Celiac disease onset and inflammatory milieu

Abstract

No one is born with celiac disease, yet some individuals are predisposed to its development, based on two main factors. The transition from gluten tolerance to onset occurs in a subset of individuals who both carry the human leukocyte antigen (HLA)-DQ2 or HLA-DQ8 alleles in their genetic makeup (30%-40%) and consume gluten. What transpires to foment evolution in approximately 3% of these individuals, and why it happens, is of interest to benefit public health. At issue is the tipping point, where deamidated gluten peptides acquire increased affinity and binding to specific HLA-DQ2/DQ8 molecules on the surface of antigen-presenting cells (APCs), thereby alerting CD4+ T-cells to inflict damage on small intestinal mucosa. Is the transition to celiac disease caused by an alteration in the special manner that gluten peptides, deamidated by transglutaminase, are bound, and how strongly they are bound on the APCs, to render them recognizable as antigen by CD4+ T-cells? Or does the transformation occur when the CD4+ T-cells are rendered immunogenic to the tightly bound and deamidated gluten peptides on the APCs? Are supplementary conditions crucial? Might the modification be reversible under certain circumstances? It is suggested that onset may be triggered by the cumulative burden of extrinsic inflammatory factors vs anti-inflammatory factors acting on a genetically susceptible host consuming gluten, rather than arising from a sole insult. A paradigm to possibly reduce the odds of developing the disease, and to reduce the time needed for recovery after diagnosis, is presented. Implications for incorporating modern gastrointestinal endoscopic techniques are discussed.

Key Words: Celiac disease; Gastrointestinal endoscopy; Gluten; Immunogenic; Inflammation

Core Tip: In this article, the relationship between celiac disease and the inflammatory milieu is discussed and contrasted. The effects of inflammation on disease onset and recovery on a gluten-free diet are described in detail. After thoroughly covering the published literature, we suggest a hypothesis-generating paradigm that may possibly reduce the odds of a susceptible person developing the disease, and reduce the time needed for recovery after diagnosis. The paradigm reflects biologic plausibility and emerging mechanistic evidence, rather than clinical endorsement. The growing relevance of these concepts to modern gastrointestinal endoscopic technique is then highlighted.

INTRODUCTION

Celiac disease is a chronic immune-mediated enteropathy, precipitated by dietary gluten in genetically predisposed individuals carrying the human leukocyte antigen (HLA)-DQ2/DQ8 haplotypes[1,2]. Although exposure to gliadin peptides and possession of these alleles are important for disease development, they are insufficient. Approximately 30%-40% of the global population expresses HLA-DQ2/DQ8[2-5], yet only a few percent of persons with this genetic predisposition develop the disease when consuming gluten[6-8]. The recent pattern of emergence, such as the Swedish surge from 1984 to 1996[9], and the gradual increase in rate during the past two decades[10-12], suggests that celiac disease may not result from isolated factors; rather it appears to involve an accumulation of environmental and other exogenous stressors, or a surge of such stressors, which collectively exceed the immune system’s capacity for tolerance[13]. Harmful environmental exposures, particularly those influencing gastrointestinal inflammation and immune balance, are increasingly recognized as being influential for tipping susceptible individuals from tolerance to disease, and in disease progression[14,15].

Historically, research on celiac disease onset has focused on identifying individual triggers or modulators, such as early-life infections, the timing and amount of gluten introduction in infancy, viral or antibiotic exposure, or alterations in the intestinal microbiota[16,17]. Alternatively, the disorder might be considered a threshold-based condition, in which the total inflammatory burden, rather than a sole insult of a particular type, regulates whether and when a genetically susceptible individual develops overt disease[18,19]. The concept can be termed inflammatory load: The cumulative effect of diverse exogenous and, in response, endogenous factors, provoking gastrointestinal and systemic inflammation, disrupting intestinal mucosal integrity, and/or destabilizing immune regulation[20,21]. Each of the insults may independently provoke mild immune activation, but collectively, and when coinciding with gluten exposure, finely tuned mechanisms that maintain oral tolerance in the gut-associated lymphoid tissue are potentially overwhelmed[22]. Herein, we review the mitigating circumstances, then introduce a novel synthesis, framework, and testable conceptual model, suggesting that prevention should include a consideration of the cumulative inflammatory burden.

This manuscript is intentionally structured as a hybrid review-framework paper, in which a comprehensive synthesis of the existing literature is utilized to motivate and contextualize a hypothesis-generating conceptual model. Although mechanistic in scope, the concepts discussed have increasing relevance to gastrointestinal endoscopy, which now enables early detection, risk stratification, and longitudinal monitoring of celiac disease beyond the presence of established villous atrophy.

PRO- VS ANTI-INFLAMMATORY CONSIDERATIONS

Various environmental and other exogenous factors affect celiac disease onset and recovery[23,24]. In the intestine, they can make individuals more, or less, susceptible to gluten’s effects[25]. They can alter the composition and function of gastrointestinal immune cells, and when detrimental, they promote a pro-inflammatory environment that predisposes individuals to celiac disease[26]. Unhealthy diets disrupt the gut microbiome balance and promote inflammation[27,28]. Systemic inflammation can intensify gut inflammation and autoimmune reactions, leading to more severe celiac disease symptoms and complications[29,30]. The degree of inflammation impacts immunogenicity[31]. The immune system’s attack on the small intestinal lining may become severe, eliciting damage to the intestinal villi, and leading to more pronounced symptoms after disease onset[32]. The overall balance between pro-inflammatory and anti-inflammatory signals governs the impact on immunogenicity[33] and its effect on celiac disease onset and recovery.

