辣椒素对内脏高敏感大鼠胃酸影响及机制研究

发布时间：2018-06-11 19:54

本文选题：辣椒素 + 内脏高敏感　；参考：《泸州医学院》2010年硕士论文

【摘要】： 目的：通过观察不同剂量的辣椒素(capsaicin, CAP)对内脏高敏感大鼠胃酸分泌、胃底腺区组织内辣椒素受体(vanilloid receptor subtype 1, VR1)及降钙素基因相关肽(calcitonin gene-related peptide, CGRP)和P物质(substance P, SP)、血浆胃泌素(GAS)和胃粘膜屏障的影响,探讨不同剂量的CAP对内脏高敏感大鼠胃酸的影响及其机制。方法：(1)实验动物分组：60只SD大鼠,雌雄不拘,体重180g～200g之间,随机分为A组(对照组)、B组(模型组)、C1组(CAP1组)、C2组(CAP2组)及C3组(CAP3组),每组大鼠12只。(2)内脏高敏感大鼠模型的建立：实验大鼠适应性饲养2w,自由进食进水。造模方法：A组腹腔注射生理盐水,1ml/鼠；B组、C1组、C2组和C3组大鼠腹腔注射鸡卵清白蛋白(chicken egg albumin), 1ml/鼠,制备内脏高敏感大鼠模型。造模过程中观察大鼠一般情况。2w后取A组全部大鼠12只组成正常对照组,随机从B组、C1组、C2组和C3组中抽取共15只大鼠组成模型对照组采用肛门直肠扩张(colorectal distention, CRD)实验以观察大鼠腹部撤离反射(abdominal withdrawal reflex, AWR),并用AWR评分评估大鼠对直肠扩张刺激的内脏感觉改变。(3)CAP的使用方法及剂量：大鼠造模成功后进行CAP治疗实验,具体方法如下：A组：自由进食进水,生理盐水10mL/kg/d分2次灌胃,连续2w；B组：自由进食进水,生理盐水10mL/kg/d分2次灌胃,连续2w；C1组：自由进食进水,CAP 1mg/kg/d分2次灌胃灌胃,容积为10mL/kg/d,连续2w；C2组：自由进食进水,CAP 5mg/kg/d分2次灌胃灌胃,灌胃容积为10mL/kg/d,连续2w；C3组：自由进食进水,CAP20mg/kg/d分2次灌胃,灌胃容积为10mL/kg/d,连续2w。(4)检测指标及方法：①造模及给药过程中观察大鼠的一般情况。②胃酸的测定：所有大鼠于实验前一夜开始禁食(不禁水),末次灌胃后1h,用质量分数为2%的戊巴比妥钠(40mg/kg)腹腔注射麻醉,打开腹腔,结扎食管胃连接部和幽门部,将整个胃取出,剪开胃并用生理盐水清洗胃内容物,收集清洗液,用酸碱滴定法测定胃酸。③胃底腺区组织内VR1和CGRP、SP的表达情况：取各组大鼠胃底腺区组织(约0.5cm×0.5cm)全层一块,立刻置于4%的多聚甲醛中充分固定2h,石蜡包埋,连续5μm切片,用免疫组化方法分别进行CGRP、SP、VR1测定。④血浆GAS：心脏采血2ml,4℃离心后提取上清液,用放射免疫法检测血浆GAS水平的变化。⑤胃粘膜屏障改变：把清洗过胃内容物之后的胃置于4%的多聚甲醛中充分固定后,肉眼观察胃粘膜形态,采用Guth标准改良评分评定胃粘膜损伤情况,并取胃粘膜组织送病理组织学检查,光镜下对HE染色后的胃粘膜进行观察。结果：(1)动物一般情况：造模过程中,A组大鼠精神状态好,行动灵活,食欲佳,皮毛光泽,大小便正常,体重增加；B组、C1组、C2组、C3组大鼠出现好斗,易激惹,互相撕咬等情况,但无死亡。随着CAP治疗时间延长,C1组、C2组、C3组大鼠好斗、激惹、互相撕咬情况减少,而B组大鼠无明显减少。在灌胃过程中C1组有5只大鼠、C2组有5只大鼠、C3组有6只大鼠曾出现呛咳,但均无窒息和死亡。(2)AWR评分结果：正常对照组注水1ml、3ml和5ml的AWR评分分别为：0.67±0.50分、1.44±0.53分、2.56±0.53分,模型对照组注水1ml、3ml和5ml的AWR评分分别为：0.53±0.52分、2.53±0.52分、3.40±0.63分。随着气囊体积的增加,正常对照组和模型对照组AWR评分值逐渐递增。模型对照组注水3ml和5ml气囊扩张时AWR评分均显著高于正常对照组(P0.05)。(3)胃液总酸度的变化：各组大鼠的胃液总酸度分别为：A组48.17±5.75mmol/L,B组45.67±3.05 mmol/L,C1组45.25±2.93 mmol/L,C2组34.42±2.28 mmol/L,C3组30.50±4.25 mmol/L.其中B组大鼠胃液总酸度低于A组大鼠,但没有统计学意义(P0.05)；C1组大鼠胃液总酸度和B组大鼠胃液总酸度比较没有统计学意义(P0.05)；随着CAP剂量增加。,C2组大鼠胃液总酸度显著低于A组(P0.05),C2组大鼠胃液总酸度显著低于B组(P0.05),C2组大鼠胃液总酸度显著低于C1组(P0.05)；并且随着CAP剂量继续增加,C3组大鼠胃液总酸度显著低于A组(P0.05),C3组大鼠胃液总酸度显著低于B组(P0.05),C3组大鼠胃液总酸度显著低于C1组(P0.05),C3组大鼠胃液总酸度显著低于C2组(P0.05)。