白藜芦醇对内毒素诱导的大鼠葡萄膜炎的治疗作用

Experimental Study of the Therapeutic Effect of Resveratrol on Endotoxin Induced Uveitis in Rats

目的：探讨眼局部使用白藜芦醇（Res）对内毒素诱导的大鼠葡萄膜炎的治疗效果及相关机制。方法：实验研究。将36只雄性Wistar大鼠按随机数字表法分为空白组、模型组、0．25％Res治疗组、0．5％Res治疗组、1％Res治疗组和0．1％地塞米松（Dex）治疗组，每组6只。将大肠杆菌细胞壁脂多糖（LPS）溶于无菌的0．9％氯化钠溶液配成1mg／ml的LPS溶液，并注射入模型组和治疗组大鼠双后足垫（每只注射200μg），空白组注射等量0．9％氯化钠溶液。各治疗组于LPS诱导前后分别给予相应浓度的Res和Dex10μ1点双眼，每2小时1次，注射LPS前后各6次。空白组和模型组给予等量的0．9％氯化钠溶液点眼。观察大鼠眼部炎症反应、临床表现评分，注射LPS24h后测房水肿瘤坏死因子-α（TNF-α）和白细胞介素-6（IL-6）浓度、眼球病理切片HE染色以及免疫组织化学检测虹膜睫状体p38丝裂原活化蛋白激酶（MAPK）和核转录因子-κB（NF-κB）p65等，探讨与分析Res的治疗效果。采用单因素方差分析、Mann．Whitneyu检验进行数据处理。结果：模型组于LPS注射后4h可见虹膜血管扩张充血，之后炎症反应逐渐加重，24h时可见前房大量点状渗出、瞳孔区纤维渗出及晶状体前囊膜渗出物沉着，模型组24h临床评分为4.3±1．2，1％Res组为2．0±0．6，1％Res组葡萄膜炎反应明显轻于模型组（P〈0．05）；1％Res组房水中TNF-α和IL-6的浓度与模型组相比均明显降低（P〈0．05）；1％Res组眼部组织病理学改变较模型组轻（P〈0．05）；1％Res组虹膜睫状体p38MAPK和NF-κB p65的核转位阳性细胞率较模型组均明显降低（P〈0．05）。结论：1％Res局部点眼能有效减轻内毒素诱导的大鼠自身免疫性葡萄膜炎的临床症状，其机制可能与抑制MAPK和NF-κB信号通路有关。

Objective： The study aims to investigate and discuss the therapeutic effect and the mechanism of topical resveratrol （Res） on rats with uveitis. Methods： In this experimental study, 36 Wistar male rats were randomly divided into six equal groups as follow： control group, model group, 0.25% Res group, 0.5% Res group, 1% Res group, and 0.1% dexamethasone （Dex） group. The endotoxin lipopolysaccharide （LPS） was dissolved in normal sterile saline （1 mg/ml） and subcutaneously injected （200μg LPS） into the hind feet of rats to induce uveitis in the model group and in the four treatment groups. Rats from the control group were injected with the equal volume of the normal sterile saline solution. For the 4 treatment groups, 10 Ixl of Res （0.25, 0.5, or 1%） or Dex （0.1%） solutions were applied topically as drops to both eyes, every 2 hours, 6 times before LPS injection and continued every 2 hours, 6 times after injection. Rats of the control and model groups received the same amount of saline. During the observation phase, the inflammation of the anterior segment was observed, and the clinical scores were evaluated 24 h after LPS injection. Anterior segment photography was performed 24 h after LPS injection. The levels of TNF-α and IL-6 in the aqueous humor were determined 24 h after LPS injection. At the end of experiment, the rats were sacrificed and the eyeballs were fixed for histopathological examination. The nuclear translocation of p38 mitogen-activated protein kinase （MAPK） and nuclear factor-kappaB （NF-kB） p65 in the iris ciliary body （ICB） were stained by immunohistochemistry. Data analysis was performed using one-way ANOVA and the Mann- Whitney Utest. Results： In the model group, iris vasodilatation was observed 4 h after LPS injection, and the inflammatory reaction gradually increased. After 24 h, massive exudation was present in the anterior chamber, and fibrous exudation was present in the pupil area and anterior capsule of the lens. 1% Res group reacted much smaller than the model group. The composite clinical score of the model group was 4.3±1.2, which was significantly greater than for the 1% Res group, 2.0±0.6 （P〈0.05）. The TNF-α and IL-6 levels in the aqueous humor of the 1% Res group were significantly lower than for the model group （P〈0.05）. Consistent with these data, the histopathological damage of the ocular tissue treated with 1% Res was less severe than that of model group （P〈0.05）. Lastly, the expression of p38 MAPK and NF-κB p65 levels in the 1% Res group were lower than in the model group （P〈0.05）. Conclusions： The topical administration of 1% Res suppressed ocular inflammation due to LPS injection in endotoxin induced uveitis （EIU） rats and decreased the levels of inflammatory cytokines associated with the inhibition of MAPK and NF-κB signaling pathway. Key words： uveitis; lipopolysaccharide; Resveratrol; p38 mitogen-activated protein kinase; nuclear transcription factor-κB p65; rats