Examples of inflammatory factors include: Polluted air[34], water[35], and ground[36], all of which may be due to the presence of toxic chemicals and elements such as cadmium[37], lead[38], and arsenic[39], microplastics[40], glyphosate[41] and other pesticides and herbicides[42], a western diet[43] including consumption of refined carbohydrates and ultra-processed foods and oils[44] such as highly salty[45] or sugary foods[46], foods with chemical additives for freshness, taste, and color[47], also alcohol[48], cigarette smoking[49], marijuana[50], other restricted or illegal drugs[51], stress and anxiety[52] caused by such issues as excessively hot temperatures[53] and congested traffic[54], surgery[55], other trauma[56], and bacterial, viral, and fungal infections[57], other pathogens[58], hypoxia, such as from high altitudes[59], poor dental hygiene[60], poor posture[61], obesity[62], sleep deprivation[63], a sedentary lifestyle[64], genetic factors[65], and other autoimmune disease[35]. Many of these factors may also contribute to gut dysbiosis, defined as an imbalance in gut bacteria and changes in inflammatory gene expression[66]. Major inflammatory factors, mechanisms, and biomarkers in celiac disease are listed in Table 1. The list is not exhaustive but shows a wide variety of insults that can influence inflammation as it pertains to celiac disease. Examples of beneficial factors which are anti-inflammatory include diets with plenty of soluble fiber as found in fruits and vegetables[67], which ferment in the gastrointestinal system to enable bacteria residing there to produce short-chain fatty acids (SCFAs)[68], whole grains[69], lean meat such as poultry[70], also fish, especially those rich in omega-3 fatty acids such as cod and salmon[71], peanuts and natural unsweetened peanut butter[72], other nuts and seeds[73], unsweetened dark chocolate high in cacao content[74], black and green tea[75], spices and condiments such as black pepper[76], cinnamon[77], honey[78], mustard[79], supplements such as resveratrol[80], antioxidant vitamins such as C and E[81], modulation by the intestinal epithelium[82], good dental hygiene[69], good posture[70], a healthy body mass index[83], sufficient sleep[63], moderate exercise[84], and fresh air and country living[85].

Table 1 Major inflammatory factors, mechanism, and biomarkers in celiac disease.

Environmental factor	Mechanism of inflammatory effect	Key biomarkers
Viral infection (e.g., reovirus, enterovirus)	TLR activation, Th1 skewing, cytokine upregulation	IL-2, IL-6, IFN-γ, CD4+ T-cell activation
Glyphosate exposure	Disruption of tight junctions, oxidative stress	Zonulin, CRP, fecal calprotectin
High-fat diet	Microbial dysbiosis, LPS translocation	LPS, TNF-α, SCFA levels, gut permeability markers
Antibiotic use	Loss of microbial diversity, immune dysregulation	Alpha diversity, Treg/Th17 ratio, fecal SCFA
Chronic psychological stress	Cortisol elevation, HPA axis modulation, IL-6 release	Cortisol, IL-6, neutrophil/Lymphocyte ratio
Air pollution (PM2.5, NOx)	Oxidative stress, epithelial inflammation	CRP, IL-8, 8-isoprostane, oxidized glutathione
Lead/arsenic in water	Heavy metal-induced ROS production, gut barrier damage	8-OHdG, malondialdehyde, IL-1β, CRP
Microplastics	Foreign particle immune activation, mucosal disruption	CRP, IL-1β, gut epithelial stress markers
C-section birth (vs vaginal)	Lack of exposure to maternal microbiome	Delayed colonization profiles, IL-10, TSLP
Ultra-processed foods	Emulsifier-induced permeability, dysbiosis	CRP, TNF-α, calprotectin, dysbiosis indices
Sedentary lifestyle	Reduced gut motility and microbiome diversity	IL-6, reduced SCFA, microbial diversity
Season of birth (winter gluten intro)	Increased infection risk during gluten intro window	Season-correlated cytokine trends, seroconversion timing
Urban vs rural environment	Lower biodiversity, increased allergen burden	IL-6, TLR ligands, LPS burden, biodiversity scores
Early gluten introduction	Possible immunological mistiming	Seroconversion timing, IL-2, immune T cell skew
Lack of breastfeeding	Delayed gut colonization, Treg suppression	Secretory IgA, transforming growth factor-β, Treg deficiency
Sleep deprivation	IL-6 elevation, impaired mucosal recovery	IL-6, TNF-α, cortisol, sleep markers

TLR: Toll-like receptor; Th1: Type 1 helper T cell; IL: Interleukin; CRP: C-reactive protein; LPS: Lipopolysaccharide; TNF-α: Tumor necrosis factor alpha; SCFA: Short-chain fatty acid; Treg: Regulatory T cells; HPA: Hypothalamic-pituitary-adrenal axis; ROS: Reactive oxygen species; 8-OHdG: 8-hydroxy-2’-deoxyguanosine; TSLP: Thymic stromal lymphopoietin; PM2.5: Fine particulate matter; Nox: Nitrogen oxides; IFN-γ: Interferon-γ.

A schematic of how detrimental exogenous agents, which are pro-inflammatory markers, exert their effect on the small intestine is shown in Figure 1. They may also contribute to gut dysbiosis so that there is a diminishment in beneficial bacteria and an increase in harmful ones[66,86]. Moreover, detrimental environmental factors often induce epigenetic changes, which alter gene expression without modifying the underlying DNA sequence, so that an epigenetic memory is formed in stem cells, leading to persistent, maladaptive responses that perpetuate chronic inflammation and impair barrier repair[87]. The harmful exogenous factors increase intestinal permeability and contribute to a leaky gut, i.e., an increased intestinal permeability with dysregulation of inflammatory gene expression[88,89]. Hence, gluten and other insults can pass through the barrier as the mucus layer and tight junctions between cells degenerate. Contrarily, an anti-inflammatory lifestyle, consisting of beneficial environmental and other exogenous factors, and comprising a nutrient-dense, high-fiber diet, good hygiene, proper posture, and healthy habits, reduces the expression of pro-inflammatory genes and actively improves the balance of gut bacteria[70]. By fostering a diverse microbiome, these habits strengthen the intestinal barrier, lower systemic inflammation, and diminish the risk of celiac disease onset.

Figure 1 Examples of exogenous factors contributing to inflammation leading to celiac disease. These factors act to increase inflammation, dysbiosis, and epigenetic changes. All contribute to increased intestinal permeability.

