Citrullinated matrisome: An emerging stromal determinant in hepatocellular carcinoma prognostication

Abstract

Accurate prognostic stratification of hepatocellular carcinoma (HCC) remains a significant clinical challenge. Although the TNM staging system and serum alpha-fetoprotein levels provide a foundational framework for risk assessment, there is an urgent need for biomarkers that reflect the intrinsic biological aggressiveness of both the tumor and its microenvironment. In this editorial, we discuss the pivotal study published in the World Journal of Gastrointestinal Oncology by Cho et al, which identifies elevated expression of citrullinated glial fibrillary acidic protein (cit-GFAP) as a novel and independent prognostic marker associated with reduced overall survival (OS) following curative hepatic resection. We situate these findings within the broader context of HCC biology, highlighting the critical contributions of the tumor microenvironment and post-translational modifications. The study successfully bridges stromal biology with protein citrullination, demonstrating that high cit-GFAP expression is significantly correlated with increased mortality, independent of conventional prognostic factors such as tumor size. We examine the biological plausibility of this association, considering the role of glial fibrillary acidic protein in hepatic stellate cell activation and the growing body of evidence linking citrullination to extracellular matrix remodeling and cancer progression. Additionally, we offer a critical evaluation of the study’s implications, acknowledge its limitations—particularly its retrospective design and the observed discrepancy between OS and recurrence-free survival—and propose a forward-looking research agenda focused on biomarker validation and translational development. The identification of cit-GFAP underscores the matrisome as a promising reservoir of prognostic biomarkers and potential therapeutic targets, thereby advancing the mechanistic understanding of HCC pathogenesis.

Key Words: Hepatocellular carcinoma; Tumor microenvironment; Citrullination; Glial fibrillary acidic protein; Biomarker; Prognosis; Hepatic stellate cells; Extracellular matrix

Core Tip: This editorial highlights the groundbreaking study by Cho et al that identifies citrullinated glial fibrillary acidic protein (cit-GFAP) as a novel stromal biomarker in hepatocellular carcinoma (HCC). The study shifts the prognostic focus from the tumor cell per se to the post-translationally modified extracellular matrix, demonstrating that elevated cit-GFAP expression is an independent predictor of poor survival following curative resection. We discuss how these findings underscore the critical role of post-translational modifications within the tumor microenvironment and propose a future research agenda for the validation of cit-GFAP and the exploration of peptidylarginine deiminase inhibition as a potential therapeutic strategy in HCC.

This editorial refers to "Clinical significance of citrullinated glial fibrillary acidic protein in predicting outcomes in hepatocellular carcinoma" by Cho et al, 2025; https://doi.org/10.4251/wjgo.v17.i12.110877.

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INTRODUCTION

Hepatocellular carcinoma (HCC) constitutes a significant global health burden, ranking as the sixth most commonly diagnosed cancer and the fourth leading cause of cancer-related mortality worldwide[1]. Its pathogenesis is uniquely associated with underlying chronic liver disease, typically characterized by persistent inflammation, fibrosis, and cirrhosis due to hepatitis B or C virus infection, alcohol misuse, or metabolic dysfunction-associated steatotic liver disease[2]. This distinct etiological background contributes substantially to the complexity of HCC management. Although the therapeutic landscape has been transformed by immune checkpoint inhibitors and molecularly targeted agents, clinical outcomes remain suboptimal for many patients, particularly those with advanced-stage disease[3,4]. Even among the 30%-40% of patients diagnosed at an early stage eligible for curative interventions such as resection or ablation, high recurrence rates—exceeding 50% within five years—significantly impair long-term survival[5].

These ongoing challenges related to tumor recurrence and poor survival outcomes have spurred intensive efforts to identify more robust prognostic tools. While the American Joint Committee on Cancer TNM staging system and the Barcelona Clinic Liver Cancer classification provide essential clinical frameworks, they often fail to fully capture the biological heterogeneity of HCC. Serum alpha-fetoprotein (AFP), despite its widespread clinical use, is limited by inadequate sensitivity and specificity[6]. Consequently, the oncology community has increasingly turned to the molecular architecture of the tumor and the tumor microenvironment (TME) in search of biomarkers that enable more precise risk stratification and improved prediction of treatment response[7,8].

