Characterization of biliary microbiota dysbiosis in extrahepatic cholangiocarcinoma

Abstract

Extrahepatic cholangiocarcinoma (CCA) accounts for 3% of digestive cancers. The role of biliary microbiota as an environment-related modulator has been scarcely investigated in CCA, and the putative impact of associated diseases has not been yet assessed. We characterized the biliary microbiota in CCA patients in order to identify a specific CCA-related dysbiosis. The biliary effluents were collected through an endoscopic retrograde pancreatic cholangiography (ERCP) examination involving 28 CCA and 47 patients with gallstones, herein considered as controls. The biliary effluents were submitted to bacterial DNA extraction and 16S rRNA sequencing, using Illumina technology. Overall, 32% of CCA and 22% of controls displayed another associated disease, such as diabetes, pancreatitis, inflammatory bowel disease, or primary sclerosing cholangitis. Such associated diseases were considered in the comparisons that were made. Principal coordinate analysis (PCoA) detected a significant disparity of biliary microbiota composition between CCA patients and controls without an associated disease. Amongst the most abundant phyla, Proteobacteria did not significantly differ between CCA patients and controls, whereas Firmicutes levels were lower and Bacteroidetes higher in CCAs’ biliary microbiota than in the controls’ microbiota. The most abundant genera were Enterococcus, Streptococcus, Bacteroides, Klebsiella, and Pyramidobacter in CCA’s biliary microbiota. Additionally, levels of Bacteroides, Geobacillus, Meiothermus, and Anoxybacillus genera were significantly higher in CCA patients’ biliary microbiota, without an associated disease, in comparison with controls. A specific CCA-related dysbiosis was identified as compared to controls independently from associated diseases. This suggests that a microorganism community may be involved in CCA pathogenesis.

The Role of Microbiota in Primary Sclerosing Cholangitis and Related Biliary Malignancies

“…The role of biliary microbiota has scarcely been investigated in CCA. However, previous studies performing 16S RNA sequencing demonstrated a significant difference of biliary microbiota composition between CCA patients and controls [162]. Initially, some authors suggested that bile fluid dysbiosis could be linked to various diseases, including biliary lithiasis.…”

“…Although the pathogenesis of CCA in PSC is largely unelucidated, inflammation-driven carcinogenesis concomitant with genetic and epigenetic alterations are thought to be the underlying factors [156][157][158]. However, the knowledge of predisposing risk factors is scarce and PSC-CCA development does not seem to be linked to duration and severity of PSC [159][160][161][162]. The leading hypothesis is that PSC causes multifocal strictures of the biliary tree, biliary infections, and accumulation of toxic bile acids, which results in an increased expression of adhesion and antigen-presenting molecules and inflammatory mediators.…”

“…In response, cholangiocytes lining the biliary tree activate a wide range of cellular processes such as hepatocellular proliferation, apoptosis, angiogenesis and fibrosis as well as immunoregulatory functions, such as upregulation of human leukocyte antigen molecules and recruitment of immune cells. However, it remains unclear if activated cholangiocytes directly activate B cell antibody production and T cell expansion [162]. Moreover, the biliary epithelial injury-induced regenerative response by peribiliary glands is also believed to be a key process involved in inflammation-induced CCA carcinogenesis [163,164].…”

“…In case of persistent stimuli, the disease may progress to high-grade dysplasia and ultimately CCA [156,157,[165][166][167]. Emerging evidence already demonstrated that cholangiocytes in explant livers from PSC-CCA patients show metaplasia and dysplasia more often than in patients with PSC without CCA [162,168,169]. Strikingly, the majority of CCA cases develop from a dominant stricture, which is defined as a stricture with a diameter < 1.5 mm in the common bile duct or <1.0 mm in the hepatic duct [155].…”

“…Although bacteria may find a more favorable growth environment in obstructed bile, this likely does not represent the only reason for differential colonization. It has been indicated that altered bile metabolism may provide metabolic advantages to certain bacteria, which might explain tumor growth and outcomes through host immune response to CCA [112,162]. Jia et al were the first to characterize intestinal microbiota, bile acids, and cytokines in patients with CCA by analysis of fecal microbiota in a series of intrahepatic CCA (iCCA).…”