Review of the mechanisms of the biliary-enteric axis in the development of cholangiocarcinoma

Abstract

Cholangiocarcinoma (CCA) is a particularly aggressive and challenging type of cancer, known for its poor prognosis, which is worsened by the complex interplay of various biological and environmental factors that contribute to its development. Recently, researchers have increasingly focused on the significant role of the biliary-enteric communication of liver-gut axis in the pathogenesis of CCA, highlighting a complex relationship that has not been thoroughly explored before. This review aims to summarize the key concepts related to the biliary-enteric communication of liver-gut axis and investigate its potential mechanisms that may lead to the onset and progression of CCA, a disease that presents substantial treatment challenges. Important areas of focus will include the microbiome's profound influence, which interacts with host physiology in ways that may worsen cancer development; changes in bile acid metabolism that can create toxic environments favorable for tumor growth; the regulation of inflammatory processes that may either promote or inhibit tumor progression; the immune system's involvement, which is crucial in the body's response to cancer; and the complex interactions within metabolic pathways that can affect cellular behavior and tumor dynamics. By integrating recent research findings from various studies, we aim to explore the multifaceted roles of the biliary-enteric communication of liver-gut axis in CCA, providing new insights and perspectives for future research while identifying promising therapeutic targets that could lead to innovative treatment strategies aimed at improving patient outcomes in this challenging disease.

Key Words: Cholangiocarcinoma; Biliary-enteric communication of liver-gut axis; Microbiome; Bile acid metabolism; Inflammatory response

Core Tip: Cholangiocarcinoma (CCA) is a type of cancer known for its poor prognosis and complex causes. Recently, researchers have focused more on the biliary-enteric axis's role in developing CCA. This review summarizes the basic concepts of the biliary-enteric axis and its potential mechanisms in CCA, including the microbiome's influence, changes in bile acid metabolism, regulation of inflammation, immune system involvement, and interactions in metabolic pathways. We integrate current research findings to explore the multidimensional roles of the biliary-enteric axis in CCA. This approach offers new perspectives for future research and identifies potential therapeutic targets.

INTRODUCTION

Cholangiocarcinoma (CCA) is a cancer that develops from the bile duct's epithelial cells, and it poses significant clinical challenges because of its poor prognosis and complicated causes. The development and progression of this aggressive cancer are influenced by the complex interactions among genetic, epigenetic, and environmental factors[1]. Recent research has increasingly focused on the role of the biliary-enteric communication of liver-gut axis, also known as the gut-liver axis, in the pathogenesis of CCA. The biliary-enteric communication of liver-gut axis is a bidirectional communication network between the gut microbiota and the liver, involving metabolic, immunological, and inflammatory pathways[2]. The concept of the gut-liver axis encompasses the complex interactions between the gut microbiota, bile acids, and the host's immune and metabolic systems. The gut microbiota, a diverse community of microorganisms residing in the gastrointestinal tract, plays a crucial role in maintaining homeostasis and modulating host physiology[3]. Dysbiosis, which refers to an imbalance in gut microbiota, has been linked to several liver diseases, including CCA. Alterations in the composition and function of the gut microbiota can influence bile acid metabolism, inflammatory responses, and immune regulation, thereby contributing to carcinogenesis[4]. Bile acids, synthesized in the liver from cholesterol and modified by gut microbiota, are essential for lipid digestion and absorption. They also act as signaling molecules, regulating metabolic and immune functions through receptors such as the farnesoid X receptor (FXR)[5]. Dysregulated bile acid metabolism can lead to an altered bile acid pool, which has been associated with liver inflammation, fibrosis, and cancer[6]. The gut-liver axis thus represents a critical interface where microbial metabolites, including bile acids, influence liver health and disease[7].

