Recent advances in the diagnosis of celiac disease

Abstract

Celiac disease (CD) is a prevalent immune-mediated disorder triggered by gluten ingestion in genetically susceptible individuals. Primarily affecting the small intestine, CD can also have adverse systemic health effects. However, the majority of affected individuals remain undiagnosed because of asymptomatic or subclinical manifestations. While the diagnostic gold standard remains the combination of positive serum immunoglobulin A antibodies against tissue transglutaminase and villous atrophy on small intestinal biopsy, an evolving understanding of CD pathogenesis has elevated various serum biomarkers to increasingly important complementary diagnostic tools. This editorial aims to outline the scientific merits and potential limitations of serum biomarker assays for CD diagnosis, alongside their established and emerging clinical applications in both initial diagnosis and long-term disease monitoring.

Key Words: Celiac disease; Diagnosis; Immunoglobulin A; Serum; Biomarker

Core Tip: Celiac disease is a chronic immune-mediated enteropathy triggered by dietary gluten in genetically predisposed individuals, causing systemic inflammation through malabsorption and extra-intestinal manifestations While duodenal biopsy is the gold standard, serum antibodies against tissue transglutaminase are crucial complementary tools. This editorial evaluates serum biomarkers as vital tools for diagnosis and monitoring, complementing traditional biopsy methods in this immune-mediated disorder.

INTRODUCTION

Celiac disease (CD) is a chronic immune-mediated enteropathy triggered by dietary gluten in genetically predisposed individuals, causing systemic inflammation through malabsorption and extra-intestinal manifestations[1,2]. Its hallmark pathological features include progressive villous atrophy, crypt hyperplasia, and increased intraepithelial lymphocytes within the small intestinal mucosa upon gluten exposure[2,3]. With an estimated global prevalence approaching 1%, CD ranks among the most prevalent lifelong autoimmune conditions, although substantial regional variations exist and a considerable proportion of cases remain undiagnosed[4,5]. Historically considered rare in China, emerging epidemiological evidence now indicates that CD is not uncommon within the Chinese population, with a reported adult incidence reaching 2.19%[6]. Although strict, lifelong adherence to a gluten-free diet (GFD) remains the only established therapy, significant challenges persist, including dietary limitations, variable mucosal healing rates, and the absence of effective non-dietary pharmaceutical interventions[7-9]. The current diagnostic gold standard mandates both seropositivity for serum tissue transglutaminase-immunoglobulin A (tTG-IgA) antibodies and histopathological evidence of villous atrophy on small intestinal biopsy[10,11]. Serological biomarker development has advanced significantly beyond traditional tTG-IgA and exploring point-of-care testing alongside biomarkers predicting severity or monitoring dietary adherence[12-14]. While these evolving serological tools offer enhanced screening and adjunctive diagnostic value, limitations in sensitivity for early disease or non-responsive CD necessitate ongoing refinement, underscoring the critical need for novel biomarkers and diagnostic strategies which form the primary focus of this editorial.

IMMUNOLOGICAL BIOMARKERS

TTG-IgA/tTG-IgG antibody

Serum tTG-IgA antibody testing has emerged as the cornerstone for screening and monitoring CD[4,15]. Current guidelines from the European Society for Paediatric Gastroenterology, Hepatology and Nutrition propose that duodenal biopsy may be omitted for diagnosing CD in children exhibiting tTG-IgA levels ≥ 10 times the upper limit of normal (ULN)[16]. This strategy effectively reduces reliance on invasive procedures while maintaining high diagnostic confidence. Supporting the extension of this paradigm, a retrospective study confirmed that an anti-tTG-IgA titer ≥ 10 × ULN provides high diagnostic sensitivity and specificity. Notably, this study further suggested that in adults younger than 50 years without concurrent gastrointestinal disorders, an anti-tTG-IgA titer ≥ 10 × ULN may be sufficient to establish a CD diagnosis, potentially obviating the need for confirmatory duodenal biopsy[17]. Collectively, these studies underscore the significant clinical utility of tTG-IgA measurement in enabling non-biopsy diagnosis of CD across both pediatric and adult populations. Beyond diagnosis, follow-up studies demonstrate the role of tTG-IgA in monitoring mucosal healing. Among treated CD patients undergoing follow-up duodenal biopsy, those achieving antibody titers below the negative cutoff (tTG-IgA < 1.2 U/mL) exhibited a significantly increased probability of normal intestinal histology[18]. This finding supports the utility of tTG-IgA serology in monitoring mucosal healing. An important consideration in interpreting tTG-IgA results is the increased prevalence of selective IgA deficiency within the CD population, which poses a risk for false-negative serology. A recent study found that total IgA levels significantly modulate the diagnostic performance of tTG-IgA cutoffs for CD, with higher IgA concentrations (≥ 245 mg/dL) favoring lower thresholds for improved sensitivity despite reduced specificity[19]. Therefore, incorporating quantitative total IgA assessment into the interpretation of tTG-IgA serology is essential not only to identify deficiency but also to optimize diagnostic thresholds, thereby refining the applicability and accuracy of biopsy-free CD diagnostic strategies.

