Structure-function effects of different pectin chemistries and its impact on the gastrointestinal immune barrier system

Abstract

The gastrointestinal immune system is crucial for overall health, safeguarding the human body against harmful substances and pathogens. One key player in this defense is dietary fiber pectin, which supports the gut’s immune barrier and fosters beneficial gut bacteria. Pectin’s composition, including degree of methylation (DM), RG-I, and neutral sugar content, influences its health benefits. This review assesses how pectin composition impacts the gastrointestinal immune barrier and what advantages specific chemistries of pectin has for metabolic, cardiovascular, and immune health. We delve into recent findings regarding pectin’s interactions with the immune system, including receptors like TLRs and galectin 3. Pectin is shown to fortify mucosal and epithelial layers, but the specific effects are structure dependent. Additionally, we explore potential strategies for enhancing the gut immune barrier function. Understanding how distinct pectin chemistries affect the gastrointestinal immune system is vital for developing preventive and therapeutic solutions for conditions related to microbiota imbalances and immune issues. Ultimately, this review offers insights into strategies to boost the gut immune barrier’s effectiveness, fostering better overall health by using specific pectins in the diet.

Keywords:

• Gastrointestinal immune system
• pectin
• barrier function
• degree of methylation
• dietary fiber

Introduction

Dietary fibers are plant-based carbohydrate structures that resist the digestive enzymes in the upper part of the human gastrointestinal tract and can reach the large intestine intact and serve as a fermentation substrate for the gut microbiota. These dietary fibers are considered to provide many health benefits (Anderson et al. 2009; Barber et al. 2020). It has been suggested that dietary fiber prevents or lowers the frequency of chronic diseases such as the risk for both type 1 and 2 diabetes and certain gastrointestinal diseases and they have been suggested to prevent or delay the development of heart failure and hypertension (Marques et al. 2017; Petruzziello et al. 2006). One such dietary fiber with health benefits is pectin. Pectin was traditionally seen as a gelling agent, thickening agent, and stabilizer in the food sector (Christensen 2020) and not so much as a health-promoting agent. In recent years it is more and more utilized as a fat replacer and health-promoting functional ingredient (Ciriminna et al. 2015; Min et al. 2010). Pectin has been shown to be instrumental in preventing gastrointestinal diseases (Fan et al. 2020; Jiang et al. 2016; Vogt et al. 2016), lower hyperglycemia (Hu et al. 2020; Liu et al. 2016), alleviate inflammatory processes (Hu et al. 2020; Vogt et al. 2016), and contribute to improved metabolic activities in cells such as by promoting mitochondrial health (Hu et al. 2020). Also, pectin is recognized for its beneficial effects on gut microbiota (Jiang et al. 2016). All these effects are highly dependent on the chemistry of the pectin applied. It has been shown that the diverse structural properties of pectin, such as degree of methylation (DM), RG-I, and neutral sugar composition, can contribute to their functional properties and final effect on the consumer’s health (Sousa et al. 2015). A major distinction in pectins is made in the DM but also other chemical characteristics as will be outlined in a dedicated section in this review have been shown to have a strong impact on its biological efficacy in promoting health. The current knowledge about this will be reviewed here.

Originally the health benefits were attributed to the possible prebiotic effects of pectin. Pectins have been shown to stimulate the growth of microbiota such as Ruminococcaceae, Succinivibrionaceae, Lachnospiraceae and Bacteroidaceae by supporting the adhesion of microbial cells to small intestinal cells (Zhao et al. 2018) and by stimulating production of fermentation products short-chain fatty acids (SCFAs) such as acetic and butyric acid (Bianchi et al. 2018; Slavin 2013). Enhanced consumption of citrus pectin with higher galacturonic acid (GalA) content (> 74%) was shown to alter the microbiota and its microbial metabolites (Li, Zhang, and Yang 2018). Especially the relative abundance of bacteroides, parabacteroides, olsenella, and bifidobacteria increased in the gut microbiota of mice on a high-fat diet, and also enhanced the production of SCFAs such as acetic acid and propionic acid (Li, Zhang, and Yang 2018). Pectin may also modulate microbiota composition by changing the ability of gut epithelial cells to allow adherence of gut bacteria. Early studies with crude pectin preparations have shown that some pectins may enhance the adhesion of Lactobacillus rhamnosus and Bifidobacterium bifidum to the gut epithelial cell line Caco-2 cells (Naqash et al. 2017; Parkar et al. 2010).

Not only pectin induced SCFA production but also pectin induced enhanced daily output of stool-neutral sterols and bile acids are considered to be health effects (Dongowski and Lorenz 2004; Reddy, Watanabe, and Sheinfil 1980). These effects are also pectin structure dependent as intestinal steroids in rats were higher with higher DM and higher molecular weights of pectin (Dongowski and Lorenz 2004). However, effects are not always dependent on the effects on gut microbiota. Also, direct effects on intestinal cells have been reported. Some pectins and in particular the lower DM pectins can stimulate the secretion of intestinal mucin by goblet cells and enhance the gut barrier function by forming a network of pectin with mucus which can exert a stronger protective effect toward toxins and pathogens (Hino et al. 2013; Liu et al. 2005). The gut immune barrier system plays a crucial role in preserving intestinal health and well-being (Bischoff 2011). During recent years more insight has become available on the interaction between pectin and different barrier layers within the gut immune barrier system. They do so by interacting with specific receptors on cells. It is now recognized that pectin interacts with different receptors such as pattern recognition receptors (PRR), galectin-3 (Gal-3) and pectin derived short chain fatty acids can bind to G protein-coupling receptors (GPCRs). In this review, we will discuss these interactions and how different types of pectins may interact with different layers of the gastrointestinal immune barrier. By reviewing the current knowledge on the impact of different pectin chemistries on the gastrointestinal immune barrier we hope to contribute to the development of customizing pectin formulations or developing products that can aid in preventing or delaying the onset of human disease.

