Newly Developed Synbiotics and the Chemotherapy-Damaged Gut

Abstract

Mucositis is a common side-effect of cancer chemotherapy and radiotherapy. Features of mucositis include erythema, ulceration, and inflammation of the gastrointestinal tract accompanied by clinical symptoms of abdominal pain and digestive disturbances. New treatment strategies are required. Experimental evidence is accumulating showing therapeutic promise for new nutraceutical agents including probiotic bacteria, probiotic-derived factors, prebiotics, and plant extracts. However, the targeted development of new combinations of these agents (synbiotics) to combat mucositis remains largely unexplored. The current review addresses the potential for these nutraceutical agents to reduce the severity of chemotherapy-damaged mucositis by strategically aligning their underlying mechanism of action with features of mucositis pathogenesis. The potential for certain plant extracts to act as prebiotics, in combination with probiotics or their derived factors, is further investigated. These unique synbiotic formulations could form the basis of a new naturally sourced adjunctive approach to cancer chemotherapy.

Cancer and Mucositis

Cancer is medically known as a malignant neoplasm that results from abnormal unregulated cell growth. ¹ These transformed cells can metastasize to the surrounding uninvolved tissues via the lymphatic system or bloodstream. ² Cancer types include bladder cancer, ³ lung cancer, ⁴ breast cancer, ⁵ colon and rectal cancer, ⁶ non-Hodgkin lymphoma, ⁷ endometrial cancer, ⁸ pancreatic cancer, ⁹ kidney cancer, ¹⁰ prostate cancer, ¹¹ leukemia, ¹² and thyroid cancer. ¹³

Treatment approaches for malignancies include surgery, chemotherapy, radiation therapy, immunotherapy, and monoclonal antibody therapy.^14–16 Chemotherapy is employed widely in cancer management. ¹⁷ However, its effectiveness is often limited by its toxicity to other normal, healthy cells and tissues. ¹⁷ The toxicity to the digestive tract is termed mucositis.

Mucositis

Chemotherapy drugs are used to limit the proliferation of tumor cells. However, these drugs coincidentally impair populations of normal cells such as enterocytes. ¹⁸ Mucositis is a common side-effect of cancer chemotherapy. It causes painful inflammation and ulceration of the mucous membranes lining the digestive tract. ¹⁹ The term alimentary mucositis describes the damage that occurs along the entire gastrointestinal tract. Mucositis can also affect the mouth, referred to as oral mucositis.^19,20 The current review focuses mainly on gastrointestinal mucositis.

A significant number of patients undergoing chemotherapy (40% to 100%) experience different stages of mucositis, depending on the dose and types of chemotherapeutic agent employed.^21,22

Intestinal damage caused by chemotherapy can manifest in several ways. For example, chemotherapy has been reported to trigger the death of normal rapidly dividing cells, especially those lining the gastrointestinal tract. ²³ Chemotherapy has also been demonstrated to change the gut microbiota communities in human individuals and intestinal mucin levels in rats.^23,24 This damage is responsible for breaches to the intestinal barrier, accompanied by increases in intestinal permeability in cancer patients undergoing chemotherapy. ²⁵ Subsequently, chemotherapy-induced intestinal impairment provides a conduit for entry of pathogenic bacteria into the systemic circulation, with pro-inflammatory cytokine release and associated inflammation. ²⁰ In addition, oxidative stress contributes to further intestinal injury. ²⁶ Considered together, damage induced by chemotherapy may lead to intestinal dysfunction, inflammation, and an impaired immune system.^22,23

Cancer patients undergoing chemotherapy usually experience symptoms that include abdominal distension, nausea, dysphagia, loss of taste, reduced oral intake, weight loss, and malnutrition.^22,27 Moreover, patients can also experience diarrhea caused by electrolyte imbalance, bleeding from intestinal ulcerations, and bacterial sepsis.^22,28 Indeed, death may even occur. ²²

