Alterations Of Intestinal Microbiota In Acute Pancreatitis And Its Association With Inflammation

Background and Aim: Acute pancreatitis(AP) is a common disease of digestive system. Generally, patients with mild acute pancreatitis(MAP) usually has a mild clinical progress and an extremely low mortality and without organ failure or complications. However, patients with severe acute pancreatitis(SAP) is characterized by organ dysfuntion in the early phase of disease, which would increase the mortality up to 36-50%. Once secondary infection occurs in the later stage, mortality will rise. Previous studies demonstrated that the intestinal microbiota has an important impact on the maintenance of the immune system and the prevention of bacterial overgrowth and secondary infectious complications.Objective: Serum and feces of AP patients were collected for detection of related inflammatory cytokines and intestinal mucosal barrier function, and 16 S r DNA sequencing analysis was performed to evaluate alterations of the intestinal bacteria and the role of intestinal microbiota dysbiosis in the progression of AP.Methods: Patients with AP who met our criteria were enrolled in this study during July 2015 to December 2015. Firstly, we collected serum of all the patients after admission and detected the related inflammatory cytokines by ELISA, including TNF-?, IL-1?, IL-18, DAO. Secondly, the V4 region of the 16 S r DNA gene was extracted from all the samples by PCR to assess the alterations of the intestinal microbiota through the Illumina Miseq. Finally, we collected the clinical data and conducted the statistical analysis: all patients were divided into three groups: MAP, MSAP and SAP group, and differences of basic information and related indexes among the three groups were compared. Patients with MSAP and SAP were divided into infection and non-infection group, differences of basic information and related indexes between the two groups were compared. We also conducted the Spearman correlation analysis between the intestinal flora and related indexes in order to understand the relationship of intestinal microbiota and inflammation.Results: 1. A total of 116 patients were enrolled in our study, including 41 patients with MAP and 52 patients with MSAP and 23 patients with SAP, averge ago51.6±14.6 years old, including 58 male and 58 female, all feces and blood samples were collected within the first week of AP onset, the mean number of days of AP onset was 2.8±1.7 days, and the mean number of days of sample collection after AP onset was 4.6±2 days. 2.Comparison of inflammatory cytokines between three groups:CRP level of MSAP and SAP group was higher than that of MAP group(P<0.05); IL-1? and TNF-? level of SAP group was higher than that of MSAP and MAP group(P<0.05), in addition, IL-1? and TNF-? level of MSAP group was higher than that of MAP group(P<0.05). PCT and DAO level of SAP group was higher than that of MAP and MASP group(P<0.05). IL-18 level of SAP group was higher than that of MAP group(P<0.05). 3. Comparison of intestinal flora between three groups: 116 samples generated 847 OUT, species diversity among the three groups had no obvious difference(P>0.05). Phylum: Actinobacteria level of SAP group was lower than that of MAP group(P<0.05). Class:Betaproteobacteria level of MSAP group was higher than that of MAP group(P<0.05). Order:Burkholderiales level of MSAP group was higher than that of MAP group(P<0.05);Bifidobacteriales level of SAP group was lower than that of MAP group(P<0.05). Family:Bifidobacteriaceae and Lactobacillaceae level of SAP group was lower than that of MAP group(P<0.05). Genus :Bifidobacterium,level of SAP group was lower than that of MSAP and MAP group(P<0.05); Lactobacillus level of SAP group was lower than that of MAP group(P<0.05); Faecalibacterium and Butyricicoccus level of MSAP group was higher than that of MAP and SAP group(P<0.05). species : Faecalibacterium prausnitzii and Butyricicoccus pullicaecorum level of MSAP group was higher than that of MAP and SAP group(P<0.05); Lactobacillus reuteri level of SAP and MSAP group was lower than that of MAP group(P<0.05). 4. Comparison between infection group and non-infection group: 24 h APACHE-II score, 48 h APACHE-II score, hospital stay, ICU stay and DAO of patients in the infection group were higher than that of non-infection group(P<0.05). Clostridium butyricum and Butyricicoccus pullicaecorum in the infection group was lower than that of the non-infection group(P<0.001). 5. Correlation between the fecal bacteria and TNF-?, IL-1?, IL-18, DAO,APACHE-II score: potentially pathogenic bacteria Coprococcus(r=0.351) was positively correlated with IL-1?. TNF-? level was positively correlated with Enterococcus(r=0.208), Proteus(r=0.300) and Pseudomonas(r=0.307). IL-18 level was positively correlated with Staphylococcus(r=0.236).Conclusions: 1. At early phase of the disease, the species diversity of intestinal microbiota between three groups has no obvious statistical difference, but when compared with the MAP and MSAP, the beneficial bacteria decreased significantly in SAP(P<0.05), indicated that the dysbiosis of intestinal microbiota in the early stages of disease may plays an important role in the progression of AP. 2. Decreased beneficial bacteria and increased potentially pathogenic bacteria may be involved in the progression of AP. This may promote transformation of MAP and MASP to SAP, eventually leading to bacterial translocation.