Clinical features and biomarker research advances in immunoglobulin 4-related digestive system disorders

Abstract

Immunoglobulin G4 (IgG4)-related disease is a systemic fibro-inflammatory condition characterized by tumefactive lesions, dense lymphoplasmacytic infiltration, and storiform fibrosis. Among its various presentations, IgG4-related digestive system disorders (IgG4-DSD) are the most prevalent, frequently mimicking malignancies and posing significant diagnostic challenges. This review synthesizes advances over the past five years in the clinical spectrum, pathogenesis, and biomarker landscape of IgG4-DSD. We detail the specific features of key manifestations, particularly type 1 autoimmune pancreatitis and IgG4-related sclerosing cholangitis, which often coexist and manifest as painless jaundice. Differentiating these from pancreatic cancer relies on identifying characteristic radiological signs, such as the “sausage-like” pancreatic enlargement, and excluding conditions like primary sclerosing cholangitis. Furthermore, we explore the immunopathogenic mechanisms driving tissue fibrosis, emphasizing the upregulation of Th2 cytokines [interleukin (IL)-4, IL-13] and the roles of T follicular helper cells and M2 macrophages. Recognizing the limitations of serum IgG4, including the prozone effect and potential false negatives, we critically evaluate novel biomarkers with higher sensitivity and specificity. Promising candidates include circulating plasmablasts (CD19⁺CD24⁻CD38^hi), peripheral helper T cells, chemokines such as thymus and activation-regulated chemokine/CCL17, and the interferon-α/IL-33 axis. Integrating these novel markers with clinical and radiological findings is essential for early diagnosis, monitoring disease activity, and implementing personalized therapeutic strategies.

Key Words: Immunoglobulin 4-related disease; Biomarkers; Digestive system; Diagnosis; Pathogenesis

Core Tip: Given that immunoglobulin-related digestive system disorders frequently mimic malignancies, leading to misdiagnosis, this review systematically maps the clinical spectrum from classical (pancreato-biliary) to rare (esophageal/mesenteric) manifestations. A key highlight is the paradigm shift from single-marker diagnosis to multidimensional biomarker assessment (e.g., plasma blasts, interferon-α-interleukin-33 axis). It focuses on the utility of these novel markers in monitoring disease activity and predicting fibrosis, providing evidence-based strategies for early recognition and personalized management.

INTRODUCTION

Immunoglobulin G4-related disease (IgG4-RD) is a systemic fibro-inflammatory condition recognized as a distinct clinical entity over the past two decades. Although it can affect nearly any organ, the digestive system is a preferential target, manifesting as autoimmune pancreatitis (AIP), sclerosing cholangitis, or hepatopathy. Pathologically, the disease is defined by a triad of dense lymphoplasmacytic infiltration, obliterative phlebitis, and storiform fibrosis, often accompanied by elevated serum immunoglobulin G4 (IgG4) levels and a dramatic response to glucocorticoid therapy.

The epidemiology of IgG4-RD exhibits significant regional and phenotypic heterogeneity. In Japan, the estimated incidence ranges from 0.28 cases to 1.08 cases per 100000 population[1]. Similarly, a recent large-scale study in the United States reported an incidence of 1.39 cases per 100000 person-years[2]. While the overall incidence appears comparable, phenotypic presentations differ markedly. International cohort analyses indicate that Asian patients more frequently exhibit pancreato-biliary involvement (the “Asian phenotype”), whereas Western cohorts show a higher prevalence of retroperitoneal fibrosis and aortitis[3].

In China, despite the absence of nationwide registry data, rapidly emerging evidence from major tertiary centers indicates a substantial disease burden. Notably, Chinese patients exhibit immunopathological profiles closely aligned with those of other East Asian populations, characterized by markers such as peripheral helper T cells (Tph) cells[4] and human epididymis protein 4 (HE4)[5], which substantiates this immunopathological similarity. This strong geographic clustering suggests a significant genetic component; for instance, HLA-DRB1*04:05 is associated with susceptibility in Asian populations, while different alleles predispose Western populations[1]. Crucially, this regional heterogeneity poses a challenge for biomarker standardization. Diagnostic cutoffs for serum IgG4 (> 135 mg/dL), originally established in Japanese cohorts[1], may fluctuate in sensitivity and specificity when applied to populations with different genetic backgrounds or lower baseline IgG4 levels. Therefore, validating novel biomarkers across multi-ethnic cohorts is essential to establish globally applicable thresholds.

The most critical clinical challenge in managing IgG4-related digestive system disorders (IgG4-DSD) lies in distinguishing them from gastrointestinal and pancreatobiliary malignancies. Misdiagnosing IgG4-DSD as a malignancy frequently leads to unnecessary radical surgeries, whereas a delayed or missed diagnosis postpones the timely administration of highly effective steroid therapies and crucial longitudinal follow-up. To address these diagnostic bottlenecks, significant strides have recently been made in endoscopic imaging and artificial intelligence (AI). Specifically, recent pioneering studies have demonstrated that integrating deep learning with advanced spectral visualization—such as emulating hyperspectral and narrow-band imaging in wireless capsule endoscopy—substantially enhances the early detection and precise classification of gastrointestinal disorders[6-8].

However, despite these remarkable technological leaps in morphological and optical assessment, a critical clinical gap remains. Current AI-assisted imaging workflows, while highly sensitive to structural and mucosal abnormalities, cannot fully capture the complex underlying immunological activity, molecular heterogeneity, or fibrotic staging that uniquely define IgG4-DSD. Consequently, relying solely on advanced imaging is often insufficient for a definitive diagnosis, particularly in seronegative or atypical presentations. This review explicitly addresses this gap by comprehensively mapping the emerging multidimensional biomarker landscape of IgG4-DSD. By delineating how novel molecular and cellular markers can be integrated as next-step tools alongside cutting-edge diagnostic workflows, this evaluation provides physicians with a distinctive decision-making advantage: Enabling the robust exclusion of malignancy, facilitating the safe, earlier initiation of targeted steroid therapy, and optimizing long-term disease monitoring.