MOLECULAR BIOMARKERS OF INFLAMMATION

When acute inflammation occurs, it elicits an inflammatory response to recruit and activate antigen-presenting cells (APCs) such as dendritic cells, which then present the antigens to T-cells, triggering a strong reaction that includes the release of inflammatory cytokines[90,91]. Genes encoding pro-inflammatory cytokines, including interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), and IL-6, are key players in inflammation, while genes encoding anti-inflammatory cytokines like transforming growth factor-β and IL-10, help to regulate and resolve inflammation[92]. Interferon (IFN)-I signaling leads to the expression of inflammatory genes and the release of other inflammatory mediators, thereby contributing to the inflammatory response[93]. The release of IFN-I is triggered by stimuli including viral infections, immune complexes, systemic inflammation, and the inflammatory cytokines[94]. The nuclear factor-κB pathway is a major regulator of inflammatory gene expression, and is activated by various inflammatory insults[95]. The expression of certain genes, such as IL-15, enhances inflammation[96] and it is a hallmark of celiac disease once onset occurs[97].

Several inflammatory biomarkers can be utilized to measure inflammation in individuals susceptible to or with celiac disease, and to determine the immune status. They include metrics which measure immune activation, including tTG-IgA[98], endomysial antibodies[99], circulating gliadin-specific CD4+ T-cells[100], and other markers comprising methods for sampling general inflammation, including: C-reactive protein, IL-2, IL-6, IL-17, TNF-α, complete blood count or complete blood count (neutrophil: Lymphocyte ratio)[101-103], and fecal calprotectin[104]. These biomarkers are metrics of microbial diversity, SCFA concentration, and pathobiont prevalence[105-107]. Additional measures include flow cytometry for T-cell subsets analysis, cytokine release assays, and metabolomics panels[108,109]. Dietary assessment indices, including the Dietary Inflammatory Index or the Empirical Dietary Inflammatory Pattern, provide scores to determine the potential of a diet to promote or reduce inflammation[110,111]. They furnish numerical scores based on the consumption of various foods and nutrients, with higher scores generally indicating a more pro-inflammatory diet and lower scores indicating a more anti-inflammatory diet. Single-cell RNA sequencing dissects the cellular heterogeneity of celiac disease by providing high-resolution, cell-type-specific transcriptomic profiles from tissue samples[31,112,113]. It maps the inflammatory landscape by identifying rare immune cell subsets, tracking evolutionary trajectories of infiltrating cells, and revealing unique molecular mechanisms, such as IFN-induced gene expression and activated T-cell cytotoxic programs, driving tissue damage in response to gluten.

MOLECULAR MIMICRY

When inflammation occurs, it contributes to molecular mimicry caused by viral infection[114,115]. When a viral protein or peptide shares structural similarities with a host protein, such as gluten, gluten peptides, or transglutaminase[116], it can trigger an immune response, with T-cell activation followed by mistaken attack on host tissue. For example, antibodies against rotavirus proteins may also recognize gluten, eliminating immune tolerance[117]. Viral infections precede the release of inflammatory cytokines as well as the activation of immune cells[118]. The inflammation can disrupt normal intestinal function, and create a more permissive environment for molecular mimicry to drive autoimmune reactions leading to celiac disease. Drugs including acetaminophen may reduce the body’s immune response to vaccines, and suppress response during infection, potentially leading to a slower recovery or increased viral shedding[119,120], and thereby enhancing the possibility of molecular mimicry. They can also reduce the activity of immune cells like T-cells and B-cells[121], which are crucial for normal antibody production and overall immune function.

THE GASTROINTESTINAL BARRIER

The gastrointestinal lining, composed of a single layer of epithelial cells, plays a crucial role in maintaining gastrointestinal barrier integrity[122-124], thereby preventing celiac disease onset, as illustrated in Figure 2. This barrier consists of tight junctions between cells, which help control the passage of substances between cells[125]. It also includes a mucus layer to provide a physical blockade and to house gut microbiota[126]. A healthy gut is semi-permeable and normally leaky, enabling the absorption of nutrients, while preventing the uncontrolled entry of harmful substances[127]. Gluten, other exogenous factors, and abnormal microbiota composition all disrupt gut tight junctions, triggering an autoimmune response[27,128] and exacerbating inflammation[129], thereby increasing intestinal permeability[130-133]. Systemic inflammation directly damages the epithelial cells lining the gut, disrupting tight junctions that normally maintain the barrier, and increasing the intestinal permeability (right panel)[134,135]. The mucosal cell alterations that occur in the intestinal barrier are a consequence of a response to gluten that targets transglutaminase, which is a focus for autoantibodies[136], leading to inflammation and barrier damage[137-139]. Symptoms of leaky gut, the end result of increased permeability, can include abdominal pain, bloating, food sensitivity, fatigue, and more serious health problems[140].

Figure 2 The intestinal barrier and factors causing its disruption. Both the mucus layer and tight junctions between cells contribute to a healthy barrier. However, disruption in the mucus layer and tight junctions between cells will result in increased intestinal permeability.

At the molecular level, pro-inflammatory cytokines, including TNF-α and IL-6, alter the expression of proteins involved in tight junctional integrity, and lead to increased permeability. Age-related increases in certain inflammatory factors such as TNF further contribute to enhanced intestinal permeability, and also cause dysbiosis[141,142]. The inflammation diminishes beneficial gastrointestinal bacteria including Faecalibacterium prausnitzii, which are important for maintaining gut health, and increases pro-inflammatory bacteria[143]. This disruption can further exacerbate systemic inflammation by releasing inflammatory molecules and increasing barrier permeability[144], leading to or worsening autoimmune reactivity, including in those individuals with celiac disease[145]. The chronic inflammation leads to persistence of villous atrophy[146] and further erosion of the barrier[147].

Damage to the intestinal lining can additionally be caused by pro-inflammatory exogenous factors, including chronic use of non-steroidal anti-inflammatory drugs[148], stress and poor sleep habits[149], chemotherapy and radiation[150], certain diseases such as HIV/AIDS[151], as well as food allergens[152]. Antacids may harm the gut lining by reducing the diversity and richness of the intestinal microbiota, and altering gut barrier function, potentially leading to a more permeable gut, whereby additional toxins and other foreign substances enter the bloodstream[153]. A disrupted barrier function caused by other autoimmune conditions, such as type I diabetes, increases the risk of developing celiac disease in genetically disposed individuals, and vice versa[154].