Pectin and chemical variation in its composition

Pectin is a complex polysaccharide found in the primary cell walls of plants, where it provides structural support and plays a role in intercellular adhesion between plant cells (Caffall and Mohnen 2009). As pectin can enhance the viscosity of solutions, it is commonly used as a thickening agent (Chan et al. 2017; Ciriminna et al. 2015). Apple pomace and citrus peels are the main sources of commercially applied pectins. Other sources that have been considered include sugar beet, sunflower seed heads, and various fruits (May 1990). Pectin from sugar beet has several disadvantages as a commercial source due to its low gelling ability compared to pectin from apple and citrus sources. All these pectins may differ in their chemical structure and have different physicochemical effects on the product or on health benefits. Independent of the source, pectins can be found throughout the plant in different conformations and structures.

Structure of pectin

Pectin is composed of multiple domains, including homogalacturonan (HG), rhamnogalacturonan-I (RG-I), rhamnogalacturonan-II (RG-II), apiogalacturonan, and xylogalacturonan (Figure 1). These domains can vary in their chemical structure and distribution within different plant organs and plant sources (Kontogiorgos 2020).

Figure 1. Schematic structure of pectin. Pectin molecules contain different regions such as homogalacturonan, rhamnogalacturonan I, rhamnogalacturonan II, xylogalacturonan, and apiogalacturonan. The primary structure that dominates the smooth pectin region is the linear, partially esterified HG backbone, accounting for approximately 60% of the total pectin. The hairy regions, on the other hand, consist primarily of highly branched RG I and RG II domains, with smaller contributions from xylogalacturonan and apiogalacturonan.

Galacturonans

HG, as the smooth area of pectin, is a linear polymer of alpha-1,4-linked D-galacturonic acid residues with some of the acid groups being esterified with methyl groups (Mohnen 2008). It is thought to be a component of all primary cell walls and is involved in calcium-mediated gel formation, which contributes to maintenance of the integrity of the cell wall (Figure 3). The GalA residues can be methyl-esterified at C-6. The de-esterification process is regulated by the action of pectin methyl esterase (PME), which removes methyl groups from the polymer (Mort, Qiu, and Maness 1993). This de-esterification process is complex but highly regulated. It results in the ability to stretch acidic residues that can associate with other HG chains through calcium cross-links (Wang, Wilson, and Cosgrove 2020). However, the functional properties of pectin are not only influenced by the degree of methyl esterification on the GalA backbone, but also by the distribution of non-esterified GalA residues which refers to the degree of blockiness (DB) or distribution of non-esterified GalA residues on the HG backbone (Willats, Knox, and Mikkelsen 2006). Also, in some plant sources such as sugar beet and spinach, the GalA residues in the HG backbone can be acetyl-esterified at O-2 and/or O-3 which has been shown to have a negative impact on the gelation property of pectin (Perrone et al. 2002; Ralet et al. 2003, 2005).

HG can also be substituted with other side chains such as β-d-apiofuranosyl residues and xylose to form apiogalacturonan and xylogalacturonan respectively, or even more complicated side chains termed A-F which contain thirteen different types of sugars attached to the GalA residues via twenty-one different glycosidic linkages which form RG-II (Kontogiorgos 2020; Mohnen 2008; Ndeh et al. 2017). RG-II is characterized by a short HG backbone with complex side chains. It is widely distributed in primary cell walls and is considered to be attached to HG (Vincken et al. 2003). RG-II is resistant to degradation by pectin-degrading enzymes, suggesting it has a distinct structural role in maintaining the integrity of the cell wall (Figure 2).

Figure 2. The gastrointestinal immune barrier. The gastrointestinal immune barrier is composed of three main layers: the mucus layer, the epithelial layer, and the lamina propria. These layers work together to protect the GI tract from harmful substances and to maintain immune homeostasis. (1) The mucus layer is the first line of defense in the GI immune barrier. It is composed of a gel-like matrix called mucus, which is produced by goblet cells. The mucus layer acts as a physical barrier, trapping pathogens, toxins, and other foreign particles. It also contains antimicrobial molecules that assist in neutralizing pathogens and prevent their entry into the underlying tissues. (2) The cellular layer is located beneath the mucus layer and consists of epithelial cells that form a tight barrier. These cells are connected by tight junctions, preventing the passage of harmful substances between them. The cellular layer also contains intraepithelial lymphocytes (IELs), specialized immune cells that monitor and protect the epithelium from invading pathogens. IELs can quickly respond to antigens and release immune mediators to eliminate threats. (3) The lamina propria is the third layer of the GI immune barrier, located beneath the cellular layer. It is a connective tissue layer containing a rich population of immune cells, such as lymphocytes, macrophages, and dendritic cells. These immune cells sample the luminal content, detect pathogens, and initiate immune responses when necessary. The lamina propria is also involved in immune tolerance, allowing beneficial nutrients and commensal bacteria to pass while preventing the invasion of harmful substances.

RG-I

The backbone may contain RG-I which is composed of alternating L-Rhamnose (Rha) and D-GalA residues. The backbone can be modified with various side chains, such as arabinan, galactan, and arabinogalactan, which contribute to the structural and functional diversity of pectin. These regions are known as the hairy area of the pectin (Kontogiorgos 2020). Most Rha residues were substituted at O-4 while less GalA residues are substituted at O-2 (Yapo 2011). The side chains various enormously based on the plant’s organ, type of plant, and development stages of the plant (Lerouge et al. 1993; Vincken et al. 2003). RG-I is highly variable in both its fine structure and distribution within the cell wall. It is composed of neutral oligosaccharides such as galactan and arabinan epitopes, but the relative proportions of these epitopes can vary within different regions of plant organs (Kontogiorgos 2020). RG-I is covalently attached to HG and possibly RG-II, but the precise nature of these connections is not well understood. RG-I has been implicated in a range of processes and health benefits (Figure 3).

Figure 3. The shifted egg-box model of the crosslinking between calcium and homogalacturonan.