(4)胃底腺区组织内VR1的表达情况：大鼠胃底腺区组织内VRl表达水平分别为：A组为0.58±0.51分,B组为1.25±0.62分,C1组为2.00±0.60分,C2组为2.58±0.52分,C3组为3.25±0.75分,其中A组VR1表达水平显著低于B组(P0.05),A组VR1表达水平显著低于C1组(P0.05),A组VR1表达水平显著低于C2组(P0.05),A组VR1表达水平显著低于C3组(P0.05)；B组VR1表达水平也显著低于C1组(P0.05),B组VR1表达水平也显著低于C2组(P0.05),B组VR1表达水平也显著低于C3组(P0.05)；随着CAP剂量的增加,C2组VR1表达水平显著高于C1组(P0.05)；C3组VR1水平也显著高于C2组(P0.05)。(5)胃底腺区组织内CGRP.SP的表达情况：大鼠胃底腺区组织内CGRP表达水平分别为：A组为1.00±0.43分,B组为1.08±0.52分,C1组为1.58±0.52分,C2组为2.17±0.58分,C3组为2.67±0.49分,其中A组CGRP表达水平与B组相比无显著性差异(P0.05),A组CGRP表达水平显著低于C1组(P0.05),A组CGRP表达水平显著低于C2组(P0.05),A组CGRP表达水平显著低于C3组(P0.05)；B组CGRP表达水平也显著低于C1组(P0.05),B组CGRP表达水平也显著低于C2组(P0.05),B组CGRP表达水平也显著低于C3组(P0.05)；随着CAP剂量的增加,C2组CGRP表达水平显著高于C1组(P0.05),C3组CGRP水平也显著高于C2组(P0.05)。大鼠胃底腺区组织内SP表达水平分别为：A组为0.50±0.52分, B组为0.83±0.58分,C1组为0.75±0.45分,C2组为0.75±0.45分,C3组为0.58±0.52分,其中B组SP表达水平较A组略有升高,但没有统计学意义(P0.05)；其余各组间SP表达水平均没有统计学意义。(6)血浆GAS的变化：大鼠血浆GAS水平分别为：A组49.50±7.51 pg/ml,B组63.25±5.99 pg/ml,C1组44.33±2.90 pg/ml,C2组37.42±3.55 pg/ml,C3组33.00±2.37pg/ml。其中A组GAS水平显著低于B组(P0.05),A组GAS高于C1组(P0.05),A组GAS高于C2组(P0.05),A组GAS低于C3组(P0.05)；B组GAS水平也显高于C1组(P0.05),B组GAS水平也显高于C2组(P0.05),B组GAS水平也显高于C3组(P0.05)；C2组GAS水平相比显著低于C1组(P0.05),而C3组的血浆GAS水平显著低于C2组(P0.05)。(7)胃粘膜屏障：A组、B组、C1组、C2组及C3组的胃粘膜损伤评分(Guth标准改良评分)比较各组间无显著性差异(P0.05)；光镜下观察胃粘膜组织病理学改变：A组、B组、C1组、C2组大鼠胃粘膜基本光滑,上皮细胞形态正常,排列整齐,未见粘膜脱落、充血、水肿、炎性细胞浸润及腺体结构紊乱和坏死。C3组大鼠胃粘膜上皮细胞轻度脱落、充血、水肿,未见腺体结构紊乱和坏死。对胃粘膜病理损伤程度进行半定量统计,病理损伤积分A组为0.25±0.45分,B组为0.17±0.39分,C1组为0.32±0.45分,C2组为0.33±0.47分,C3组为0.83±0.72分。其中C3组病理损伤积分显著高于A组(P0.05),C3组病理损伤积分显著高于B组(P0.05),C3组病理损伤积分显著高于C1组(P0.05),C3组病理损伤积分显著高于C2组(P0.05)。结论：1.腹腔注射鸡卵清白蛋白可成功建立大鼠内脏高敏感模型。2.CAP1mg/kgd、5mg /kg/d及20mg/kg/d连续灌胃2w可抑制内脏高敏感大鼠胃酸分泌,并且随着灌胃剂量的增加,抑酸效应增强。3.CAP影响内脏高敏感大鼠的胃酸分泌可能通过激活胃壁内VRl而发挥作用。4.CAP影响内脏高敏感大鼠的胃酸分泌可能与CGRP有关,与SP无明显相关。5.CAP影响内脏高敏感大鼠的胃酸分泌可能与GAS有关6. CAP 1mg/kg/d和5mg/kg/d连续灌胃对正常和内脏高敏感大鼠的胃粘膜屏障无损伤作用,而CAP 20mg/kg/d连续灌胃2w可造成胃粘膜轻微的损伤。
[Abstract]:Objective : To investigate the effects of different doses of capsaicin ( CAP ) on gastric acid secretion , gastrin ( GAS ) and gastric mucosal barrier in rats with visceral hypersensitivity . Methods : ( 1 ) Group B ( model group ) , group C1 ( CAP1 group ) , C2 group ( CAP2 group ) and C3 group ( CAP3 group ) were randomly divided into group A ( control group ) , group B ( model group ) , C1 group ( CAP1 group ) , C2 group ( CAP2 group ) and C3 group ( CAP3 group ) , and the rats were randomly divided into group A ( control group ) , group B ( model group ) , group C1 ( CAP1 group ) , C2 group ( CAP2 group ) and C3 group ( CAP3 group ) .
