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Review

Exploring Perianal Fistulas: Insights into Biochemical, Genetic, and Epigenetic Influences—A Comprehensive Review

by
Maciej Przemysław Kawecki
1,
Agnieszka Marianna Kruk
1,
Mateusz Drążyk
1,*,
Zygmunt Domagała
2 and
Sławomir Woźniak
2
1
Clinical and Dissecting Anatomy Students Scientific Club, Wroclaw Medical University, 50-367 Wroclaw, Poland
2
Division of Anatomy, Wroclaw Medical University, 50-367 Wroclaw, Poland
*
Author to whom correspondence should be addressed.
Gastroenterol. Insights 2025, 16(1), 10; https://doi.org/10.3390/gastroent16010010
Submission received: 7 January 2025 / Revised: 21 February 2025 / Accepted: 4 March 2025 / Published: 7 March 2025
(This article belongs to the Section Gastrointestinal Disease)
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Abstract

:
The development of perianal fistulas leads to a significant decrease in the quality of patients’ lives. The onset of this condition is dependent on many factors, including inflammation or trauma. In the occurrence of Crohn’s disease-associated fistulas, numerous molecular factors and metabolic pathways are involved. To integrate the current knowledge on the biochemical, genetic, and epigenetic factors taking part in the development of perianal fistulas, we conducted a literature review. We gathered and analyzed 45 articles on this subject. The pathophysiology of fistulas associated with Crohn’s disease (CD) involves epithelial–mesenchymal transition (EMT) and matrix remodeling enzymes, with key regulators including transforming growth factor β (TGF-β), tumor necrosis factor α (TNFα), and interleukin-13 (IL-13). Genetic factors, such as mutations in receptor-interacting serine/threonine-protein kinase 1 (RIPK1), interleukin-10 receptor (IL-10R), and the MEFV gene, contribute to the onset and severity of perianal fistulas, suggesting potential therapeutic targets. Understanding the complex interplay of molecular pathways and genetic predispositions offers insights into personalized treatment strategies for this challenging condition. Further research is necessary to elucidate the intricate mechanisms underlying the pathogenesis of perianal fistulas and to identify new therapeutic interventions.

1. Introduction

Fistulas are connections formed between two epithelial surfaces. They can occur, for example, between the intestinal loops (enteroenteric), between the intestine and the skin (enterocutaneous), or between the rectum and the skin of the buttocks (perianal) [1,2].
Patients with perianal fistulas develop symptoms that significantly reduce their quality of life, such as anal pain, unpleasant leakage, and sometimes fecal incontinence [1,3,4,5]. It is reported that they have a negative impact on many aspects of life, including relationships, social life, and work life, and they may even lead to an inability to take part in physical activities [3].
The classic form of a perianal fistula is a late, chronic stage of a rectal abscess. Such an abscess, through overgrowth, forms a connection with the skin of the gluteal region, so that a narrow channel connecting the rectum to the skin surface is formed. These lesions are usually initiated in the crypt glands (cryptoglandular), at the level of the dentate line [4,5].
One of the most significant fistula classification systems is that proposed by the American Gastroenterology Association [6], which divides perianal fistulas into simple and complex types. A simple fistula, in this system, involves a connection between the rectum and a single opening in the skin, without being associated with an abscess. A complex fistula, on the other hand, passes through or over the anal sphincter muscles, and is associated with multiple end openings in the skin, as well as abscesses or anal strictures. The term “complex fistula” is also applied to fistulas that are connected to other internal organs (e.g., bladder, vagina, bowel) [1].
Based on current knowledge, EMT and matrix remodeling enzymes appear to play a special role in the pathophysiology of fistulas associated with CD. The EMT process produces transitional cells (TCs). In this case, epithelial cells transform into mesenchymal cells (like myofibroblasts) while gaining the ability to migrate and penetrate adjacent tissues. A regulatory factor crucial to the development of EMT is TGF-β. Its concentration increases in the zone between transitional and epithelial cells. The other important factor is TNF, which stimulates both the EMT process and the production of TGF-β by fibroblasts. Other molecules that are upregulated in the TCs of fistula ducts in CD include IL-13, IL-13 receptor, ETS1, and DKK1 [2,7].
According to data from a survey conducted in 2023 by A. Spinelli et al., Crohn’s perianal fistulas have a great negative impact on patients’ everyday life [3]. The goal of this paper is to identify the scientific areas that require further research in order to better understand the development of fistulas, ultimately leading to more effective treatments.