RG-II

RG-II is characterized by a highly complex and branched structure. Similar to RG-I, it consists of a backbone of alternating GalA and Rha residues. However, RG-II has additional structural features that distinguish it from RG-I, including unusual sugar residues, such as 2-keto-3-deoxy-d-lyxo-heptulosaric acid (KDO), apiose, and specific glycosidic linkages. RG-II can have various side chains attached to its backbone, including arabinan, arabinogalactan, and other sugar residues. These side chains can be further modified with ester and glycosidic linkages, forming intricate networks within the cell wall matrix. RG- II is known as a bioactive component as it attenuates proliferation of tumor cells (Ai et al. 2018; Park et al. 2013).

Apiogalacturonan and xylogalacturonan

Apiogalacturonan is primarily composed of linear chains of GalA residues and substituted with mono- or disaccharide apioduranosyl (Schols and Voragen 1996). Apiogalacturonan molecules can be linked together through HG regions or form cross-links with other pectic polysaccharides, such as RG-I and RG-II, to provide structural integrity to the cell wall. Xylogalacturonan is a complex polysaccharide that contains both GalA and xylose residues. It is a component of hemicellulose, which is another major constituent of the plant cell wall. Xylogalacturonan chains can be branched or linear, and their arrangement can vary between different plant species. Both apiogalacturonan and xylogalacturonan are involved in various biological processes, including cell growth, cell adhesion, and responses to environmental stresses (Popov and Ovodov 2013).

The connections and interactions within the pectic network are likely to be complex and multifaceted (Cardoso, Coimbra, and Lopes da Silva 2003). In addition to covalent attachment to other macromolecules in the cell wall, pectic polysaccharides have the potential to interact with ions and low-molecular-weight compounds. For example, acidic domains can bind cations such as calcium and aluminum, which can influence the conformational states and properties of the pectic network (Decreux and Messiaen 2005). Polyamines can also compete for calcium-binding sites on HG, which can affect the supramolecular assembly of pectins and the action of PMEs.

Molecular genetic approaches have been applied to provide insight into the structural chemistry of pectin. This includes the manipulation of pectin structure in planta, the isolation of mutants, and the elucidation of biosynthetic pathways. The identification and characterization of genes involved in pectin biosynthesis and modification has been instrumental for the development of e.g., specific monoclonal antibodies (Hu et al. 2017). However, the complexity and heterogeneity of pectin, as well as the indirect relationship between synthesis genes and carbohydrate structure, make the functional analysis of pectin a challenging task.

Gastrointestinal immune barrier system

Before reviewing current knowledge on how pectins impact the gastrointestinal immune barrier, it is needed to review the composition and organization of this crucial protective layer that protects the human body from undesired infectious agent present in the lumen of the intestine. The intestinal tract is one of the largest surfaces between the outside environment and inside of the body (Kagnoff 1993). A major function of the intestinal tract is to absorb nutrients and to transport them into the circulation. Another very important function is to serve as the gatekeeper for the human body. Many pathogens and toxins enter the human body via the gastrointestinal tract. The gastrointestinal immune barrier prevents these substances from entering the human system (Johansson, Larsson, et al. 2011; König et al. 2016). The gastrointestinal tract is the largest active immune organ of humans. It is 795.5 ± 129 cm in length and harbors the largest number of immune cells which is about 80% of all immune cells in the lamina propria (Hounnou et al. 2002; Mowat and Agace 2014). Combined with the lamina propria (LP), the gastrointestinal immune barrier is composed of an immune active part that serves to fight intruders such as pathogens, toxins and possible allergens and a barrier that is the true gatekeeper of the human body. From the lumen to the LP the gastrointestinal immune barrier is composed of three layers (Figure 2).

The first layer is composed of a mucus layer, which plays a crucial role in the gastrointestinal immune barrier. This layer acts as a protective shield, trapping potential pathogens, toxins, and other harmful substances, preventing them from directly interacting with the epithelial cells (Allaire, Crowley, et al. 2018; Bourlioux et al. 2003). This layer is composed of a highly glycosylated polymeric protein called mucin, which forms a gel-like structure overlying the intestinal epithelium. The mucosal layer serves as a chemical barrier, limiting the contact between the microbiome and epithelial cells. The intestinal mucus layer serves as a barrier but is also a feeding layer for the gut microbiota (Paone and Cani 2020; Szentkuti et al. 1990). The mucus layer is different in composition in the small and large intestine (Ermund et al. 2013; Johansson, Ambort, et al. 2011). In the small intestine, the mucus layer is thinner (∼20 μm) and less dense compared to the large intestine (thickest in the colon, ∼830 μm) (Atuma et al. 2001). The tinner layer in the small intestine facilitates efficient absorption of nutrients and expedites the movement of digested food through the intestinal tract. The mucus layer in the large intestine is thicker and composed of a two-layered mucus system and is more densely packed. It contains a higher concentration of mucins, antimicrobial peptides, and other immune-related molecules. This thicker mucus layer provides enhanced protection and serves as a habitat for the gut microbiota, which plays a vital role in gut health and immune regulation (Johansson, Ambort, et al. 2011; Johansson, Larsson, et al. 2011).

