Unveiling Colitis: A Journey through the Dextran Sodium Sulfate-induced Model

Abstract

Animal models of inflammatory bowel disease (IBD) are valuable tools for investigating the factors involved in IBD pathogenesis and evaluating new therapeutic options. The dextran sodium sulfate (DSS)-induced model of colitis is arguably the most widely used animal model for studying the pathogenesis of and potential treatments for ulcerative colitis (UC), which is a primary form of IBD. This model offers several advantages as a research tool: it is highly reproducible, relatively easy to generate and maintain, and mimics many critical features of human IBD. Recently, it has also been used to study the role of gut microbiota in the development and progression of IBD and to investigate the effects of other factors, such as diet and genetics, on colitis severity. However, although DSS-induced colitis is the most popular and flexible model for preclinical IBD research, it is not an exact replica of human colitis, and some results obtained from this model cannot be directly applied to humans. This review aims to comprehensively discuss different factors that may be involved in the pathogenesis of DSS-induced colitis and the issues that should be considered when using this model for translational purposes.

Introduction

The incidence of inflammatory bowel disease (IBD) is high in Western countries (eg, approximately 1.3% in the United States), and millions of cases are found in Europe and Northern America.^1,2 Although IBD is low-endemic in Eastern countries, its incidence in such countries has dramatically increased in the past 25 years.³ This trend poses a global health challenge; unfortunately, we lack a range of safe and effective medication for this disease.^4,5 Therefore, tremendous efforts have been made to investigate the pathogenesis of and potential treatments for IBD. In the context of such efforts, animal models of IBD are valuable and indispensable tools that provide options for characterizing the complexity of IBD pathogenesis and evaluating potential human therapeutics.^6,7

Among the numerous animal models that have been developed, chemically induced models of IBD are most commonly used because of their simplicity, controllability, and low cost.^8,9 Oxazolone, trinitrobenzene sulfonic acid (TNBS), and dextran sodium sulfate (DSS) are widely used to induce IBD in model animals.^8,9 The mucosal inflammation induced by these chemicals has been used to explore IBD pathogenesis and perform preclinical therapeutic studies. Unlike oxazolone and TNBS, which are small molecules, DSS is a sulfated polysaccharide that can have a variable molecular weight (MW). Therefore, the DSS-induced colitis model involves a pathogenic mechanism different from those induced by hapten reagents such as oxazolone and TNBS. Studies have shown that DSS forms complexes with medium-chain-length fatty acids (MCFAs) in the colonic lumen to generate nanovesicles, which fuse with colonocyte membranes to exert deleterious effects principally in the distal colon.¹⁰

The DSS model of colitis is also more flexible than other chemically induced animal models of colitis. For example, by changing the concentration and dosing frequency of DSS administration, researchers can produce acute, chronic, or relapsing models.¹¹ In the chronic phase of DSS-induced colitis, model animals exhibit dysplasia and thus closely mimic the clinical course of human UC.^12,13 Dextran sodium sulfate–based induction is often combined with other treatments to generate additional models, such as the use of azoxymethane (AOM) injection for studying colitis-associated cancer (CRC)¹⁴ (a topic beyond the scope of this review). In translational studies, DSS models have been applied to evaluate different therapeutic agents in preclinical studies. Subsequent comparisons with clinical data obtained using these agents showed that DSS-induced colitis can be used as a model that is highly relevant to human UC.¹⁵

A large body of literature involves the use of DSS-induced colitis models for evaluating therapeutics. However, this is not an exact replica of human colitis. For example, T and B cells are deeply involved in the development of human UC but are not required for the development of DSS-induced colitis.⁴ Therefore, the usefulness of the DSS-induced acute colitis model is limited to studying how the innate immune system contributes to the development of intestinal inflammation. The present review aims to discuss different factors that may be involved in the pathogenesis of DSS-induced colitis and the issues that need to be taken into account when using this model.