Patients with selective IgA deficiency pose a continual challenge in the screening and diagnosis of CD. Studies indicate that tTG-IgG testing may be useful for identifying those with IgA deficiency who also have CD, and it can help inform the decision to perform a small intestinal biopsy[20]. However, among 178 patients who were tTG-IgG positive but tTG-IgA negative, only six were confirmed to have CD by intestinal biopsy, suggesting that the diagnostic performance of tTG-IgG alone is limited[21]. Furthermore, duodenal biopsy findings revealed no significant correlation between anti-tTG-IgG levels and the degree of pathological damage, whereas a clear relationship was observed between mucosal injury and anti-tTG-IgA titers[22]. Therefore, tTG-IgG may be better utilized in combination with tTG-IgA rather than as a standalone test in patients with IgA deficiency.

Deamidated gliadin peptide antibodies (deamidated gliadin peptide-IgG/IgA)

The World Gastroenterology Organization Global Guidelines on Celiac Disease recommend that in patients with total IgA deficiency, testing for deamidated gliadin peptide (DGP)-IgG antibodies should be performed. A diagnosis of CD is confirmed when a positive DGP-IgG result is combined with histological evidence of villous atrophy on small intestinal biopsy[23]. In pediatric populations, the combined use of tTG-IgA and DGP-IgG antibodies significantly enhances diagnostic sensitivity for CD, rising from 50% to 92%[24]. Notably, among children under two years of age, seroconversion to DGP-IgG positivity preceded tTG-IgA seroconversion in 80% of cases[24]. Furthermore, a meta-analysis suggested that incorporating DGP-IgG testing, particularly in patients with a high clinical suspicion of CD, might enhance diagnostic sensitivity[25]. Collectively, these findings indicate that in clinical practice, the combined serological assessment of tTG-IgA and DGP-IgG warrants consideration for diagnosing CD in highly suspected children under two years of age. Maheshwari et al[26] reported notably high diagnostic performance for DGP antibodies in young children: Both sensitivity and specificity reached 100% and 94%, respectively, in children < 4.0 years. In contrast, within the ≥ 4.0 years age group, DGP-IgA and DGP-IgG demonstrated sensitivities of 60% and 88%, and specificities of 95% and 96%, respectively. A separate meta-analysis further confirmed the robust diagnostic performance of serum DGP-IgA testing for CD, reporting a pooled sensitivity of 83.8% and specificity of 92.1%[27]. However, divergent evidence exists regarding the incremental value of DGP testing. Pacheco et al[28] reported that DGP-IgG measurement does not significantly augment the diagnostic efficacy of tTG-IgA alone for CD. Consequently, the precise clinical utility of DGP-IgG/IgA antibodies in CD diagnosis and monitoring necessitates further validation through rigorously designed prospective trials.