Pathogenesis of Mucositis

To reduce the severity of mucositis, it is important to understand the pathophysiology of this condition. Sonis ²⁹ in 2004 summarized the 5 stages of chemotherapy-induced mucositis. Briefly, the first stage is an “initiation phase,” associated with an increased level of reactive oxygen species and DNA damage to intestinal cells and tissues. The second stage is the “upregulation and message generation phase” characterized by the production of cell transcription factors such as nuclear factor κ-light-chain-enhancer of activated B cells. Together with apoptosis, the mucosa becomes more friable and the patient begins to feel pain. The third stage is the “signaling and amplification phase.” During this stage, levels of tumor necrosis factor-α are increased, together with increased levels of nuclear factor κ-light-chain-enhancer of activated B cells, resulting in further cell apoptosis and tissue injury beneath the mucosal surface. This is followed by the “ulceration and inflammation phase,” which is associated with bacterial colonization and the production of pro-inflammatory cytokines such as interleukin-1 and tumor necrosis factor-α. Together this can result in bacteremia, sepsis, and breached mucosa integrity. The final stage is a healing phase, whereby the intestinal epithelium begins to recover and the normal homeostasis is restored in the gut microbiota. ²⁹

Treatment of Mucositis

Interest in studying oral mucositis has grown steadily^27,30; however, in contrast, research into treatments for gastrointestinal mucositis has received less attention. To date, treatment approaches for gastrointestinal mucositis have mainly focused on the identification of agents with the potential to protect the mucosa and promote the repair process, without compromising the cytotoxic effects of chemotherapy. Several growth factors such as milk growth factors and insulin-like growth factor-I^28,31,32 have exhibited the potential to promote differentiation and proliferation of epithelial cells affected by chemotherapy. Moreover, new nutraceuticals have recently been tested in experimental models for their effects against mucositis. These include probiotic preparations such as VSL#3, ³³ Streptococcus thermophilus, ³⁴ Lactobacillus spp., and Escherichia coli Nissle 1917 ³⁵ together with prebiotics such as fructo-oligosaccharide. ³⁶ Moreover, recently, probiotic-derived factors have begun to show promise at preventing mucositis in vivo and in vitro.^35,37 In addition, there are now early indications that antioxidant and anti-inflammatory constituents in plant extracts, such as Iberogast ³⁸ and grape seed extract, ³⁹ and animal-sourced oils, such as Lyprinol ⁴⁰ and Emu oil, ⁴¹ could have utility in preventing mucositis.

In this review, certain probiotic strains, probiotic-derived factors, and plant extracts will be explored for their potential to ameliorate mucositis. The different nature and origins of these agents would suggest that the severity of mucositis could be reduced through different pathways. Thus, the strategy of using strategically defined combinations of these nutraceuticals, such as combinations of probiotics/probiotic-derived factors and plant extracts, could be more effective than the individual agents in protection from mucositis.

Probiotics and Mucositis

Probiotics are living microorganisms that, when administered in a sufficient amount, confer health benefits to the host. ⁴² Lactobacillus and bifidobacterium genera have been found to be the most effective probiotics. Lactococcus, streptococcus, and enterococcus species, as well as some nonpathogenic strains of Escherichia coli and some bacilli and yeast strains, may also meet this definition. ⁴³

The beneficial properties of probiotics have been studied over decades. It is believed that probiotics have the potential to maintain a healthy gut microbiota, and most importantly reduce the severity of certain disorders, which include gastrointestinal diseases such as inflammatory bowel disease, diabetes, and atopic diseases.^44–46 Indeed, Escherichia coli Nissle 1917 has been prescribed widely for the treatment of inflammatory bowel disease in some countries, including Germany. ⁴⁷

The mechanisms of probiotic action have been summarized by Howarth in 2010. ⁴⁸ Briefly, certain probiotics have the ability to adhere to the intestinal–lumen interface, compete with pathogens for nutrients and binding sites, enhance mucosal barrier function, modulate cell kinetics by adjusting the proliferation to apoptosis ratio, and promote innate and adaptive immune responses. Based on these features, it is possible to speculate that probiotics, especially some newfound probiotic strains, could hold the potential to reduce gut damage from chemotherapy administration.

Prisciandaro et al reviewed evidence supporting the use of probiotics for the potential prevention and treatment of chemotherapy-induced mucositis. ⁴⁹ The current review extends this strategy to include recently identified probiotics and the added potential for these bacteria to be more effective when administered in combination with certain antioxidant plant extracts.