To provide a rigorous synthesis of current evidence, we performed a comprehensive literature search to identify relevant studies discussing the clinical characteristics and biomarkers of IgG4-DSD. Major electronic databases, including PubMed/MEDLINE, EMBASE, and Web of Science, were searched from inception to November 2025. The search strategy employed a combination of Medical Subject Headings and free-text terms. Key search terms included “IgG4-related disease”, “autoimmune pancreatitis”, “IgG4-related sclerosing cholangitis”, “pathogenesis”, “biomarkers”, and “diagnosis”. We prioritized original articles, systematic reviews, and meta-analyses published in the English language. Inclusion criteria focused on studies providing specific data on diagnostic markers, pathogenesis, or clinical manifestations of IgG4-RD involving the gastrointestinal tract, pancreas, or biliary system. We excluded conference abstracts without full text and non-English publications. Two authors independently screened the titles and abstracts to ensure the relevance and quality of the selected literature, with the graded evidence further synthesized in the following sections.

CLINICAL FEATURES OF IGG4-RELATED DIGESTIVE SYSTEM DISORDERS

IgG4-related sialadenitis

IgG4-related sialadenitis (IgG4-RS) predominantly affects middle-aged males, with a prolonged disease course. Clinically, it presents as unilateral or bilateral submandibular gland swelling, with firm consistency and clear demarcation[9]. Notably, the underlying skin color remains normal[10]. Although localized, IgG4-RS often serves as a diagnostic clue for underlying pancreatic involvement. Differential diagnosis from Sjögren’s syndrome is necessary, as it is characterized by dry mouth, dry eyes, glandular destruction, and elevated antinuclear antibody levels[11]. Pathologically, approximately 70% of infiltrating CD4⁺ T cells are follicular helper T cells[12]. When minor salivary glands are involved, storiform fibrosis and obliterative vasculitis are not apparent[13], which can confound the diagnosis. Fine-needle aspiration cytology cannot adequately assess tissue architecture, often necessitating a core needle or excisional biopsy to confirm the full diagnostic triad.

IgG4-related esophagitis

In 2010, Lopes et al[14] reported the first case of IgG4-related esophagitis, which led to unnecessary surgery due to ineffective pharmacotherapy; pathological examination confirmed the diagnosis. By 2020, Padniewski et al[15] had reviewed 17 cases in the literature. The disease, albeit rare, manifests as IgG4-RD involving the esophagus, with symptoms such as dysphagia, odynophagia, and weight loss. Persistent refractory reflux and dysphagia warrant consideration of esophageal involvement by IgG4-RD[16]. Endoscopic findings may include masses, strictures, or ulcers. Histology often reveals IgG4-positive plasma cell infiltration and significant fibrosis. These presentations can easily be mistaken for esophageal malignancies or reflux esophagitis. Superficial endoscopic biopsies must be deep enough to capture the characteristic subepithelial fibrosis to provide a definitive diagnosis.

IgG4-related gastric disease

IgG4-RD involving the stomach most commonly affects the gastric body, followed by the antrum, fundus, and pylorus, with initial nonspecific symptoms such as abdominal pain, weight loss, and anemia[17]. CT findings may demonstrate isolated gastric lesions, including masses, focal wall thickening, or ulcers[18], which differ from IgG4-RD in other locations[19]. A multicenter study in 2025 found that IgG4-related gastropathy exhibits extensive plasma cell infiltration in the mucosal lamina propria. This differs from Helicobacter pylori-induced gastritis, which, while also presenting with substantial plasma cell infiltration in the mucosa, confines this infiltration solely to the mucosal surface (Figure 1). However, a critical diagnostic challenge is the “overlapping characteristics” with mucosa-associated lymphoid tissue lymphoma. Both conditions exhibit dense lymphoplasmacytic infiltration, and clinicians must utilize immunohistochemistry and flow cytometry to differentiate the polyclonal nature of IgG4-RD from the monoclonal expansion of B-cells in lymphoma to avoid catastrophic misdiagnosis. Diagnosis is challenging due to nonspecific clinical presentations and imaging findings, but endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) may improve the diagnostic yield[20].

Figure 1 Presents differences in the location of plasma cell infiltration between gastritis caused by Helicobacter pylori and immunoglobulin G4-related gastrointestinal disease. IgG4: Immunoglobulin G4.

IgG4-associated autoimmune hepatitis

IgG4-associated autoimmune hepatitis (IgG4-AIH) is a complication of IgG4-RD involving the pancreas or bile ducts, leading to liver damage, including interlobular bile duct injury[21]. Clinically and pathologically, IgG4-AIH resembles classical AIH, but it responds more quickly and effectively to glucocorticoid therapy[22]. IgG4-AIH generally satisfies classical AIH diagnostic criteria, coupled with elevated serum IgG4 levels and IgG4-positive plasma cell infiltration. Hepatic IgG4 levels may serve as post-therapeutic markers of efficacy[23]. Cases of bile duct injury in IgG4-AIH call for larger cohort studies to verify associations between IgG4-related sclerosing cholangitis (IgG4-SC) bile duct stricture and interlobular bile duct damage[24]. Liver biopsies may capture interface hepatitis but frequently miss deeper diagnostic hallmarks, requiring careful synthesis with biliary imaging and systemic serology.

IgG4-SC

IgG4-SC refers to IgG4-RD involving the biliary tract, characterized by inflammatory thickening of the bile duct wall, ductal strictures, IgG4-positive plasma cell infiltration, and fibrosis, extending to the gallbladder wall. Approximately 90% coexist with type 1 AIP (AIP-1) or IgG4-related pancreatitis[25,26]. Symptoms include obstructive jaundice, upper abdominal discomfort, and weight loss[27].