It is against this backdrop that the study by Cho et al[9], published in World Journal of Gastrointestinal Oncology, offers a timely and significant contribution. The authors propose citrullinated glial fibrillary acidic protein (cit-GFAP) as a novel biomarker emerging from the underexplored intersection of stromal biology and specific post-translational modifications. This editorial discusses the implications of their findings, exploring how citrullination of a key stromal protein may enhance prognostic accuracy and open new avenues for understanding the biological mechanisms driving HCC progression.

THE TME AND HEPATIC STELLATE CELLS: BEYOND A PASSIVE BYSTANDER

The concept that HCC arises solely from the autonomous transformation of hepatocytes has been conclusively refuted. It is now well established that the tumor TME actively contributes to hepatocarcinogenesis, disease progression, and metastasis[10]. The HCC TME comprises a complex ecosystem consisting of cancer cells, immune cells, endothelial cells, cancer-associated fibroblasts (CAFs), and a dynamically remodeled extracellular matrix (ECM).

Within this reactive stromal compartment, hepatic stellate cells (HSCs) play a central and functionally critical role[11]. In response to chronic liver injury, quiescent HSCs—characterized by their vitamin A storage—undergo activation and differentiate into proliferative, fibrogenic, and contractile myofibroblasts. Activated HSCs are the primary cellular source of excessive ECM deposition, a hallmark of liver fibrosis and cirrhosis, which constitute the predominant pathological substrates for HCC development. Beyond their pivotal role in fibrogenesis, HSCs directly promote tumorigenesis by secreting a wide array of growth factors (e.g., transforming growth factor-β, platelet-derived growth factor), pro-angiogenic mediators (e.g., vascular endothelial growth factor), and inflammatory cytokines that collectively enhance cancer cell proliferation, invasion, and survival[12,13]. Emerging evidence also demonstrates that physical cues from the TME, such as increased matrix stiffness, can activate HSCs, which subsequently promote HCC cell migration via TGM2/ITGB1-mediated ECM remodeling and intercellular mitochondrial transfer, revealing a mechano-oncogenic axis in HSC function[14]. A substantial body of evidence consistently demonstrates that increased abundance of activated HSCs—commonly identified by alpha-smooth muscle actin (α-SMA) expression—in HCC tissues is strongly associated with adverse prognosis[15,16]. Moreover, recent studies have uncovered marked functional heterogeneity among HSC subpopulations, with distinct subpopulations exhibiting divergent roles in tumor promotion, ECM remodeling, and immune regulation, further refining our understanding of HSC biology in HCC[17].

It is within this pathophysiological context that glial fibrillary acidic protein (GFAP) emerges as a relevant biomarker. Although primarily recognized as a specific marker for astrocytes in the central nervous system, GFAP is also an established marker for quiescent and early-activated HSCs in the liver[18,19]. Its expression is considered indicative of the initial phase of HSCs activation in response to liver injury, potentially preceding the full myofibroblastic transition marked by α-SMA. Carotti et al[19] demonstrated that GFAP-positive HSCs are associated with early-stage fibrosis and vascular remodeling in recurrent hepatitis C following liver transplantation, suggesting a functional role in modifying the tissue microarchitecture—a process that may predispose to neoplastic transformation. Notably, comparative proteomic studies have revealed that GFAP expression is significantly upregulated—approximately six-fold—in HCC tissues compared with adjacent non-tumor liver, further implicating GFAP in hepatocarcinogenesis[20].

CITRULLINATION: REMODELING THE PROTEIN LANDSCAPE IN CANCER

Citrullination is a critical post-translational modification catalyzed by a family of calcium-dependent enzymes known as peptidylarginine deiminases (PADs)[21]. This process involves the hydrolytic deimination of arginine residues to generate citrulline, resulting in the loss of a positive charge. Although chemically subtle, this modification has substantial functional consequences: It can disrupt intra- and intermolecular hydrogen bonding, alter protein conformation, modulate protein–protein interactions, and influence the susceptibility of proteins to proteolytic degradation[22].