2. Methodology

This literature review aims to explore the current understanding and management of perianal fistulas, focusing on biochemical, genetic, and epigenetic influences in their formation.
We used the keyword “perianal fistula” and checked the first 100 papers on PubMed, Embase, Web of Science sites, and Google Scholar. When selecting research papers, we adhered to two primary principles: the selected papers had to address the etiology, pathogenesis, management, or impact of perianal fistulas, with a specific focus on CD-related fistulae; and preference was given to papers published from 2010 to 2024, to ensure the incorporation of up-to-date information and latest advancements in the field. In this way, we chose 7 articles on which to base our paper [1,2,4,5,7,8,9]. These selected articles collectively offer a comprehensive perspective on perianal fistulas, molecular insights, and pathogenesis.
With this foundation, we began expanding the scope by seeking additional studies to delve deeper into the biochemical, genetic, and epigenetic aspects of fistula etiopathogenesis. We used additional keywords to expand our search field (Figure 1).
This gave us 1160 results. We included papers that were published for broad, free access and were written in English. We were interested only in research conducted on human models; thus, we excluded those based on animals and cell line cultures. We did not have a preference for the type of article, and we included case studies, original works, and review papers. As stated previously, papers had to cover the etiology, pathogenesis, management, or impact of perianal fistulas, with a specific focus on CD-related fistulae.
Next, we scanned papers on the basis of their abstracts and full texts using the same standards. Those directly addressing the role of the factors of interest in other diseases and not contributing to the focus of this study, e.g., those where the topic of fistulas was only briefly mentioned in the text, were not included.
All of these criteria allowed us to shorten our list of results to 115 papers. From these papers, we excluded duplicates, and we were left with 31 publications.
Because some topics were not sufficiently explored in our search results, we created new search keywords: “fistula impact on life quality”, “RIPK1 role in inflammation”, “TGF- β signaling pathway”, “β-caterin signaling pathway”, “SATB1 regulation β-caterin”, and “β6 integrin activation”. For each of these, we found additional scientific papers that expanded knowledge on each of the mentioned topics [3,10,11,12,13,14]. This allowed us to build a better foundation for investigating two important aspects: the epidemiology of perianal fistulas in CD, and the biochemistry of signaling pathways in the pathogenesis of fistulas.
At that point, we gathered studies on which to base our study: papers that expanded upon the topic of fistulas, and additional papers that expanded upon and clarified the data collected in this study; this totaled 44 papers, to which we refer in this study.
We gathered data from these papers and organized them into sections covering similar topics, which we also used to divide our paper. First, we focused on the risk factors and pathogenesis of fistulas, which gave us insight into the factors that may provoke fistula formation. Based on this, we created separate sections in this work, into which we placed information extracted from the papers, referring consecutively to EMT, RIPK1, IL-10R, MEFV, TGF-β, TNF-α, IL-13, DKK-1, matrix remodeling enzymes, and bacterial remnants. Using these sections as a basis, we wrote this paper by merging knowledge from articles on similar topics. If any biochemical or epidemiological aspects were not sufficiently explained, we used additional papers to clarify and expand our understanding of the subject (Figure 2).