The second layer of the barrier is the intestinal epithelial barrier, which is a crucial component of the gastrointestinal immune system that serves to protect the body from harmful substances and pathogens that enter the gut. Similar to the mucus layer, the composition and characteristics of the intestinal epithelial barrier also differ between the small and large intestine. The intestinal cellular layer is composed of a single epithelial layer and intraepithelial cells which includes M cells, goblet cells, Paneth cells, and enteroendocrine cells (Goto and Kiyono 2012). M cells, also known as microfold cells, are specialized epithelial cells found in the Peyer’s patches (PP) of the small intestine. They play an important role in the immune system by sampling antigens from the gut lumen and presenting them to immune cells in the underlying lymphoid tissue (Man, Prieto-Garcia, and Nicoletti 2004). Goblet cells, named for their distinctive shape, are responsible for the production and secretion of mucus which lubricates the gut and protects the mucosa from damage and infection (Allaire, Crowley, et al. 2018). Paneth cells, located at the base of the small intestine, produce a variety of antimicrobial peptides that defend against bacterial infections (Goto and Kiyono 2012). Paneth cells are uncommon in the colon, and the presence of intraepithelial lymphocytes is even scarcer compared to the small intestine. (Mowat and Agace 2014). Enteroendocrine cells are scattered throughout the gut and secrete hormones that regulate digestion and absorption. Columnar epithelial cells, which form the bulk of the intestinal epithelium, are involved in absorbing nutrients and electrolytes and in producing mucus. Epithelial cells form a continuous, polarized monolayer that functions as a physical barrier, facilitated by tight connections between each cell (Takiishi, Fenero, and Câmara 2017). These connections are formed by tight junction proteins, which seal the paracellular pathway and conduct gate and fence functions to restrict the passage of pathogenic molecules and bacteria.

The barrier function refers to the mucosal barrier, epithelial cell layer and the tight junctions between them. The permeability of the mucosal barrier is depending on the tight junction barrier function when the epithelial cell layer is intact. The intestinal epithelial cells serve as mediators between the mucosal immune system and luminal materials, which is pivotal for maintaining mucosal homeostasis and preventing diseases (Turner 2009). The intestinal epithelial cell layer not only provides a physical barrier but also serves as a biochemical barrier that maintains intestinal homeostasis between host tissue and commensal bacteria (Peterson and Artis 2014). Intestinal epithelial cells also maintain the pathways for the delivery of antigens to the antigen-presenting cells located in the lamina propia (Peterson and Artis 2014). Barrier dysfunction has been found in many diseases such as inflammatory bowel diseases, type 2 diabetes and type 1 diabetes, obesity, and celiac disease (Everard and Cani 2013; König et al. 2016; Schleimer and Berdnikovs 2017). Some studies showed that epithelial barrier disruption in IL10^-/^- mice increased the production of IFN-γ and TNF-γ, which may be considered a significant contributing factor in the development of colitis (Arrieta et al. 2009). Genetic factors could potentially play a role in the impairment of intestinal barrier function in families affected by inflammatory bowel disease as some studies showed that the presence of specific NOD2/CARD15 mutations genetically linked to barrier defects implies that immune activation could be responsible for early barrier disruptions observed in healthy relatives and in patients prior to disease reactivation (Buhner et al. 2006; Edelblum and Turner 2009). This might lead to barrier dysfunction and could potentially activate immunoregulatory responses.

The third defense barrier is the LP, which, together with the mesenteric lymph nodes (MLN) and PP are referred to as the gut-associated lymphoid tissue (GALT) which contains a large proportion of the gastrointestinal immune cells (Mörbe et al. 2021; Spahn and Kucharzik 2004). GALT contains both CD4⁺ and CD8⁺ T cells. Immune cells are another important component of the intestinal epithelial barrier, as they play a crucial role in the immune response and the tolerance of the host against external substances (Mörbe et al. 2021). These immune cells, including dendritic cells, T cells, B cells, and macrophages, work together with the epithelial cells and the mucosal layer to protect the body from potential threats (Forchielli and Walker 2005). The complex interplay between these different components of the gastrointestinal immune system is essential for maintaining the health and integrity of the intestinal barrier.

Gastrointestinal immune barrier and gut microbiota

The human gut is home to trillions of microorganisms, which together with the host form a superorganism in which energy and metabolites are exchanged and by which homeostasis is maintained in the immune system. The gut microbiota plays a crucial role in the development and function of the mucosal immune system, particularly during early life, and can influence the balance between pro-inflammatory and regulatory host responses (Lee and Mazmanian 2010). Alterations in the composition of the microbiota, known as dysbiosis, have been linked to the development of inflammatory disorders such as inflammatory bowel diseases (IBDs) (Nishida et al. 2017).

As outlined in the preceding section, the gut immune system is composed of both innate and adaptive immune mechanisms that cooperate to protect the host and maintain intestinal homeostasis (Faderl et al. 2015; Kayama and Takeda 2012). Epithelial cells, which form the first line of defense against the microbiota, express receptors for pathogen-associated molecular patterns (PAMPs) and produce antimicrobial products and chemokines in response to PAMPs signals (Kelly and Mulder 2012; R. Marques and Boneca 2011). The gut microbiota also stimulates the production of microbicidal peptides and secretory IgA by the host, which contain the microbiota within the intestinal lumen and neutralize PAMPs (Neu, Sharma, and Young 2010).

While some members of the microbiota have adopted peace-keeper activities to colonize the intestine, others are pro-inflammatory and can stimulate the production of effector immune responses by the host. One such group is segmented filamentous bacteria, which stimulate the postnatal maturation of immune responses in the mouse gut and are necessary for the control of colonization by the invasive pathogen Citrobacter rodentium (Cerf-Bensussan and Gaboriau-Routhiau 2010). The destruction of the microbiota following treatment with antibiotics can jeopardize innate immune responses in the gut and promote colonization by pathogens (Kim, Covington, and Pamer 2017; Thaiss et al. 2016).

Intestinal dysbiosis has been observed in patients with IBDs such as Crohn’s disease and ulcerative colitis (Maloy and Powrie 2011). Dysbiosis can arise from pro-inflammatory bacteria, such as enteroinvasive E. coli strains, or from reduced levels of resident Firmicutes spp. and Bacteroides spp. and an overgrowth of proteobacteria. However, it is difficult to assign a causative role for dysbiosis in IBD as host-predisposing factors and the reciprocal nature of the regulation of the immune system and microbial community structures must be considered (Kamada et al. 2013).