Competition With Pathogens for Binding Sites

It has been reported that mucin binding proteins can be identified from probiotics such as Bifidobacteria bifidium species and Lactococcus lactis ssp. lactis BGKP1, contributing to binding properties in vivo and in vitro.^50,51 In addition, a few Lactobacillus spp. and Bifidobacterium spp. were found to inhibit the growth of certain intestinal pathogens, including Escherichia coli and Salmonella spp., in laboratory experiments.^52,53 Probiotics may therefore compete with pathogens for binding sites and nutrients, potentially decreasing the likelihood of pathogen infection and secondary infections associated with chemotherapy.

Probiotic Versus Microbiota

Chemotherapy tends to alter the composition of the microbiota toward a more pathogenic community. For example, decreases of Clostridium spp., Lactobacillus spp., and Streptococcus spp. and an increase in Escherichia spp. were found in the jejunum of 5-fluorouracil-injected rats. ²⁴ Administration of probiotics (Bifidobacteria and Lactobacillus), for 7 days after elective laparoscopic radical surgery for colorectal cancer, significantly modified the fecal microbiota by increasing numbers of Bifidobacterium, Lactobacillus, and Enterococcus and decreasing Escherichia coli and Staphylococcus aureus. ⁵⁴ Thus, certain probiotics could potentially restore and normalize the chemotherapy-affected gut microbiota.

Maintenance of the Gut Barrier

Breach of the intestinal barrier has always been a serious issue for cancer patients undergoing chemotherapy. ²⁵ Disturbed mucin secretion, goblet cell production, and enterocyte tight junction proteins amplify the risk of bacterial infection and translocation into surrounding tissues.^24,29 Some probiotic strains such as Lactobacillus plantarum spp., and the probiotic mixture VSL#3, have demonstrated capacities to modulate tight junction proteins such as occludin, zonula, occludens-1, claudin-1, claudin-2, claudin-4, junction adhesion molecule-A, and F-actin, which are important in building the physical connections between epithelial cells for normal gut permeability.^55–57 VSL#3 increased expression of the epithelial tight junction protein, occludin, and downregulated expression of claudin-2, which attenuated increased gut permeability in mice with experimentally induced Crohn’s disease. ⁵⁵ Moreover, certain probiotics have the ability to maintain intestinal mucin levels, which may be influenced by chemotherapy.

The capacity for probiotic strains such as Lactobacillus spp. and Clostridium tyrobutyricum to adjust mucin secretion could therefore play a pivotal role in restoring microbiota composition and mucosal immunity in chemotherapy-treated patients.^{58 –60} Furthermore, certain probiotics have the potential to modulate cell kinetics by adjusting homeostasis between apoptosis and proliferation. For example, Lactobacillus rhamnosus regulated cellular proliferation and migration and mitogen-activated protein kinase pathways, responsible for cell proliferation, differentiation, and cytoprotection.^61,62 In contrast, some other probiotic mixtures such as kefir were found to reduce the incidence of apoptosis in heart tissues by controlling the activities of pro-apoptotic proteins such as Bax, bad, cytochrome c, and caspase-3 in ovalbumin-treated rats. ⁶³ Considered together, an imbalance in the ratio of enterocyte apoptosis to proliferation, induced by chemotherapy, could potentially be restored by the intake of certain probiotics.

Probiotics and Immunity

Administration of certain probiotics can enhance innate immunity by increasing numbers and activities of immune cells and mediating inflammation. ⁴⁵ In addition, probiotics may improve adaptive immune function by modulation of antigen-specific antibodies. ⁵⁴ For example, numbers of circulating natural killer cells and immature T cell subsets increased in elderly individuals ingesting Lactobacillus delbrueckii subsp. bulgaricus 8481 for 6 months. ⁴⁵ Moreover, the immune risk phenotype, characterized by an inverted CD4/CD8 ratio, an increase of CD8+CD28^null T cells, and cytomegalovirus infection was also amplified in the same study.^45,64 Furthermore, administration of Clostridium tyrobutyricum depressed expression of pro-inflammatory cytokines such as tumor necrosis factor-α and interleukin-18 in the descending colon of dextran sodium sulfate–treated mice. ⁵⁹ Similar results have also been described in cytokine-mediated gastrointestinal diseases, ⁶⁵ alcohol-induced inflammation,^66,67 and even chemotherapy-induced mucositis. ³⁴ Besides, administration of probiotics (Jinshuangqi tablets) for 7 days significantly improved levels of IgA, IgG, IgM, interelukin-2, and CD4+ in the serum of colorectal cancer patients, demonstrating the capacity for probiotics to modulate adaptive immunity. ⁵⁴

Probiotic-Derived Factors

Probiotic-derived factors are proteins or other molecules released from living probiotics, with the potential to confer health benefits to the host. Investigation of these factors could be used to achieve therapeutic benefits while avoiding risks related to the administration of live bacteria. ⁴⁸ Identification of specific probiotic-derived factors also provides an opportunity to better understand mechanisms of probiotic action.

