亚东,等. 白藜芦醇对糖尿病大鼠晶状体混浊的防治作用及其机制[J]. 中华实验眼科杂志,2023,41(06)：545-553.
DOI:10.3760/cma.j.cn115989-20200922-00660版权归中华医学会所有。
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