Introduction

Methods to clear CBDS carry varying risk of adverse effects while leaving them in situ can lead to complications secondary to retained stones.^1–3 Thus, the modality of choice reasonably affects the assessment of whether an attempted removal of asymptomatic CBDS is feasible and adequate. In Sweden, available methods to clear common bile duct stones identified on intraoperative cholangiography during cholecystectomy are flushing/manipulating with cholangio catheter, trans cystic stone extraction, intra-operative endoscopic retrograde cholangiopancreatography (IOERCP), postoperative ERCP, laparoscopic choledochotomy, open choledochotomy and leaving the stone in situ.

Aims & Methods

This observational cohort study was based on the national Swedish Register for Gallstone Surgery and ERCP (GallRiks) with validated prospectively assembled data for 90% of all cholecystectomies and ERCP procedures in Sweden from 2009 to 2023.⁴ The aim was to assess efficacy and safety of the respective methods of clearing CBDS by estimating their odds ratio (OR) of failing to reach textbook outcome. Leaving CBDS in situ was considered reference method.
Textbook outcome was defined as cholecystectomy and CBDS clearance laparoscopically completed in one session without immediate or 30-day surgical complications (bleeding from wound or GI tract, wound dehiscence, wound infection, sepsis, bile duct injury with or without leakage or bleeding), readmissions, reinterventions, stent placement or prolonged postoperative hospitalization beyond three days.
The size of CBDS was categorized as small (<4mm) intermediate (4-8 mm) and large (>8mm). Patients were considered asymptomatic if no previous or current sign of bilirubinemia, cholangitis or pancreatits was recorded.

Results

Among the 192 203 patients in the register between 2009 and 2023, 25 295 patients presented with a CBDS during cholecystectomy. Failure to achieve textbook outcome was seen for 12 147 (48%) across all stone sizes and management options. Odds ratio of failing textbook outcome when adjusting only for CBDS size was 0.7 (95%CI 0.63-0.77) for intraoperative ERCP and 0.63 (95% CI 0.55-0.71) for transcystic extraction while flushing and manipulating the stone had OR 0.5 (95% CI 0.44-0.57). Laparoscopic choledochotomy had OR 14.36 (95% CI 8.83- 25.09) to fail textbook outcome. All compared to leaving the stone in situ.
For a subgroup of asymptomatic stones smaller than 4mm the flushing manipulating option was the only one beneficial compared to leaving them in situ with unadjusted OR 0.73(95% CI 0.59-0.90) of failing textbook outcome.

Conclusion

This study introduces a novel composite outcome measure—textbook outcome—to define successful, complication-free clearance of common bile duct stones (CBDS) encountered during cholecystectomy. Using this measure, the greatest benefit was seen in the flush/manipulate group, which was also the only approach that provided a clear advantage for small asymptomatic stones compared to leaving them in situ.
Balancing safety and efficacy of CBDS clearance methods remains essential, given the potential for complications associated with interventions in the common bile duct.
Importantly, small asymptomatic stones did not seem to benefit from more invasive removal techniques beyond flushing and manipulation with the cholangiocatheter,
when evaluated using this composite outcome measure. However, these results need to be interpreted with caution due to the risk of selection bias.

References

1 Lammert F, Gurusamy K, Ko CW, Miquel J-F, Méndez-Sánchez N, Portincasa P, et al. Gallstones. Nat Rev Dis Primers [Internet]. Nature Publishing Group; 2016 Apr 28 [cited 2023 Nov 6]; 2: 1–17. Available from: https://www.nature.com/articles/nrdp201624
2 Saito H, Kadono Y, Shono T, Kamikawa K, Urata A, Nasu J, et al. Remaining issues of recommended management in current guidelines for asymptomatic common bile duct stones. World J Gastroenterol [Internet]. 2021 May 14 [cited 2023 Oct 24]; 27: 2131–2140. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8117734/
3 Berg LS, Friis-Andersen H, Zinther NB, Öztoprak M, Gotschalck KA. Feasibility and outcome of transcystic laparoscopic common bile duct exploration as first-line treatment for common bile duct stones: a retrospective cross-sectional study. Surg Endosc [Internet]. 2025 Feb 11 [cited 2025 Feb 17]; 1–11. Available from: https://link.springer.com/article/10.1007/s00464-025-11587-7
4 Enochsson, L. S G. GallRiks Annual Reports 2018 [Internet]. 2019 [cited 2019 Oct 3]. Available from: https://www.ucr.uu.se/gallriks/fou/arsrapporter/arsrapport-2018-2/viewdocument/837

Introduction

Crohn’s disease (CD) is associated both (i) with polymorphisms in the card15 gene, coding for the cytosolic NOD2 innate immunity receptor, recognizing bacterial peptidoglycan (PG), and (ii) with a decrease in Faecalibacterium, an anti-inflammatory intestinal bacterium¹. Oral administration of Faecalibacterium could represent a novel therapeutic strategy for CD patients, which is currently under investigation with F. prausnitzii EXL01 strain (human clinical trial #NCT05542355). Indeed, Faecalibacterium exhibits anti-inflammatory effects both in vitro and in vivo in colitis models^1-6 and we recently discovered that it directly induced the production of Interleukin-10 (IL-10) in CD14+ human monocytes⁷. However, the bacterial molecules, the host receptors and signaling pathways involved are still poorly understood. Therefore, the aim of the current study was to decipher the role of the interaction between Faecalibacterium, its PG and NOD2 in health and CD.