Crucially, differentiating IgG4-SC from primary sclerosing cholangitis (PSC) is essential. Radiologically, IgG4-SC typically presents with long, continuous, and symmetrical strictures (often in the lower common bile duct), whereas PSC is characterized by short, band-like, and “beaded” strictures affecting both intrahepatic and extrahepatic ducts[28]. Furthermore, PSC is commonly associated with inflammatory bowel disease (IBD) and occurs in a younger demographic[29]. A “pathological mimicry” exists, as some pancreatobiliary malignancies can trigger a prominent IgG4-positive plasma cell infiltrate in the peritumoral stroma, potentially misleading a diagnosis if based solely on IgG4 cell counts. Diagnosis relies on a multimodal approach. While biopsy of the major duodenal papilla is the preferred histological method for patients with concurrent AIP, endoscopic retrograde cholangiopancreatography with intraductal biopsy is warranted in indeterminate cases to exclude cholangiocarcinoma[30]. However, clinicians must be wary of “biopsy pitfalls”. Current consensus guidelines emphasize that while endobiliary brush cytology is a standard procedure for evaluating biliary strictures, its primary utility is in ruling out cholangiocarcinoma. For IgG4-SC, brush cytology and forceps biopsy often yield superficial mucosal tissue, which may lack the characteristic storiform fibrosis or obliterative vasculitis found in deeper layers, potentially leading to false-negative results. For relapsing or refractory cases, B-cell depletion therapy using rituximab (anti-CD20) has proven effective. Recently, novel agents such as inebilizumab, a CD19^-targeting monoclonal antibody, are under investigation to broadly deplete B-cell lineages (including plasmablasts) and inhibit IgG4-related autoimmune responses more accurately[26].

IgG4-related AIP

AIP is categorized into two distinct subtypes: AIP-1 and type 2 AIP (AIP-2), with AIP-2 being more localized and potentially coexisting with other autoimmune disorders, particularly IBD. In contrast, AIP-1 is the pancreatic manifestation of IgG4-RD[31], often affecting individuals around 60 years old, predominantly males, initially presenting with obstructive jaundice[27]. Although pancreatic inflammation is present, IgG4-related AIP is usually painless[32], with jaundice resulting from biliary strictures. Chronic fibrotic-inflammatory pancreatic responses may impair islet cell function, with 33%-78% of patients typically developing diabetes[33]. Radiologically, a normal pancreas appears “feather-like” on CT/MRI, whereas IgG4-RD involvement yields a smooth, diffuse, or focal enlargement termed a “sausage-like” appearance[34]. Differentiating AIP from pancreatic cancer (PC) is critical. PC often shows low-density contrast enhancement due to hypovascularity and the “double duct sign”, reflecting concurrent dilation of the pancreatic and common bile ducts.

As summarized in Table 1, the differentiation between AIP and PC extends beyond simple radiological findings. Diagnostic uncertainty remains a significant hurdle. According to the International Consensus Diagnostic Criteria and large multicenter cohorts, up to 15%-30% of patients with AIP-1 remain seronegative for serum IgG4. Conversely, based on international clinical guidelines and extensive cohort analyses, approximately 7%-10% of patients with pancreatic ductal adenocarcinoma (PDAC) may exhibit elevated serum IgG4 levels, frequently leading to misdiagnosis, and a subset of PC tissues can meet the histological cutoff for IgG4-positive cells per high-power field.

Table 1 Differential diagnosis and diagnostic pitfalls: Type 1 autoimmune pancreatitis vs pancreatic cancer.

Feature	AIP-1	PC	Diagnostic pitfalls & uncertainty
Clinical presentation	Often painless obstructive jaundice; multi-organ involvement	Painless jaundice; weight loss; rapid progression	Overlap: Both can present with “painless jaundice” and weight loss in elderly patients
Serum IgG4	Significantly elevated (> 2 × ULN is highly suggestive)	Usually normal	Pitfall: Approximately 10% of PC patients show mild IgG4 elevation. Normal IgG4 does not rule out AIP (seronegative cases)
Imaging (CT/MRI)	“Sausage-like” enlargement; delayed enhancement; capsule-like rim	Focal mass; hypo-vascular (low-density) enhancement	Uncertainty: Atypical mass-forming AIP can perfectly mimic the focal appearance of PC
Ductal signs	Long or multiple strictures without upstream dilation	“Double duct sign” (abrupt cutoff with upstream dilation)	Trap: AIP can occasionally cause distal ductal dilation, simulating malignancy-induced obstruction
Histology (biopsy)	Dense lymphoplasmacytic infiltrate; storiform fibrosis; obliterative vasculitis	Atypical cells; disorganized glandular structure (adenocarcinoma)	Biopsy pitfall: EUS-FNA/FNB samples are small; “storiform fibrosis” is often missed. False positive: Peritumoral stroma in PC can show high IgG4+ cell counts
Treatment response	Rapid radiological resolution with steroids	No response to steroids (disease progresses)	Risk: A “steroid trial” should only be performed after rigorous exclusion of malignancy to avoid delaying surgery

AIP: Autoimmune pancreatitis; PC: Pancreatic cancer; IgG4: Immunoglobulin G4; ULN: Upper limit of normal; CT: Computed tomography; MRI: Magnetic resonance imaging; EUS-FNA/FNB: Endoscopic ultrasound-guided fine-needle aspiration/fine-needle biopsy.

Endoscopic ultrasound-guided biopsy is essential for diagnosing AIP and excluding PC, as PC typically lacks the systemic characteristics of IgG4-RD. However, the “gold standard” of histology is often limited by the small size of EUS-FNA samples, which may not capture the focal distribution of storiform fibrosis. Therefore, a diagnosis should never rely on a single pathological metric but must be a synthesis of clinical, serological, and radiological evidence, often requiring a “wait-and-see”, carefully monitored diagnostic steroid trial in highly selected cases.