Therapeutic interventions targeting the microbiota, such as antibiotics, probiotics, and fecal microbiota transplantation, have shown promise in the treatment of IBD (Walker and Lawley 2013). Antibiotics can be used to eradicate the microbiota and reduce inflammation, but the long-term effects of antibiotic treatment on the microbiota and host immune system are unknown. Probiotics, which are live microorganisms that are similar to the beneficial microorganisms found in the human gut, have demonstrated some efficacy in the treatment of IBD but their mechanisms of action are not fully understood. Fecal microbiota transplantation, in which feces from a healthy donor are transplanted into the gut of a recipient, has shown promising results in the treatment of IBD and other diseases but there are concerns about the safety and long-term effects of the procedure (Rossen et al. 2015; Wang et al. 2019).

In conclusion, the gut immune barrier system plays a crucial role in the regulation of the gut microbiota and the maintenance of intestinal homeostasis (Goto and Kiyono 2012; Kamada et al. 2013; Okumura and Takeda 2018). Dysbiosis has been linked to the development of IBDs, and therapeutic interventions targeting the microbiota show promise in the treatment of these diseases. However, further research is needed to fully understand the complex interactions between the immune barrier system, microbiota, and host immunity in health and disease.

Possible mechanisms by which specific pectin types might support gastrointestinal health

As a dietary fiber, pectin has different functions in the digestion system. Due to its gelling ability, pectin can retain water and resist digestive enzymes in the small intestine, which could be a possible mechanism for how pectin slows down the absorption of glucose and lipid (Kontogiorgos 2020) and support metabolic health. By forming a coating on the mucosal surface, pectin effectively slows down the interaction between glucose and the mucosa, thereby delaying sugar absorption and improving glycemic control (Wikiera, Irla, and Mika 2014). This interaction with mucus is accomplished as follows. Pectin can bind with divalent cations such as Mg²⁺, Ca²⁺, Ba²⁺, or Zn²⁺. Pectin and divalent cations can form a strong inter-chain by cation bonding according to the so-called ‘shifted egg-box’ model (Braccini and Pérez 2001). This process is the basis of the gel formation (Christensen 2020; Thakur, Singh, and Handa 2009). These gels integrate into the mucus and lower glucose absorption. However, we have recently shown that this is not the only mechanism by which specific pectins can lower glucose absorption. Specific pectins such as DM19HB, DM18LB, DM84LB and DM88HB can also lower apical expression of glucose transporters and thereby lower glucose absorption (Tang et al. 2023).

While being present on mucus, pectin can also contribute to metabolic and immune health by serving as a substrate for gut microbiota. As mentioned above pectin cannot be digested in the small intestine but can be fermented by gut microbiota in the large intestine. Pectinolytic organisms in the colonic tract, such as the phyla Bacteroidetes and Firmicutes (Baylisss and Houston, 1984; Chung et al. 2016; Dongowski, Lorenz, and Anger 2000; Elshahed et al. 2021; Salyers et al. 1977) can ferment pectin into beneficial metabolites, which are SCFAs. SCFAs can be used by host cells to support metabolism and lower the Ph and by that inhibit adhesion and growth of pathogenic bacteria (Koh et al. 2016). It has been suggested that pectin supplementation influences the production and composition of SCFAs. The production of acetate and butyrate was increased after 6 h pectin exposure and was mediated by clostridium cluster XIV bacterial species (Bang et al. 2018).

Chemical structure

The chemical structure of pectin plays an important role in its fermentability and functionality. This includes characteristics such as RG-Ι content and DM (Mao et al. 2019; Tian et al. 2017; Y. Zhao et al. 2021). In a study involving 177 healthy individuals, it was found that carrot-derived RG-I (cRG-I) (1.5 g per day) exhibited significant immunomodulatory properties. It was shown that RG-I had the most potent effect on alleviating symptoms caused by rhinovirus 16 infection. Notably, RG-I facilitated a quicker interferon-induced response, stimulated the expression of EIF2AK2, a crucial antiviral gene, expedited viral clearance, and resulted in a reduction of both symptom severity (by 20%) and duration (by 25%) (Lutter et al. 2021). RG-I has also been studied for its gastroprotective activity. The administration of rhamnogalacturonan (RGal) was found to effectively inhibit the activity of MPO, which serves as a marker for neutrophil infiltration, as well as an agent that reduce TNF-α levels.(Maria-Ferreira et al. 2014) These findings indicate that RGal treatment reduces the migration of neutrophils to the ulcer site, thereby promoting an improvement in the gastric inflammatory process induced by acetic acid (Maria-Ferreira et al. 2014). Interestingly, citrus pectin with low DM fermented faster and more efficiently than high DM pectin in vitro and in vivo (Dongowski, Lorenz, and Proll 2002; Tian et al. 2017). However, M Gulfi et al. suggested that higher DM apple pectin fermented more thoroughly with slightly higher production of SCFAs than lower DM apple pectin (Gulfi, Arrigoni, and Amadò 2006). Thus, the fermentability of pectin could also be influenced by the source of pectin. Also, for soy and sugar beet pectin differences in production of SCFAs were observed (Tian et al. 2016). Acetate, lactate, formate, butyrate, and propionate are the main SCFA products and intermediates (Macfarlane and Macfarlane 2003). All these SCFAs are also recognized for their immunomodulatory potential. They are considered intermediaries that facilitate communication between the intestinal microbiome and the immune system.

Receptors involved in pectin signaling

Pectins or their fermentation products might bind and signal via different types of receptors and by that partially or completely exert their health benefits (Figure 4). The fermentation products of pectin such as SCFAs have been shown to exhibit multiple mechanisms of action. SCFAs can e.g., inhibit histone deacetylase (HDAC), serve as an energy source and bind to GPCRs (Kootte et al. 2012; Lattimer and Haub 2010; Soliman 2019; Tolhurst et al. 2012). HDAC inhibition by pectin-derived SCFAs has been suggested as a potential therapeutic strategy in a variety of diseases (Blanco-Pérez et al. 2021). Also binding of pectin-derived SFCA to GPCRs has been suggested to be involved in many health benefits (Williams, Scott, and Wood 2019). via GPCRs SCFAs can activate signaling pathways that are involved in the resolution of inflammation in the gut (Lattimer and Haub 2010; Tolhurst et al. 2012). SCFAs can initiate the signal transmission by binding to Free fatty acids receptors (FFARs) such as FFAR2, FFAR3, hydroxycarboxylic acid receptor 2 (HCA2), Olfr-87(mice), and OR51E2 (humans) (Bolognini et al. 2016; Ohira, Tsutsui, and Fujioka 2017; Ratajczak et al. 2019). These receptors are not only distributed on the digestive tract epithelium but also on immune cells and smooth muscle cells of blood vessels (Ratajczak et al. 2019). The activity of the immune and inflammatory response is influenced by signaling through FFARs and HDAC activity (Kim and Betz 2018). SCFAs are believed to be responsible for maintaining the regulatory T cell (Treg) pools and its activity, which exert their effect by inhibiting HDAC (Figure 4) (Priyadarshini et al. 2021; Kim and Betz 2018).