Initially studied extensively in the context of rheumatoid arthritis—where citrullinated proteins function as autoantigens—and neurodegenerative disorders, citrullination has recently emerged as a significant contributor to oncogenesis[23]. The “citrullinome” in cancer is highly diverse, encompassing nuclear proteins such as histones—which can regulate gene expression—cytoplasmic proteins, and, importantly, components of the ECM—collectively referred to as the “matrisome”.

Citrullination of ECM proteins, including fibronectin and collagen I, has been showed to profoundly alter cell–ECM interactions. For instance, citrullinated fibronectin enhances breast cancer cell invasion by disrupting integrin clustering and destabilizing focal adhesions, thereby promoting cellular motility[24]. In colorectal cancer, PAD4-mediated citrullination of the ECM has been identified as a key driver of liver metastasis, facilitating the formation of a pre-metastatic niche that supports tumor cell colonization and proliferation[25]. These findings establish a strong precedent: Citrullination serves as a potent mechanism for ECM remodeling that actively promotes tumor progression and dissemination.

These observations from other malignancies underscore protein citrullination, particularly within the ECM, as a pro-tumorigenic mechanism. However, it is essential to distinguish this general principle from the specific concept of a systematically modified “citrullinated matrisome” in HCC. To date, large-scale proteomic studies—such as those employing mass spectrometry with citrulline-specific enrichment—have not been conducted to comprehensively map the repertoire of citrullinated stromal proteins in HCC tissues. Therefore, while cit-GFAP and citrullinated fibronectin provide compelling evidence that such modifications occur and are biologically relevant, they represent isolated, non-systematic findings within what may be a much broader landscape. The term “citrullinated matrisome” in HCC should thus be interpreted as a heuristic framework—a hypothesis positing that the functional state of the stroma may be regulated by a coordinated program of citrullination—rather than a fully characterized biological entity. Its validation awaits future citrullinome-wide profiling efforts.

SYNTHESIZING THE CONCEPTS: THE RATIONALE FOR CIT-GFAP IN HCC

The study by Cho et al[9] is built upon an elegant integration of two key biological concepts: The central role of HSCs in the HCC tumor TME and the tumor-promoting potential of protein citrullination. GFAP is a known substrate for PADs, particularly PAD2, and its citrullinated form has been detected in pathological conditions such as Alzheimer’s disease[26]. Given the expression of GFAP in HSCs and the well-established contribution of these cells to HCC pathogenesis, the authors hypothesized that cit-GFAP may hold clinical significance.

Their prior research provided a critical foundation by demonstrating the accumulation of cit-GFAP in α-SMA-positive HSCs in a mouse model of bile duct ligation-induced liver fibrosis[27]. This finding established a direct link between HSC activation and GFAP citrullination in the liver. The logical next step—now realized in their current work—was to investigate the presence and prognostic relevance of cit-GFAP in human HCC tissues.

APPRAISING THE FINDINGS: STRENGTHS AND INTRIGUING NUANCES

The findings of Cho et al[9] reveal a novel and clinically relevant association that warrants careful evaluation. In their cohort of 169 patients with surgically resected HCC, high expression of cit-GFAP—observed in nearly half of the cases—was a strong and independent predictor of reduced overall survival (OS), with a hazard ratio of 2.753. This prognostic impact was comparable to that of large tumor size, a well-established adverse clinical factor. The strength of the study lies in its rigorous methodology, including the use of tissue microarrays, a previously validated antibody, independent pathological review, and appropriate multivariate statistical analysis adjusted for key clinicopathological variables.

Independent prognostic value

Notably, cit-GFAP retained statistical significance in the multivariate model, whereas AFP did not. This indicates that cit-GFAP provides prognostic information beyond conventional serological markers, suggesting its potential as a novel biomarker for refining risk stratification in HCC. These preliminary data positions cit-GFAP as a promising candidate for further evaluation, although its incremental value over established and emerging multiparameter models remains to be determined.

