Effects And Mechanisms Of Volatile Oil Of Amomi Fructus On Drug-Induced Intestinal Mucositis

Object:Intestinal mucositis is an major side effect caused by chemotherapy,such as 5-fluorouracil(5-FU)irinotecan,methotrexate and so on.Amomi Fructus,dry and mature fruit of Amomum villosum Lour.,A.villosum Lour.var.xanthioides T.L.Wu et Senjen and A.longiligulare T.L.Wu,is recorded and used in traditional Chinese medicine system as a excellent crude drug for the treatment of digestive system disorders and has more than 1300 years of medicinal and edible history.The objective of the study was to evaluate the effect of volatile oil of A.villosum Lour.(VOAV)and its main component,bornyl acetate(BA),on intestinal mucositis induced by 5-FU.In the meantime,the potential role of VOAV and BA on regulation of intestinal microflora,adjustment of inflammatory process and oxidative stress,maintaining intestinal permeability are investigated in order to provide more theoretical and scientific foundation for its clinic usage.Method:72 SD rats were randomly divided into normal group(CON),5-FU+saline group(MOD),5-FU+live combined Bifidobacterium and Lactobacillus tables(CBL),5-FU+VOAV(VOAV.H,VOAV.M,VOAV.L),5-FU+BA(BA.H,BA.M,BA.L),8 rats in each group.Chemotherapy lasted for 5 days,and drug intervention were continued for 12 days.Diarrhea occurrence,body weight and food intakes were daily recorded.Intestinal histology and inflammatory cytokines such as IL-6,ROS,NF-κB,TNF-α were tested to evaluate the effect of VOAV and BA on inflammation.Western blotting was used to detected the expression of p38 MAPK in intestinal tissue.The intestinal tight junction protein occludin and ZO-1 were measured by flow cytometry to future evaluate the improvement of VOAV and BA on intestinal barrier function.The levels of lipopolysaccharide was also tested.Based on determination of varied region V4 in 16 S rDNA,the regulation role of VOAV and BA in the intestinal microecology system was illustrated.Results:VOAV prevented inflammatory state in intestinal mucositis rats induced by 5-FU.Our study found that treatment with VOAV and its representative constituent,BA,could prevent diarrhea and reverse weight and food intake loss,which are important symptoms of intestinal mucositis induced by 5-FU.5-FU induced a significant decrease in villus height,an increase in the crypt depth,while treatment with VOAV and BA significantly relieved the histopathological changes.VOAV could effectively reduce the damage of small intestine caused by 5-FU and promote the repair of injury,so as to protect the stability of intestinal morphology in a dose dependent mannar.VOAV improved the injure of intestinal mucosal barrier in intestinal mucositis rats.VOAV and BA could significantly decrease the expression of NF-κB which was induced by 5-FU,and significantly reduce the content of inflammatory markers,such as ROS,IL-6 and TNF-α,in blood and small intestinal.The effect was similar with the positive drug and showed a certain dose relationship.VOAV and BA also could significantly inhibit the increase of MPO in rats induced by 5-FU,which representing an improvement of intestinal inflammation state.Both VOAV and BA could effectively reduce the expression of p38 MAPK and caspase-3 showed a certain dose relationship.Meanwhile,our study showed that VOAV and BA could increase the expression of intestinal tight junction protein ZO-1 and occludin,improve the intestinal mucosal barrier,and inhibit the translocation of LPS and protect the integrity of intestinal barrier function.VOA regulated intestinal microbial balance in intestinal mucositis rats induced by 5-FU.The OTUs cluster analysis showed that the group of BA was close the the CON,suggesting that it had a significant regulatory of these OTUs.Pearson correlation analysis showed that most of OTUs,which belong of Bacteroidetes,Frimicutes and Proteobacteri,was positively associated with inflammatory factors,such as ROS,MPO,LPS and Caspase-3.16 S rDNA sequencing results showed that there was no significant difference in the number of OUT among all groups.At the level of phylum,VOVA and BA could significantly reduce the relative abundance of Proteobacteria,which was usually considered as potential pathogen.VOAV and BA could reduce the relative abundance of some genera of Proteobacteria phylum,such as Helicobacter and Escherichia.They also reduced Bactetoides of Bacteroides phylum and Ruminococcus、Parabacteroides、Clostridum of Firmicutes phylum.Meanwhile,VOAV and BA effectively increased the abundance of Lactobacillus.BA could increase the abundance of Bifidobacterium and VOAV could increase the abundance of Roseburia,Coprococcus,which were considered as probiotics to improved intestinal microecological.VOAV and BA reduced the relative abundance of Helicobacter and Escherichia,the abundance of Bactetoides in Bacteroides phylum,and reduce the relative abundance of some genera of Firmicutes phylum,such as Ruminococcus、Parabacteroides、Clostridum,which are considered as potential pathogenic bacteria,to improve the intestinal microflora.At the same time,VOAV and BA both increased the relative abundance of Lactobacillus in Firmicutes phylum.In addition,BA could increase the relative abundance of Bifidobacterium in Actinomycete phylum and VOAV could increase the relative abundance of Roseburia,Coprococcus in Firmicutes phylum,which are considered as probiotics bacteria,to improve the intestinal microflora.Conclusion:VOAV and its main component BA could inhibit the production of ROS which was considered as precursor material of intestinal mucosa,and inhibit the NF-κB pathway to improve the state of inflammation.VOAV and BA could inhibit MAPK and caspase-3 signaling and suppressed the MPO activity,further decreased apoptosis,inhibit the translocation of LPS and protected the integrity of mucosal.At same time,VOAV and BA could regulate the intestinal microbial imbalance induced by 5-FU to stable intestinal mucosal mechanical barrier.