3. Results

3.1. Risk Factors

The most important factors associated with fistulation are male sex and the presence of ischiorectal or intersphincteric abscesses. The presence of inflammatory bowel disease (IBD) is also considered one of the main factors that increases the risk of fistula occurrence [15].
According to data from 2015, the incidence of IBD is 0–39.4 per 100,000 in North America and 0.9–37.0 in Europe. Susceptibility to IBD is increased by a family history of IBD; stress; an unhealthy diet, for example, one that is low in fiber, high in omega-6, low in omega-3 PUFAs, and low in vitamin D; infections of the gastrointestinal tract; intake of NSAIDs, aspirin, and oral contraceptives; and undergoing postmenopausal hormone therapy [16]. Other sources also indicate obesity (BMI > 25 kg/m2), high daily salt intake, diabetes, hyperlipidemia, dermatosis, anorectal surgery, smoking, alcohol intake, a sedentary lifestyle, excessive intake of spicy and high-fat food, infrequent participation in sports, and prolonged sitting on the toilet for defecation [17]. Preventing the development of IBD may be one of the strategies for helping to prevent the formation of perianal fistulas.
It has been shown that 0.5% to 4.3% of all perianal abscesses (PAs) or fistulas occur during infancy, especially among males [18,19]. The progression of these conditions among infants tends to resolve spontaneously in most instances, while older children exhibit patterns of PAs and perianal fistulas similar to those seen in adults.
There are two theories regarding the formation of perianal fistulas in infants. According to the first theory, congenital abnormalities result in deeper crypts of Morgagni, which leads to blockage in their narrowest parts. In this way, a PA forms, which, if it drains through the skin, creates a perianal fistula. The second theory suggests that congenital perianal fistulas progress to a PA and then develop into an epithelial lining of the fistula. Due to these discrepancies, the pathogenesis of PA/perianal fistulas remains elusive [19].

3.2. Pathogenesis

More than 90% of perianal fistulas are cryptoglandular fistulas [4]. They can arise as a complication after hidradenitis suppurativa, trauma, cancer, or tuberculosis [5]. The second most common type is CD-associated fistulas. It is important to note that due to differences in pathogenesis, the treatment of cryptoglandular and CD-related fistulas differ significantly. CD-related fistulas require a combination of drug therapy and surgical interventions, while cryptoglandular fistulas are mainly treated surgically [4,5,8].
CD is a chronic immune-mediated inflammatory condition that can affect any segment along the length of the entire gastrointestinal tract. It is often complicated by intestinal strictures and fistulas [1]. It represents, along with ulcerative colitis (UC), the main form of IBD [2]. CD can have one of three types of course: inflammatory, constrictive, and penetrating, with the penetrating form often leading to fistulas, although any type of CD can progress and change over time [1,9,20]. Single nucleotide polymorphisms (SNPs) in gene loci of ZMIZ1, LOC105373831, and KSR1 can increase the probability of development of constrictive or penetrating disease phenotypes. SNPs within the gene loci TNFSF15 and CEBPB-PTPN1 tend to protect against progression to these phenotypes of the disease (Table 1) [21].
A complete understanding of the pathophysiology of CD-associated fistulas has not yet been obtained. Two mechanisms seem to have a major role: EMT and matrix remodeling enzymes.