Endomysial antibodies (endomysial-IgA)

Currently, endomysial (EMA)-IgA testing serves primarily as a secondary confirmatory assay for patients exhibiting equivocal tTG-IgA results, including individuals at elevated risk for CD, to establish a definitive diagnosis[29,30]. Specifically, when tTG-IgA antibody titers ≥ 10 × ULN and total serum IgA levels are within the normal range, EMA-IgA testing may be unnecessary. In contrast, EMA-IgA testing is warranted when tTG-IgA antibody titers are low or borderline. A positive EMA-IgA result necessitates further diagnostic evaluation via small intestinal biopsy to confirm CD[31]. A significant limitation inherent to EMA-IgA testing is the requirement for specialized expertise for result interpretation, rendering the assay inherently subjective. Recent investigations suggest that machine learning models offer a promising approach for the rapid and precise automated analysis of EMA-IgA test patterns, potentially streamlining interpretation[32]. Nevertheless, the clinical merits and robustness of such automated approaches require further validation through additional prospective trials and datasets. While serum EMA-IgA testing retains established clinical utility within the diagnostic algorithm for CD, its predictive value for assessing the degree of villous atrophy or confirming mucosal healing during follow-up remains unproven[33]. Consequently, rigorous clinical trials and studies remain imperative to validate the utility of EMA-IgA in monitoring gluten-free dietary adherence and predicting histological outcomes.

NON-IMMUNE BIOMARKERS

Intestinal fatty acid binding protein

Intestinal fatty acid binding protein (I-FABP), a cytosolic protein abundantly expressed in mature enterocytes, functions as a sensitive biomarker for intestinal epithelial injury[34,35]. Following damage to the intestinal epithelium, I-FABP is rapidly released into the systemic circulation. It exhibits particularly high expression within the villous enterocytes of the distal jejunum, which is the primary site of early mucosal damage in CD[36,37]. Emerging evidence indicates that serum I-FABP quantification in CD patients reflects alterations in intestinal permeability and offers a non-invasive method for monitoring mucosal structural integrity[38]. Supporting this, a comparative study of 26 healthy volunteers and 13 biopsy-confirmed CD patients demonstrated significantly elevated plasma I-FABP levels in untreated CD patients relative to controls[39]. Further validation arises from a prospective study involving 90 children with elevated tTG-IgA titers, positive human leukocyte antigen-DQ2/DQ8 haplotypes, and indication for diagnostic duodenal biopsy (test group), alongside 80 children with normal tTG-IgA titers (controls). Serial plasma I-FABP assessments over a 6-month follow-up period revealed elevated levels in 67.8% of the test group. Critically, I-FABP concentrations decreased to control-equivalent levels following 6 weeks of GFD initiation. These findings collectively indicate that plasma I-FABP measurement may facilitate non-invasive CD diagnosis in pediatric populations. The observed post-GFD decline further suggests its potential utility in monitoring disease activity following dietary intervention[40]. Consequently, plasma I-FABP quantification represents a promising biomarker with dual diagnostic and therapeutic monitoring applications in CD. Nevertheless, robust validation through additional large-scale prospective clinical trials is imperative prior to its adoption in routine clinical practice.

Citrulline

Citrulline, a nonprotein amino acid synthesized by small intestinal enterocytes, serves as a functional biomarker of intestinal synthetic capacity and, consequently, absorptive competence[41,42]. A meta-analysis established a robust inverse correlation between plasma citrulline concentrations and the severity of intestinal pathology[43]. Reported diagnostic sensitivity and specificity of plasma citrulline for detecting intestinal damage were 80% and 84%, respectively. Studies consistently demonstrate significantly reduced citrulline levels in treatment-naive CD patients compared to healthy controls. Notably, levels increase following GFD implementation and concomitant mucosal recovery[43], suggesting its utility in monitoring therapeutic response. A recent investigation evaluated plasma citrulline alongside plasma I-FABP and serum regenerating islet-derived protein 1α in treatment-naive CD patients vs controls. Subsequent validation within an intestinal injury prediction model and a prospective cohort identified a plasma citrulline threshold of ≤ 30 μmol/L as predictive of intestinal villous abnormalities, yielding a diagnostic sensitivity of 79.5% and specificity of 83.1%[44]. Collectively, this evidence strongly positions plasma citrulline as a promising non-invasive biomarker for assessing intestinal damage in CD. However, translation into routine clinical practice necessitates further validation through large-scale, prospective multicenter trials.