The rats were randomly divided into group B , C1 , C2 and C3 . The rats were randomly divided into group B , C1 , C2 and C3 . The rats were randomly divided into group B , C1 , C2 and C3 . The rats were randomly divided into group B , C1 , C2 and C3 .
Group B : Free food intake , normal saline 10 mL / kg / day divided into 2 times , continuous 2w ;
C1 group : free food intake , CAP 1mg / kg / day divided into two times , the volume was 10 mL / kg / d , continuous 2w ;
Group 2 : Free food intake , CAP 5mg / kg / day divided into two times , the volume of gastric lavage was 10 mL / kg / d , continuous 2w ;
The gastric mucosal barrier was determined by using the Guth standard modified score to assess the gastric mucosa injury .
The AWR scores of group B , C1 , C2 , C3 were significantly higher than those in normal control group ( P 0.05 ) .
There was no significant difference between the total acidity of gastric juice and the total acidity of gastric juice in group C ( P0.05 ) .
The total acidity of gastric juice in C2 group was significantly lower than that in group A ( P0.05 ) , and the total acidity of gastric juice in C2 group was significantly lower than that in group C ( P0.05 ) .
The total acidity of gastric juice in group C was significantly lower than that in group C ( P0.05 ) . The total acidity of gastric juice in group C was significantly lower than that in group C ( P0.05 ) .
The level of expression in group B was also significantly lower than that in group C ( P0.05 ) , and the level of expression in group B was significantly lower than that in group C ( P0.05 ) , and the level of expression in group B was significantly lower than that in group C ( P0.05 ) .
With the increase of CAP dose , the level of expression level in C2 group was significantly higher than that in C1 group ( P0.05 ) .
The level of CGRP in group C was significantly higher than that in group C ( P0.05 ) . The level of CGRP in group A was 1.00 卤 0.43 , group B was 1.08 卤 0.52 , group C was 1 . 58 卤 0.52 , group C was 2 . 17 卤 0.58 , C3 group was 2.67 卤 0.49 , group A was significantly lower than that in group C ( P0.05 ) , CGRP expression level in group A was significantly lower than that in group C ( P0.05 ) .
The level of CGRP in group B was significantly lower than that in group C ( P0.05 ) , and the level of CGRP in group B was significantly lower than that in group C ( P0.05 ) .
The level of CGRP in group C was significantly higher than that in group C ( P0.05 ) . The levels of CGRP in group C were 0.50 卤 0.52 , 0.83 卤 0.58 in group A , 0.75 卤 0.45 in group C , 0.75 卤 0.45 in group C , 0.58 卤 0.52 in group C , and 0.58 卤 0.52 in group C , but not statistically significant ( P0.05 ) .
The levels of GAS in plasma GAS were 49.50 卤 7.51 pg / ml , 63.25 卤 5.99 pg / ml in group B , 44.33 卤 2.90 pg / ml in group B , 37.42 卤 3.55 pg / ml in group C , 30.00 卤 2.37pg / ml in C3 group .
The GAS levels in group B were also significantly higher than those in group C ( P0.05 ) , and GAS levels in group B were significantly higher than those in group C ( P0.05 ) .
The level of GAS in group C was significantly lower than that in group C ( P0.05 ) , while the level of plasma GAS in C3 group was significantly lower than that in group C ( P0.05 ) . ( 7 ) There was no significant difference between group A , group B , C1 , C2 and C3 group ( P0.05 ) .
The pathological changes of gastric mucosa in C3 group were significantly higher than those in group A ( P0.05 ) . CAP 1mg / kg / day and 5mg / kg / d continuous gastric lavage had no damage to the gastric mucosa barrier of rats with normal and visceral hypersensitivity , while CAP 20mg / kg / day could cause slight injury to gastric mucosa .
【学位授予单位】：泸州医学院
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R363

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