IgG4-related mesenteritis

IgG4-RD involving the mesentery represents a rare chronic condition characterized by sclerosing mesenteritis, with localized or diffuse fibrosis and inflammation of the small bowel mesentery. Patients commonly experience nonspecific abdominal discomfort, such as pain, distention, diarrhea, and sometimes ischemia. Studies indicate that IgG4-RD-diagnosed females more frequently exhibit lacrimal and submandibular gland swelling, whereas males show increased digestive system symptoms, including abdominal pain, nausea, vomiting, and jaundice[35]. Imaging studies reveal soft-tissue masses encasing mesenteric vessels due to fibrosis and inflammation. It is often misdiagnosed as neoplasms[36,37] due to its aggressive radiological appearance. Minimally invasive biopsies are notoriously difficult to obtain from the mesentery, often necessitating a surgical biopsy for a definitive diagnosis.

The “great masquerader”: Navigating diagnostic uncertainty and biopsy pitfalls

The clinical reality of IgG4-DSD is characterized by frequent diagnostic overlaps with malignancy. The reliance on pathological “cutoffs” (e.g., > 10 IgG4⁺ cells/HPF) is often insufficient because these markers occur in various inflammatory and neoplastic environments. High-quality diagnosis requires recognizing the limitations of small-core biopsies, which may fail to show the diagnostic triad of IgG4-RD. In cases of persistent uncertainty, clinical pathways should emphasize longitudinal monitoring and, where appropriate, the “diagnostic steroid trial”, provided that malignancy has been rigorously screened via multimodal imaging and multidisciplinary discussion.

BIOMARKERS FOR IGG4-RD WITH A FOCUS ON DIGESTIVE MANIFESTATIONS

While IgG4-RD is a systemic condition, the clinical priority for gastroenterologists lies in distinguishing digestive manifestations, including AIP and IgG4-SC, from pancreatobiliary malignancies. This section synthesizes the relevance of emerging biomarkers within these digestive clinical pathways.

Circulating plasmablasts: A key tool for pancreatobiliary differentiation

Circulating plasmablasts have emerged as dynamic indicators of disease activity within the digestive tract. Studies utilizing flow cytometry, such as those by Lin et al[38], have consistently demonstrated expanded subsets in untreated IgG4-RD patients (n = 42). The levels of this subset positively correlated with serum IgG4 levels, number of affected organs, and IgG4-RD response index, and decreased following glucocorticoid therapy. Ciccocioppo et al[39] studied 19 patients with AIP-1, 10 with type 2 or other AIP, 17 with PC, chronic pancreatitis (n = 20), and intraductal papillary mucinous neoplasms or chronic asymptomatic hyperlipidemia (n = 21). Peripheral blood total plasma cell counts and CD45⁺CD19⁺CD38^hiCD20^-CD24^-CD27⁺IgG4⁺ cell counts were measured by flow cytometry. Results showed that total plasma cell quantification at a cutoff of 4500 cells/mL could distinguish AIP-1 from all other pancreatic diseases with a sensitivity of 47% and specificity of 81%. At a cutoff of 210 IgG4⁺ cells/mL, sensitivity was 80%, and specificity was 97% (area under the curve = 0.879). Plasma cell counts were significantly higher in patients with ≥ 3 organ involvement compared to those with ≤ 2 organ involvement (7370/mL vs 3435/mL, P = 0.01). In 12 patients treated with RTX for 3-6 months, the median plasma cell count decreased from 6356 to 1419/mL (P < 0.01), but serum IgG4 levels did not decrease significantly (P = 0.12). The IgG4⁺ plasma cell count is considered a potentially useful biomarker for distinguishing type 1 and type 2/NOS AIP from other pancreatic diseases. In active IgG4-RD patients, peripheral plasmablasts remain significantly elevated even with normal serum IgG4 levels and may rise again before clinical relapse, serving as a diagnostic marker independent of serum IgG4. Therefore, detecting peripheral plasma cells in patients suspected of IgG4-RD can serve as a marker, helping to avoid unnecessary major surgeries such as Whipple procedures.

Immune cells

Tph: Given their established role in sustaining B-cell activation (section: T cell differentiation and cytokine signaling), circulating Tph cells may outperform serum IgG4 as indicators of disease activity. Ji et al[4] identified that the frequency of circulating Tph cells (CD4⁺CXCR5^-PD-1^hi) is markedly expanded in patients with active, untreated IgG4-RD. Importantly, Tph cell counts correlate more strongly with the IgG4-RD responder index (RI) and the number of involved organs than serum IgG4 levels do. Furthermore, Tph levels significantly decrease following remission, suggesting that quantifying circulating Tph cells could serve as a superior indicator for monitoring disease flares and treatment response compared to traditional serological markers. Given that the RI used in Ji et al’s study[4] heavily weighs pancreatic and biliary involvement, these findings translate directly into a robust tool for monitoring the inflammatory status of the digestive organs.

Follicular regulatory T cells: Ito et al[40] utilized flow cytometry to characterize follicular regulatory T cells (Tfr), T follicular helper (Tfh), and B cells in co-cultured tonsil or submandibular gland tissue samples from 49 IgG4-RD patients and 53 healthy controls. Real-time quantitative polymerase chain reaction (PCR) was used to measure interleukin (IL)-10, transforming growth factor β (TGF-β), and cytotoxic T lymphocyte antigen-4 expression. Tfr cells were significantly increased in IgG4-RD patients, and the proportion of CD25⁺CD127^-Tfr cells in peripheral blood correlated positively with serum IgG4 levels and the number of affected organs. This marker holds potential for monitoring residual inflammation in the pancreatic parenchyma during maintenance therapy, although further validation in isolated pancreatobiliary cohorts is needed.