3.3. Epithelial–Mesenchymal Transition

Approximately 27% of CD fistulas exhibit epithelial cells lining the inner surface (epithelialized fistulas), while the remaining are classified as non-epithelialized fistulas covered by myofibroblast-like TCs. A distinct region has been identified within most patients’ latter fistulas where epithelial cells appear to transition into TCs, suggesting EMT involvement in CD-associated fistula development [2].
Specialized epithelial cells undergo a change whereby they transition into mesenchymal-like cells, gaining the capability to migrate and infiltrate neighboring tissues. In these TCs, genes of epithelial cell markers, such as cytokeratin 8 and cytokeratin 20, are expressed [2,22]. Other biosynthesized factors include mesenchymal markers, such as vimentin and smooth muscle actin (α-SMA). They also downregulate the expression of adhesion molecules, such as E-cadherin [2,4]. TCs are upregulating transcription factors, such as SNAIL1, SLUG (also known as SNAIL2), and β6-integrin [2,4,23]. TGF-β is a pivotal regulator of EMT, inducing IL-13 expression and an EMT-like phenotype in intestinal epithelial cells. IL-13, in turn, promotes the expression of invasion-associated genes, suggesting a synergistic role of TGF-β and IL-13 in fistula pathogenesis [24]. Other molecules that are upregulated in the TCs of fistula tracts in CD include IL-13 receptor, ETS1 (also known as protein C-ets-1) and DKK1 (dickkopf-related protein 1), protein kinase (RNA), PKR-like endoplasmic reticulum kinase (PERK), and nuclear factor B (NFkB) [4,24]. Translocation of β-catenin to the nucleus for gene regulation and increased β6-integrin expression are also considered to be EMT markers [24]. We will look further into these markers in the following sections of this paper.
SNAIL1 expression is notably elevated in the nuclei of TCs lining fistula tracts and crypts adjacent to these tracts, suggesting the involvement of SNAIL1 in fistula pathogenesis. Although SLUG expression is lower than SNAIL1 in TCs, it is present in fibrotic areas around CD fistulas. This suggests its potential role in fibrosis development, but a rather minor role—if any—in EMT onset leading to CD-associated fistulas [2].
The main differences between idiopathic and CD-related perianal fistulas are observed on the level of the IL-12 and IL-1RA/IL-1β ratio, both measured at the internal opening. Other cytokine levels are similar between both types of fistulas. Based on these findings, it is possible that CD-related and cryptoglandular perianal fistulas are more immunologically similar than previously assumed. If so, biological therapies used for CD-related perianal disease could be used in the treatment of cryptoglandular fistulas [4].

3.4. The Role of RIPK1

It has been observed that mutations in RIPK1 may lead to the development of perianal fistulas. These patients may already exhibit severe intestinal inflammation, as well as skin lesions and perianal inflammation, in the first months of life. Studies on samples taken from patients with PAs have allowed the identification two mutations causing RIPK1 protein dysfunction: c.1934C>T in exon 11 and c.580G>A in exon 4 [25].
RIPK1 mutations lead to deficits in inflammatory regulation, which contribute to the development of fistulas through chronic inflammation and tissue damage in the perianal region. One of the pathways found to be upregulated in RIPK1-deficient macrophages is inflammasome activation [25]. Inflammasome induces the secretion of the pro-inflammatory cytokines IL-1β and IL-18 in response to various stimuli, including lipopolysaccharides (LPSs) [26]. Impaired inflammasome activation has been reported in other monogenic disorders associated with IBD [25]. Before 2022, 14 patients with deleterious RIPK1 mutations were reported in the literature, and 12 of them presented with PAs or fistulas (Table 2) [27,28,29,30].
Deficiency in RIPK1-dependent signaling may contribute to inflammatory and degenerative diseases, such as severe colitis and perianal fistulas, particularly in young patients [10,25]. This is because RIPK1 participates in mediating apoptosis and inflammation [10]. However, current studies remain insufficient to fully determine the role of RIPK1 in the regulation of intestinal immune responses [25].

3.5. The Significance of IL-10R

Monogenic defects play a crucial role in the development of IBD in infants, and thus, these mutations may be factors that increase the risk of fistula formation [15,31]. The first monogenic defects identified in children with infantile IBD were mutations in IL-10R-inducing IL-10 signaling pathways. Such mutations lead to abnormal suppression of IL-6 secretion in response to LPS/IL-10 co-stimulated peripheral blood mononuclear cells [31,32]. These mutations can disrupt the IL-10-induced signaling pathway, resulting in improper phosphorylation of Transcription Factor 3 and increased levels of TNF-α and other pro-inflammatory cytokines, such as IL-6, IL-1, etc. [31,32].
It is hypothesized that without an IL–10–mediated anti-inflammatory reaction, the existence of intestinal commensal bacteria triggers a vigorous immune response, leading to excessive inflammation and subsequent tissue injury. This could promote higher levels of intestinal bacteria migration and lead to persistent intestinal lymphadenopathy or, potentially, organ-specific abscesses [32]. Abscesses like this, as previously stated, can evolve into fistulas (Table 2) [5].
Individuals harboring IL-10R mutations typically exhibit treatment-resistant enterocolitis and severe perianal conditions during infancy, including PAs, fissures, and fistulas [33]. There is no difference in the course of the disease between the IL-10RA, IL-10RB, and IL-10 mutations [34].