Survival-recurrence dissociation

A particularly intriguing observation is the discrepancy between the significant association with OS and the nonsignificant trend observed for recurrence-free survival (RFS). Although patients with high cit-GFAP expression consistently exhibited higher recurrence rates at 1 year, 3 years, and 5 years, the difference did not reach statistical significance (P = 0.125). The authors appropriately attribute this to limited statistical power due to sample size. However, this finding may also reflect more complex underlying biological mechanisms. It is plausible that cit-GFAP influences OS through pathways independent of tumor recurrence. This hypothesis is supported by established knowledge of HSCs biology. First, the association between HSCs activation or abundance and progressive hepatic functional decline is well documented; intratumoral HSCs are known to exacerbate fibrosis and parenchymal dysfunction, both of which can independently increase mortality risk of mortality[11,15]. Consequently, a cit-GFAP-rich stromal microenvironment may serve as a histological indicator of a functionally impaired liver remnant. Second, HSCs are potent regulators of the tumor immune microenvironment. They can recruit regulatory T cells and myeloid-derived suppressor cells and express immune checkpoint ligands, thereby contributing to the formation of an immunosuppressive niche that may impair anti-tumor immunity and potentially reduce the efficacy of immunotherapies[15,28]. Recent evidence has further demonstrated that activated HSCs shape the ECM-disorganized and immunosuppressive microenvironment via the CCL11/CCR3 axis in HCC, particularly under lenvatinib treatment, providing a more detailed molecular framework for HSC-mediated immune modulation[29]. While direct evidence linking cit-GFAP to these specific immunomodulatory functions remains lacking, its expression within the HSCs compartment renders such mechanisms biologically plausible and warrants further investigation. This dissociation highlights the need for validation in larger, prospective cohorts.

Nuclear localization

The predominantly nuclear localization of cit-GFAP, as reported, is unexpected and requires careful interpretation. Canonical GFAP is a cytoplasmic intermediate filament protein, and its presence in the nucleus is atypical. Citrullination-induced structural modifications may facilitate nuclear translocation, potentially enabling non-canonical functions such as modulation of gene transcription. While this observation could be incidental, it may also represent a novel mechanistic insight. In either case, confirmation through complementary techniques and further investigation are necessary to determine its functional relevance.

A CRITICAL APPRAISAL: STRENGTHS, LIMITATIONS, AND UNANSWERED QUESTIONS

While the findings of Cho et al[9] are undoubtedly novel and biologically plausible, a rigorous evaluation within the context of clinical-translational biomarker development necessitates a balanced assessment of both strengths and inherent limitations. The study’s retrospective, single-center design and modest sample size (n = 169) represent fundamental constraints. Although the cohort is well characterized, the absence of a reported sample size calculation or power analysis limits the ability to determine whether the study was adequately powered to detect the observed effect sizes—particularly for the secondary endpoint of RFS, where only a nonsignificant trend was observed. This may partially explain the notable discrepancy between the robust association with OS and the lack of statistical significance in recurrence outcomes.

The statistical modeling, while appropriate, warrants further scrutiny. The multivariate Cox regression identified cit-GFAP and tumor size as independent prognostic factors. However, several established prognostic variables commonly included in HCC resection models—such as microvascular invasion, surgical margin status (R0 vs R1), and receipt of adjuvant therapies—were not incorporated into the analysis. While this omission does not invalidate the observed cit-GFAP signal, it underscores the need to validate its truly independent prognostic value in future studies using more comprehensive multivariable models.

From a methodological perspective, the dichotomization of cit-GFAP expression using a 50% threshold based on staining intensity, although practical for initial analysis, is inherently arbitrary. The field would benefit from data-driven approaches—such as receiver operating characteristic (ROC) curve analysis relative to survival outcomes—in larger cohorts to establish an optimal and biologically meaningful cutoff. Moreover, the unexpected nuclear localization of cit-GFAP presents a compelling observation that challenges the conventional understanding of GFAP as a cytoplasmic intermediate filament protein. The study relies solely on immunohistochemistry for subcellular localization. Independent validation using complementary techniques—such as subcellular fractionation followed by Western blotting or immunofluorescence confocal microscopy—is essential to confirm this finding and exclude potential technical artifacts. If substantiated, this could open a novel avenue of research into potential non-canonical, intranuclear functions of citrullinated GFAP.