While increased intestinal permeability is a characteristic of untreated celiac disease, it typically resolves after several months on a gluten-free diet[126,155]. Adhering to a strict gluten-free diet therefore enables, but does not guarantee, healing of the intestinal lining[126,155]. Factors including gluten exposure duration, symptom severity, and individual genetic factors influence the extent of mucosal damage and the time to recovery[156]. Various additional strategies are sometimes added to address leaky gut in celiac disease, including a FODMAP dietary regimen, supplementation with prebiotics and probiotics[157], and stress reduction techniques[158,159]. The overall recovery process for the gastrointestinal barrier after onset of the gluten-free diet typically takes 3-6 months in children[160], and 2 years or more in adults[161]. As the villi are restored and the barrier heals, intestinal permeability can return to normal. However, continued adherence to a gluten-free diet is essential to prevent the return of leaky gut and the associated symptoms of celiac disease[162]. The degree of intestinal permeability and recovery can be assessed with both in vivo and in vitro testing methods[163]. In vivo testing typically involves measuring the passage of inert markers through the intestinal lining.

Although avoidance of gluten ingestion from wheat, rye, or barley prevents or ameliorates immune response and gastrointestinal barrier damage in susceptible individuals, care must also be taken to avoid the detrimental effects of a poor gluten-free diet in the form of nutritional deficiency, ingestion of heavy metals from the consumption of replacement grains, lack of anti-inflammatory foods, and gravitating toward foods with unhealthy ingredients[164,165]. A poor gluten-free diet can increase cardiovascular risk and increase the risk of obesity, metabolic syndrome, and negatively impacts blood sugar and lipid metabolism[166-168].

THE GASTROINTESTINAL MICROBIOME

In the human body, 40 trillion bacteria live in and on skin, intestines, mouth, genitals, and elsewhere, and are emitted in one’s microbial signature to the point where people having been in a room can be detected and distinguished based on the aerial bacteria[169]. The gut microbiome, as a collection of microorganisms in the digestive tract, likely has a major role in celiac disease development[170]. Inflammation and nutrient malabsorption due to persistent villous atrophy impact gut microbiome balance, and potentially contribute to the persistence of dysbiosis. Imbalance of intestinal flora may also be caused by a poorly planned gluten-free diet[27]. Bacterial genes and the gastrointestinal tract have a synergistic relationship[171]. Beneficial gut bacteria assist in breaking down foods, making nutrients available, and producing beneficial vitamins and anti-inflammatory compounds. A less diverse microbiome struggles to perform its essential functions, including protecting the gut from harmful organisms. The gut microbiota can also influence the immune response to gluten[172].

At parturition, the birth canal may provide and inoculate the infant’s intestinal biome to enhance the baby’s immune system, and even healthy wombs contain a small but measurable number of bacteria which may be transferred to the baby[173]. In breastfed babies, Bifidobacteria and Lactobacilli, which are beneficial bacteria contained in and thriving on breast milk, become the dominant species in the infant gut during the first few months of life[174]. They produce natural antibiotics (i.e., antimicrobial peptides) that kill harmful bacteria[132,175]. They stimulate the baby’s Th1 immune response and reduce the Th2 response, thus lowering the risk of allergies[176]. They strengthen the infant gut wall by triggering the growth of new epithelial cells lining the walls, thereby making it more difficult for invaders to enter the bloodstream[177]. They also stimulate the production of mucus, which adds another barrier to the gut lining, feeds beneficial microbes, and stops pathogenic strains of bacteria such as Escherichia coli from clinging to the lining[178].

An increased abundance of pro-inflammatory or autoimmune-linked taxa, such as Clostridium difficile, Prevotella, and Enterococcus is present in infants who later develop celiac disease[179]. There is also a reduced abundance of anti-inflammatory taxa such as Bifidobacteria and Lactobacillus in those predisposed to the disease[180-183]. Decreased production of butyrate, a SCFA produced by beneficial gut bacteria from a healthy diet that includes whole grains, and other gut microbiota imbalances, can bring about increased systemic inflammation[184] and it may spur celiac disease onset[185]. From healthy to pre-celiac disease states, SCFA profiles alternate, suggesting that a root cause is a functional disruption in microbiota metabolism[186,187]. The shifts occur prior to seroconversion (positive tTG-IgA level)[188], which hints that dysbiosis precedes immune activation in celiac disease onset. The altered environment can favor the growth of harmful bacteria and suppress the growth of beneficial bacteria[189].

When dysbiosis increases intestinal permeability, it enables gluten peptides to cross the barrier and trigger autoimmunity, leading to the intestinal damage characteristic of celiac disease[86,155]. Dysbiosis can also contribute to the progression of celiac disease, and the persistence of its symptoms, by additionally disrupting the intestinal barrier, altering gluten metabolism, and modulating immune response[144]. Changes in gastrointestinal bacterial composition and function affect the digestion of gluten, potentially exacerbating the effects of inflammation[190]. Specific beneficial bacterial species, including Firmicutes and Actinobacteria, may decrease in abundance in celiac disease, while Proteobacteria, which are harmful, may increase[191]. Alterations in NOD-like receptor activity, a key part of the innate immune system, are influenced by dysbiosis, with the potential to further promote immune response to gluten in individuals with celiac disease[192,193].

Gut dysbiosis, when it occurs, may persist for years after gluten-free diet onset in individuals with celiac disease[86]. This persistence is related to genetic predisposition, the specific composition of the gut microbiome, and food types consumed on the gluten-free diet[194]. The chronic systemic inflammation that results unsettles the delicate balance of the gut microbiota, leading to additional dysbiosis, and can disrupt immune tolerance, causing the production of autoantibodies and furthering inflammation[185]. Studies have shown a strong link between chronic inflammation and the development of autoimmune disease in general[195].

