原花青素改善尿路感染所致膀胱损伤的机制研究

Mechanism of procyanidins in improving bladder injury caused by urinary tract infection

目的探讨原花青素改善尿路感染所致膀胱损伤的机制。方法通过膀胱注射大肠埃希菌液构建尿路感染模型。实验大鼠分为对照组、模型组(尿路感染模型)和低、中、高剂量实验组(尿路感染模型,分别给予25,50和100 mg·kg^(-1)原花青素治疗)以及高剂量+脂多糖(lipopolysaccharides,LPS)实验组(尿路感染模型,给予100 mg·kg^(-1)原花青素+LPS治疗)。用蛋白质印迹法测定膀胱组织中Toll样受体4(Toll-like receptor-4,TLR4)/核因子-κB(nuclear factor-κB,NF-κB)信号蛋白表达差异,测定膀胱指数和尿白细胞数目,用酶联免疫吸附试验法检测尿液N-乙酰-β-D-氨基葡萄糖苷酶(N-acetal-β-D-glucosaminidase,NAG)和血清中C反应蛋白(C-reactive protein,CRP)、CD4/CD8水平,检测外周血白细胞数目。结果对照组、模型组、高剂量实验组和高剂量+LPS实验组的TLR4蛋白表达水平分别为0.22±0.02,0.52±0.04,0.23±0.02和0.35±0.03;p-p65/p65蛋白表达水平分别为0.27±0.03,0.78±0.07,0.31±0.02和0.47±0.04;膀胱指数分别为(0.04±0.01)%,(0.13±0.01)%,(0.05±0.01)%和(0.10±0.01)%;尿白细胞数目分别为(22.25±1.93)×10^(2),(136.74±10.37)×10^(2),(50.22±4.32)×10^(2)和(115.42±10.66)×10^(2) cell·mL^(-1);NAG含量为(30.28±2.24),(60.47±4.53),(34.62±1.59)和(48.76±2.55)U·L^(-1);外周血白细胞数目分别为(5.78±0.37)×10^(9),(13.18±1.22)×10^(9),(6.56±0.11)×10^(9)和(9.84±0.42)×10^(9) cell·L^(-1);CRP含量分别为(8.59±0.53),(17.20±1.63),(9.54±1.03)和(13.43±1.45)μg·mL^(-1);CD4/CD8分别为(1.12±0.16)%,(9.63±1.09)%,(1.67±0.14)%和(5.91±0.64)%。模型组的上述指标和对照组相比,差异均有统计学意义(均P<0.05);高剂量实验组的上述指标和模型组相比,差异均有统计学意义(均P<0.05);高实验+LPS组的上述指标与高剂量实验组相比,差异均有统计学意义(均P<0.05)。结论原花青素改善尿路感染所致膀胱损伤,抑制炎症,调节免疫,作用机制与降低TLR4/NF-κB信号激活有关。

Objective To investigate the mechanism of procyanidins in improving bladder injury caused by urinary tract infection.Methods Urinary tract infection model was established by bladder injection of Escherichia coli.The experimental rats were divided into control group,model group(urinary tract infection model),experimental-L,-M,-H groups(urinary tract infection model,treated with 25,50,100 mg·kg^(-1) procyanidins,respectively)and experimental-H+lipopolysaccharides(LPS)group(urinary tract infection model,treated with 100 mg·kg^(-1) procyanidins+LPS).Western blot was used to determine the Toll-like receptor-4(TLR4)/nuclear factor-κB(NF-κB)signaling protein expression in bladder tissue;bladder index and urinary leukocyte number were determined;enzyme-linked immunosorbent assay method was used to detect urine N-acetal-β-D-glucosaminidase(NAG)and serum C-reactive protein(CRP),CD4/CD8 levels,and peripheral blood leukocyte counts were detected.Results The expression levels of TLR4 protein in the control group,model group,experimental-H group and experimental-H+LPS group were 0.22±0.02,0.52±0.04,0.23±0.02,0.35±0.03;the expression levels of p-p65/p65 protein were 0.27±0.03,0.78±0.07,0.31±0.02,0.47±0.04;bladder indexes were(0.04±0.01)%,(0.13±0.01)%,(0.05±0.01)%,(0.10±0.01)%;the number of urinary leukocytes was(22.25±1.93)×10^(2),(136.74±10.37)×10^(2),(50.22±4.32)×10^(2),(115.42±10.66)×10^(2) cell·mL^(-1);the NAG content were(30.28±2.24)U·L^(-1),(60.47±4.53)U·L^(-1),(34.62±1.59)U·L^(-1),(48.76±2.55)U·L^(-1);the number of peripheral blood leukocytes was(5.78±0.37)×10^(9),(13.18±1.22)×10^(9),(6.56±0.11)×10^(9),(9.84±0.42)×10^(9) cell·L^(-1);CRP contents were(8.59±0.53),(17.20±1.63),(9.54±1.03),(13.43±1.45)μg·mL^(-1);CD4/CD8 was(1.12±0.16)%,(9.63±1.09)%,(1.67±0.14)%,(5.91±0.64)%.The differences of above indicators between the model group and the control group were statistically significant(all P<0.05);the differences between the experimental-H group and the model group was statistically significant(all P<0.05);the differences between the experimental-H+LPS group and the experimental-H group were statistically significant(all P<0.05).Conclusion Procyanidins can improve bladder injury caused by urinary tract infection,inhibit inflammation,and regulate immunity,the mechanism of action was related to reducing the activation of TLR4/NF-κB signaling.