3.6. Mediterranean Fever Gene (MEFV)

Within the inflammasome, there are several components known to have modulated activity when pyrin production is disturbed. These components are cryopyrin, caspase-1, and its substrate, pro-IL-1β. It is noteworthy that the gene responsible for the pyrin production is MEFV, and apart from these inflammasome modifications, MEFV mutation may also result in a systemic increase in IL-1β [35,36].
These changes, while contributing to the course of familial Mediterranean fever (FMF), in 70% of FMF cases, also cause gastrointestinal symptoms similar to those of IBD (CD and UC) and intestinal Behçet’s disease (BD) (Table 2) [35,37].
The combination of CD and FMF raises the risk of the stricturing disease pattern and the development of extraintestinal symptoms; however, FMF is not considered a risk factor for CD susceptibility (the prevalence of CD in this case is estimated at 28.6%) [38,39].
Table 2. The mutations that elevate the risk of fistula development.
Table 2. The mutations that elevate the risk of fistula development.
Reference and Number of
Patients		Mutated ProteinAffected MechanismEffect
Sultan et al., 2022 [25]—14 people with the mutation, 12 of them with PAs or fistulasRIPK1Inflammasome activation—increased secretion of IL-1β and IL-18, decreased production of IL-6Development of fistulas through severe intestinal inflammation and tissue damage in perianal region
Glocker et al., 2009 [32]—4
Hung et al., 2021 [31]—1
Kotlarz et al., 2012b [33]—10
Pigneur et al., 2013b [34]—16
n = 31		IL-10RSuppression of IL-6 secretion in response to LPS/IL-10, increased levels of TNF-α and other pro-inflammatory cytokines, such as IL-6 and IL-1Excessive inflammation of intestinal tract
Asakura et al., 2018 [35]—1
Baran et al., 2018 [36]—1
Fidder et al., 2005 [39]—18
n = 20		MEFVReduced amounts of pyrin produced or production of malformed pyrin proteinInappropriate or prolonged inflammatory response

3.7. The Role of TGF-β, TNF-α, and IL-13

The epithelial-to-mesenchymal transition (EMT) is mainly activated by TGF-β. The production of TGF-β may increase due to TNF-α; thus, it is also capable of inducing EMT via secondary messengers: Smad2, Smad3, and Smad4 [11,23,24]. Subsequently the expression of TGF-β target genes reduces E-cadherin levels. E-cadherin activity is also inhibited by SNAIL1, which results in decreased intercellular adhesion [23,40].
Within patients with fistulizing CD, TGF-β may also stimulate IL-13 secretion in fibroblasts via β-catenin messenger, which may then result in raised levels of β6-integrin and the transcription factor SLUG (Figure 3) [23,24,40]. Feedback mechanisms regulating cell migration and invasion of CD-associated perianal fistulas also involve TNF-α, TGF-β, and IL-13 [23].

3.8. The Significance of DKK-1

The mRNA levels of DKK-1 were found to be elevated in intestinal tissue samples from patients with active CD and UC compared to samples from non-IBD control patients or patients with CD in remission [23]. Additionally, DKK-1 has a potential role in the mediation of inflammatory response [23], and is recognized as a potent antagonist of the canonical Wnt pathway [12,23].
It has been found that TGF-β increases the expression of DKK-1 in epithelial cells, but at the same time, it decreases it in fibroblasts derived from fistulas in CD patients, and in control fibroblasts from patients without IBD [23]. When exposed to TGF-β, epithelial cells undergo EMT and produce more DKK-1 [23]. Higher DKK-1 levels in these cells can limit cell activity by blocking Wnt signaling, which slows down cell growth, movement, and invasion. It has been found that DKK-1 is also needed for TGF-β to induce IL-13 expression [13,23]. This suggests that DKK-1 plays a dual role: it can inhibit TGF-β/Wnt signaling, but it also helps to activate IL-13, promoting cell invasion. Moreover, during EMT, TGF-β boosts IL-13 production, which, in turn, suppresses DKK-1 expression. This creates a complex feedback loop in which TGF-β induces DKK-1, while IL-13 inhibits it (Figure 4). In inflammatory conditions like chronic intestinal inflammation, excessive TGF-β secretion can lead to overproduction of IL-13, which suppresses DKK-1 and promotes EMT. As shown previously, IL-13 then triggers the expression of genes involved in cell invasion, such as β6-integrin, leading to the formation of fistulas (Figure 3) [24,25]. This effect can be reversed in all the cell types studied using anti-IL-13 antibodies. These findings support the hypothesis that inhibiting IL-13 can prevent the formation of fistulas (Table 3) [23,24].