Aims & Methods

The PG structure of F. prausnitzii EXL01 was determined by ultra high-performance liquid chromatography and mass spectrometry. NOD2 activation was evaluated using HEK-human NOD2 reporter cell line. Following stimulation with either bacteria or PG, secretion of IL-10 and TNF-a by i) WT and NOD2 KO THP1 human monocytic cell line and ii) by dendritic cells (DC) from WT and NOD2^-/- mice and from human patients with CD harboring 0 or 2 NOD2 loss-of-function frameshift mutations were evaluated by ELISA.

Results

Faecalibacterium PG presents an amino acid substitution in the minimal motif recognized by NOD2 (data patented). PG of F. prausnitzii activates NOD2, but not NOD1, in a dose-dependent manner and has a higher ability to activate NOD2 than the one from other Gram positive bacteria. We showed that the production of IL-10 by THP1 monocytes and murine DC upon stimulation either with F. prausnitzii or its PG was decreased in absence of NOD2, indicating that secretion of IL-10 by human and murine immune cells upon Faecalibacterium stimulation is dependent on NOD2. Interestingly, no significant effect was observed for TNF-a secretion, leading to a significant decrease of the IL-10/TNF- a ratio in NOD2 KO cells. In addition, secretion of IL-10 upon stimulation with F. prausnitzii or its PG was lower in DC from CD patients with 2 NOD2 loss-of-function frameshift mutations compared to DC from CD patients without NOD2 mutation. Levels of secreted TNF-a were higher in DC from CD patients with 2 NOD2 loss-of-function frameshift mutations compared to DC from CD patients without NOD2 mutation, leading to decreased IL-10/TNF- a ratio in absence of functional NOD2. The pathways downstream of NOD2 as well as the effects of Faecalibacterium and its PG on NOD2 in vivo are currently under investigation.

Conclusion

Our study showed that Faecalibacterium and its PG activate NOD2, and therefore the production of IL-10, in a different way than other Gram-positive bacteria. These results provide new insight into the molecular pathways underlying the immunomodulatory effects of F. prausnitzii, and highlight NOD engagement as a key immunological feature. Identification of PG as a molecule responsible for the therapeutic effects paves the way for the next generation of Faecalibacterium-based drugs, whether whole-cell or component-driven, in inflammatory diseases such as CD.

References

1. Sokol H, Pigneur B, Watterlot L, et al. Faecalibacterium prausnitzii is an anti-inflammatory commensal bacterium identified by gut microbiota analysis of Crohn disease patients. Proc Natl Acad Sci U S A. 2008; 105: 16731-6.
2. Lenoir M, Martin R, Torres-Maravilla E, et al. Butyrate mediates anti-inflammatory effects of Faecalibacterium prausnitzii in intestinal epithelial cells through Dact3. Gut Microbes. 2020; 12: 1-16.
3. Quevrain E, Maubert MA, Michon C, et al. Identification of an anti-inflammatory protein from Faecalibacterium prausnitzii, a commensal bacterium deficient in Crohn's disease. Gut. 2016; 65: 415-25.
4. Rossi O, van Berkel LA, Chain F, et al. Faecalibacterium prausnitzii A2-165 has a high capacity to induce IL-10 in human and murine dendritic cells and modulates T cell responses. Sci Rep. 2016; 6: 18507.
5. Sarrabayrouse G, Bossard C, Chauvin JM, et al. CD4CD8 lymphocytes, a novel human regulatory T cell subset induced by colonic bacteria and deficient in patients with inflammatory bowel disease. PLoS Biol. 2014; 12: e1001833.
6. Touch S, Godefroy E, Rolhion N, et al. Human CD4+CD8alpha+ Tregs induced by Faecalibacterium prausnitzii protect against intestinal inflammation. JCI Insight. 2022; 7.
7. https://www.biorxiv.org/content/10.1101/2024.10.06.616495v1

Disclosure

HS reports lecture fees, board membership, or consultancy from Carenity, AbbVie, Astellas, Danone, Ferring, Mayoly Spindler, MSD, Novartis, Roche, Tillots, Enterome, BiomX, Takeda, Biocodex, has stocks from Enterome and is co-founder of Exeliom Biosciences. PL report lecture fee, board membership, or consultancy from Biose, Biostime, Boiron, Bonduelle, BMS, Bromatech, IPSEN, iTaK, Lallemand, Lesaffre, L’Oréal, Mayoli, Merck, Procter and Gamble, Second Genome, Therascience and URGO and is co-founder of Exeliom Biosciences. Other authors declare no competing interests. DS, LH, PR and BH are employees of Exeliom Biosciences.