Observation On The Efficacy Of Wujiajian Zhengqi Powder In The Treatment Of Rotavirus Enteritis With Cold Dampness And The Mechanism Of Regulating PI3K/AKT/mTOR Autophagy Pathway

Objective:1.To observe the clinical curative effect of Wujiajian Zhengqi Powder in the treatment of cold-damp rotavirus enteritis through a randomized controlled trial.2.Based on the PI3K/AKT/mTOR autophagy pathway this study studied the mechanism of action of Wujiajian Zhengqi Powder in the treatment of rotavirus enteritis.Methods:1.116 cases of children with rotavirus enteritis(RVE)were taken as the research objects,59 cases in the experimental group and 57 cases in the control group.Both groups were given oral rehydration salts(ORS)for basic treatment.The control group was given Montmorillonite powder orally,and the experimental group was given Wujiajian Zhengqi Powder orally.Objectively evaluate the treatment of rotavirus by Wujiajian Zhengqi Powder by observing the scores of main and secondary symptoms and total score,symptom efficacy,time to return to normal stool characteristics,stool RV negative rate,serotype indicators,and comprehensive efficacy of the two groups of patients.2.Taking human cloned colon adenocarcinoma cells(Caco-2)as the vector and rotavirus P strain(G3P8)as the research object,firstly,we studied whether rotavirus infection can cause autophagy in host cells,and verified by using Autophagy flow kit and transmission electron microscope were used to observe the effect of virus infection on autophagy,and q RT-PCR was used to detect the expression of VP7 and NSP4 in rotavirus genome to study the relationship between autophagy and rotavirus replication.Then 404-week-old male Wistar rats were randomly divided into a high-dose group of Wujiajian Zhengqi Powder,a middle-dose group of Wujiajian Zhengqi Powder,a low-dose group of Wujiajian Zhengqi Powder,a blank control group,10 animals in each group.After 3 days of administration,the drug-containing serum of the rats was taken.Autophagy flow kit and transmission electron microscope were used to observe the effect of drug-containing serum on autophagy of infected cells.The expressions of autophagy-related proteins Beclin 1,P62,and PI3K/AKT/mTOR signaling pathway-related proteins PI3 K,p-AKT,and p-mTOR were detected by Western Blot.The expression of ATG5 and ATG12 m RNA was detected by q RT-PCR.Result:1.Wujiajian Zhengqi Powder has good curative effect in the treatment of rotavirus enteritis.Before treatment,there was no significant difference in symptom scores and physicochemical indexes between the experimental group and the control group(P>0.05).After treatment,both the experimental group and the control group effectively relieved the main and secondary symptoms such as diarrhea,abdominal pain,bowel sounds,nausea and vomiting.However,the remission effect of Wujiajian Zhengqi Powder is significantly better than control group(P<0.01),and the total score of TCM symptoms was significantly lower than that of the control group(P<0.01).In terms of antidiarrheal time,rotavirus negative rate,and inflammatory indexes,Wujiajian Zhengqi Powder was more effective than montmorillonite(P < 0.01).Comparing the comprehensive efficacy between the two groups,the experimental group of Wujiajian Zhengqi Powder was significantly better than the control group(P<0.01).2.At the early stage of rotavirus infection,the autophagy level of infected cells increased continuously.At 4h,the autophagy level was the highest.However,the autophagy level of infected cells with the addition of autophagy inhibitor 3-MA was significantly inhibited.The expression levels of NSP3 and VP7 in the rotavirus genome were detected by q RT-PCR,and it was found that the levels of viral NSP3 and VP7 were significantly decreased after the introduction of autophagy inhibitors to inhibit autophagy(P<0.01).After the addition of drug-containing serum,the results showed that the number of autophagic vesicles in the high,medium and low dose groups was significantly less than that in the control group,and the number of autophagic vesicles in the high dose group was the least.In addition,the levels of autophagy-related proteins Beclin 1 were significantly decreased,the m RNA expressions of ATG5 and ATG12 were also significantly decreased,and the level of P62 protein was significantly increased.The levels of PI3K/AKT/mTOR signaling pathway related proteins PI3 K,p-AKT,p-mTOR and PI3 K,AKT,mTOR,m RNA were significantly decreased(P<0.01).Conclusion:1.Wujiajian Zhengqi Powder significantly alleviated the clinical symptoms of diarrhea,abdominal pain,fever,nausea and vomiting in children with rotavirus enteritis,shortened the recovery time of stool traits,increased the negative rate of virus,reduced inflammatory reaction,and there were no adverse reactions.2.The mechanism of Wujiajian Zhengqi Powder on rotavirus infection may be related to its ability to regulate PI3K/Akt/mTOR signaling pathway and autophagy.