3.9. Matrix Remodeling Enzymes

Matrix metalloproteinases (MMPSs) facilitate fistula progression by degrading the extracellular matrix (ECM) and promoting cell migration [4]. Increased MMPS activity has been observed in both experimental studies and humans with IBD [7,40]. In the fistula tracts of individuals with CD, a significant increase in the production of MMPSs has been observed, particularly the MMPS3 protein, as well as elevated mRNA levels in mononuclear cells and fibroblasts [7].
Due to an increased amount of mechanosensitive proteins and collagen fibers in the ECM, the levels of integrins, MMPs, and HA in the fistula were found to be noticeably raised in comparison to the surrounding regions [41]. Also, excessive expression of TNFAIP6 was noticed, contributing to an overexpression of mechanosensitive genes, such as SNAI1, ITGA4, ITGA2B, and PTK2B. This led to an inordinate release of collagen, due to the creation of an abnormal HA scaffold with the recombinant product of the TNFAIP6 gene—the TSG-6 protein—in the ECM (Figure 5). Both abnormal changes in the ECM and activation of mechanosensitive proteins trigger EMT, which may lead to a fistulization process [41].

3.10. Bacterial Remnants

Bacterial remnants (e.g., LPS) cause disorder in ECM integrity and trigger EMT, which may result in the development of chronic inflammatory lesions and, consequently, the release of pro-inflammatory cytokines and other molecules. Additionally, EMT may also lead to the fistulization process by enhancing the migration of epithelial cells [4].
Recent evidence suggests that in CD patients, defects in the epithelial lining—caused by inflammation or injury—lead to the development of pathogen-associated patterns [4,42]. These patterns trigger various pathways involving TNF-α, TGF-β, IL-13, MMPSs, and integrin αvβ. These pathways promote EMT, allowing cells to invade and migrate, leading to the formation of a penetrating fistula lined with TCs [4].
Activation of NOD2 and subsequent NFκB signaling may induce the production of pro-inflammatory cytokines, including IFN-γ, IL-13, and TNF, leading to an inappropriate immune response to bacterial antigens, such as muramyl dipeptide (MDP), during CD, and contributing to the failure of intestinal barrier function [22]. EMT may be initiated by the aforementioned cytokines as a result of wound-healing processes during inflammation. Expression of Ets-1, SNAIL1, and SLUG (transcription factors tied to EMT and cell invasiveness) is induced during the inflammatory process (Figure 6) [2,22]. Simultaneously, the expression of molecules involved in invasive cell growth, such as β6-integrin, is also upregulated, which may ultimately contribute to the formation of CD-associated fistulas [22].
The transcription factor ETS-1, a proto-oncogene that mediates the activation of the αvβ6-integrin receptor via its β6-integrin subunit, is upregulated by MDP and TNF in HT29 epithelial cells, fistula fibroblasts, and control fibroblasts from CD patients [14,22]. Moreover, TNF enhances β6-integrin expression through ETS-1, suggesting that ETS-1 is a crucial regulatory element in bacterially induced EMT and fistula formation, originating from bacterial stimulation of TNF expression [22].