These limitations do not diminish the study’s pioneering contribution but rather delineate a clear path forward for subsequent research. They emphasize that the current work should be regarded as a hypothesis-generating discovery cohort. Its primary value lies in identifying cit-GFAP as a promising candidate biomarker, whose clinical utility and underlying biological mechanisms now require rigorous validation and further investigation. Table 1 provides a systematic comparison of cit-GFAP with other established stromal biomarkers in HCC, highlighting its unique features and current evidence level.

Table 1 Comparison of citrullinated glial fibrillary acidic protein with other stromal biomarkers in hepatocellular carcinoma.

Biomarker	Cellular source	Posttranslational modification	Prognostic value (HR)	Key feature	Current evidence level
α-SMA	Activated HSCs/CAFs	None	Approximately 1.5-2.5	Most widely used; marks myofibroblastic activation	Extensive validation
GFAP (total)	Quiescent/early HSCs	None	Not established	Marks early HSC activation; less prognostic	Limited
Desmin	HSCs	None	Approximately 1.6-2.0	HSC cytoskeletal marker	Moderate
cit-GFAP	Activated HSCs/CAFs	Citrullination	2.75	Posttranslational modification; independent of AFP	Hypothesisgenerating (single study)
Citrullinated fibronectin	ECM	Citrullination	Not reported in HCC	Promotes invasion in breast cancer	Preclinical only

HR: Hazard ratio; HSC: Hepatic stellate cell; α-SMA: Alpha-smooth muscle actin; CAF: Cancer-associated fibroblast; GFAP: Glial fibrillary acidic protein; cit-GFAP: Citrullinated glial fibrillary acidic protein; AFP: Alpha-fetoprotein; ECM: Extracellular matrix; HCC: Hepatocellular carcinoma.

OUR PERSPECTIVE: WHAT THIS STUDY TRULY ADDS—AND WHAT IT DOES NOT

From our perspective, the most valuable contribution of Cho et al[9] is conceptual rather than immediately clinical. The study convincingly shows that citrullination of a stromal protein (GFAP) can be detected in HCC tissues and correlates with patient survival. This finding shifts the paradigm from simply quantifying stromal cell abundance toward interrogating the functional state of the stroma via chemical modifications, opening a new line of inquiry into the “citrullinated matrisome” as a determinant of HCC progression.

In our positive evaluation, the study has notable strengths: It addresses a genuinely novel question; the methodology is rigorous (tissue microarrays, validated antibody, independent scoring); cit-GFAP remained significant in multivariate analysis while AFP did not, suggesting independent prognostic value; and the authors transparently acknowledge most limitations. Moreover, the consistent directional trend toward higher recurrence rates in the high-expression group—though not statistically significant—merits further investigation in larger cohorts.

However, the study falls short of establishing cit-GFAP as a ready-to-use clinical biomarker. The retrospective, single-center design and modest sample size (n = 169) raise legitimate concerns about statistical power, particularly for the secondary endpoint of RFS. The non-significant RFS trend (P = 0.125) could reflect type II error rather than a true biological dissociation.

The most clinically nuanced observation is the OS-RFS discrepancy. While the authors attribute this to insufficient power, we argue that alternative biological explanations deserve equal consideration. Activated HSCs—the likely cellular source of cit-GFAP—contribute not only to tumor progression but also to progressive liver fibrosis and parenchymal dysfunction. A cit-GFAP-rich stroma may indicate a functionally impaired liver remnant, increasing mortality risk from hepatic failure independently of tumor recurrence. Moreover, HSCs recruit regulatory T cells and myeloid-derived suppressor cells, suggesting that cit-GFAP could influence survival through immune-mediated pathways. These hypotheses warrant dedicated investigation.