Competition With Pathogens

Bacteriocins, such as reuterin and other proteinaceous molecules, have been shown to inhibit the adhesion and viability of known enteric pathogens.^48,68 Lactobacillus acidophilus ATCC 4356 releases a proteinaceous molecule that inhibited the activity of 8 of the human Campylobacter jejuni strains affecting Caco-2 cells. ⁶⁹ Moreover, other strains of Lactobacillus spp. produced non–lactic acid molecules in their cell-free cultured supernatants, which were responsible for killing activity against pathogenic Salmonella enterica serovar Typhimurium SL1344.^70,71 Together, these probiotic-derived factors both restricted the activity and killed certain pathogens, thereby decreasing the likelihood of infection, potentially reducing bacterial invasion caused by chemotherapy.

Maintenance of the Gut Barrier

Modulation of the mucus layer, tight junctions, and gut permeability by probiotic-derived factors play an important role in the maintenance of gut integrity. These factors could therefore potentially ameliorate the gut damage induced by chemotherapy. ⁴⁸ Administration of Lactobacillus rhamnosus supernatants to alcohol-treated mice altered intestinal tight junction proteins by increasing ileum mRNA levels of claudin-1. ⁶⁷ Moreover, mRNA levels of intestinal trefoil factor, P-glycoprotein, and cathelin-related antimicrobial peptide were also restored by Lactobacillus rhamnosus supernatant pretreatment. ⁶⁷ In addition, Lactobacillus rhamnosus supernatants prevented barrier dysfunction of Caco-2 cell monolayers, induced by alcohol. ⁷² Certain proteinaceous factors extracted from Clostridium butyridium CGMCC0313-1 have been reported to augment the expression of A20 in HT-29 cells, which is important in the maintenance of barrier function. ⁷³ Together, these in vitro results demonstrate the potential for probiotic-derived factors to maintain the intestinal barrier following chemotherapy-induced gut damage in vivo and in vitro.

Cell Kinetics

Two novel proteins, p75 (75 kDa) and p40 (40 kDa), released from Lactobacillus rhamnosus, have been reported to promote the propagation of epithelial cells and prevent apoptosis of epithelial cells induced by tumor necrosis factor-α. ⁶⁵ Moreover, a recent study by Lebeer et al found that p75, renamed as Msp1, secreted by Lactobacillus rhamnosus, is an O-glycosylated protein. ⁷⁴ Glycosylation of Msp1 plays an important role in communication between the microbe and the host. ⁷⁴ In addition, Prisciandaro et al recently reported that supernatants from Lactobacillus rhamnosus and Escherichia coli Nissle 1917 significantly decreased caspase 3/7 activity after a challenge by the chemotherapy antimetabolite, 5-fluorouracil, in the IEC-6 cell line, showing their potential ability to prevent or inhibit enterocyte apoptosis induced by chemotherapy. ³⁵

Immunity

Probiotic-derived factors have been shown to influence pathogen-induced or oxidative stimuli–induced inflammation. Several metabolites (<3000 Da), released from Bifidobacterium breve and Streptococcus thermophilus, have demonstrated the ability to inhibit tumor necrosis factor-α secretion from lipopolysaccharide-affected peripheral blood mononuclear cells or the THP-1 cell line. ⁷⁵ In addition, these factors (metabolites) were reported to suppress lipopolysaccharide-FITC (a fluorescent marker by flow cytometry) binding to THP-1 cells and also to inhibit nuclear factor-κB activation. Moreover, Lactobacillus reuteri–formed biofilms have been shown to decrease tumor necrosis factor production in lipopolysaccharide-activated monocytoid cells. ⁷⁶ Caco-2 cells pretreated with spent culture supernatants of Lactobacillus plantarum 2142 for 1 hour decreased interleukin-8 synthesis and, in addition, induced expression of Heat-shock protein (Hsp) 70. ⁷⁷ More recently, probiotic Lactobacillus paracasei CNCM I-4034 cell-free supernatants reduced pro-inflammatory tumor necrosis factor-α and chemokine MCP-1 in human dendritic cells challenged with enteropathogenic Salmonella. ⁷⁸ Interestingly, these supernatants produced similar beneficial effects to their living “parent” probiotic, Lactobacillus paracasei CNCM I-4034.