Figure 4. Pectin and its currently known receptors in the gastrointestinal tract. Pectins and their fermentation products have various health benefits and can interact with different types of receptors. Short-chain fatty acids (SCFAs) derived from pectin fermentation can inhibit histone deacetylase (HDAC) and bind to G protein-coupled receptors (GPCRs), potentially providing therapeutic benefits for disease prevention and promoting the resolution of inflammation in the gut. SCFAs can activate signaling pathways through receptors such as FFAR2, FFAR3, HCA2, olfr-87 (in mice), and OR51E2 (in humans). Pectin can also interact with galectin-3 (gal-3) receptors, which play a role in various biological processes and have implications in preventing cancer progression and metastasis. Specific pectin chemistries can interact with pattern recognition receptors (PRRs) like TLR2 and TLR4, influencing immune responses. Pectins can also impact cholesterol metabolism and bile acid excretion. The effects of pectins on receptors and immune responses depend on their structure and composition. Pectin oligosaccharides can either enhance or downregulate inflammatory activities and immune activation, while low-DM citrus pectins can inhibit proinflammatory pathways while leaving tolerogenic pathways unaffected.

Pectin might also directly interact with receptors involved in metabolism and/or immunity. One of the best-recognized receptors for pectin is Gal-3. Gal-3 has a significant impact on a wide range of biological processes and appears to be implicated in various physiological and pathophysiological conditions (Dumic, Dabelic, and Flögel 2006). These conditions include development of organs, immune responses, as well as neoplastic transformation and metastasis (Dumic, Dabelic, and Flögel 2006). The initial discovery of Gal-3-pectin interaction by Gunning, A.P. et al. demonstrated the recognition of galactan components in pectin by the recombinant form of Gal-3 (Gunning, Bongaerts, and Morris 2009). This finding suggests potential implications in reduction of pectin on progression and metastasis formation of cancer (Gunning, Bongaerts, and Morris 2009). A few studies demonstrate that the supplementation of 1% modified citrus pectin (commercial pectin) resulted in a reduction of Gal-3 signaling which was accompanied by decreased renal fibrosis, attenuated macrophage infiltration, lower expression of pro-inflammatory cytokines, and reduced apoptosis (Kolatsi-Joannou et al. 2011). Studies in vivo demonstrate pro-inflammatory effects of Gal-3 which can be attenuated by specific pectins such as low-DM pectins (Hsu et al. 2000) Also, lower DM pectins such as lemon pectin DM5 can reduce oxidative and cytokine stress in insulin-producing beta-cells through a Gal-3 dependent mechanism (Hu et al. 2020). Potato and citrus pectins showed anticancer activity by specific binding to the human Gal-3 and impacting cell-cycle genes lowering metastasis of cancer cells (Gunning, Bongaerts, and Morris 2009).

Pectins also interact directly with immune receptors and by that participate in regulating immune responses. Our previous study shows that specific pectin chemistries allow interactions with PRR such as TLR (Beukema et al. 2021). Stimulation with lemon pectins induces activation of TLR2, TLR4 and NF-κB in a DM and DB dependent fashion (Beukema et al. 2021; Vogt et al. 2016). Pectin was also found to improve the 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA) and hepatic activity of cholesterol 7α-hydroxylase activity by impacting fecal bile acid excretion (Garcia-Diez et al. 1996).

The immunological targets of pectin might be via several receptors. Current insight is that ability to bind to these receptors or a combination thereof is highly dependent on the structure of the applied pectin. Pectin oligosaccharides have been shown to bind to TLR4, which activates nuclear factor-kappa B (NF-κB) and COX-2 signaling pathways. They are associated with inflammatory activities and enhanced immune activation (Ishisono, Yabe, and Kitaguchi 2017). However, pectin oligosaccharides can also downregulate the expression of NF-κB and COX-2 during active inflammation leading to antioxidant and anti-inflammatory effects and supporting recovery from injury or immune attack. The binding between TLR4 and pectin oligosaccharides affects the activation of the TLR4/NF-κB/COX-2 signaling cascade (Chanput et al. 2010; Li et al. 2014; Ishisono, Yabe, and Kitaguchi 2017; Jia et al. 2018). Other pectins however are predominantly regulatory and prevent too strong immune responses. Such a characteristic is attributed to lower DM citrus pectins with a DM from 7–25 that inhibit the proinflammatory TLR2-1 pathway while leaving the tolerogenic TLR2-TLR6 pathway unaffected. They interact with TLR2 via electrostatic forces (Sahasrabudhe et al. 2018). This anti-inflammatory effect is TLR2-1 dependent and unrelated to SCFAs produced by microbiota, suggesting that low-DM pectins may reduce inflammation by directly interacting with TLR2-TLR1 receptors (Beukema et al. 2021; Sahasrabudhe et al. 2018).