4. Conclusions

The conducted analysis indicates a significant role of genetic factors, epigenetic factors, and proteins in the pathogenesis of perianal fistulas. Based on numerous studies, EMT and matrix remodeling enzymes are crucial in the development of fistulas associated with CD. Both EMT and matrix remodeling enzymes facilitate the transformation of epithelial cells into mesenchymal-like TCs, promoting invasion and tissue infiltration that contribute to fistula formation.
The TCs lining the fistulas express factors that upregulate the EMT process, such as TGF-β, ETS1, vimentin, α-SMA, SNAIL1, PERK, NFκB, and DKK1 [2,4,7,41]. TNF-α stimulates TGF-β production, which then enhances EMT processes and promotes cell migration and invasion in the development of CD-associated fistulas [2,7,11,23]. TGF-β also induces EMT by activating receptors that phosphorylate Smad2 and Smad3, which then form a complex with Smad4 and translocate to the nucleus to regulate target genes, reducing E-Cadherin levels and promoting the nuclear relocation of β-catenin (Table 3). The use of anti-TNF-α antibody therapy is widespread and supported by randomized trials; it has been proven to provide a 36% chance of permanent closure of the fistula, compared to a 19% chance of full recovery while on a placebo [43,44].
Mutations in RIPK1, IL-10R, and MEFV contribute to fistula development by disrupting immune regulation, which leads to chronic inflammation and tissue damage. RIPK1 mutations result in impaired inflammasome activation and dysregulated immune responses, promoting inflammation and necroptosis, which can cause fistulas. IL-10R mutations prevent the anti-inflammatory signaling pathway, resulting in excessive immune reactions to commensal bacteria, and causing tissue injury, abscess formation, and fistula development. MEFV mutations increase IL-1β production through dysregulated inflammasome activity, leading to inflammation that can mimic IBD and contribute to fistula formation [25,31,32,41]. This suggests that, especially in cases of infantile fistulizing features, genetic differentiation may be necessary to select the appropriate therapy. MEFV mutations treated with colchicine therapy show great results for co-existing fistulas [36], and fistulas due to mutations in the IL-10R or IL-10 genes may be treated with allogeneic hematopoietic stem cell transplantation. Bacterial remnants, such as LPS and MDP, contribute to fistula formation in CD by triggering EMT and disrupting ECM integrity. These remnants activate immune pathways and induce pro-inflammatory cytokines and growth factors, leading to increased epithelial cell migration, tissue invasion, and the formation of fistulas lined with TCs, due to abnormal immune response and impaired intestinal barrier function [4,42]. This indicates that antibiotics could be used for their prophylactic effects against infections and abscesses, or as adjuncts in medical treatments, such as in combination with infliximab; further research is needed to confirm these uses [43]. The exact mechanisms of fistula formation remain unknown, and require further investigation, particularly regarding the role of genes associated with these conditions.
This paper primarily aims to highlight the biochemical and genetic aspects of fistula formation. These topics are still not sufficiently researched, and more data are needed in order to develop effective treatment methods. The aim was to suggest which areas of science should be explored further to better understand the mechanisms of fistula development. We hope that by gathering all the factors contributing to fistula formation in one paper, we have created an anchor point for other researchers to begin their work and to avoid missing any crucial aspects that may influence new approaches to fistula treatment and prevention.

Funding

This research received no external funding.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

No new data were created or analyzed in this study. Data sharing is not applicable to this article.

