Exploration Of The Bile Duct Microenvironment In Cholangiolithiasis Patients With Sphincter Of Oddi Laxity:From The Perspective Of The Microbiome And Metabolome

Background:Cholangiolithiasis is a common and severe disease in China. With the features of high residual stone rate, high recurrence rate and high reoperation rate, it has troubled surgeons for decades. Bile duct microenvironment plays key roles in cholangiolithiasis occurrence. Sphincter of Oddi laxity (SOL) is associated with cholangiolithiasis, probably due to enhanced reflux of intestinal contents that changes the microenvironment. However, the microenvironment has not been comprehensively investigated.Methods:Patients with cholangiolithiasis were consecutively recruited from February 2014 to January 2015.The demographic information, medical history, preoperative auxiliary examination and antibiotics application were prospectively collected, and their bile was sampled intraoperatively for high-throughput experiments. Pyrosequencing of 16S ribosomal RNA gene was performed to characterize the microbiota in the bile. A liquid chromatography-mass spectrometry-based method was used to profile bile composition. Clinical manifestation, microbiome, and bile composition were compared between patients with and without SOL.Results:18 patients with SOL and 27 patients without SOL were finally included. Clinically, patients with SOL showed higher white blood cell count (10.6 vs.6.5; Unit, 109/L; P=0.005), proportion of neutrophils (75.6%vs.65.8%; P=0.050) and serum C-reactive protein (91.5vs.32.4; Unit, mg/L; P= 0.022). Bacteria in the bile duct were overwhelmingly aerobes and facultative anaerobes. Proteobacteria (average abundance 80.76%) and Firmicutes (average abundance 10.47%) were the most widespread phylotypes, especially Enterobacteriaceae (average abundance 10.47%). Compared with those without SOL, patients with SOL possessed more varied microbiota. In the SOL group, pathobionts, such as Bilophila (2.09% vs.0.04%; P=0.011) and Shewanella algae (2.77 vs.0.67%; P=0.002) had richer communities while harmless bacteria, such as Bifidobacterium (0.002% vs.0.048%; P=0.034) and Listeriaceae (0.001% vs.0.016%;P=0.025) were poorer. Metabolomics analysis showed the differences in bile composition between groups were mainly distributed in lipids and bile acids. The contents of lysopnosphatidylethanolamines, monounsaturated fatty acids and glycine-conjugated bile acids were higher in the SOL group while the contents of phosphatidic acids and taurine-conjugated bile acids were lower in the SOL group. Particularly, the higher abundance of Bilophila involved in taurine metabolism was associated with lower contents of taurine derivatives in the bile of patients with SOL.Conclusions:A bile duct microenvironment with more severe bacterial infection and stronger lithogenicity was found in patients with SOL. The findings suggest that protection of sphincter of Oddi and prevention of reflux of intestinal contents should be paid enough attention to in the treatment of cholangiolithiasis.