History of Using DSS to Induce Colitis

The use of DSS to induce colitis is not based on a serendipitous finding. Back in the 1960s, it was reported that adding extracts of red seaweeds to the drinking water of guinea pigs, rabbits, rats, and mice caused ulcerative disease in the colons of these lab animals.¹⁶ Chemical characterization showed that the red seaweed extracts contained carrageenan, which is a sulfated polysaccharide of 100-800 kDa.¹⁷ Further studies showed that hydrolysis-degraded products of carrageenan (molecular weight [MW] approximately 30 kDa) are more ulcerogenic than native or undegraded carrageenan and retain their original sulfate content and polyanionic properties.¹⁸ Other studies suggested that similar high-MW sulfated polysaccharides, such as sulfated amylopectin, sodium lignosulphonate, and DSS, may cause similar lesions in the colons of animals.

In 1985, Ohkusa’s group first demonstrated that administration of DSS to hamsters led to the development of colitis symptoms similar to those seen in human IBD, such as inflammation of the colon and diarrhea.¹⁹ In 1990, the same group reported that providing DSS to mice in their drinking water for a short period of time induced a very reproducible acute inflammation that was limited to the colon and characterized by erosions/ulcers, loss of crypts, and infiltration of granulocytes.¹¹ This DSS-induced mouse model of colitis provided a powerful tool for studying IBD. Since the initial report, this model has been widely adopted by other researchers and has been used in many studies to investigate the pathogenesis of IBD and evaluate the efficacy of new therapeutics.⁶

Chemical Characterization, Tissue Uptake, and Biodistribution of DSS

Dextran sodium sulfate is a negatively charged polysaccharide with a highly variable molecular weight (MW) that typically ranges from 5 to 1400 kDa.²⁰ Structural analysis showed that DSS is an α-D-(1→6) linear glucan with 2 sulfate groups on each α-glucose unit (as depicted in Figure 1A), and elemental analysis showed that its degree of sulfation is 16% to 19%.²² Molecular modeling revealed that in aqueous solution, DSS exhibits a 3D structure best described as helical (Figure 1B and 1C).²¹ The presence of disulfate-substituted side chains affects only the spatial dimensions of DSS, without altering its main helical structure.²¹

The MW of DSS governs the dimensions of its main chain, which are crucial for its ability to penetrate the intestinal mucus layer. The typical pore size of the intestinal mucus layer is around 200 nm.²³ Consequently, DSS with lower MW (MW <40 K) can penetrate the mucosal layer more easily than its higher MW counterparts (ie, >100 K). To illustrate, a medium-sized DSS with MW 40K has a length of approximately 50 nm and a radius of gyration of about 100 nm, while large-sized DSS with MW 100 K measures around 100 nm in length and has a radius of gyration of about 200 nm in aqueous solution (see Figure 1D).²¹ Thus, large DSS molecules are more likely to encounter obstacles while crossing the mucus layer.

Dextran sodium sulfate molecules have also been observed to form complexes with microbe-derived MCFAs, such as butyrate or dodecanoate, which are abundant in the colonic region. Large-sized DSS is more prone to undergoing complexation with MCFAs, and this has been identified as a significant factor in the tendency of large DSS to damage colonic epithelial cells after penetrating the mucus layer. However, a DSS-lipid complex has a much larger diameter (2-4 times larger) than the corresponding noncomplexed DSS, which further hinders the ability of large-sized DSS to penetrate the mucus layer.

Once across the mucus layer, DSS interacts with epithelial intestinal cells, where its 3D structure contributes to its ability to induce colitis. Simulation modeling showed that the DSS helix has a diameter at least as wide as an α-hemolysin nanopore.²¹ This unique 3D structure has been suggested to contribute to the pathogenesis of DSS-induced colitis by creating channels within the cell membrane that allow the passage of large molecules, including pathogenic polypeptides and proteins. In the case of large-sized DSS, the DSS-MCFA complex fuses with colonocyte membranes to trigger inflammatory signaling in the distal colon. Therefore, the severity of colitis formation reflects a complex interplay of multiple factors, including the pore size of the mucus layer, the DSS size, the concentrations and locations of MCFAs in the GI tract, and the ability of the DSS to fuse with cell membranes.

