川陈皮素基于AMPK/eNOS信号通路通过下调DRD2改善大鼠慢传输型便秘的作用机制研究OA
Research of mechanism of nobiletin improving slow transit constipation in rats by downregulating DRD2 based on the AMPK/eNOS signaling pathway
目的 探讨川陈皮素(NOB)通过多巴胺受体D2(DRD2)对慢传输型便秘(STC)大鼠的作用及其机制.方法 70只SD大鼠随机分为空白组、模型组、低、中、高剂量实验组、AAV9-NC组和AAV9-DRD2组,每组10只.除空白组外,各组均用15 mg·kg-1复方地芬诺酯灌胃建立STC模型.低、中、高剂量实验组分别用 0.1、0.2、0.4 g·kg-1 NOB 灌胃,AAV9-NC 组和 AAV9-DRD2 组在中剂量实验组基础上分别给予尾静脉注射AAV9-NC和AAV9-DRD2100 μL.检测大鼠体质量和粪便指标,用实时定量反转录聚合酶链式反应检测DRD2表达水平,用酶联免疫吸附实验法检测血管活性肠肽(VIP)和一氧化氮(NO)水平,用蛋白质印迹法检测腺苷酸活化蛋白激酶(AMPK)/内皮型一氧化氮合酶(eNOS)信号通路相关蛋白水平.结果 空白组、模型组和低、中、高剂量实验组的粪便含水量分别为(35.24±5.25)％、(24.76±3.47)％、(28.38±2.77)％、(32.76±3.49)％和(33.32±4.34)％;粪便数量分别为(53.20±5.25)、(42.40±4.55)、(47.10±5.36)、(49.50±5.10)和(50.10±5.04)粒;模型组与空白组比较,低、中、高剂量实验组与模型组比较,上述指标在统计学上差异均有统计学意义(P＜0.05,P＜0.01,P＜0.001).DRD2 mRNA 的相对表达水平分别为 1.00±0.12、2.95±0.47、2.73±0.49、2.17±0.35和2.08±0.32,中、高剂量实验组的DRD2 mRNA相对表达水平较模型组均显著降低(均P＜0.001),但2者比较在统计学上差异无统计学意义(P＞0.05),故选用中剂量实验组进行后续实验.空白组、模型组、中剂量实验组、AAV9-NC 组和 AAV9-DRD2 组 VIP 水平分别为(43.15±4.52)、(64.42±7.64)、(51.68±5.37)、(49.25±4.44)和(56.84±6.41)ng·L-1,NO水平分别为(6.84±0.71)、(11.68±1.37)、(9.28±0.93)、(9.35±0.97)和(10.45±1.03)μmol·L-1,AMPK 蛋白相对表达水平分别为 1.00±0.15、2.42±0.34、1.68±0.17、1.55±0.18 和 1.94±0.21,eNOS 蛋白相对表达水平分别为 1.00±0.11、3.18±0.35、2.28±0.23、2.35±0.27和 2.95±0.33,模型组与空白组比较,中剂量实验组与模型组比较,AAV9-DRD2组与AAV9-NC组比较,上述指标在统计学上差异均有统计学意义(均P＜0.05).结论 NOB能够改善大鼠STC,可能是通过下调DRD2表达,调节VIP和NO水平,抑制AMPK/eNOS信号通路实现的.
Objective To investigate the effects and mechanism of nobiletin(NOB)on slow transit constipation(STC)in rats through dopamine receptor D2(DRD2).Methods A total of 70 SD rats were randomly divided into blank group,model group,experimental-L,M,H group,AAV9-NC group and AAV9-DRD2 group,with 10 rats in each group.STC model was established with 15 mg·kg-1 compound diphenoxylate by intragastric administration except in blank group.In experimental-L,M,H group,0.1,0.2,0.4 g·kg-1 NOB was administered intragastricly.On the basis of experimental-M group,AAV9-NC group and AAV9-DRD2 group were given AAV9-NC and AAV9-DRD2100 μL,respectively.Weight and fecal indexes of rats were detected.The expression levels of DRD2 was detected by quantitative real time polymerase chain reaction.Vasoactive intestinal peptide(VIP)and nitric oxide(NO)levels were detected by enzyme-linked immunosorbent assay.Levels of AMP-activated protein kinase(AMPK)/endothelial nitric oxide synthase(eNOS)signaling pathway related proteins were detected by Western blot.Results The fecal water content in blank,model,and experimental-L,M,H groups were(35.24±5.25)％,(24.76±3.47)％,(28.38±2.77)％,(32.76±3.49)％and(33.32±4.34)％,respectively;the fecal quantity were(53.20±5.25),(42.40±4.55),(47.10±5.36),(49.50±5.10)and(50.10±5.04)grains,respectively.Compared model group with blank group,and compared experimental-L,-M,-H groups with model group,the above indicators showed statistically significant differences(P＜0.05,P＜0.01,P＜0.001).The relative expression levels of DRD2 mRNA in blank,model and experimental-L,M,H groups were 1.00±0.12,2.95±0.47,2.73±0.49,2.17±0.35 and 2.08±0.32,respectively.The relative expression levels of DRD2 mRNA in the experimental-M and experimental-H groups were significantly lower than those in the model group(all P＜0.001),with no significant difference between the two groups(P＞0.05);therefore,experimental-M was selected for subsequent experiments.The VIP levels of blank group,model group,experimental-M group,AAV9-NC group and AAV9-DRD2 group were(43.15±4.52),(64.42±7.64),(51.68±5.37),(49.25±4.44)and(56.84±6.41)ng·L-1,respectively;NO levels were(6.84±0.71),(11.68±1.37),(9.28±0.93),(9.35±0.97)and(10.45±1.03)μmol·L-1,respectively;the relative expression levels of AMPK protein were 1.00±0.15,2.42±0.34,1.68±0.17,1.55±0.18 and 1.94±0.21,respectively;the relative expression levels of eNOS protein were 1.00±0.11,3.18±0.35,2.28±0.23,2.35±0.27 and 2.95±0.33,respectively.Compared model group with blank group,compared experimental-M group with model group,and compared AAV9-DRD2 group with AAV9-NC group,the above indicators all showed statistically significant differences(all P＜0.05).Conclusion NOB can improve STC in rats,possibly by down-regulating DRD2 expression,regulating VIP and NO levels,and inhibiting the AMPK/eNOS signaling pathway.
魏晓广;王权;孙文娟;李敏;程玲
南阳市第一人民医院消化内科,河南南阳 473000南阳市第一人民医院消化内科,河南南阳 473000南阳市第一人民医院消化内科,河南南阳 473000南阳市第一人民医院消化内科,河南南阳 473000南阳市第一人民医院消化内科,河南南阳 473000
医药卫生
川陈皮素慢传输型便秘多巴胺受体D2腺苷酸活化蛋白激酶/内皮型一氧化氮合酶血管活性肠肽
nobiletinslow transit constipationdopamine receptor D2AMP-activated protein kinase/endothelial nitric oxide synthasevasoactive intestinal peptide
《中国临床药理学杂志》 2026 (5)
664-670,7
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