Probiotic-derived factors from Bifidobacterium bifidum LMG13195 could be a potential immunoregulator in vitro. These soluble factors, after being previously cocultured with HT29 cells, enhanced numbers of CD4⁺CD25^high cells expressing chemokine receptor Treg markers in human peripheral blood mononuclear cells. ⁷⁹ Probiotic-derived factors could therefore exert similar effects to their parent probiotic counterparts by modulating immune cells to reduce inflammation and restore immunity affected by chemotherapy.

Prebiotics

Prebiotics are indigestible compounds that “selectively stimulate the growth and/or activity of microbial species in the gut microbiota and confer health benefits to the host.” ⁸⁰ Prebiotics must be neither hydrolyzed nor absorbed in the proximal gastrointestinal tract, must be selectively fermented by one or a limited number of beneficial bacteria in the intestine, and be able to alter the colonic microbiota toward a healthier composition.^80,81 Prebiotic compounds include inulin, ⁸² a plant derived fructan; lactulose ⁸³ ; galacto-oligosaccharide ⁸⁴ ; and fructo-oligosaccharide. ⁸⁵ Other compounds studied for their prebiotic potential include arabinoxylan-oligosaccharides, ⁸⁶ chito-oligosaccharides, ⁸⁷ epilactose, ⁸⁸ germinated barley foodstuff, ⁸⁹ mannan-oligosaccharides, ⁹⁰ sialyl-oligosaccharides, ⁹¹ xylo-oligosaccharides, ⁹² and β-1,4-mannobiose. ⁹³

The potential role of prebiotics in mucositis treatment has recently (2012) been reviewed comprehensively by Wang et al. ⁹⁴ Briefly, prebiotics have the ability to modulate probiotic bacteria by selective colonization of the intestinal microbiota, thereby decreasing chemotherapy-induced intestinal dysbiosis. Prebiotics could therefore maintain homeostasis of the gut microbiota following induction of mucositis. Moreover, certain prebiotics play an important role in the digestion and absorption of nutrients in the gut ⁹⁵ and modulate intestinal barrier function through effects on mucin expression ⁹⁶ and modification of mucosal immune responses. ⁹⁷ Prebiotics could therefore become new nutraceuticals in terms of the amelioration of mucositis. Howarth recently proposed the importance of inflammasomes in mediating inflammation by prebiotics. However, further studies are required to determine the underlying mechanism of prebiotic action in the prevention and/or treatment of mucositis. ⁹⁸

Plant Extracts and Their Active Constituents

Many natural plant extracts have demonstrated medicinal uses at inhibiting the viability of pathogens. ⁹⁹ These include Cat’s claw (Uncaria tomentosa), Maca (Lepidium meyenii), and Dragon’s blood (Croton lechleri). However, in this review, we focus on the potential of readily accessible plant extracts, such as grape seed extract and Iberogast (and their commonly shared active constituents), to reduce the severity of gastrointestinal mucositis.

Phenolic compounds or polyphenols are widely distributed in the plant regime. ¹⁰⁰ They are characterized by the presence of multiple phenol structures. Flavonoids are one type of phenolic compounds, which can be classified into flavonols, flavanols, and many others. Flavonol comprises a ketone group (RC(=O)R′); flavanol or flavan-3-ols (catechins) belong to the flavonoid classification and are structurally similar to flavonol, but they are in the non-ketone group (Table 1). ¹⁰¹

Table 1.

Classification and Chemical Structure of Phenolic Compounds.

Classification	Chemical Structure
Plant	Example
Phenolic compounds	(Simple phenols)
Flavonoids and many others	(Flavonoids)
Flavonols (ketone group)	(Flavonols; ketone group)
Flavanols (non-ketone group)	(Flavanols; non-ketone group)
Proanthocyanidins (high-molecular-weight polymers)	(Proanthocyannidins)
Free radical scavenging
Antioxidant
Antibacterial and antiviral
Anti-inflammatory
Anti-carcinogenic
Anti-allergenic
Vasodilatory

Adapted from Fine AM. Oligomeric proanthocyanidin complexes: history, structure, and phytopharmaceutical applications. Altern Med Rev. 2000;5:144-151.

