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非甾体抗炎药相关大鼠小肠上皮屏障损伤发生机制及其药物干预效果

Mechanism of nonsteroidal antiinflammatory drugs-induced injury in rats small intestinal epithelial barrier and the effects of drug interfering

摘要:

目的:探讨 NSAID 相关大鼠小肠上皮屏障损伤的评价方法及机制,并研究黏膜保护剂和抑酸剂对上皮屏障的保护作用。方法将96只大鼠分成形态学观察组和机制研究组,每组48只。每组再分成8个亚组,分别为健康对照组、模型组(采用吲哚美辛造模)、替普瑞酮预防组、雷贝拉唑预防组、治疗对照组、替普瑞酮治疗组、雷贝拉唑治疗组,以及替普瑞酮、雷贝拉唑联合治疗组(简称联合治疗组),每个亚组6只。应用共聚焦激光显微内镜在体检测各亚组大鼠小肠脱落细胞间隙密度。采用ELISA 法检测各亚组大鼠血清 TNF-α水平,采用 Western 印迹法检测各亚组空肠组织 caspase-3, NF-κB,闭锁小带蛋白1和闭合蛋白水平。统计学处理采用 LSD-t 检验或 Hamhane′s T2检验。结果替普瑞酮预防组和雷贝拉唑预防组大鼠空肠组织的脱落细胞间隙密度分别为(57.43±24.55)/1000和(59.80±21.14)/1000,均低于模型组的(110.93±50.58)/1000,差异均有统计学意义(t =53.50、54.13,P 均<0.01)。联合治疗组的脱落细胞间隙密度为(40.53±15.39)/1000,低于治疗对照组的(93.80±40.65)/1000,差异有统计学意义(t =44.27,P <0.01)。替普瑞酮预防组和雷贝拉唑预防组的血清 TNF-α水平分别为(25.80±8.97)和(22.74±7.15)ng/L,均低于模型组的(44.48±7.42)ng/L,差异均有统计学意义(t=18.68、21.74,P 均<0.01)。联合治疗组 TNF-α水平为(13.66±4.98)ng/L,低于治疗对照组的(24.67±6.70)ng/L,差异有统计学意义(t=9.02,P <0.01)。替普瑞酮预防组和雷贝拉唑预防组的空肠组织 caspase-3水平分别为1.47±0.35和1.58±0.34,NF-κB 水平分别为1.27±0.14和1.21±0.10,与模型组(2.44±0.45和1.69±0.13)相比,差异均有统计学意义(t =0.97、0.86、0.42、0.48,P 均<0.01)。联合治疗组的 caspase-3和 NF-κB 水平分别为0.66±0.06和0.44±0.21,低于治疗对照组,差异均有统计学意义(t=0.34、0.56,P 均<0.01)。替普瑞酮预防组和雷贝拉唑预防组的闭合蛋白水平分别为0.69±0.16和0.74±0.11,闭锁小带蛋白1水平分别为0.81±0.08和0.84±0.12,与模型组(0.45±0.22和0.64±0.07)相比,差异均有统计学意义(t=0.24、0.29、0.17、0.21,P 均<0.01)。联合治疗组的闭合蛋白和闭锁小带蛋白1水平分别为2.50±0.46和1.76±0.18,高于治疗对照组,差异均有统计学意义(t=1.50、0.76,P 均<0.01)。结论脱落细胞间隙密度或可作为预测炎性反应程度、上皮屏障通透性,以及评价药物疗效的有价值的指标。替普瑞酮和雷贝拉唑对 NSAID 相关大鼠小肠损伤有较好的预防和治疗作用。

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abstracts:

Objective To investigate the assessment methods and mechanisms of nonsteroidal antiinflammatory drugs (NSAID)-induced injury in rat small intestinal epithelial barrier,and to explore the protective effects of mucosal protective agents and antacids on it.Methods A total of 96 rats were evenly divided into the morphologic observation group and the mechanism research group,and 48 in each group.Then each group was evenly divided into eight subgroups:the healthy control group,the model&nbsp;group (model established with indomethacin),the teprenone prevention group,the rabeprazole prevention group,the treatment control group,the teprenone treatment group,the rabeprazole treatment group and the teprenone and rabeprazole combined group (combined group),six in each group.Exfoliated cells gap density of small intestine of each subgroup was determined by confocal laser endomicroscopy.Serum level of tumor necrosis factor-α (TNF-α)was measured with enzyme-linked immunosorbent assay (ELISA). The expression of nuclear factor-κB (NF-κB),caspase-3,zonula occludens-1 (ZO-1 )and occludin at protein level was detected by Western blotting.The LSD-t test or Hamhane′s T2 test was performed for statistical analysis.Results The exfoliated cells gap densities of the teprenone prevention group and the rabeprazole prevention group were (57.43 ± 24.55 )/1 000 and (59.80 ± 21 .14 )/1 000,respectively, which were both lower than that of the model group ((110.93±50.58)/1 000),and the differences were statistically significant (t= 53.50 and 54.13,both P < 0.01 ).The exfoliated cells gap density of the combined group was (40.53 ±15 .39)/1 000,which was lower than that of the treatment control group ((93.80±40.65 )/1 000 ),and the difference was statistically significant (t =44.27,P <0.01 ).The serum levels of TNF-α of the teprenone prevention group and the rabeprazole prevention group were (25 .80±8.97)ng/L and (22.74 ±7.15 )ng/L,repsectively,which were both lower than that of the model group ((44.48 ± 7.42 )ng/L),and the differences were statistically significant (t = 18.68 and 21 .74,both P <0.01 ).The serum level of TNF-αof the combined group ((13.66 ±4.98)ng/L)was lower than that of the treatment control group ((24.67±6.70)ng/L),and the difference was statistically significant (t = 9.02,P < 0.01 ).The caspase-3 levels of teprenone prevention group and rabeprazole prevention group were 1 .47 ±0.35 and 1 .58 ±0.34,and the NF-κB levels of these two groups were 1 .27±0.14 and 1 .21 ± 0.10,respectively.Compared with those of model group (2.44 ± 0.45 and 1 .69±0.13),the differences were statistically significant (t =0.97,0.86,0.42 and 0.48,respectively, all P <0.01 ).The levels of caspase-3 and NF-κB of the combined group were 0.66±0.06 and 0.44 ± 0.21 ,respectively,which were lower than those of the treatment control group,and the differences were statistically significant (t=0.34 and 0.56,both P <0.01).The expressions of occludin at protein level of the teprenone prevention group and the rabeprazole prevention group were 0.69 ±0.16 and 0.74 ±0.11 , and the levels of ZO-1 were 0.81 ± 0.08 and 0.84 ± 0.12.Compared with those of the model group (0.45 ±0.22 and 0.64±0.07 ),the differences were statistically significant (t =0.24,0.29,0.17 and 0.21 ,respectively,all P <0.01 ).The levels of occludin and ZO-1 of the combined group were 2.50 ± 0.46 and 1 .76±0.18,which were higher than those of the treatment control group,and the differences were statistically significant (t =1 .50 and 0.76,both P <0.01 ).Conclusions The exfoliated cells gap density may be a valuable indicator to predict the degree of inflammation response and permeability of epithelial barrier as well as to evaluate efficacy of medication.Teprenone and rabeprazole have prevention and treatment effects on NSAID-induced injury in rat small intestine.

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