Current investigations into the effect of the microbiome on celiac disease are typically limited to stool samples investigation, which are not equivalent to analyzing the intestinal biome[196]. Emerging non-invasive techniques for improved characterization of the intestinal biome include capsule sampling[197], breath tests[198], and secretome profiling[199,200]. Another limitation in the studies published thus far is in the presentation of self-reported questionnaires[201], which require validation to increase reliability[202]. Digital phenotyping and objective biomarkers from wearable sensors and continuous metabolomic monitoring can potentially reduce error[203]. Metabolomic monitoring improves understanding of celiac disease by identifying distinct, non-invasive metabolic signatures in blood, urine, or saliva that reflect active inflammation, gut microbiota alterations, and malabsorption[107,204]. It enables the real-time tracking of disease progression and responses to a gluten-free diet by revealing metabolic changes even before intestinal damage occurs. Some studies suggest that modulating the gut microbiota with probiotics can have a positive impact on celiac disease[128,143,205]. However, subsequent work will be needed to enhance comprehension of the intestinal microbiome and the best investigative techniques, including modeling early predictive biomarkers from stool SCFAs, microbial diversity, zonulin, and tTG-IgA kinetics[143,206]. It might also be helpful to design an interventional trial for high-risk infants[207], targeting, for example, microbiome preservation and the regulation of inflammatory load. Another assistive measure would be the development of a quantitative risk stratification algorithm that combines genetics, the microbiome, metabolomics, and environmental data[208,209].

IMMUNE SYSTEM FORERUNNERS OF CELIAC DISEASE

When gluten is ingested, it is then partially decomposed in the gastrointestinal system to gliadin, inducing adaptive and innate immune responses[210,211]. The steps by which gluten proteolysis is recognized as antigenic in celiac disease are shown in Figure 3. The deamidation of gluten peptides by tissue transglutaminase, as illustrated in the figure, improves affinity for HLA-DQ2/8 molecules in susceptible individuals through a process of negatively charged glutamic acid residues binding to positively charged pockets within the HLA-DQ molecules[90,212]. This increased affinity triggers a more vigorous T-cell response, a key factor in the onset of celiac disease. APCs, such as dendritic cells, contain major histocompatibility complexes (MHCs), whereby deamidated gluten peptides (DGPs) are attached and presented to CD4+ T-cells[90,213]. The increased binding affinity leads to a more robust presentation. Activation of the T-cells triggers a cascade of events including the release of inflammatory cytokines[167], which damages the intestinal lining[214]. The persistent inflammation and scarring of the small intestine cause the characteristic immune response[215,216].

Figure 3 Molecular-level steps by which gluten is recognized as antigenic in celiac disease. Gluten from wheat, rye, or barley is broken down into gluten peptides. Tissue transglutaminase deaminates them, triggering a cascade that leads to intestinal villous atrophy. HLA: Human leukocyte antigens; APC: Antigen-presenting cell; IFN-γ: Interferon-γ; IL: Interleukin.

Reprogramming of natural memory T-cell behavior, including their contribution to inflammation and cytotoxicity, can signal disease onset and progression[217]. The CD4+ T-cells play a crucial role in activating gluten-specific B cells, which then produce anti-transglutaminase and other antibodies[218]. Activated B cells also form complexes with gluten peptides, leading to the production of antibodies specific for DGPs[219]. Toll-like receptor (TLR) signaling, activated by gluten peptides, contributes to inflammation, immune responses, and disease pathogenesis[220-222]. It can trigger the release of pro-inflammatory cytokines and chemokines, which lead to intestinal inflammation and scarring. Specifically, TLR3, TLR4, TLR7, and TLR9 have been implicated in the mechanisms, with altered expression levels being observed in affected individuals[223]. The released cytokines advance inflammation and destruction of the intestinal villi, hindering nutrient absorption[81]. Specific transcription factors and signaling pathways, including mitogen-activated protein kinases and nuclear factor-κB, induce the expression of genes encoding the pro-inflammatory cytokines and other inflammatory mediators[95,224].

PATHOGENIC STEPS LEADING TO CELIAC DISEASE

Gluten is the trigger or antigen in celiac disease onset, and specifically, gliadin peptides that resist digestion, which interact with tissue transglutaminase enzyme present at the epithelial surface and/or when the antigens pass through a leaky intestinal membrane to the lamina propria[90]. This leads to deamidated gliadin that then activates CD4+ T-cells in HLA-DQ2/8 positive individuals[124]. Hence the question: Why not preemptively begin a gluten-free diet in all susceptible individuals? Such a measure would prevent future onset. However, a lifelong gluten-free diet is both nutritionally and socially burdensome[225]. Furthermore, since 97% of HLA-DQ2/8 carriers never develop celiac disease, broad restriction would not be an appropriate choice but rather, an overtreatment[226]. Moreover, gluten is important for immune training and tolerance in early life, and its complete absence might even cause harm in some contexts[227]. There is evidence that early gluten introduction, at age 4-6 months, does not increase celiac disease risk in the genetically at-risk pediatric population[17,228,229]. Yet a recent presentation from the Celiac Disease Genomic, Environmental, Microbiome, and Metabolomic Study (CDGEMM) group[183] found that gluten intake between 7-15 months is linked to an increased risk of celiac disease. Hence, timing and amount of gluten alone are likely influential, but not the passkey, for disease onset.

The recognized immunologic steps leading to celiac disease[230,231] are illustrated in Figure 4. Stage 1, termed genetic susceptibility, is defined by core variables HLA-DQ2/DQ8 haplotypes[232]. It is possible that other genetic modifiers are involved, specifically non-HLA loci including IL-2, IL-21, and SH2B3[233]. Some gliadin peptides, specifically p31-43, can also trigger an innate immune response in individuals with celiac disease, even without HLA-DQ2/8 alleles[234]. This step is a necessary and innate predisposition, but it is not alone predictive of onset. Stage 2, entitled environmental priming and microbiome development, may occur in the first two months of life[235]. Several variables contribute to this stage including mode of birth (vaginal vs C-section)[236], early diet (breastmilk vs baby formula)[237], exposure to antibiotics which affect the gastrointestinal biome[238], and hygiene including the presence of household pets and siblings, as well as interaction with other children and activities away from the home[239]. The gastrointestinal microbiome features of normal individuals include Bifidobacteria, Lactobacilli, and SCFA producers, whereas in dysbiosis, there is a prevalence of Enterococcus, Clostridium, and Prevotella[240]. At this stage, the window for preventing onset is likely to be very high. Based on prior knowledge, breastfeeding and reduction of unnecessary antibiotics should be encouraged[238,241]. Microbes present in vaginal fluid may possibly colonize the gut of newborns when they pass through the birth canal, though this is still uncertain[242,243]. Diet, health status, and stress during pregnancy are perhaps transferred from the mother to her offspring via the microbiome[244].