Cytokines

In the management of digestive IgG4-RD, cytokines serve as more than just mechanistic players; they are functional tools for the clinician to bridge the gap between indeterminate imaging and definitive diagnosis. This section focuses on cytokines that address the critical needs of distinguishing IgG4-DSD from malignancy and monitoring therapeutic response.

Thymus and activation-regulated chemokine: Serum thymus and activation-regulated chemokine (TARC) (CCL17) serves as a systemic indicator of multiorgan inflammatory burden characteristic of IgG4-RD. In a cohort of 29 treatment-naïve patients (21 males, 8 females), Umeda et al[41] observed that serum TARC concentrations were significantly higher in IgG4-RD (486 pg/mL) compared to healthy controls (254 pg/mL). For the gastroenterologist, the value of TARC lies in its strong correlation with the number of involved organs and the IgG4-RD RI (r = 0.41, P = 0.03). Because AIP-1 and IgG4-SC are typically manifestations of a systemic disease, whereas pancreatobiliary malignancies are predominantly localized, a significantly elevated TARC level provides a systemic “red flag” supporting an IgG4-related etiology. Crucially, TARC levels are independent of serum IgG4. In cases of “seronegative” AIP where imaging is suspicious, but serology is normal, elevated TARC can offer vital evidence to avoid unnecessary surgical resection for suspected cancer.

Interferon-I and IL-33: Building on its established pathogenic role (section: Innate immune activation and antigen presentation), the clinical utility of the interferon (IFN)-α/IL-33 axis lies in monitoring therapeutic response. Minaga et al[42] demonstrated that serum concentrations of these cytokines decrease rapidly following prednisolone treatment, mirroring the radiological resolution of pancreatic swelling and biliary strictures. For patients with indeterminate pancreatic masses, a rapid decline in IFN-α and IL-33 levels following a steroid trial can confirm a biological response to therapy. This is particularly valuable in seronegative phenotypes, providing a non-invasive confirmation of the diagnosis and guiding the clinician in the decision to begin steroid tapering[42,43].

Eotaxin-3/CCL26: Eotaxin-3/CCL26 has recently been identified via high-throughput proteomics as a novel serum biomarker specifically associated with IgG4-RD lymphadenopathy[44]. This association is clinically relevant because lymphadenopathy represents a high-activity/high-relapse phenotype in IgG4-RD. Patients in the lymph node-positive group exhibited the IgG4 level, IgG4/IgG ratio, and eosinophil count were significantly elevated (P < 0.001); they exhibited a greater number of organ involvement sites (mean 4.6 vs 2.5, P < 0.001); and demonstrated a higher recurrence rate with monotherapy corticosteroids (57% vs 27%, P = 0.03). High expression persisted in patients with fewer organs involved but with lymphadenopathy, suggesting a stronger association with lymph node inflammation than with widespread organ involvement. This study provides the first systematic evidence that “lymphadenopathy” represents a high-activity, high-relapse phenotype in IgG4-RD, independent of organ involvement. For the gastroenterologist, identifying this high-relapse phenotype via Eotaxin-3 can guide the early introduction of steroid-sparing agents, preserving long-term pancreatic exocrine function and preventing biliary cirrhosis.

IgG4/IgG RNA ratio

The IgG4/IgG RNA ratio has garnered significant attention as a potential molecular biomarker for diagnosing IgG4-RD, yet its clinical application remains controversial with notable regional variations. de Vries et al[45] reported in 2020 that IgG4/IgG RNA testing does not effectively diagnose or differentiate IgG4-RD from primary biliary cholangitis or malignant tumors. Schulte et al[46] reported in 2020 that, considering factors such as molecular technology stability, their study employed multiplex digital droplet PCR to simultaneously detect IgG4 and total IgG RNA levels in IgG4-RD patients. Analysis indicated that the multiplex digital droplet IgG4/IgG RNA ratio showed no significant advantage over conventional quantitative PCR and might even yield false positives. Serum IgG4 levels outperformed the IgG4/IgG RNA ratio in differentiating IgG4-RD. When distinguishing malignant from benign non-IgG4-RD patients, serum IgG4 levels demonstrated a lower false-positive rate than the IgG4/IgG RNA ratio. The study concluded that the IgG4/IgG RNA ratio may have limited clinical utility in the diagnosis and management of IgG4-RD.

IgG subclasses

Up to 20%-30% (some cohorts report up to 40%) of patients with AIP-1 remain seronegative; analysis of IgG subclasses and laboratory artifacts is critical to avoid misdiagnosis. Chan et al[47] observed that serum IgG2 levels (cutoff > 5.3 g/L) are significantly elevated in IgG4-RD, even when IgG4 is normal. In the context of indeterminate biliary strictures (IgG4-SC), measuring IgG2 can help confirm an IgG subclass-related etiology, potentially sparing the patient from invasive hepatic resection. Large-scale analysis indicates that while mild IgG4 elevations occur in various conditions, levels exceeding 5 × the upper limit of normal (ULN) have a positive predictive value of 75.4% for confirmed IgG4-RD[48]. A vital clinical consideration for gastroenterologists is the “prozone effect”, where extremely high antigen concentrations lead to falsely low serum IgG4 readings[49]. If clinical and imaging features (e.g., “sausage-like” pancreas) strongly suggest AIP but serology is normal, clinicians must request laboratory dilution to rule out this technical error.

Other biomarkers

HE4: Yan et al[5] proposed HE4 as a novel biomarker for fibrosis and long-term prognosis. Unlike inflammatory markers that fluctuate with steroid use, HE4 may help clinicians assess the degree of established biliary or pancreatic scarring. This is crucial for managing “burnt-out” IgG4-SC, where biliary strictures persist due to fibrosis rather than active inflammation, helping to avoid unnecessary prolonged steroid exposure.