We therefore argue that Cho et al’s study[9] should be classified as a hypothesis-generating discovery report. Its primary value is to justify larger, prospective, multicenter validation studies with pre-specified statistical plans, optimized cut-offs, comprehensive covariate adjustment, and orthogonal validation of subcellular localization. Until then, cit-GFAP remains a promising but unvalidated candidate biomarker. Its true clinical utility will be determined not by this single retrospective study but by the rigor and reproducibility of subsequent validation efforts.

NAVIGATING THE LIMITATIONS AND FUTURE DIRECTIONS

As a pioneering study, the work by Cho et al[9] has inherent limitations, which the authors transparently acknowledge. The retrospective, single-center design necessitates validation in prospective, multicenter studies. Furthermore, the arbitrary 50% threshold used to define “high” expression should be refined through data-driven approaches such as ROC curve analysis in future investigations. Looking ahead, this discovery opens a rich and multifaceted research agenda.

Validation and standardization

The immediate priority is to validate the prognostic value of cit-GFAP in independent cohorts that are larger and more diverse with respect to ethnicity and underlying etiology. This effort should be accompanied by the standardization of the immunohistochemical assay and scoring criteria to ensure reproducibility across institutions and facilitate future clinical application. As outlined in the latest AASLD practice guidance, rigorous validation and standardization are essential prerequisites for the clinical adoption of any novel biomarker in HCC management[30].

Mechanistic elucidation

A critical question remains: Is cit-GFAP merely a passive marker of an aggressive, citrullination-prone TME, or does it actively contribute to HCC progression? Functional studies using cell line models and genetically engineered animal systems are required to determine whether cit-GFAP enhances the tumor-promoting functions of HSCs or directly influences cancer cell behavior. Elucidating its mechanistic role is essential for understanding its biological significance.

Integration with other biomarkers

How does cit-GFAP compare to or interact with other emerging biomarkers in HCC, such as glypican-3, protein induced by vitamin K absence/antagonist-II, or circulating tumor DNA derived from liquid biopsies? Beyond these, YKL-40, an ECM-associated glycoprotein, has recently been investigated as an early biomarker for HCC recurrence after liver transplantation, further illustrating the diverse landscape of emerging serum markers that may complement cit-GFAP in multimodal panels[31]. The development of a multimodal biomarker panel incorporating cit-GFAP may enhance risk stratification and improve prognostic accuracy.

Therapeutic implications

The identification of cit-GFAP also opens a speculative yet biologically plausible avenue for therapeutic exploration targeting the enzymes responsible for citrullination—PADs. It is essential to frame this proposition within a clear conditional framework: PAD inhibition could emerge as a novel stromal-targeting strategy only if cit-GFAP (and potentially a broader program of matrisome citrullination) is functionally validated as an active driver of HCC progression, rather than merely serving as a correlative biomarker. This hypothesis is supported by proof-of-concept studies demonstrating that PAD4 inhibition attenuates liver fibrosis in rodent models, as well as by the availability of small-molecule PAD inhibitors[32]. Several of these inhibitors have already entered early-phase clinical trials for non-oncological conditions, including rheumatoid arthritis and cardiovascular disease, highlighting their pharmacological feasibility while underscoring the current lack of oncology-specific data[33]. Recent structural and mechanistic studies on PAD2 and PAD4 have provided deeper insights into the molecular determinants of isoform selectivity and substrate recognition, which may inform the rational design of next-generation PAD inhibitors for oncology applications[34]. Future research must prioritize functional validation of citrullination within the HCC stroma, development of selective inhibitors, and rigorous evaluation in clinically relevant HCC models to assess both efficacy and safety before any translation into clinical practice can be justified.