Figure 4 Stages in the pathogenic change from normal to the celiac disease state. The description of these stages is provided in the text. At each subsequent stage, the window for prevention of celiac disease onset/reversibility is diminished. HLA: Human leukocyte antigens; CD: Celiac disease.

At stage 3 (4-12 months of age), there is immune priming and gluten introduction[227,245]. The variables include the precise timing, dose, and matrix of gluten introduction, the co-existence of any infections (particularly viral), and gut permeability, which depends in part on zonulin activity. It is currently thought that gluten exposure in the context of dysbiosis or viral activation may breach oral tolerance[176]. Thus there is the possibility of celiac disease onset, at least transiently. However, prevention is likely attainable by controlling gluten exposure, supporting the epithelial barrier, perhaps with an abundance of SCFAs present, and the possibility of vaccine development against relevant viruses[126,246].

With further exposure to environmental insult, the likelihood of celiac onset increases dramatically. At stage 4, there is both dysbiosis and metabolomic drift in the gastrointestinal tract[247,248]. The key changes observed in early CDGEMM studies include reduced gastrointestinal SCFAs, increased pro-inflammatory metabolites, altered bile acid and tryptophan metabolism, and increased microbial genes related to oxidative stress and lipopolysaccharide synthesis[183,249]. These events have functional impact in reducing regulatory T-cell (Treg)-inducing signals and increasing epithelial stress and antigen presentation[250,251]. It may be difficult but not impossible to prevent disease onset at this stage, depending on microbiome modulation, based on fecal microbiota transplantation, diet, targeted probiotics and emerging microbiota-derived postbiotics, or metabolite mimetics[247,252,253].

At stage 5 there is immunologic activation and seroconversion[12]. The activation of gliadin-specific CD4+ T-cells occurs (which is HLA-restricted, the human version of the MHC), along with transglutaminase modification of gliadin resulting in deamidated gliadin peptides, followed by production of transglutaminase-IgA and epithelial membrane antigen[254]. The outcome may be transient seroconversion with immune quiescence[255-258], or otherwise, persistent autoimmunity leading to mucosal injury. The window for intervention prior to celiac onset is now likely narrow, if at all extant. It may yet be mitigated by immune tolerance induction therapy, currently under research investigation[259], and support for intestinal barrier repair and inflammation resolution[260].

Finally at stage 6 there is the onset of clinical celiac disease[261]. The window for prevention is closed, and the only viable therapy known to date is a strict and lifelong gluten-free diet[162]. The harm is irreversible without a gluten-free diet, with manifestations possibly including small intestinal villous atrophy and crypt hyperplasia, malabsorption of nutrients, diarrhea, growth failure, iron deficiency anemia, and perhaps extra-intestinal symptoms such as rash, fatigue, and neurological disorders[262]. Disease-specific antibodies typically decrease within weeks of gluten-free diet commencement[263]. A gluten-free diet can induce rapid changes in memory T-cells, shifting them towards a non-destructive state, although gluten-specific CD4+ T-cells persist for years[264].

INFLAMMATORY LOAD AS PERPETRATOR IN CELIAC DISEASE ONSET

Inflammatory load, defined equationally as the weighted sum of inflammatory vs anti-inflammatory factors contributing to autoimmunity in celiac disease, can be understood as a conceptual and a relative construct, rather than being a single quantitative variable with a universal threshold. It reflects the cumulative convergence of heterogeneous stressors, vs ameliorating influences, that on balance and collectively determine whether immune tolerance capacity is exceeded. The precise threshold is expected to vary across individuals, depending on genetics, age, microbiome configuration, and immune history, and thus it is best studied longitudinally rather than being defined a priori. Endoscopic and biopsy-derived metrics potentially serve as downstream integrative readouts of this cumulative burden, and enable testability, even when upstream contributors cannot be individually quantified.

Two leading hypotheses describe the immunologic trigger prompting celiac disease onset, and they are related to inflammatory load. The quantitative/affinity threshold hypothesis (peptide binding model, termed Model 1)[265,266], suggests that when the binding strength and density of DGP-HLA complexes reach a threshold, they cross-activate T-cells which previously ignored these complexes due to their low expression or insufficient affinity. The transition could be due to increased deamidation, for example, resulting from inflammation-induced transglutaminase expression, APC structural modification, increased presentation efficiency in response to inflammatory stress, and/or enhanced intestinal permeability, which eventually leads to leaky gut and enables additional peptides to reach the lamina propria. Hence, disease onset would result from a strong and sustained APC presentation of DGPs exceeding a critical threshold, causing T-cell activation[267]. Whereas, the T-cell threshold hypothesis (loss of tolerance model, termed Model 2)[268,269] suggests that the DGP-HLA complex is always presented effectively in HLA-DQ2/8 carriers, but that T-cells initially tolerate the antigen via anergy (functional immune unresponsiveness), active Treg suppression, or the lack of co-stimulation[270,271]. At the tipping point, the tolerance mechanisms fail, whereupon T-cells experience a gain of effector function, potentially triggered by either viral infection which induces bystander activation or molecular mimicry, a microbiota shift to alter the IL-15/IL-21cytokine milieu and break tolerance, or stressors that impair Treg function or enhance APC activation via TLR or other signaling[222,272,273]. Celiac disease onset may in fact require both mechanisms, i.e., an increase in effective presentation of DGPs via inflammation, higher peptide load, and/or APC activation, and additionally, the failure of immune regulation from impaired Treg suppression[271] or cytokine skewing[274]. Table 2 summarizes the key factors involved in two-signal T-cell activation[265-274]. It shows the molecular component, the normal state, and what transpires at disease onset. Both signals contribute to T-cell activation. This two-step process is crucial for preventing autoimmunity, as T-cells must receive confirmation of a threat from APCs via co-stimulation. Sometimes included in the model is a signal 3, which refers to cytokines (e.g., IL-2, IL-12, and IFN-α/β) released by APCs or T-cells themselves to further enhance proliferation and differentiation. Other molecules, such as CTLA-4, regulate or terminate T-cell responses, providing crucial immune checkpoints.