Systemic immune-inflammation index: Karadeniz et al[50] evaluated systemic immune-inflammation index (SII), integrating platelet, neutrophil, and lymphocyte counts, in 30 IgG4-RD patients, 38 with granulomatosis with polyangiitis, 46 with sarcoidosis, and 27 controls. In digestive practice, SII may aid in differentiating AIP from other localized inflammatory conditions or atypical sarcoidosis. Incorporating SII into predictive models supports earlier diagnosis in patients presenting with non-specific abdominal symptoms before the onset of obstructive jaundice.

Critical appraisal and clinical applicability of emerging biomarkers

While numerous novel biomarkers have been identified, a critical appraisal of the current evidence reveals significant gaps between discovery and clinical application. First, most studies cited (e.g., on Tph cells and TARC) are single-center, retrospective investigations with relatively small sample sizes, limiting their statistical power and generalizability. For instance, the identification of Tph cells as a pathogenic subset relied on single-cell sequencing from only three patients[4], necessitating validation in larger, diverse cohorts. Second, external validation is largely missing. Except for serum IgG4 and circulating plasmablasts, few markers have been validated across different ethnic populations or independent cohorts. The reproducibility of the IgG4/IgG RNA ratio, for example, has been challenged by conflicting results and methodology-dependent variations[45,46]. Third, methodological standardization is lacking. Techniques like flow cytometry for plasmablasts are difficult to standardize across clinical laboratories compared to routine serological assays. Consequently, while markers like the IFN-α/IL-33 axis and HE4 show mechanistic promise, they remain in the exploratory phase. Currently, only serum IgG4 is considered an “established” marker, albeit with known specificity issues.

Real-world implementation and analytical challenges of emerging biomarkers

While emerging biomarkers such as circulating plasmablasts, Tph cells, and specific cytokines (TARC, IFN-α, IL-33) show immense potential for diagnosing and monitoring IgG4-DSD, their transition from research settings to routine gastrointestinal clinics requires overcoming significant analytical and logistical hurdles.

Assay availability and standardization: Currently, most cytokine and chemokine assays (e.g., TARC, IL-33) rely on enzyme-linked immunosorbent assay or multiplex bead-based technologies (Luminex). In many regions, these kits are restricted to (RUO) and lack FDA or CE-IVD clearance for in vitro diagnostics, limiting their immediate availability in standard hospital laboratories. For cellular markers, flow cytometry is the gold standard. However, the clinical utility of plasmablasts and Tph cells is frequently hampered by inter-laboratory variability. There is currently a lack of internationally harmonized gating strategies. For instance, while plasmablasts are generally defined as CD19⁺CD20^-CD27⁺CD38^hi, subtle differences in antibody clones, fluorochrome choices, and manual gating subjectivity can lead to significant variations in absolute counts and percentages across different centers.

Pre-analytical concerns: The viability of plasmablasts and Tph cells drops rapidly ex vivo. Whole blood samples must typically be processed within 12-24 hours of collection, requiring strict temperature control (ambient temperature) to prevent cellular degradation. Markers like IFN-α and IL-33 are susceptible to proteolytic degradation. Blood samples require prompt centrifugation, and the resulting serum or plasma must be immediately aliquoted and frozen at -80 °C. Repeated freeze-thaw cycles significantly compromise the reliability of the cytokine readouts, demanding a rigorous cold-chain transport system.

Feasible adoption pathways in gastrointestinal clinics: To bridge the gap between advanced immunology and everyday gastrointestinal practice, a “hub-and-spoke” laboratory model is recommended. In this framework, local GI clinics (the “spokes”) focus on initial screening using standard, widely available tests (e.g., serum IgG4, IgG2) and standardized pre-analytical sample collection. For highly complex tests requiring fresh cell processing or multiplex cytokine arrays, samples are expedited via dedicated transport networks to centralized, specialized reference laboratories (the “hubs”). Furthermore, establishing automated analysis pipelines and standardized external quality assessment programs among these central hubs will be essential to minimize inter-laboratory bias and validate specific clinical cutoff values before these biomarkers can be incorporated into formal diagnostic guidelines.

Translational integration in digestive practice: From biomarkers to clinical decisions

While IgG4-RD is a systemic condition, its gastrointestinal manifestations, including AIP and IgG4-SC, present unique diagnostic challenges. To better achieve the goal of translational integration, we propose a scenario-based integration strategy (summarized in Table 2) that directly correlates prevalent clinical scenarios (e.g., indeterminate pancreatic mass, isolated biliary stricture, suspected relapse, and fibrotic/burnt-out disease) with a prioritized sequence of diagnostic tests.

Table 2 Translational integration of diagnostic modalities across prevalent clinical scenarios.

