高尿酸血症大鼠的肠道尿酸排泄研究

Intestinal uric acid excretion in rats with hyperuricemia

目的观察果糖诱导的高尿酸血症大鼠肠道尿酸排泄量的改变,从肠道转运蛋白腺苷三磷酸结合转运蛋白(ABCG2)的表达探讨高尿酸血症的病理机制。方法将24只雄性SD大鼠按体质量随机分为正常组与模型组,正常组给予清水,模型组给予10%果糖饮水建立高尿酸血症模型。每10 d动态检测大鼠血清尿酸(SUA)、粪便尿酸(FUA)水平。在体肠灌流结合高效液相检测各组大鼠肠道尿酸排泄量(IUC)的变化。免疫组化染色观察转运体ABCG2在大鼠各肠段的表达位点,并进行蛋白表达半定量分析。蛋白质免疫印迹法(Western blot)进一步检测各组大鼠各肠段转运体ABCG2的蛋白表达。逆转录实时荧光定量PCR(RT-q PCR)检测各组大鼠各肠段转运体ABCG2的mRNA表达水平。结果实验第20~40天,与正常组比较,模型组的SUA水平显著升高(P<0.05)。实验第10~30天,与正常组比较,模型组的FUA水平显著降低(P<0.05或P<0.01)。模型组大鼠肠道IUC显著低于正常组(P<0.05)。免疫组化实验结果显示,ABCG2蛋白主要表达于各肠段的肠绒毛及肠腺中,模型组空肠与回肠的ABCG2蛋白表达显著低于正常组。Western blot蛋白分析结果亦显示模型组空肠和回肠的ABCG2蛋白表达显著低于正常组。RT-q PCR结果显示,与正常组比较,模型组空肠、回肠的ABCG2 mRNA表达显著减少。结论 10%果糖饮水可成功诱导大鼠高尿酸血症模型,该模型存在肠道尿酸排泄障碍,可能与下调空肠和回肠ABCG2 mRNA表达,抑制其蛋白表达有关。

Objective To explore the pathological mechanism of hyperuricemia by observing the alteration of intestinal uric acid excretion and intestinal transporter ABCG2 expression in rats with hyperuricemia induced by fructose. Methods SD male rats（ n = 24） were randomly divided into normal group and model group according to the weight. The rats in the normal group were given water,and the rats in model group were given 10% fructose solution to establish hyperuricemia model. Levels of serum uric acid（ SUA） and fecal uric acid（ FUA） were measured every 10 days. The intestinal urate excretion（ IUE）level was detected by using in situ perfusion method and HPLC. The location and expression of transporter ABCG2 in the intestine was analyzed with immunohistochemical staining and semi-quantitative analysis of protein was conducted. Protein expression of transporter ABCG2 in each part of intestine was further detected by using Western blot. The expression of mRNA of ABCG2 in both groups was determined with RT-q PCR. Results From the 20 thto the 40 thdays,SUA in the model group was significantly higher thanthat in normal group. From the 10 thto the 30 thdays,the FUA in model group was significantly lower than that in normal group. The IUC in model group was significantly lower than that in normal group. Immunohistochemical staining showed that the intestinal transporter ABCG2 was located in the intestinal villus and gland,and the analysis of staining showed that the protein expression of ABCG2 in the jejunum and ileum was significantly lower than those in the normal group. Western blot tests exhibited the same results. RT-q PCR indicated that compared with the normal group,expression of mRNA of ABCG2 in the jejunum and ileum in the model group were markedly lower. Conclusion Fructose could successfully induce hyperuricemia in rats. Hyperuricemia induced by fructose presents dysfunction in the intestinal uric acid excretion,which is probably related to the down-regulation of the ABCG2 mRNA level and inhibition of ABCG2 protein level in the jejunum and ileum.