Inflammation is a key driver of CCA, with chronic inflammatory conditions such as primary sclerosing cholangitis (PSC) being significant risk factors[8]. The bbiliary-enteric communication of liver-gut axis modulates inflammatory responses through microbial products like lipopolysaccharides (LPS) and short-chain fatty acids (SCFAs), which can trigger immune responses and promote a pro-inflammatory environment conducive to tumorigenesis[9]. Understanding the inflammatory pathways and their regulation by the gut microbiota is crucial for elucidating the mechanisms underlying CCA development[10]. The immune system plays a pivotal role in the surveillance and elimination of cancer cells. However, in the context of CCA, immune evasion mechanisms are often activated, allowing tumor progression[11]. The biliary-enteric communication of liver-gut axis influences immune responses through the modulation of immune cell populations and the production of immunomodulatory metabolites[12]. Exploring the interactions between the gut microbiota, bile acids, and the immune system can provide insights into potential therapeutic strategies for CCA[13].

In summary, the gut-liver axis plays a multifaceted role in the pathogenesis of CCA, involving the gut microbiota, bile acid metabolism, inflammatory responses, and immune regulation. This review aims to provide a comprehensive overview of the current understanding of the gut-liver axis in CCA, highlighting the potential mechanisms and therapeutic targets. By integrating findings from recent studies, we seek to offer new perspectives and directions for future research in this emerging field.

BIOLOGICAL BASIS OF THE BILIARY-ENTERIC COMMUNICATION OF LIVER-GUT AXIS

Composition and function of bile

Bile is a complex fluid produced by the liver and stored in the gallbladder, playing a crucial role in digestion and absorption of lipids. It is composed of bile acids, cholesterol, phospholipids, bilirubin, and electrolytes. Bile acids, the primary component, are synthesized from cholesterol in the liver and conjugated with glycine or taurine to form bile salts, which are more soluble and effective in emulsifying fats[14]. The emulsification process facilitates the action of pancreatic lipase, aiding in the breakdown of dietary fats into fatty acids and monoglycerides, which are then absorbed in the small intestine. Additionally, bile serves as a route for the excretion of bilirubin, a byproduct of hemoglobin breakdown, and excess cholesterol. The enterohepatic circulation of bile acids, where they are reabsorbed in the ileum and returned to the liver, is essential for maintaining bile acid homeostasis and efficient digestion[15].

Composition and role of gut microbiota

The gut microbiota is a diverse group of microorganisms, such as bacteria, archaea, viruses, and fungi, that live in the gastrointestinal tract. These microorganisms are crucial for host health. They help with digestion, synthesize vitamins, and regulate the immune system. The composition of the gut microbiota is affected by factors like diet, age, genetics, and environmental influences[16]. In healthy individuals, the gut microbiota mainly consists of two phyla: Firmicutes and Bacteroidetes. There are also smaller amounts of Actinobacteria, Proteobacteria, and Verrucomicrobia. The gut microbiota contributes to the metabolism of complex carbohydrates, producing SCFAs like acetate, propionate, and butyrate, which serve as energy sources for colonocytes and have anti-inflammatory properties[17]. An imbalance in the gut microbiota, known as dysbiosis, has been associated with several diseases, such as inflammatory bowel disease, obesity, and cancer[18].

Signaling mechanisms of the biliary-enteric communication of liver-gut axis

The biliary-enteric communication of liver-gut axis includes intricate signaling processes that control bile acid metabolism and the composition of gut microbiota (Figure 1). Primary bile acids are synthesized by cholesterol in liver cells and secreted into bile by the transporter bile salt export pump. When eating, the gallbladder contracts, releasing bile into the duodenum, where it forms mixed micelles with phospholipids, fatty acids, cholesterol, and fat-soluble vitamins, which are wrapped in amphiphilic conjugated bile acids. Enables it to be absorbed before being transported back to the liver via portal circulation via high-affinity transporters on the top and basolateral sides of intestinal cells. Every day, hundreds of milligrams of bound bile acids leave the enterohepatic circulation and enter the large intestine, where they are quickly unbound by the bacterial bile brine hydrolysis enzyme, releasing taurine or glycine and free bile acids. When bound bile acids reach the end of the ileum, they are transported to intestinal cells via ileal sodium-bile acid cotransporter, bound to recombinant human fatty acid binding protein 6 and transported to the portal circulation via the basolateral expression of intestinal cells OSTα and OSTβ.