Pectin’s health effects in different organs

Several studies have been shown that pectin is beneficial for health, such as for not only gastrointestinal health but during recent years there are more reports demonstrating also beneficial effects on cardiovascular health, digestive health, stabilizing blood sugar levels and for weight management (Blanco-Pérez et al. 2021; Brouns et al. 2011; Brown et al. 1999; Larsen et al. 2019; Liu et al. 2016; Wu et al. 2019). During the past years it has become recognized that also on an organ level specific chemistries of pectins are responsible for these health benefits. These chemistries include variations in DM, HG and RG-I content. More researchers are now focusing on the health-related effects of pectin’s structures. The subsequent subsections will delve into the effects that are dependent on this structural aspect and discuss impact on an organ level.

Gastrointestinal health

Gastrointestinal health was shown to improve by consumption of marsh cinquefoil Comarum palustre L. pectins (57% GalA residues) at doses of 50 and 100 mg/kg. This pectin attenuated tissue myeloperoxidase production and stimulated mucin secretion. By this, it alleviated experimental colitis in mice (Popov et al. 2006). Also, some studies show that specific pectins plays a vital role in maintaining the integrity and the functionality of the mucus layer. Especially, low DM citrus pectin stimulates the production of mucin 2 (Hino et al. 2013; Liu et al. 2005; Xie et al. 2020). Another study indicates that feeding pigs with 5% apple pectin leads to increased gene expression of mucin 2 and TFF3 in both the ileum and cecum. Additionally, this effect might associate with interactions between specific bacterial genera, particularly those involved in butyric acid production, and the modulation of mucosal immunity (Wu, Zheng, et al. 2020). RG-I enhanced the promotion of mucosal enterocytes and mucus-secreting goblet cells, alongside the preservation of collagen balances and augmentation of cell proliferation in Caco-2 cells (Maria-Ferreira et al. 2018).

Pectin offers additional health benefits beyond its protective effects on the mucus layer. It helps maintain the permeability of the epithelial cell layer by upregulating tight junctions and preserving their integrity (Beukema, Faas, and De Vos 2020). From our own previous study, we showed that lemon pectin DM19 and DM43 with high DB rescue A23187-induced intestinal epithelial barrier disruption (Tang et al. 2023). Mice with DSS-induced colitis, which received pretreatment of L102 pectin (DM 38.09 ± 0.78) at doses of 100 mg/kg and 200 mg/kg, exhibited increased expression of ZO-1 (Fan et al. 2020). Interestingly, one study found that in the cecum, low DM pectin increased the expression of claudin-1 and ZO-2 at both the protein and mRNA levels (Wu et al. 2019). However, in the colon, low DM pectin did not have the same effects on claudin-1 or occludin (Wu et al. 2019). Some studies showed that 5% (wt/wt) low DM pectin supplementation in female NOD/LtJ mice enhanced the barrier integrity and the length of the villus in non-obese diabetic mice (Wu et al. 2019). Apple-derived pectin (unknown composition) supplementation (5% wt/wt) for six weeks in rats improved the expression of intestinal alkaline phosphatase and the function of the gut epithelial barrier, specifically in the maintenance of tight junctions. This might contribute to the prevention of intestinal inflammatory disorders (Jiang et al. 2016). Compared to artichoke pectins, citrus pectin treatment in mice increased the expression of MUC-3, occludin and ZO-1, and decreased the expression of intercellular adhesion molecules 1 and inducible nitric oxide synthase, in colitis mice induced by dextran sulfate sodium (DSS), to maintain the barrier function (Sabater et al. 2019).

Pectin also exhibits benefits on immune cells in the LP. Some studies show that pectic polysaccharide fragments from Biophytum petersianum have immunomodulating activity on PP and macrophages by increasing IL-6 production, which also stimulates the growth of bone marrow cells (Grønhaug et al. 2011). It has been reported that an intact lemon pectin backbone induces the activation of TLR2, TLR4 and NF-κB in a DM-dependent way, and also shows epithelial barrier protective effects (Vogt et al. 2016). Emerging new insight indicate that RG-I in pectin exhibit significant activities that are associated with better human health (Wu et al. 2019; Cui et al. 2020; Ho et al. 2016; Dong et al. 2010). RG-I from Nerium indicum flowers contain high levels of arabinose and galactose and effectively stimulate macrophages to produce nitric oxide (NO), thereby facilitating immune system activation (Dong et al. 2010). High molecular RG-I containing polymers from elderflowers have high complement fixing activity and macrophage stimulating activities (Ho et al. 2016).

Also, clear chemistry dependent effects have been reported for effects of pectins on gut microbiota. A study found that exposing adult human microbiota to cRG-I resulted in increased levels of acetate, propionate, and butyrate (Abbeele et al. 2021). Certain bacteria, such as Bacteroides dorei and Prevotella species, played a key role in breaking down cRG-I, while Bifidobacterium longum fermented only specific components of cRG-I (Abbeele et al. 2021). The authors found that carrot RG-I supplementation can reduce interpersonal compositional differences in the gut microbiota by stimulating specific bacteria such as Bacteroides dorei/vulgatus and Bifidobacterium longum (Abbeele et al. 2023). Besides the carrot-derived RG-I, pepper-derived RG-I has been found to exhibit immunomodulatory effects on phagocytosis and to also have a modulatory effect on the microbiota, resulting in increased production of SCFAs (McKay et al. 2021). Pectin can be fermented by enzymes derived from various bacterial genera such as from Enterococcus, Bifidobacteria, Lactobacilli (Blanco-Pérez et al. 2021; Chung et al. 2016; Gómez et al. 2014). It has been reported that pectins selectively modulate the gut’s microbial communities, which is related to the presence of amide groups, the composition of neutral sugars, and the distribution of HG and rhamnogalacturonan fractions, but the DM plays a crucial role as the primary regulator of microbiota composition (Larsen et al. 2019). The effectiveness of low-DM pectin L13 (resource unknown) enhanced microbiota diversity and promoted the abundance of Clostridiaceae and Lachnospiraceae at the family level, and Bacteroides and Lachnospira at the genus level and supported gastrointestinal health (Huang et al. 2022).