Prevalent clinical scenario	Prioritized sequence of diagnostic tests	Expected interpretations & decision logic	Evidence level & basis
Indeterminate pancreatic mass	Imaging: CT/MRI	Imaging: Look for “sausage-like” enlargement, capsule-like rim	High (imaging, IgG4, EUS-FNB limitations: Based on ICDC & large multicenter cohorts). Moderate (Tph, plasmablasts: Requires broader validation)
	Serology: Serum IgG4, IgG2	Serology: IgG4 > 2 × ULN. Normal IgG4 does not rule out AIP (beware of the prozone effect)
	Cellular markers: Plasmablasts, Tph cells. Targeted biopsy: EUS-FNA/FNB	Cellular: Highly expanded plasmablasts/Tph cells support IgG4-RD
		Biopsy: Storiform fibrosis and lymphoplasmacytic infiltrate; standard EUS-FNA may miss focal fibrosis
Isolated biliary stricture	Imaging: MRCP/ERCP	Imaging: Long, continuous, symmetrical strictures (lower CBD)	High (imaging, cytology limitations: Supported by current consensus guidelines). Moderate (IgG2, plasmablasts)
	Serology: Serum IgG4, IgG2	Serology: Elevated IgG4 and IgG2 (> 5.3 g/L)
	Cellular markers: Plasmablasts	Cellular: Plasmablasts elevated independent of serum IgG4
	Targeted biopsy: Endobiliary brush cytology/forceps biopsy	Biopsy: Primary utility is ruling out cholangiocarcinoma. High false-negative rate for IgG4-SC as superficial brushings often miss deep-seated storiform fibrosis
Suspected relapse	Imaging: PET-CT/MRI	Imaging: Detects early subclinical organ recurrence	Moderate (plasmablasts & Tph predictive value; longitudinal cohorts). Low/exploratory (eotaxin-3, TARC)
	Serology: Eotaxin-3, TARC, IgG4	Serology: Eotaxin-3 identifies high-relapse lymphadenopathy phenotype; TARC evaluates systemic burden
	Cellular markers: Plasmablasts, Tph cells	Cellular: Significant rise in Plasmablasts/Tph cells before clinical or biochemical (LFTs) worsening
	Targeted biopsy: Usually deferred unless new atypical lesions appear	Biopsy: N/A
Fibrotic/“burnt-out” disease	Imaging: CT/MRI	Imaging: Persistent residual fibrotic masses/strictures without active swelling	Moderate (IFN-α/IL-33 axis response). Low (HE4: Based on exploratory translational studies)
	Serology: HE4, IFN-α/IL-33 axis	Serology: HE4 elevated (marker of established fibrosis); IFN-α/IL-33 low (no active inflammation)
	Cellular markers: Plasmablasts	Cellular: Normal or decreased post-therapy
	Targeted biopsy: Deep core biopsy (if needed)	Biopsy: Dense storiform fibrosis with sparse plasma cells. Logic: Differentiate residual scarring from active disease to prevent unnecessary prolonged steroid exposure

CT: Computed tomography; MRI: Magnetic resonance imaging; EUS-FNA/FNB: Endoscopic ultrasound-guided fine-needle aspiration/fine-needle biopsy; ICDC: International Consensus Diagnostic Criteria; Tph: Peripheral helper T cells; MRCP: Magnetic resonance cholangiopancreatography; ERCP: Endoscopic retrograde cholangiopancreatography; IgG4: Immunoglobulin G4; ULN: Upper limit of normal; AIP: Autoimmune pancreatitis; IgG4-RD: Immunoglobulin G4-related disease; TARC: Thymus and activation-regulated chemokine; IgG4-SC: Immunoglobulin G4-related sclerosing cholangitis; PET: Positron emission tomography; N/A: Not applicable; IFN: Interferon; IL: Interleukin; HE4: Human epididymis protein 4; LFTs: Liver function tests; CBD: Common bile duct.

First, regarding the differentiation of pancreatobiliary malignancy, distinguishing AIP-1 from PDAC or IgG4-SC from cholangiocarcinoma is the most critical clinical hurdle. Here, IgG4⁺ plasmablasts offer superior specificity compared to serum IgG4. When imaging reveals an indeterminate pancreatic mass or biliary stricture, elevated plasmablasts strongly support a benign IgG4-related etiology, potentially sparing patients from unnecessary Whipple procedures or hepatic resections. Second, in defining clinical pathways for “seronegative” digestive phenotypes, approximately 30%-40% of AIP type 1 patients present with normal serum IgG4. In this context, utilizing serum IgG2 or Tph cells serves as a vital safety net to reduce missed diagnoses in patients presenting with obstructive jaundice or pancreatic swelling. Finally, regarding fibrosis and prognosis in the digestive tract, chronic inflammation can lead to irreversible pancreatic atrophy or sclerosing cholangitis. Monitoring the IFN-α/IL-33 axis provides real-time data on the resolution of pancreatic inflammation, while HE4 may help clinicians assess residual biliary fibrosis, guiding the decision to taper steroids or introduce steroid-sparing agents to preserve exocrine/endocrine function. To further enhance the practicality and clinical utility of this article, we propose a straightforward algorithmic flowchart for pancreatobiliary differentiation (Figure 2). This step-by-step decision tree guides clinicians from the initial presentation of indeterminate masses or strictures through standard evaluations and translational biomarker testing. Crucially, it delineates the logic for excluding malignancy via targeted tissue acquisition before safely initiating a monitored diagnostic steroid trial.

Figure 2 Algorithmic flowchart for pancreatobiliary differentiation: Integrating translational biomarkers to exclude malignancy. IgG4: Immunoglobulin G4; CT: Computed tomography; MRI: Magnetic resonance imaging; ULN: Upper limit of normal; TARC: Thymus and activation-regulated chemokine; Tph: Peripheral helper T cells; IgG4-DSD: Immunoglobulin G4-related digestive system disorders; EUS-FNA/FNB: Endoscopic ultrasound-guided fine-needle aspiration/fine-needle biopsy; ERCP: Endoscopic retrograde cholangiopancreatography; MDT: Multidisciplinary team; IFN: Interferon; IL: Interleukin.

ADVANCES IN PATHOGENESIS RESEARCH

The pathogenesis of IgG4-RD involving the digestive system is a multifaceted process characterized by the breakdown of immune tolerance, aberrant innate immune activation, and a polarized adaptive immune response. This cascade ultimately leads to the hallmark pathological findings: Lymphoplasmacytic infiltration and storiform fibrosis.

Innate immune activation and antigen presentation

The initiation of IgG4-RD involves a complex interplay between innate and adaptive immunity. M2-polarized macrophages play a pivotal role in sustaining chronic inflammation and fibrosis. Studies have shown that IL-13 regulates macrophage activation toward the M2 phenotype, a mechanism validated in LATY136F knock-in mice[51]. These activated M2 macrophages secrete pro-fibrotic factors such as TGF-β and PDGF[52] and can be induced via the toll-like receptor 7 signaling, which contributes to IL-33 production[53]. Recent advances highlight the IFN-α/IL-33 axis. Minaga et al[42,54] demonstrated that plasmacytoid dendritic cells (pDCs) produce IFN-α and IL-33, which mediate chronic inflammation and fibrosis in AIP. Depletion of pDCs or inhibition of this axis significantly delayed disease development in experimental models.

