氧化应激-NLRP3-TGF-β_(3)通路参与糖尿病胃轻瘫大鼠胃平滑肌细胞外基质的重构

Oxidative stress-NLRP3-TGF-β_(3) pathway is involved in remodeling of extracellular matrix of gastric smooth muscle in diabetic gastroparesis rats

目的:观察糖尿病胃轻瘫(DGP)大鼠胃平滑肌细胞外基质(ECM)的变化,探讨氧化应激-核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)-转化生长因子β_(3)(TGF-β_(3))通路在该变化中的作用。方法:SPF级4周龄雄性SD大鼠40只。将20只大鼠以链脲佐菌素诱导建立糖尿病模型,并随机分为2组(diabitic 1、2组),每组各10只;另20只正常大鼠随机分为2个正常对照组(NC 1、2组),每组各10只。检测胃内色素残留率与胃肠推进率,确定DGP大鼠模型建立。从diabitic 1、2组成功建立DGP模型的大鼠中选取10只作为DGP组,从NC 1、2组中随机选取10只作为NC组。生化检测胃平滑肌组织总超氧化物歧化酶(T-SOD)和过氧化氢酶(CAT)活性及丙二醛(MDA)含量;Masson染色和免疫组织化学染色观察胃平滑肌ECM及I型胶原蛋白(Col I)和III型胶原蛋白(Col III)表达变化;Western blot检测Col I、Col III、caspase-1(CASP-1)、TGF-β_(3)、NLRP3、基质金属蛋白酶2(MMP-2)和金属蛋白酶组织抑制物1(TIMP-1)的蛋白表达;ELISA检测胃平滑肌白细胞介素1β(IL-1β)含量。结果:DGP组较NC组T-SOD和CAT活性降低,MDA含量增加(P<0.01),ECM含量和Col I表达减少(P<0.01),Col III表达变化不显著(P>0.05),NLRP3、CASP-1、TGF-β_(3)和MMP-2表达均升高(P<0.01或P<0.05),TIMP-1表达下降(P<0.01),IL-1β含量增加(P<0.01)。结论:DGP大鼠胃平滑肌存在ECM重构,且氧化应激-NLRP3-TGF-β_(3)通路变化是该重构发生的原因之一。

AIM:To observe the effects of diabitic gastroparesis(DGP)on the extracellular matrix(ECM)of rat gastric smooth muscle.METHODS:Streptozotocin was applied to construct the diabetic model in SD rats.Twenty model rats were randomly divided into 2 groups(diabitic 1 and 2 groups,n=10),while 20 normal rats were randomly di⁃vided into normal control(NC)groups(NC 1 and 2 groups,n=10).The DGP was confirmed by the residual rate of pig⁃ment in the stomach and the rate of gastrointestinal propulsion.Ten rats were randomly selected from diabitic 1 and 2 groups as DGP group,and 10 rats were randomly selected from NC 1 and 2 groups as NC group.The total superoxide dis⁃mutase(T-SOD)and catalase(CAT)activity and the malondialdehyde(MDA)content in gastric smooth muscle tissue were measured by biochemical detection.The changes of gastric smooth muscle ECM and expression of collagen type I(Col I)and collagen type III(Col III)were observed by Masson staining and immunohistochemical staining.The expres⁃sion levels of Col I,Col III,caspase-1(CASP-1),transforming growth factor-β_(3)(TGF-β_(3)),nucleotide-binding oligomer⁃ization domain-like receptor protein 3(NLRP3),matrix metalloproteinase-2(MMP-2)and tissue inhibitor of metallopro⁃teinase-1(TIMP-1)in gastric smooth muscle tissue were detected by Western blot.The content of interleukin-1β(IL-1β)in gastric smooth muscle was detected by ELISA.RESULTS:Compared with NC group,the activity of T-SOD and CAT decreased,while the content of MDA increased(P<0.01)in DGP group.The ECM amd the expression of Col I decreased(P<0.01),and the expression of Col III did not change(P>0.05).The expression of NLRP3,CASP-1,TGF-β_(3) and MMP-2 all increased(P<0.01 or P<0.05),the expression of TIMP-1 protein decreased(P<0.01),and the content of IL-1βincreased(P<0.01)in DGP group.CONCLUSION:Remodeling of ECM is observed in the gastric smooth muscle of DGP rats,and the change of oxidative stress-NLRP3-TGF-β_(3) pathway is one of the reasons for this remodeling.