In chemical structure, single-molecule flavan-3-ol adds hydroxyls (–OH) becoming catechin or its isomer, epicatechin. Procyanidins, members of the proanthocyanidins, are concentrated polymers formed from the monomers catechin and epicatechin. ¹⁰² Therefore, catechin trimers become oligomeric proanthocyanidin complexes and higher order polymers become anthocyanidins (Table 1). ¹⁰²

The most significant effect of procyanidins/proanthocyanidins is that they possess reactive oxygen species free radical scavenging ability. This results from antioxidant activities in many plant-sourced extracts including grape seed extract, Cat’s Claw, Maca, and Dragon’s Blood.^103–106 The antioxidant ability of plant extracts against reactive oxygen species is elucidated in the following sections.

Effects of Active Constituents of Plant Extracts on Reactive Oxygen Species

Reactive oxygen species are chemically reactive molecules (free radicals) containing oxygen such as oxygen ions and peroxides. Oxidative stress plays an important role in the pathogenesis of disorders, which include rheumatoid arthritis, asthma, psoriasis, and contact dermatitis. ¹⁰⁵ Chemotherapy generates reactive oxygen species, which can initiate a series of biological actions. ²⁹ For example, reactive oxygen species induce neutrophil infiltration and pro-inflammatory cytokine production, such as tumor necrosis factor-α. Increased tumor necrosis factor-α in the submucosa activates and amplifies transcription factors such as nuclear factor-κB. This is thought to induce cell apoptosis, leading to breakdown of the intestinal epithelial monolayer, intestinal barrier damage, pathogen invasion, and clinical manifestations of mucositis.^29,107,108 Furthermore, reactive oxygen species potentially reacts with macromolecules of the gut mucosa, such as lipids, proteins, and nucleic acid, and causes lipid peroxidation, which has a role in destruction of the intestinal epithelium.^39,109 Therefore, overproduction of reactive oxygen species plays an important role in the pathogenesis of chemotherapy-induced gut damage.

Many studies have reported the antioxidative properties of phenolic compounds. Phenolic substances such as procyanidins interact with the polar head group of phospholipids in cell membranes to exert their antioxidant effects. ¹¹⁰ Verstraeten et al ¹¹¹ showed that dimer and trimer procyanidins protected the bilayer from oxidant-induced stress by interacting with membrane phospholipids. Facino et al ¹¹² reported that a procyanidin-enriched diet fed to rats was associated with increased antioxidant activity in the plasma and reduced cardiac damage. Similar results were obtained by Busserolles et al, ¹¹³ who fed rats procyanidin-rich extracts for 8 weeks. Plasma antioxidant capacity was higher in rats than those fed the control diet, indicating the antioxidant effect of plasma procyanidins in vivo. ¹¹³

Malondialdehyde level is a presumptive marker for lipid peroxidation that results from lipid-free radical interaction. ¹⁰⁶ Kalin et al reported the ability of proanthocyanidins to decrease malondialdehyde levels. ¹⁰⁶ This has also been demonstrated by Gulgun et al, ¹⁰⁹ who showed that proanthocyanidins decreased mucosal damage and oxidant stress from the chemotherapy drug methotrexate through decreasing lipid peroxidation. Additionally, proanthocyanidins significantly inhibited toxicity from cancer chemotherapeutic drugs (Idarubicin and 4-hydroxyperoxycyclophosphamide) through increased expression of the anti-apoptotic protein, Bcl-2. ¹¹⁴ Furthermore, Kalin et al ¹⁰⁶ concluded that proanthocyanidins decreased the induction of adhesion molecules such as ICAM-1, VCAM-1, and E-selectin, which are responsible for damage to the vascular endothelium.