Figure 1 Signaling mechanisms of the biliary-enteric communication of liver-gut axis. FXR: Farnesoid X receptor; FGF: Fibroblast growth factor; TGR: Takeda G protein-coupled receptor; GLP: Glucagon-like peptide 1.

Bile acids function as signaling molecules. They activate nuclear receptors, including the FXR and the G protein-coupled bile acid receptor 1 (GPBAR1 or TGR5). FXR is primarily expressed in the liver and intestine and regulates genes involved in bile acid synthesis, transport, and detoxification[19]. When FXR is activated in the ileum, it promotes the expression of fibroblast growth factor 19. This factor inhibits bile acid synthesis in the liver, helping to maintain bile acid homeostasis. TGR5, expressed in various tissues including the intestine, modulates energy expenditure and glucose metabolism by stimulating the release of glucagon-like peptide-1 from enteroendocrine cells[20]. The ability of bile acids to activate TGR5 was different, and the order from high to low was lithic acid, deoxycholic acid, goose deoxycholic acid, ursodeoxycholic acid and cholic acid. Additionally, bile acids affect the composition of gut microbiota by exerting antimicrobial effects, which shape the structure of the microbial community[21]. The ability of the gut microbiome to produce secondary bile acids is an important factor in the regulation of inflammation and the recruitment, differentiation, and activation of innate and adaptive immune cells. The interplay between bile acids and gut microbiota creates a feedback loop that impacts both host metabolism and microbial ecology[22] (Figure 1).

Role of liver in Bilio-intestinal junction

Current studies believe that if the intestinal mucosal barrier function is damaged, the permeability of the intestinal mucosa is changed, thus causing bacteria and their products (such as endotoxins, etc.) in the intestinal tract to enter the liver through the blood circulation (usually the portal vein system). It causes the innate immune system of liver, such as Kupffer cells (KC), to be activated by intestinal products and release a series of inflammatory factors, which can further cause damage to intestinal mucosa and distant organs. Therefore, the liver plays an important role in the bilioenteric axis. The liver is a multifunctional organ responsible for a range of key functions in the body. Its main roles include detoxifying harmful substances in the blood, metabolizing drugs and hormones, and storing essential nutrients. In addition, it produces bile, which is important for the digestion and absorption of fats and fat-soluble vitamins. Produced in the liver and stored in the gallbladder, bile acids play an integral role in the digestion and absorption of fat. When released into the gut, bile acids also interact with the gut microbiome. This interaction not only contributes to nutrient absorption, but also affects the composition of the microbial community. Any disruption in bile acid regulation can have knock-on effects on gut microbiota and liver function. For example, an imbalance of bile acids can lead to liver diseases such as cholestasis, which is reduced bile flow that can lead to liver damage.

THE ROLE OF MICROBIOTA IN CCA

Association between dysbiosis and CCA

Dysbiosis, which refers to an imbalance in gut microbiota, is increasingly recognized as a key factor in the development of several cancers, including CCA. Research indicates that CCA patients often have different microbial profiles than healthy individuals. For example, a study by Saab et al[23] showed that the biliary microbiota of CCA patients had a higher abundance of Bacteroides and a lower abundance of Firmicutes compared to controls[23]. This dysbiosis can create a pro-inflammatory environment that promotes carcinogenesis. The altered microbiota composition can disrupt the mucosal barrier, leading to increased permeability and systemic inflammation, which are known risk factors for cancer development[24]. Furthermore, the presence of specific pathogenic bacteria, such as Enterococcus and Klebsiella, in the biliary tract is associated with CCA progression, indicating that these microorganisms may play a direct role in tumorigenesis[25].

Carcinogenic mechanisms of specific microorganisms

Certain microorganisms are known to directly cause cancer in CCA. For example, Helicobacter species, particularly Helicobacter pylori, have been implicated in the development of biliary tract cancers. These bacteria induce chronic inflammation and produce carcinogenic compounds like nitrosamines. These compounds can cause DNA damage and mutations[26]. Another example is Fusobacterium nucleatum, which promotes cancer cell growth and movement by activating the TLR4/NF-κB and IL-6/STAT3 signaling pathways[27]. These pathways are essential for sustaining the inflammation that fosters tumor growth and metastasis. Additionally, these bacteria can suppress the host immune response, which further promotes cancer progression[25].