T cell differentiation and cytokine signaling

CD4⁺ T cells are the central orchestrators of the immune response. Upregulated Th2 cytokines are a hallmark of the disease. IL-4 participates in the STAT6 signaling pathway to drive IgG4 class switching[55]. Tfh cells secrete IL-21, which activates the JAK-STAT3 pathway, a critical signal for B cell differentiation and immunoglobulin class switching[56]. Furthermore, Ji et al[4] identified a novel pathogenic subset of peripheral helper T (Tph) cells (CD4⁺CXCR5^-PD-1^hi) that express high levels of CXCL13 and IL-21. Tph cells amplify local B-cell responses via CXCL13 and IL-21. IL-10, primarily secreted by regulatory T cells (Tregs) and Tfr, plays a dual role: It suppresses inflammation but also paradoxically promotes IgG4 production[57]. Ito et al[40] found that Tfr cells are significantly increased in IgG4-RD and correlate with organ involvement. Additionally, IL-35 may exert anti-inflammatory effects by regulating Tregs[58].

B cell expansion and class switching

The hallmark of IgG4-RD is the oligoclonal expansion of B cells and their differentiation into antibody-secreting plasmablasts. Under the influence of IL-4 (Th2) and IL-21 (Tfh/Tph), B cells undergo class-switch recombination to produce IgG4[55,56]. IL-10 supports IgG4-producing plasma cell differentiation[57]. Although IgG4 antibodies themselves are generally non-inflammatory, their overproduction reflects the intense underlying immunological activity driven by these cytokine networks.

Fibrosis and chemokine networks

The characteristic “storiform fibrosis” is driven by the activation of fibroblasts and myofibroblasts through multiple pathways. Activation of the IL-33/ST2 axis promotes Treg expansion and IL-13 secretion, further driving fibrosis[59]. IL-33 also acts directly on mast cells and Th2 cells to propagate type 2 immune responses[60]. TARC (CCL17) may contribute to immune cell aggregation at digestive lesion sites[41,61]. The CCL18-CCR8 axis is also markedly upregulated, with CCL18 inducing chemotaxis and fibrosis; blocking this axis attenuates inflammation in animal models[58]. SDF-1 (CXCL12) and its receptors (CXCR4/7) are elevated in lesion tissues, potentially participating in angiogenesis and fibrosis[62]. Ultimately, TGF-β secreted by M2 macrophages serves as a key driver of collagen deposition[52]. A schematic of immune circuits driving fibrosis in IgG4-DSD is shown in Figure 3.

Figure 3 Schematic diagram of the immunopathogenic cascade in immunoglobulin G4-related digestive system disorders. The inflammatory process is initiated by antigen presentation involving M2-polarized macrophages and plasmacytoid dendritic cells (pDCs). Activated pDCs secrete interferon-α and interleukin (IL)-33, establishing a pro-fibrotic microenvironment. Naïve CD4⁺ T cells differentiate into distinct pathogenic subsets: Th2 cells (producing IL-4/IL-13), T follicular helper cells (producing IL-21), and the recently identified peripheral helper T cells (CD4⁺CXCR5^-PD-1^hi). These T cell subsets promote the massive expansion of B cells and their class-switching to IgG4-secreting plasma cells via IL-4 and IL-21 signaling. Finally, storiform fibrosis is orchestrated by transforming growth factor-β (secreted by M2 macrophages) and the IL-33/ST2 axis, which activates fibroblasts and mast cells. Chemokines such as thymus and activation-regulated chemokine (CCL17) and CCL18 further recruit lymphocytes, perpetuating the cycle of chronic inflammation. IgG4: Immunoglobulin G4; IgG4-RD: Immunoglobulin G4-related disease; TARC: Thymus and activation-regulated chemokine; IL: Interleukin; TGF-β: Transforming growth factor β; Treg: Regulatory T cells.

CONCLUSION

IgG4-DSD represents a complex fibro-inflammatory spectrum where accurate diagnosis remains heavily reliant on multidisciplinary integration to rule out malignancy. While recent advancements in biomarker discovery are promising, the ultimate request for multi-ethnic prospective validation is critical. To better align with the discovery-to-clinic continuum, the field must systematically advance emerging markers (e.g., circulating plasmablasts and Tph cells) from their current “validation” phases into formal clinical guidelines, while guiding “exploratory” markers (e.g., HE4, TARC, and Eotaxin-3) through rigorous multi-center testing.

Achieving this clinical translation requires a structured and highly detailed study strategy. We propose that future research must adhere to standardized minimal reporting criteria: Precise cohort phenotype definitions that explicitly distinguish between isolated AIP-1, IgG4-SC, and multi-organ systemic involvement; ethnically adjusted cutoff selections, utilizing multi-level thresholds (e.g., > 2 × ULN and > 5 × ULN) to balance sensitivity and specificity; and transparent assay methodologies, such as standardized flow cytometry gating strategies and serological dilution protocols to rule out the prozone effect.

Furthermore, we recommend prioritizing specific study designs to bridge current evidentiary gaps. These include prospective diagnostic accuracy studies with external validation for patients presenting with indeterminate pancreatic masses or biliary strictures; head-to-head comparisons between novel cellular/cytokine markers and conventional serum IgG4 to demonstrate incremental diagnostic yield in seronegative cohorts; and longitudinal outcome studies to track markers associated with impending clinical relapse or irreversible fibrosis. Ultimately, adopting these rigorous reporting standards and study designs will successfully bridge the discovery-to-clinic continuum, transforming the management of IgG4-DSD from empirical steroid trials to precision-driven, personalized care.

ACKNOWLEDGEMENTS

We are grateful for Professor Guan-Liang Cheng's help and guidance.