This dynamic interplay leads to an optimal range of DSS MW that is most potent in inducing colitis. In the murine model, medium-sized DSS with MW 40K demonstrated a heightened capacity to induce colitis. Research has shown that providing BALB/c mice with drinking water containing 40 to 50-kDa DSS resulted in the most severe murine colitis,¹¹ whereas mice administered with 5-kDa DSS developed milder symptoms, and those treated with 500-kDa DSS presented no colon lesions²² (Figure 1E). Histochemical examination of tissue uptake showed that 500-kDa DSS failed to cross the mucus membrane, explaining why this high-MW DSS has a limited ability to induce colitis.²²

The MW of the utilized DSS also affects the location of colitis: Mice treated with 40-kDa DSS develop diffuse colitis in the middle and distal sections of the colon, whereas those treated with 5-kDa DSS present relatively patchy lesions in the cecum and upper colon.²² This phenomenon can be explained by the fact that 40-kDa DSS molecules can form DSS-lipid complexes with MCFAs, which are abundant in the colonic region.¹⁰ These complexes advance toward the middle and distal colon, causing tissue damage. In contrast, the smaller 5-kDa DSS molecules are more adept at penetrating the mucus membrane but less likely to form lipid complexes. Consequently, they interact directly with epithelial cells, exhibit less fusion capability, and yield milder symptoms.²¹

Biodistribution studies of DSS based on histochemical analyses showed that approximately 40-kDa DSS could penetrate the mucous membrane in the intestine.²⁴ After a one-time DSS treatment, small amounts of DSS were found in colonic macrophages, liver Kupffer cells, and mesenteric lymph nodes at 24 h and in splenic macrophages at 72 h. At day-7 after a 5-day DSS treatment, DSS was found in the kidney and small intestinal macrophages.²⁴ It is worth mentioning that it is difficult for the body to clear DSS from the liver and spleen: DSS can be observed in the liver Kupffer cells at 8 weeks after treatment cessation and will accumulate in the spleen during the chronic phase of colitis. Interestingly, DSS cannot be retained in the stomach or duodenum, nor does it distribute to other organs, such as the brain, heart, lung, or thymus.^24,25

Dextran sodium sulfate is reportedly stable under pH conditions of 4.0 to 7.5 and resistant to degradation by ex vivo–cultured gut microbiota in an anaerobic environment.²⁶ The 2 major excretion routes of DSS are via urine and feces. Dextran sodium sulfate was reported to be present in the proximal tubule epithelial cells after 7 days of treatment, indicating that the kidney is the major excretion organ for DSS.^24,27

Pathogenesis of DSS-induced Colitis

The exact mechanism through which DSS induces colitis is not completely understood. Johansson’s studies showed that the sulfate groups from DSS could disturb the mucus layers and make them more permeable to bacteria and other pathogens.^28,29 Chassaing et al demonstrated that DSS impairs epithelial cells and causes damage to the epithelial monolayer lining³⁰ (Figure 2). Both of these studies suggested that DSS affects barrier integrity to allow pro-inflammatory bacteria and/or intestinal contents to penetrate into underlying tissues. These pathogens stimulate the innate lymphoid response to secrete pro-inflammatory cytokines and chemokines and thereby trigger an influx of inflammatory macrophages and neutrophils. Notably, DSS-induced colitis is localized more in the colon than in the small intestine (SI) of the gastrointestinal tract.²² Laroui et al suggested that prior to the induction of colitis, DSS associates with MCFAs of the colonic lumen to form nano-sized DSS/MCFA complexes that act as vesicles to fuse with colonocyte membranes.¹⁰ Because MCFAs are present at high concentrations in the colonic lumen, this proposed mechanism could potentially explain why DSS-induced colitis is localized in the colon. Additional factors that contribute to the relatively reduced susceptibility of the SI to DSS include the following: (1) The SI has a shorter transit time compared with the colon, as demonstrated in previous studies,³¹ which translates to a limited duration of DSS exposure in the SI. (2) Unlike the large intestine, the SI houses significantly fewer microbiota,³² and the absence of MCFAs further diminishes the probability of triggering inflammation through pathogenic activation.