Microbiota as potential biomarkers

The unique microbial signatures linked to CCA present opportunities for creating new biomarkers that aid in early diagnosis and prognosis. Some microorganisms are known to have direct carcinogenic effects on CCA. For example, Helicobacter species, particularly Helicobacter pylori, have been implicated in the development of biliary tract cancers. Helicobacter species can induce chronic inflammation and produce carcinogenic compounds like nitrosamines, leading to DNA damage and mutations[26]. Another example is the role of Fusobacterium nucleatum, which has been shown to promote cancer cell proliferation and migration through the activation of the TLR4/NF-κB and IL-6/STAT3 signaling pathways[27]. These pathways are crucial in maintaining the inflammatory environment that supports tumor growth and metastasis. Additionally, the presence of these bacteria suppresses the host immune response, which further facilitates cancer progression[25].

For instance, the increased abundance of Veillonella species in the gut microbiota of CCA patients has been proposed as a potential biomarker for distinguishing CCA from other liver diseases[27]. Similarly, the depletion of beneficial bacteria such as Ruminococcus gnavus in CCA patients could serve as an indicator of disease progression[28]. Identifying these microbial markers may enable the creation of non-invasive diagnostic tools, like fecal microbiota analysis, which could greatly enhance early detection rates and improve patient outcomes[24]. Furthermore, monitoring changes in the microbiota composition could also provide insights into the effectiveness of therapeutic interventions and help in tailoring personalized treatment strategies[25].

BILE ACID METABOLISM AND ITS RELATIONSHIP WITH CCA

Bile acid synthesis and metabolic pathways

The liver synthesizes bile acids from cholesterol through a series of enzymatic reactions, mainly involving the cytochrome P450 enzyme family. Humans primarily synthesize two bile acids: Cholic acid and chenodeoxycholic acid. These acids are conjugated with glycine or taurine to form bile salts, which are then secreted into the bile ducts. These bile salts are essential for emulsifying and absorbing dietary fats in the small intestine. After facilitating fat digestion, bile salts are reabsorbed in the ileum and returned to the liver through enterohepatic circulation for reuse. This efficient recycling process means that only a small amount of bile acids is lost in the feces, requiring ongoing synthesis to maintain the bile acid pool[29]. Disruptions in bile acid synthesis or metabolism can lead to an accumulation of toxic bile acids, contributing to liver and biliary diseases, including CCA. Bile acid metabolism is closely related to intestinal flora, some intestinal bacterial groups participate in human intestinal bile acid metabolism, as shown in Table 1.

Table 1 Bacterial groups involved in bile acid metabolism in the human gut.

Bacterial group	Enzymatic	Substrate	Product
C. scindens, Clostridium hylemonae, Holdemania filiformis, Methanosphaera, Olsenella	C12 oxidation or differential isomerization (epitope pathway)	Cholic acid, deoxycholic acid	12-OxoCDCA, 12-OxoLCA, epiCA
P. hiranonis, C. leptum, Proteocatella sphenisci, Faecalicatena contorta	C7- dehydroxylation	Ursodeoxycholic acid, goose deoxycholic acid, cholic acid	Lithocholic acid, deoxycholic acid
Clostridium sporogenes, C. paraputrificum, Clostridium sordellii	C3- dehydroxylation	3-sulfocholic acid	Iso Lithocholic acid
Bacteroides spp, R. gnavus	C12- dehydroxylation	Cholic acid, deoxycholic acid	CDCA from CA, LCA from DCA
C.hylemonae, Parabacteroides spp, Holdmania hathewayi, Alistipes spp	Heterogenic bile acid pathway	Lithocholic acid, deoxycholic acid, isoDCA, isoLCA	AlloDCA, isoDCA, alloLCA, isoDCA