Urine lactulose/mannitol ratio

Lactulose is a disaccharide with a molecular weight of 342 Da that is absorbed paracellularly via the tight junctions between epithelial cells in the small intestine[45,46]. It is non-toxic, not metabolized by the body, and is rapidly excreted unchanged in the urine following absorption, allowing for its accurate quantification[45,47]. Mannitol, a monosaccharide with a molecular weight of 182 Da, is primarily absorbed via the transcellular route through the epithelial membrane[48,49]. Like lactulose, mannitol is not metabolized; after being absorbed intestinally, both compounds enter the bloodstream and are ultimately excreted in the urine. Intestinal permeability testing is a non-invasive method based on the urinary recovery rates and the lactulose/mannitol ratio (LMR)[48,49]. Research has demonstrated that intestinal permeability accurately mirrors the extent of intestinal mucosal damage and serves as a robust indicator for assessing gut barrier integrity, which is closely linked to the pathogenesis of CD[45]. Specifically, the urinary LMR is significantly elevated in patients with CD compared to healthy controls, though it remains lower than that observed in individuals with Crohn’s disease[50]. Furthermore, the sensitivity of this ratio was reported to be 87% in screening contexts and 81% in clinical settings, supporting its utility as an effective tool for screening CD in the general adult population[51]. In one study involving 22 CD patients who were positive for anti-gliadin antibody (AGA) and underwent GFD treatment, 40.9% remained AGA-positive[52]. The urinary LMR was significantly higher in these AGA-positive patients than in their AGA-negative counterparts, and both groups showed higher ratios than healthy controls. These results indicate that the urinary LMR correlates more closely with clinicopathological features than do AGA, suggesting its potential importance in monitoring disease progression in CD patients.

Blood urea nitrogen

Blood urea nitrogen (BUN) is a metabolic waste product filtered by the kidneys, commonly used to assess renal function and hydration status. While traditionally viewed as a marker of kidney health, emerging evidence suggests BUN levels may reflect broader systemic disturbances, including nutritional deficiencies and chronic inflammatory states. The study by Li et al[53], published in the World Journal of Gastroenterology, presents compelling evidence that BUN serves as a novel and independent predictor of diagnostic delays in CD, revealing an unexpected intersection between renal metabolic markers and gastrointestinal autoimmunity. The retrospective cohort study of 166 biopsy-confirmed CD patients at a major referral center in Xinjiang, representing the largest single-center CD cohort in China. Their findings paint a concerning picture: 42.2% of patients experienced diagnostic delays exceeding 2 years from symptom onset, with 18.7% enduring over 5 years before diagnosis. Crucially, elevated BUN (> 4.3 mmol/L) demonstrated a dose-dependent association with delayed diagnosis (odds ratio = 1.39, 95% confidence interval: 1.04-1.99), even after adjusting for confounders. The clinical implications of identifying BUN as a predictor of CD diagnostic delay are profound. Firstly, it provides clinicians with a readily available, inexpensive, and routinely measured biomarker that can serve as a red flag. Secondly, the proposed mediation by folate deficiency and anemia underscores the complex interplay between CD-induced mucosal damage, resultant nutritional malabsorption, and systemic metabolic consequences. Elevated BUN in this context likely reflects not only potential subtle renal involvement associated with autoimmunity but also the downstream effects of chronic inflammation and malnutrition on protein metabolism and urea cycle dynamics. Incorporating BUN into risk stratification models could help pierce this veil. Looking forward, this finding necessitates validation in larger, multi-ethnic prospective cohorts and exploration of the underlying mechanisms linking BUN elevation to delayed CD recognition.

CONCLUSION

Diagnostic approaches for CD are undergoing progressive refinement towards enhanced sensitivity, reduced invasiveness, and improved cost-effectiveness. Although both immunological (e.g., tTG-IgA, EMA-IgA) and non-immunological biomarkers (e.g., I-FABP, citrulline, LMR) associated with CD pathogenesis are actively investigated for their diagnostic and monitoring utility, histological confirmation of villous atrophy via small intestinal biopsy remains the indispensable gold standard for definitive diagnosis in most patients. This requirement persists despite the established role of positive serological testing for tTG-IgA antibodies as a primary screening tool. Consequently, future clinical research must prioritize large-scale, multicenter validation studies to rigorously evaluate the clinical performance of emerging serum biomarkers. Such efforts are critical to precisely delineate their diagnostic thresholds, individual limitations, and synergistic potential when integrated with conventional serological and histological methods. The ultimate objective is to establish optimized biomarker panels that augment diagnostic accuracy, reduce reliance on invasive procedures, and improve patient stratification, collectively advancing precision medicine in celiac disease management.