Table 2 Two-signal T-cell immunologic activation.

Signal	Component	Normal state	Disease onset
Signal 1	DGP-HLA complex on APC	Present, below threshold	Highly expressed, strongly bound
Signal 2	Co-stimulation, loss of regulation	Tregs dominant, T-cells anergic	Tregs suppressed, cytokines upregulated

DGP: Deamidated gliadin peptides; HLA: Human leukocyte antigens; APC: Antigen-presenting cell; Tregs: Regulatory T cells.

Since most of the factors involved in both immunologic models 1 and 2 above have an inflammatory, and therefore immunologic, load as a root cause, this load can be considered the common denominator linking both antigen presentation threshold and loss of tolerance. Inflammatory load is therefore an important and perhaps essential driver or modifier of the immune response to gluten ingestion. Hence, the onset of celiac disease might be thought of as an immune threshold event[275] affected by a rising inflammatory load that alters the immune system’s capacity to tolerate dietary gluten. Two principal and likely complementary mechanisms contribute to this transition. For the quantitative/affinity threshold model, stress via infection, microbiome dysbiosis, chemical exposure, or the appearance of other exogenous factors, all pro-inflammatory, increases the load and affinity of deamidated gliadin peptides presented by HLA-DQ2/8 molecules. When the peptide-MHC density and stability surpass a critical level[276], previously inert CD4+ T-cells awaken, respond, and are activated. For the loss of tolerance model, chronic inflammatory stimuli may impair Treg function, upregulate pro-inflammatory cytokines such as IL-2, IL-15, and IL-21, and drive naive or anergic gluten-specific T-cells toward an effector phenotype[269]. This immune reprogramming thereby reduces oral tolerance to initiate a sustained response to dietary gluten. In each case, the inflammatory load serves as the central amplifier, tipping the balance toward autoimmune activation. The following flow equations summarize the effects of inflammatory load on two-signal T-cell activation:

IL → APC activation → peptide density/affinity (signal 1);

IL → cytokine skewing (IL-15, IL-21) → Treg dysfunction (signal 2).

As the load increases, which can be sourced via multiple and often synergistic environmental insults, it creates a local cytokine milieu and an antigen presentation profile that exceed the threshold for immune tolerance[277]. Once this threshold is crossed, CD4+ T-cells become immunogenic to gluten antigens, leading to production of autoantibodies, epithelial cytotoxicity, and ultimately, villous atrophy[278].

Some reversibility in these cascades may be possible depending on the restoration of intestinal barrier integrity, including the downregulation of zonulin[279], inactivation or apoptosis of memory T-cells specific to gliadin[280], resolution of the pro-inflammatory cytokine patterns[272], re-establishment of tolerogenic signaling (e.g., transforming growth factor-β, IL-10)[281], and realignment of the gut microbiome toward anti-inflammatory dominance[66]. In some individuals, anti-transglutaminase IgA titers rise temporarily without sustained exposure to gluten or progression to villous atrophy[282]. Such cases may reflect a window during which immune re-equilibration is still possible (Figure 4), particularly if inflammatory load, both gluten and non-gluten-derived, is attenuated. Importantly, however, even on a gluten-free diet, complete elimination of gluten exposure is rarely achieved[283], and persistent symptoms may reflect continued inflammatory insults from minute gluten and non-gluten sources[216]. The essence of the inflammatory load framework for both disease onset and post-diagnosis symptom perpetuation is shown in diagrammatic form in Figure 5. The inflammatory load threshold to activate the quantitative/affinity vs loss of tolerance models may vary (top panel). On the left side of the panel, the inflammatory load threshold for the quantitative affinity model is high whereas it is low for the loss of tolerance model. In the middle portion of the panel, the inflammatory load threshold is intermediate for both models, and at the right in the panel, the inflammatory load threshold is low for the quantitative/affinity model vs high for the loss of tolerance model. Anywhere along the curves, increasing the inflammatory load when in the tolerant state can lead to disease onset (bottom panel). It is important to note that Figure 5, as well as Figures 3 and 4, highlight a conceptual synthesis regarding celiac disease onset and the possibility of early-stage reversibility.

Figure 5 A model of celiac disease onset. The top panel shows that various combinations of inflammatory load thresholds for the quantitative/affinity vs loss of tolerance models may occur. As inflammatory load increases in the tolerant individual (lower panel), regardless of the threshold levels (top panel) the balance will be tipped toward disease onset, so that oral tolerance may be breached.

PARADIGM FOR ASSESSING INFLAMMATORY LOAD

Measures of inflammation such as those described in the molecular biomarkers of inflammation section above, can be helpful to assess inflammatory load, but individually or collectively, they are not reliably correlated to celiac disease onset[98]. We suggest, rather, that a weighted contribution of inflammatory vs anti-inflammatory environmental and other exogenous factors is crucial to predicting celiac disease onset, and for ameliorating the symptoms and the pathologic response after onset of the disease. In the section above entitled pro- vs anti-inflammatory considerations, we list examples of these factors and influences. Major factors to be included in an inflammatory load equation should include diet (using assessment indices), exercise, body weight, oral health, ongoing infections, viral history, surgical and other trauma, posture, sleep quality, air and water pollution levels, lifestyle toxins including heavy metals and microplastics, vices such as alcohol, smoking, and illicit drugs, psychological health, genetic factors, and presence of other autoimmune disease. The equation could be implemented as a weighted summation of each major factor based on the balance of pro-inflammatory influences (positive value) vs anti-inflammatory influences (negative value). Initializing the weighting at unity, a more positive overall score would indicate a greater inflammatory load and the possibility of onset/continuance of symptoms. The inflammatory load equation would then be updated in terms of influences contributing to each factor and the major factor weighting using statistics methods (e.g., logistic regression analysis) and artificial intelligence methods. An inflammatory load equation used for assessment may have relevance to other disorders, such as atrial fibrillation, in which healthy heart tissue becomes fibrotic in part due to inflammatory response[284].