Contextualizing the prognostic signal

To appreciate the potential clinical significance of cit-GFAP, it is instructive to contextualize it within the broader landscape of stromal activation in HCC. Studies utilizing multi-gene expression signatures have consistently demonstrated the robust prognostic value of CAFs and HSCs activity. For example, a prognostic model based on CAF-related genes has confirmed that elevated stromal activity is independently associated with adverse patient outcomes, thereby reinforcing the pivotal role of the TME in HCC progression[35]. The hazard ratio for cit-GFAP (2.75) reported by Cho et al[9] is therefore comparable to—and may even exceed—that of non-citrullinated GFAP and other established activation markers such as desmin[16]. This suggests that the post-translational modification detected by cit-GFAP may identify a distinct subset of activated HSCs or a specific stromal phenotype with particularly detrimental biological properties, thus providing incremental prognostic information beyond the mere quantification of stromal cellularity.

THE “CITRULLINATED MATRISOME”: A FRAMEWORK IN DEVELOPMENT

Figure 1 presents a conceptual diagram of this framework, illustrating the sequence from chronic liver injury to HSC activation, PAD-mediated citrullination, ECM remodeling, immune modulation, and ultimately poor prognosis in HCC. The study by Cho et al[9] invites the field to consider a broader conceptual framework: The “citrullinated matrisome” in HCC. This concept proposes that the functional properties of the tumor stroma may be critically shaped by a distinct pattern of protein citrullination, analogous to how phosphorylation networks regulate cellular signaling. Currently, this remains a compelling but speculative hypothesis rather than an experimentally mapped biological reality. Several key questions raised by this work remain unanswered: Has a systematic, omics-level inventory of citrullination sites on stromal proteins in HCC been performed? How does the extent and specificity of stromal citrullination compare to other prevalent PTMs, such as phosphorylation or glycosylation? Preliminary evidence from other cancers suggests that citrullination may be highly selective and context-dependent. Therefore, rather than overinterpreting its current definition, the value of the “citrullinated matrisome” concept lies in its capacity to guide future research directions. It generates testable hypotheses—that PAD enzymes target defined stromal substrates, that distinct citrullination patterns correlate with specific stromal phenotypes (e.g., immunosuppressive vs fibrogenic), and that therapeutic modulation of this modification could contribute to normalization of the TME. The identification of cit-GFAP as a prognostic marker provides the first direct evidence that investigating this framework in HCC is a scientifically justified and promising avenue of inquiry.

Figure 1 Conceptual framework of the citrullinated matrisome in hepatocellular carcinoma progression. Chronic liver injury activates hepatic stellate cells (HSCs), which upregulate glial fibrillary acidic protein (GFAP). Peptidylarginine deiminases (PADs), particularly PAD2, citrullinate GFAP and other extracellular matrix (ECM) proteins, generating the “citrullinated matrisome”. This leads to ECM remodeling, altered integrin signaling, and recruitment of regulatory T cells (Tregs) and myeloid-derived suppressor cells, ultimately promoting HCC progression, metastasis, and poor prognosis. HBV: Hepatitis B virus; HCV: Hepatitis C virus; MASLD: Metabolic dysfunction-associated steatotic liver disease; HSC: Hepatic stellate cell; α-SMA: Alpha-smooth muscle actin; GFAP: Glial fibrillary acidic protein; PAD: Peptidylarginine deiminase; ECM: Extracellular matrix; cit-GFAP: Citrullinated glial fibrillary acidic protein; HCC: Hepatocellular carcinoma.

CONCLUSION

The study by Cho et al[9] represents a significant advancement in our understanding of HCC prognostication. By identifying cit-GFAP as a novel and independent biomarker of poor survival, the authors have effectively redirected attention toward the post-translationally modified stromal matrisome. Although constrained by the limitations inherent to retrospective biomarker discovery studies, this work successfully shifts the diagnostic focus toward the dynamically altered stromal compartment. Its true impact will be measured by the rigor of subsequent validation efforts and the depth of mechanistic research it inspires. This study challenges the conventional emphasis on the cancer cell genome and calls for increased attention to the chemically dynamic protein landscape within the TME, proposing the systematic investigation of the “citrullinated matrisome” as a key to understanding stromal dysfunction. While further validation and functional characterization are required, cit-GFAP emerges as a promising candidate for refining clinical risk stratification models. More broadly, it highlights protein citrullination and the PAD enzymes as a compelling new axis for therapeutic exploration in the ongoing fight against HCC.