Study On The Effect And Mechanism Of Tannic Acid On C57BL/6N Mice Infected By Clostridium Difficile

Objective:Clostridioides difficile(C.difficile)is a gram-positive anaerobic bacillus.Clostridioides difficile infection(CDI)is the main cause of antibiotic-associated diarrhea(AAD),and the symptoms of patients are mostly manifested as varying degrees of diarrhea during antibiotic treatment.At present,antibiotics are still the preferred treatment for CDI.We urgently need to find therapeutic drugs that have a mild effect on the intestinal flora.In the early stage of our laboratory,it has been proved that tannic acid,the effective ingredient of Chinese gallnut,can inhibit the growth of C.difficile by in vitro tests.This topic explores the effect of tannin acid on CDI through in vivo tests.The therapeutic effect and mechanism of action provide reference value for the application of tannin acid.Methods:4～5 weeks old C57BL/6N male mice were selected and treated with100mg/kg tannic acid,200mg/kg tannic acid,vancomycin,and the mixed treatment group,namely 200mg/kg tannic acid and vancomycin,simultaneously for 5 days after infection.After intragastric administration of1×10~8 CFU/ml C.difficile bacteria suspension in mice,observation was performed for 30 days.Survival rate,weight change,5-day mouse clinical score,and ELISA were used to detect serum Tcd A,Tcd B and IL-10 content,histopathological analysis of the cecum,comparison of the cecal content coating for the count of C.difficile load and spore formation,to evaluate the effect of tannin acid treatment.Results:1.The survival rate of mice with Clostridioides difficile infection treated with 100 mg/kg tannic acid was(88%),and the survival rate of mice with Clostridioides difficile infection treated with 200 mg/kg tannic acid(89%)was higher than that in the infected group(25%).2.On the 5th day of treatment,the weight of the cecum of vancomycin(1.33±0.35g)was significantly different from that of the normal group(0.35±0.11g)(P<0.01);the weight of the cecum of the mixed treatment group(1.65±0.44 g)Compared with the normal group(0.35±0.11g),it was significantly increased,and the difference was statistically significant(P<0.001);the weight of the cecum after 5 days of 200mg/kg tannin acid treatment was(0.63±0.33g),100mg/kg The weight of the cecum after 5 days of kg tannin acid treatment was(0.58±0.39g)compared with the normal group(0.35±0.11g),the difference was not statistically significant(P>0.05).3.The colony count of Clostridioides difficile infection in the cecal contents after 5 days of 200mg/kg tannic acid treatment was(16±14cfu/mg),and the colony count of C.difficile in the 100mg/kg tannic acid treatment group(11±9cfu/mg)compared to infection Group(98±28cfu/mg),can significantly inhibit the growth of C.difficile,(P<0.001);200mg/kg tannic acid treatment group spore count(12±9cfu/mg),100mg/kg tannic acid treatment group(4±3cfu/mg)compared with the infection group(59±23cfu/mg),significantly inhibited spore formation(P<0.01),(P<0.001).4.The histopathological analysis of the caecum found that tannic acid can improve the intestinal inflammation in mice.After tannic acid treatment,the infiltration of inflammatory cells was reduced and the glands were relatively intact.5.After 5 days of treatment with 200 mg/kg tannic acid,the serum Tcd A content was(24.93±4.12ng/ml),compared with the infection group(32.82±4.13ng/ml),the Tcd A content was significantly reduced after treatment(P<0.001);The Tcd A content of the 100 mg/kg tannic acid treatment group(25.57±3.36 ng/ml)significantly reduced the Tcd A content after treatment(P<0.01).However,the serum Tcd B levels after 5 days of treatment with200mg/kg and 100mg/kg tannin acid were(13.68±1.67ng/ml),and there was no statistics between(12.57±3.55ng/ml)and the infection group(13.38±2ng/ml).Academic difference(P>0.05).6.After 5 days of treatment with 200 mg/kg tannic acid,the serum IL-10content(1862.2±335.53pg/ml)increased the serum IL-10 content compared with the infection group(1410.125±202.06pg/ml),and it was statistically significant(P<0.05);Serum IL-10 content(2083.25±386.98pg/ml)after100mg/kg tannic acid treatment for 5 days increased the serum IL-10 content of the infection group(P<0.001).Conclusions:1.Tannic acid treatment of Clostridium difficile-infected mice improved the survival rate compared with the infected group,and there was no recurrence and death after drug withdrawal.2.This study found that tannic acid can improve the severity of Clostridium difficile infected by inhibiting the growth of C.difficile,spore formation,and reducing the expression of enterotoxin,that is,Tcd A,and increasing the content of serum IL-10.3.Tannic acid treatment can improve intestinal inflammation.