IMPLICATIONS FOR MODERN GASTROINTESTINAL ENDOSCOPY

Measurements to assess the inflammatory load framework and its predictive capabilities have direct implications for modern gastrointestinal endoscopy. Endoscopy increasingly plays a role in early detection, risk stratification, and longitudinal disease monitoring[285]. Within this context, endoscopic evaluation of the small intestine can help to identify mucosal changes that precede irreversible architectural damage and correspond to rising cumulative inflammatory load prior to celiac disease onset. Advances in high-definition white-light endoscopy[286], digital chromoendoscopy[287], and narrow-band imaging[288] raise the possibility of detecting subtle mucosal abnormalities associated with early epithelial stress, including altered vascular patterns, focal scalloping, or patchy villous blunting[289]. These findings may occur before classical Marsh 3 lesions, and could correspond to early immune activation predicted by inflammatory load accumulation. Emerging technologies such as confocal laser endomicroscopy and endocytoscopy[290] further enable real-time visualization of the epithelial barrier integrity, tight-junction disruption, and intraepithelial lymphocyte density, offering in vivo assessment of processes traditionally inferred from histology alone[291].

Endoscopy-guided biopsies provide an opportunity to move beyond presently used binary diagnostic thresholds. Quantitative histologic and molecular markers obtained during endoscopy with biopsy, including gradients of intraepithelial lymphocytes, crypt hyperplasia, cytokine expression, microbiome composition, and markers of intestinal permeability, may serve as endoscopic correlates of inflammatory load. Integration of these measures potentially help distinguish transient immune activation from progressive loss of tolerance in individuals at the borderline or with seronegative celiac disease[292]. Finally, endoscopy remains central to evaluating mucosal healing after initiation of a gluten-free diet. Persistent villous atrophy, despite dietary adherence, may reflect continued gluten or non-gluten inflammatory burden. Within the inflammatory load framework, endoscopy functions not merely as a diagnostic tool, but as an integrative phenotyping modality, capturing the cumulative downstream effects of immune activation, epithelial stress, and microbial perturbation.

LIMITATIONS AND FUTURE DIRECTIONS

The preventive strategies discussed herein are hypothesis-generating, although they are not yet supported by large-scale, long-term interventional trials. Their inclusion reflects biologic plausibility and emerging mechanistic evidence rather than clinical endorsement. Earlier foundational studies are retained in this review to preserve both historical and mechanistic continuity, while recent work is incorporated to reflect current technical advances. At present, no validated endoscopic standards exist that directly quantify inflammatory load. Hence, the purpose of this review and framework is to propose biologically grounded correspondences between emerging endoscopic markers and cumulative immune activation, rather than to define prescriptive clinical thresholds. Future longitudinal studies combining endoscopic findings with systemic inflammatory markers and microbiome profiling may clarify the mechanisms of onset and incomplete recovery in celiac disease, and guide individualized surveillance strategies.

Pertinent to future investigations, some metrics that can be used to gauge immunologic response in celiac disease onset as it pertains to affirming an immunologic load equation are identified in Figure 6. The topics of future research might include single-cell RNA-sequencing of mucosal CD4+ T-cells from at-risk children at seroconversion[293], circulating DGP-specific T-cell detection via MHC tetramers pre- and post-onset[294], longitudinal microbiome + IL-2/IL-21 profiling in the CDGEMM or TEDDY cohorts[204], and in vitro APC + gluten peptide binding affinity assays under inflammatory conditions[295]. Measuring gluten immunogenic peptides in the urine of patients is a non-invasive method to monitor gluten free diet compliance and transgressions[296]. Personalized risk scoring[297] would combine the HLA genotype, early microbiome composition, metabolomics, and immune profiles. Preventive trials of microbiome-targeted therapies[170], including prebiotics, probiotics, synbiotics (i.e., a mixture of probiotics and prebiotics), and even fecal microbiota transplantation, should be done in at-risk infants. Longitudinal immunophenotyping[298] can be assistive in defining the immunologic point of no return. Artificial intelligence-assisted endoscopic image analysis represents an additional avenue for detecting subtle inflammatory patterns associated with early or incomplete disease states[299].

Figure 6 Examples of metrics that can be used to gauge immunologic response in celiac disease onset. Such metrics act as confirmatory measures when developing an inflammatory load equation. AI: Artificial intelligence.

CONCLUSION

Celiac disease onset is traditionally framed as a deterministic outcome of HLA-DQ2/8 genetics and gluten exposure. However, there are many genetically at-risk individuals who never develop the disease. There is also growing evidence of environmental and immunologic modulation, and a global increase in disease rates. Inflammatory load, which is increasing in many individuals, particularly in industrialized nations, and the cumulative immunologic stress arising from microbial, nutritional, environmental, and psychosocial factors, act to amplify both antigen presentation and immune dysregulation. Herein, a comprehensive review of the role of inflammatory load in the onset and pathophysiology of celiac disease was presented, which integrated genetic susceptibility, environmental stressors, and immune threshold dynamics. A paradigm was proposed to assess the odds of a susceptible person developing the disease, and to reduce the time needed for recovery after diagnosis. The inflammatory load paradigm presented herein is a testable conceptual mechanism for how stressors vs beneficial factors quantitatively or qualitatively converge to tip the balance at the intestinal immune interface. The growing relevance of these concepts to modern gastrointestinal endoscopic technique was then highlighted. Further investigation via the implementation of a longitudinal measurement of environmental and other exogenous data, with inclusion of modern endoscopic technique, would be beneficial. A completed and weighted profile could potentially provide advantageous information for prevention, treatment, and early intervention.