Role of bile acids in the tumor microenvironment

Bile acids play a role in shaping the tumor microenvironment (TME) of CCA. They influence key cellular processes such as inflammation, apoptosis, and cell proliferation. For instance, bile acids can activate nuclear receptors such as the FXR and the pregnane X receptor, which regulate genes involved in bile acid homeostasis, lipid metabolism, and detoxification pathways. Dysregulation of these receptors can create an inflammatory TME that promotes tumor growth and progression[30]. Additionally, bile acids induce oxidative stress and DNA damage, which further promotes carcinogenesis. The interaction between bile acids and gut microbiota also plays a significant role in shaping the TME, as microbial metabolism of bile acids can produce secondary bile acids with pro-inflammatory and carcinogenic properties[31].

Carcinogenic mechanisms of abnormal bile acid metabolism

Abnormal bile acid metabolism is a critical factor in the pathogenesis of CCA. Elevated levels of bile acids can lead to chronic inflammation and bile duct injury, creating a pro-carcinogenic environment. This chronic inflammatory state is characterized by the activation of inflammatory cytokines and signaling pathways such as NF-kB, which can promote cell proliferation and inhibit apoptosis[32]. Furthermore, bile acids can promote oncogene expression and reduce tumor suppressor gene activity, leading to malignant transformation. For example, the bile acid deoxycholic acid has been shown to promote the expression of cyclooxygenase-2, an enzyme involved in inflammation and carcinogenesis[33]. Additionally, bile acids can modulate the immune response by affecting the recruitment and function of immune cells within the TME, thereby facilitating immune evasion by tumor cells[34].

THE ROLE OF INFLAMMATORY RESPONSE IN CCA

Chronic inflammation and CCA

Chronic inflammation is a known risk factor for several types of cancer, including CCA. Chronic inflammatory conditions like PSC and liver fluke infections significantly increase the risk of developing CCA[35]. These inflammatory conditions create a microenvironment that supports carcinogenesis by continuously releasing pro-inflammatory cytokines, growth factors, and ROS. These factors induce DNA damage, promote cellular proliferation, and inhibit apoptosis, thereby facilitating malignant transformation[36]. Additionally, chronic inflammation activates signaling pathways like NF-κB and STAT3, which are essential for sustaining inflammation and promoting tumor development[37].

Inflammatory response triggered by the biliary-enteric communication of liver-gut axis

The biliary-enteric communication of liver-gut axis plays a pivotal role in modulating inflammatory responses that contribute to CCA development. Disruptions in the gut microbiota, or dysbiosis, can lead to increased intestinal permeability, allowing translocation of bacterial products LPS into the portal circulation[38]. These bacterial products can trigger hepatic inflammation by activating toll-like receptors on KC and other liver-resident immune cells, leading to the production of pro-inflammatory cytokines such as IL-6, TNF-α, and IL-1β[39]. This inflammatory cascade not only promotes a pro-tumorigenic microenvironment but also enhances the recruitment of immune cells that further sustain inflammation and fibrosis, creating a vicious cycle that facilitates CCA progression[40].

Impact of inflammatory mediators on the TME

Inflammatory mediators significantly influence the TME in CCA. Cytokines such as IL-6 and TNF-α play dual roles by promoting both inflammation and immunosuppression within the TME[41]. IL-6, for instance, has been shown to enhance tumor cell proliferation, angiogenesis, and metastasis while also modulating the immune response to favor tumor growth[42]. Additionally, tumor-associated macrophages (TAMs) within the TME can polarize towards an M2 phenotype under the influence of these cytokines, further promoting tumor progression and immune evasion[43]. The interplay between these inflammatory mediators and immune cells creates a supportive niche for CCA cells, enhancing their survival and resistance to therapies[44].

