电针对胃癌前病变大鼠坏死性凋亡RIPK1/RIPK3/MLKL信号通路的影响OA

Objective To observe the effect of electroacupuncture(EA)on gastric mucosal injury and necroptosis receptor-interacting protein kinase(RIPK)1/RIPK3/mixed lineage kinase domain-like(MLKL)signaling pathway in rats with precancerous lesions of gastric cancer,so as to explore its possible mechanism underlying improvement of gastric precancerous lesions(GPL).Methods A total of 32 male SD rats were randomly divided into blank control,model,EA,and sham EA groups,with 8 rats in each group.The GPL model was established by free drinking of N-methyl-N'-nitro-N-nitrosoguanidine solution combined with intermittent fasting for 20 weeks.Starting from the 13th week of modeling,EA(2 Hz/15 Hz,0.3-0.5 mA)was applied to bilateral"Zusanli"(ST36)and"Zhongwan"(CV12)for rats of the EA group,and applied to non-acupoints(5 mm away to ST36 and CV12)for rats of the sham EA group.The treatment was conducted for 20 min,once a day,5 times a week for 8 weeks.During the experiment,the general conditions and body weight changes were recorded.Histopathological changes of the gastric tissues were observed by hematoxylin-eosin(H.E.)staining and alcian blue-periodic acid schiff(AB-PAS)staining,separately.The ELISA was used to detect the contents of tumor necrosis factor alpha(TNF-α),interleukin 1 beta(IL-1β)and transforming growth factor beta 1(TGF-β1)in the serum.Immunohistochemistry(IHC)was used to detect the levels of caudal type homeobox transcription factor(CDX)2 in the gastric tissue.The mRNA expression levels of Ki67,alpha-methylacyl-CoA racemase(AMACR),sex determining region Y box protein 2(SOX2),CDX1,RIPK1,RIPK3,MLKL,Caspase-8,Caspase-3 and Caspase-7 in the gastric tissues were detected by using real-time PCR.The protein expression levels of CDX2,AMACR,phospho-RIPK1(p-RIPK1),RIPK1,phospho-RIPK3(p-RIPK3),RIPK3,phospho-MLKL(p-MLKL),MLKL,Caspase-8,and Caspase-3 in the gastric tissues were detected by using Western blot.Results In contrast to the blank control group,the model group had a significant decrease in the body weight gain rate,average daily food intake,expression levels of SOX2 mRNA,and Caspase-3 and Caspase-8 mRNAs and proteins in the gastric tissues(P<0.01,P<0.05),and a considerable increase in the contents of serum TNF-α,IL-1β,TGF-β1,and positive expression of CDX2,expression levels of Ki67,AMACR,CDX1,RIPK1,RIPK3 and MLKL mRNAs,and p-RIPK1/RIPK1,p-RIPK3/RIPK3 and p-MLKL/MLKL ratios in the gastric tissues(P<0.01,P<0.05).The gastric mucosal glands were deeper in the opening,and had an apparent atrophy with intestinal metaplasia of the gastric epithelium.Compared with the model group,the rats in the EA group(not the sham EA group)showed improved mental state,and increased body weight gain rate and average daily food intake(P<0.01,P<0.05).The pathological conditions of gastric mucosal atrophy and intestinal metaplasia were improved.In addition,modeling-induced increases of the contents of serum TNF-α,IL-1β,TGF-β1,and positive expression of CDX2,expression levels of Ki67,AMACR,CDX1,RIPK1,RIPK3 and MLKL mRNAs,and p-RIPK1/RIPK1,p-RIPK3/RIPK3 and p-MLKL/MLKL ratios,and as well as the decreases of the expression levels of SOX2 mRNA,and Caspase-3 and Caspase-8 mRNAs and proteins were all reversed by EA(P<0.05,P<0.01),not by sham EA.H.E.staining showed that the glandular arrangement in the gastric mucosa of the rats in the sham EA group remained disordered,and the pathological histological recovery was poor.Conclusion EA can improve the degree of gastric mucosal lesions in GPL rats,which may be related to its functions in reducing the inflammatory response of the gastric mucosa and inhibiting the necroptosis RIPK1/RIPK3/MLKL signaling pathway of the gastric mucosa.