Procyanidins/proanthocyanidins also exert anti-inflammatory and anti-ulcerogenic effects.^39,109 Bak et al ¹¹⁵ showed that procyanidins from wild grape seeds exhibit a chemopreventive character, due to an increase in nuclear factor E2–related factor expression. Nuclear factor E2–related factor is related to the antioxidant response element, which mediates expression of phase II detoxifying/antioxidant enzymes, such as NAD(P)H: quinone oxidoreductase-1 (NQ01) and heme oxygenase-1 (HO-1). These enzymes play a vital role in cell protection and cancer prevention. Furthermore, it is believed that proanthocyanidins exert an antithrombotic effect and can improve mucosal blood flow, which may contribute to antiulcer effects. ¹⁰⁹ In an in vivo study, proanthocyanidins exerted antiulcer effects on the gastric mucosa, through significantly inhibiting myeloperoxidase activities (as an inflammatory indicator) and stimulating superoxide dismutase activities (as an antioxidative indicator) in rats. ¹¹⁶ It was reported that proanthocyanidin decreased leukocyte infiltration and tumor necrosis factor-α and interleukin-1β (pro-inflammatory factors), CINC-1 (an important mediator that promotes migration of neutrophils), and nitrite levels in the pleural exudation, which occurred after a carrageen-induced acute inflammatory reaction in rats. ¹¹⁷ These results further supported the anti-inflammatory effects of proanthocyandins.

In summary, phenolic compounds possess antioxidant, anti-inflammatory, and anti-ulcerogenic properties. These properties of procyanidins/proanthocyanidins are partly contributed by their free-radical scavenging characteristics, modulating apoptotic regulatory genes such as bcl-2 and p53, thereby influencing the production of pro-inflammatory factors such as tumor necrosis factor-α and interleukin-1β.^109,114 Plant extracts, which contain these active constituents, could therefore be of benefit in treating or preventing mucositis.

Plant Extracts and Intestinal Mucositis

Very few studies have tested the effects of plant extracts on mucositis. Cheah et al ³⁹ studied the potential for grape seed extract to protect the rat intestine from 5-fluorouracil-induced intestinal damage in vitro and in vivo. These investigators reported that grape seed extract had the ability to restore IEC-6 cell viability following damage by 5-fluorouracil, indicating that grape seed extract was partially effective against this chemotherapy drug in vitro. Additionally, grape seed extract showed the ability to reduce 5-fluorouracil-induced intestinal myeloperoxidase activity, an indicator of inflammation. Furthermore, grape seed extract was found to remain active in the large bowel. ³⁹ Moreover, Gulgun et al showed promising results for proanthocyanidins in improving methotrexate-induced intestinal damage in rats. ¹⁰⁹ Proanthocyanidins reduced methotrexate-induced inflammation and associated ulceration and normalized the activities of superoxide dismutase and glutathione peroxidase. This indicated that proanthocyanidins amplified the defense system against oxidative stress and also decreased lipid peroxidation following methotrexate-induced damage. ¹⁰⁹

Iberogast, also known as STW5, is a mixture of 9 different herbal extracts containing bitter candytuft (Iberis amara), angelica root (Angelicae radix), milk thistle fruit (Cardui mariae fructus), celandine herb (Chelidonii herba), caraway fruit (Carvi fructus), liquorice root (Liquiritiae radix), peppermint herb (Menthae piperitae folium), balm leaf (Melissae folium), and chamomile flower (Matricariae flos). ¹¹⁸ Iberogast modulates gastrointestinal motility and restricts gastric acid production. It also possesses anti-inflammatory, antioxidative, and free radical–inhibiting properties. The active compounds in Iberogast, especially flavonoids, are believed to contribute to its pharmacological properties in the therapy of several gastrointestinal conditions such as functional dyspepsia and irritable bowel syndrome. ¹¹⁸ Indeed, recently, Iberogast has been tested for its therapeutic efficacies on 5-fluorouracil-induced mucositis in rats. ³⁸ Iberogast improved the histopathological features of mucosa, which were damaged by 5-fluorouracil injection. Villus height and crypt depth were increased significantly in Iberogast-fed rats treated with 5-fluorouracil. ³⁸ However, other indicators of mucositis were unaffected by Iberogast in 5-fluorouracil-treated rats. ³⁸ These results indicate that Iberogast may possess the potential to reduce the severity of chemotherapy-induced mucositis; however, further studies on dose and frequency of administration are warranted.