Cardiovascular health

For cardiovascular health it has been shown that especially administration of high DM citrus pectin (DM 70) reduced low-density lipoprotein 6–7% (Brouns et al. 2011). A meta-analysis of a trial with 277 individuals that received 34 days of treatment of pectin demonstrated that consuming pectin-rich diets significantly decreased both total and low-density lipoprotein cholesterol levels (Brown et al. 1999).

Metabolism health

Also, metabolism health was shown to be improved by pectin consumption. Consumption of citrus pectin treatment (from 500 mg to 2000 mg/kg) with nonreported structure improved the glucose tolerance and reduced insulin resistance in diabetic rats (Liu et al. 2016). It was postulated that this might occur by the impact of pectin on PI3K/Akt signaling (Liu et al. 2016). Ren et al. corroborated this finding in a study with ginseng acidic polysaccharide which contains HG and RG-I-rich pectin and arabinogalactan enriched pectin which improved lipid metabolism in type 2 diabetic rats by regulating the gut microbiota composition by increasing SCFA production and enhancing the secretion of peptide tyrosine and glucagon-like peptide-1(Ren et al. 2023).

Immune health

Many have shown improved immune health after the consumption of pectin. In a study by Nascimento, G. E et al. it was shown that administration of 300 µg/mL DM36 citrus pectin can attenuate the production of the pro-inflammatory cytokines TNF-α and IL-1β induced by lipopolysaccharide (LPS) while promoting the production of the regulatory cytokine IL-10. It was shown that this effect was dependent on the degree of methyl esterification, molecular weight and presence of side chain (Nascimento et al. 2017). Sergey V.P et al. showed that lower DM citrus pectin can decrease the adhesion of peritoneal granulocytes and their ROS production and was found to improve the survival rate of mice subjected to a lethal dose of LPS (Popov et al. 2013). Lower DM 7 citrus pectin was shown to be more efficient in blocking TLR2-1 in both immune cells and in vivo in an ileitis mice model. It therewith alleviated intestinal inflammation (Sahasrabudhe et al. 2018). A study in non-obese type 1 diabetic mice showed that 5% low DM pectin (nonreported structure) supplementation suppress the activation of NOD like receptor protein 3 (NLRP3) inflammasome and increase the gene expression of the immune modulatory cytokine TGF-β1 (Wu et al. 2019). Gal-3 is pivotal in various cellular events and a key contributor to cancer cell apoptosis (Girotti et al. 2020). Pectin’s anticancer properties depend on its binding with Gal-3. Arabinoxylan/hemicellulose in pectin, containing the RG-I domain, serves as a natural ligand inhibiting the biological activity of Gal-3 (Prado et al. 2019).

General discussion and further perspectives

Strengthening the gut immune barrier has become a hot topic for not only gut health but also for immune health. This review discusses the current knowledge about the gut immune barrier, dietary fibers pectin and the interaction between them. It has become widely accepted that not just intake of more pectin as dietary fiber supports health but that specific chemistries must be consumed. The structure determines its function. The bioactivities of pectin are often associated with various side chains, such as by galactan and arabinogalactan, as well as by backbones like rhamnogalacturonan and HG, along with the effects of substituted groups, and their synergistic interactions. Different structural characteristics of pectins such as molecular weight, RG-I, DM and DB on the HG backbone, can contribute to the benefits to immune health and gut immune barrier function (Baggio et al. 2017.; Fan et al. 2020; Gunning, Bongaerts, and Morris 2009; Hu et al. 2020; Nascimento et al. 2013; Vogt et al. 2016; Wu et al. 2021). In our research we found that low DM or/and high DB lemon pectins have TLR2-1 dependent anti-inflammatory effects, and also had a protective effect on the gut barrier function by increasing the integrity of the gut barrier (Beukema et al. 2021; Vogt et al. 2016). The extent to which the protective effects of pectin on immune health or gut epithelial barrier, are applicable to other sources of pectin has not been extensively investigated but we found that the effects are rather dependent on DM and/or DB and to a lesser extend to the source when comparing orange and lemon pectins with similar chemical structures (Beukema et al. 2021).

Pectin, functioning as a soluble fiber, plays a crucial role in the fermentation process that occurs in the cecum and large intestine. This fermentation process results in the production of fermentation products that can stimulate the production of a thick, protective mucus layer, enhance bacterial adhesion, and promote the integrity of the gut epithelial barrier. As a result, pectin helps to strengthen the function of the intestinal barrier.

Pectin also has been shown to beneficially impact microbiota composition and to support production of beneficial fermentation products such as SCFAs. Dysbiosis of microbiota is known to be involved in disease severity in both diabetes and IBD (Metwaly, Reitmeier, and Haller 2022). Specific pectins, especially the higher DM that will be fermented have been shown to act as mediators in increasing the abundance of beneficial microbiota. However, impact of RG decoration and of e.g., molecular weight has not been studied in much detail and might be even more instrumental in supporting microbiota function.

The knowledge reviewed here demonstrates that specific pectin chemistries might have specific health benefits. It has been shown that with tailoring pectins with specific chemistries gastrointestinal, cardiovascular, metabolic and immune health might be stimulated and diseases might be prevented (Ho et al. 2016; Popov et al. 2013; Soliman 2019; Vogt et al. 2016). Although not studied yet, pectins might also be beneficial for brain or mental health as they support mediators such as SCFA but also indoles (Beukema et al. 2022) that are known to support brain health (Pappolla et al. 2021; Rajamma, Krishnaswami, and Kandasamy 2022). This is all dependent on specific chemistries pectins. Unfortunately, not all studies have measured the composition of the applied pectins. This is necessary to allow side-by-side comparison of data sets and to understand which chemistries need to be used in specific target groups. Current knowledge indicates that administration of pectins and dietary fibers in general is not a one-size fits all principle but that we need to use specific chemistries for specific target groups. This review hopefully contributes to expediting the process for tailoring specific pectin chemistries for supporting and preventing specific health issues.

Disclosure statement

The authors report there are no competing interests to declare.

Additional information

Funding

This work was supported by the China Scholarship Council. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No872019.