Acknowledgments

The authors would like to thank wonderful technicians of our department Zbigniew Staszewski and Mirosław Łukaszun for their support.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. The types of perianal fistulas.
Figure 1. The types of perianal fistulas.
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Figure 2. Methodology.
Figure 2. Methodology.
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Figure 3. Factors leading to fistulas (created by the author).
Figure 3. Factors leading to fistulas (created by the author).
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Figure 4. The role of TGF-β, IL-3, and DKK-1 in correct EMT and chronic EMT (created by the author).
Figure 4. The role of TGF-β, IL-3, and DKK-1 in correct EMT and chronic EMT (created by the author).
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Figure 5. The consequences of excessive TNFAIP6 expression (created by the author).
Figure 5. The consequences of excessive TNFAIP6 expression (created by the author).
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Figure 6. The role of bacterial remnants in the development of chronic inflammatory lesions through EMT (created by the author).
Figure 6. The role of bacterial remnants in the development of chronic inflammatory lesions through EMT (created by the author).
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Table 1. Mutations that predict disease progression in CD patients [21].
Table 1. Mutations that predict disease progression in CD patients [21].
MutationEffect
ZMIZ1 (Zinc Finger MIZ-Type Containing 1)ZMIZ1 is associated with transcription regulation and interaction with other transcription factors. SNPs (single nucleotide polymorphisms) in this gene may affect its function, which could have implications for inflammatory and immune processes related to CD.
LOC105373831LOC105373831 is a gene with a less well-known function, that is considered a region susceptible to genetic changes associated with IBD. SNPs in this locus may influence the expression of nearby genes or the functioning of signaling pathways.
KSR1 (Kinase Suppressor of Ras 1)KSR1 is involved in the Ras/MAPK signaling pathways, which regulate cell growth and inflammatory responses. SNPs in KSR1 may alter its function, potentially affecting inflammatory mechanisms related to CD.
TNFSF15 (Tumor Necrosis Factor Superfamily Member 15)TNFSF15, also known as TL1A, is an inflammatory factor that plays a role in regulating immune and inflammatory responses. SNPs in this gene can affect its level or function, potentially leading to increased inflammation and progression of CD.
CEBPB-PTPN1 (CCAAT/Enhancer Binding Protein Beta–Protein Tyrosine Phosphatase Non-Receptor Type 1)CEBPB is a transcription factor that regulates inflammatory response, and PTPN1 is a phosphatase protein that can modulate inflammatory signaling. SNPs in this region may affect the expression or function of these genes, which could impact the development and progression of CD.
Table 3. The role of proteins in fistula formation.
Table 3. The role of proteins in fistula formation.
ReferenceProteinRole of Protein in Fistula FormationEffect
M. Scharl et al., 2013 [24]TGF-βReduces level of E-Cadherin, induces IL-13 expressionPromotes EMT
S.M. Frei et al., 2013 [22]TNF-αStimulates production of TGF-βPromotes EMT
M. Scharl et al., 2013 [24]IL-13Increases expression of β6-integrin and transcription factor SLUGTriggers expression of genes involved in cell invasion, leading to formation of fistulas
S.M. Frei et al., 2013 [22]DKK-1Antagonist of Wnt/β-catenin signaling pathway, mediator of inflammationInhibits TGF-β/Wnt signaling, but also helps to activate IL-13, which promotes cell invasion
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Kawecki, M.P.; Kruk, A.M.; Drążyk, M.; Domagała, Z.; Woźniak, S. Exploring Perianal Fistulas: Insights into Biochemical, Genetic, and Epigenetic Influences—A Comprehensive Review. Gastroenterol. Insights 2025, 16, 10. https://doi.org/10.3390/gastroent16010010

AMA Style

Kawecki MP, Kruk AM, Drążyk M, Domagała Z, Woźniak S. Exploring Perianal Fistulas: Insights into Biochemical, Genetic, and Epigenetic Influences—A Comprehensive Review. Gastroenterology Insights. 2025; 16(1):10. https://doi.org/10.3390/gastroent16010010

Chicago/Turabian Style

Kawecki, Maciej Przemysław, Agnieszka Marianna Kruk, Mateusz Drążyk, Zygmunt Domagała, and Sławomir Woźniak. 2025. "Exploring Perianal Fistulas: Insights into Biochemical, Genetic, and Epigenetic Influences—A Comprehensive Review" Gastroenterology Insights 16, no. 1: 10. https://doi.org/10.3390/gastroent16010010

APA Style

Kawecki, M. P., Kruk, A. M., Drążyk, M., Domagała, Z., & Woźniak, S. (2025). Exploring Perianal Fistulas: Insights into Biochemical, Genetic, and Epigenetic Influences—A Comprehensive Review. Gastroenterology Insights, 16(1), 10. https://doi.org/10.3390/gastroent16010010

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