Other plant extracts such as glycolipid extracts from spinach, which comprise high levels of glycoglycerolipids, have revealed antioxidative and anti-inflammatory effects in the 5-fluorouracil setting in vivo and in vitro. Mucosal injury, such as villous atrophy and misaligned crypts in the jejunum of rats affected by 5-fluorouracil, were ameliorated significantly in rats fed glycolipid extracts. ¹¹⁹ Moreover, glycolipid extracts reduced mRNA expression of inflammatory cytokines such as interleukin-1α and tumor necrosis factor-α caused by 5-fluorouracil injection in the same group of rats. In addition, the active constituents of spinach extract, such as monogalactosyl-diacylglycerol and diglactosyl-diacylglycerol, inhibited the production of reactive oxygen species in phorbol ester challenged Caco-2 cells. ¹¹⁹ Furthermore, following 5-fluorouracil administration, hamsters fed Eriobotrya japonica seed extract showed no epithelial tissue defects or bacterial infections at the local site of mucositis in the oral cavity. ¹²⁰ Plasma lipid peroxide levels were also decreased significantly in the same group. ¹²⁰ Moreover, in a clinical trial, the severity of radi-ation-induced mucositis was attenuated in patients, as measured by significantly decreased serum interleukin-6 levels, following gargling and ingestion of indigowood root (Isatis indigotica Fort.). ¹²¹

In summary, certain plant extracts such as grape seed extract and Iberogast have revealed anti-inflammatory and antioxidant effects on chemotherapy-induced mucositis. These plant extracts could be a potential new prophylactic treatment strategy for intestinal mucositis, ³⁹ although further research is required. Further studies should focus on identifying the bioactive constituents involved together with the identification of further, as yet untested plant extracts.

Combinations of Probiotics/Probiotic-Derived Factors and Plant Extracts

Synbiotics are defined as strategically-identified symbiotic combinations of probiotics and prebiotics. Synbiotics may be exhibit greater potency compared to the sum of each individual agent. Shimizu et al recently elucidated the importance of synbiotics in the maintenance of gut health and treatment of critical illnesses such as septic complications. ¹²² Fermented milk supplemented with 2 probiotic strains, Bifidobacterium lactis Bi-07 and Lactobacillus acidophilus NCFM, and a prebiotic, isomaltooligosaccharide, administered to healthy adults and mice improved intestinal health as indicated by increases in fecal bifidobacteria and lactobacilli and decreases in enterobacilli. ¹²³ Humoral and cell-mediated immunity have also been improved by the same tested synbiotics in mice. ¹²³ The combination of Lactobacillus acidophilus and ginger extract improved ulcer index, mucus secretion, oxidative stress, and histopathological parameters, when compared with the individual agents, in a gastric ulcer setting in rats. ¹²⁴

Very few studies have described the effects of synbiotics on reducing the severity of mucositis. Smith et al tested the combination of Lactobacillus fermentum BR11 and the prebiotic, fructo-oligosaccharide, in 5-fluorouracil-injected rats. ³⁶ These investigators found that Lactobacillus fermentum BR11 partially reduced the inflammation caused by 5-fluorouracil in the small intestine in rats. However, the combination did not provide additional therapeutic benefit for mucositis. Further studies are required to test the effects of combinations of different probiotics and prebiotics on reducing the severity of mucositis.

The prebiotic potential of virtually all plant extracts identified to date remains essentially unexplored. New nutraceuticals, including probiotics, probiotic factors, prebiotics, and plant extracts, hold the potential to decrease gut damage through different mechanisms, thereby increasing the likelihood of achieving clinically measurable net benefit. For example, certain probiotics could combat mucositis through effects on bacterial composition in the bowel whereas certain plant extracts would likely be more potent at counteracting oxidative stress created by chemotherapy. Therefore, indicated combinations of probiotics (or probiotic factors) and plant extracts could potentially reduce the severity of mucositis to a greater extent than each of the agents used independently, thereby potentially achieving measurable clinical improvement.

As the majority of prebiotics are derived from plants, future studies could focus on determining the prebiotic properties of a wider range of plant extracts. Future challenges will include mechanisms to identify specific combinations of probiotics and prebiotics that exert synergistic benefits in mucositis treatment. Determining dosing regimens, frequency of administration, the specific bioactive factors involved, and their associated pharmacodynamics will represent further challenges before these promising new nutraceutical formulations can be recommended for mucositis treatment.