THE IMMUNE SYSTEM IN BILIARY-ENTERIC SHAFT WITH BILE DUCT CARCINOMA IN INTERACTION

Immune cells in CCA

Immune cells play a critical role in the TME of CCA, influencing both tumor progression and response to therapy. Tumor-infiltrating lymphocytes (TILs), such as CD8⁺ cytotoxic T cells and regulatory T cells (Tregs), play a crucial role in this context. CD8⁺ T cells are typically linked to antitumor immunity. In contrast, Tregs often create an immunosuppressive environment that promotes tumor growth. Research indicates that the presence of TILs, especially CD8⁺ T cells, is associated with a better prognosis for CCA patients[45].However, the immunosuppressive environment created by Tregs and other immune cells, such as TAMs, can reduce the effectiveness of immune responses[1]. Recent studies emphasize the need to understand the balance between these immune cell populations to create effective[46].

Regulation of immune response by the gut-liver axis

The connection between the gut and liver plays a significant role in the immune response to CCA. The liver is constantly exposed to antigens and microbial products from the gut through the portal vein, which can affect liver immunity[47]. An imbalance in gut microbiota can increase the gut barrier's permeability, allowing microbial products to enter the liver and trigger inflammation and immune responses[48]. This interaction can either enhance the body's ability to fight tumors or create an environment that suppresses immune responses, depending on the type of microbial products and the condition of the gut microbiota[49]. Understanding these mechanisms is essential for creating strategies to adjust the gut-liver connection and boost immune responses against CCA[50].

Potential directions for immunotherapy research

Immunotherapy is a promising treatment for CCA, and multiple strategies are currently being studied. Immune checkpoint inhibitors (ICIs) targeting PD-1/PD-L1 and CTLA-4 have shown potential, although their efficacy is often limited by the immunosuppressive TME[51]. Combining ICIs with other treatments, like chemotherapy or targeted therapy, may improve their effectiveness. Additionally, agents that modulate the gut microbiota could also play a role[52]. Furthermore, adoptive cell therapies, such as CAR-T cells and TILs, are being investigated for their ability to target specific tumor antigens[53]. Future research should prioritize identifying biomarkers that predict immunotherapy responses and developing combination therapies to overcome the immunosuppressive barriers in CCA[43]. Understanding how the gut and liver interact with immune responses is crucial for optimizing these treatment strategies[54].

CONCLUSION

In summary, the biliary-enteric communication of liver-gut axis is crucial in the development and progression of CCA, a cancer known for its poor prognosis and complex causes. Through a comprehensive analysis of current research, we have elucidated various mechanisms by which the biliary-enteric communication of liver-gut axis influences CCA. These mechanisms involve changes in the gut microbiota, bile acid metabolism, inflammatory responses, immune system interactions, and metabolic pathways. Recent studies have highlighted the dysbiosis of the gut microbiota as a significant factor in CCA development, with particular microbial species being implicated in carcinogenesis. The gut microbiota not only serves as a potential biomarker but also influences bile acid metabolism. Abnormal bile acid metabolism, in turn, creates a tumor-promoting microenvironment by affecting cellular signaling pathways and inducing chronic inflammation. The bile-gut axis worsens chronic inflammation, a known risk factor for CCA, creating an environment that promotes tumor growth and progression. The immune system's interaction with the bile-gut axis adds another layer of complexity. Immune cells play crucial roles in the TME, and their functions are modulated by signals originating from the gut microbiota and bile acids. Understanding these interactions opens new avenues for potential immunotherapy approaches in the treatment of CCA. However, despite these advancements, the current research landscape still faces several limitations. Many studies are still in preliminary stages, often limited by small sample sizes and the need for more robust, longitudinal data. There is also a significant gap in translating these findings into clinical practice. Future research should focus on large-scale, multi-center studies to validate these mechanisms and explore their clinical applicability. Clinically, the bile-gut axis offers promising therapeutic targets. Strategies aimed at modulating the gut microbiota, normalizing bile acid metabolism, and controlling inflammation could provide new treatment avenues for CCA. Potential interventions could include probiotics, bile acid sequestrants, anti-inflammatory agents, and new immunotherapy options.

In conclusion, the biliary-enteric communication of liver-gut axis represents a multifaceted and influential player in the development and progression of CCA. While current research has laid a strong foundation, there remains much to uncover. Future studies should aim to bridge the gap between basic research and clinical application, ultimately improving diagnostic and therapeutic strategies for this challenging malignancy.