The presence of administered DSS in the liver, spleen, kidney, and mesenteric lymph nodes suggests its ability to traverse mucosal epithelial layers. However, it remains unclear whether this crossing of DSS occurs via a transcellular or paracellular (via tight junctions) mechanism. Studies showed that after 1 day of DSS treatment, the mouse colon exhibits downregulation of zonula occludens-1 (ZO-1; a component of the tight junction complex) and upregulation of tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-1β, and IL-12 (pro-inflammatory cytokines).³³ After 3 days of DSS treatment, impairment of epithelial barrier function was found, as evidenced by a significant increase in permeability visualized by the Evans blue assay.³³ Histological analysis showed increased macrophage infiltration and loss of basal crypts in the colonic mucosa of mice with acute-stage DSS-induced colitis. Other features, such as increased apoptosis among epithelial cells and downregulation of tight junction proteins (occludin, ZO-1, and claudins) suggested that there is an imbalance between the apoptosis and proliferation of colonic epithelial cells in animals experiencing DSS-induced colitis.

Under DSS treatment, the impairment of colonic epithelial barrier function allows luminal bacteria, viruses, and associated antigens to enter the mucosa. Together with the entry of DSS itself, these pathogens could trigger inflammatory responses. Such responses typically involve numerous inflammatory mediators, including cytokines, chemokines, eicosanoids, reactive oxygen and nitrogen species, inducible nitric oxide synthase (iNOS), nitric oxide, and other components that activate the complement system.^34,35 At this acute stage of colitis, inflammation develops in the absence of T cell-mediated adaptive immunity, as shown in SCID4 and Rag1^−/− mice, indicating that inflammatory mediators produced via mechanisms of innate immunity are sufficient to cause acute colitis and tissue damage.³⁶ As DSS treatment is continued, the secretion of these inflammatory mediators increases progressively. While a steady upregulation of Th1 cytokines (TNF-α, IFN-γ, IL-1, and IL-12) characterizes the acute phase of colitis,³⁷ the chronic phase of DSS-induced colitis involves Th2-mediated inflammatory mediators (IL-4, IL-6, and IL-10).³⁸

Activation of innate immunity by DSS colitis was found to involve Toll-like receptors (TLRs)³⁹ and dectin receptors.⁴⁰ For example, studies showed that deletion of genes related to the TLR signaling pathway led to an exacerbation of DSS colitis mouse models.⁴¹ The genes encoding TLR2, TLR4, and Myd88 were shown to participate in promoting epithelial cell proliferation and colonic epithelial progenitor cell survival and improving barrier restoration, which is important for efficient wound healing from DSS injury. Deleting the TLR4 and Myd88 genes also decreased neutrophil infiltrates, partially because the absence of these proteins reduced the levels of key neutrophil-recruiting chemokines in macrophages.⁴¹ Administration of TLR2-targeting ligands or activation of epithelial TLR4 may also lessen the severity of DSS injury: the former was found to upregulate anti-inflammatory IL-10, while the latter increased the secretion of granulocyte-macrophage colony-stimulating factor (GM-CSF) by colonic epithelial cells.^42,43 These observations uncovered several important factors that contribute to the recovery stage of DSS-induced colitis.

A mechanistic discussion of DSS-induced colitis must also address the NLRP3 inflammasome response, triggered by TLR activation.⁴⁴ This inflammasome, which is a multiprotein complex including caspase 1, NACHT/LRR/PYD domain-containing protein-3 (NLRP3), and apoptosis-associated speck-like protein (ASP) (Figure 3), critically regulates the innate immune system and inflammatory signaling. Activation of TLR signaling pathways leads to cross-talk with NLRP3 inflammasome responses; this leads to the release of caspase 1, which cleaves pro-IL-1β to mature IL-1β, a key pro-inflammatory cytokine in DSS colitis.⁴⁵

Although DSS-induced acute colitis is believed to be caused primarily by disruption of the epithelium and activation of innate immunity (macrophages and neutrophils), studies showed that T-cell responses can aggravate the inflammatory response to acute DSS administration in wild-type mice with intact innate and adaptive immunity.⁴⁶ These mice present polarized Th1 responses in acute colitis and mixed Th1/Th2 responses in chronic-phase DSS colitis induced by repeated cycles of DSS administration.⁴⁷ Both Th1 and Th1/Th2 responses can promote the production of pro-inflammatory cytokines, such as TNF-α and IL-6, in macrophages during DSS colitis.⁴⁸

Single-cell RNA sequencing (scRNA-Seq) has emerged as a transformative tool and shed new light on the molecular underpinnings of colitis to offer valuable insights into potential therapeutic targets. It is worth noting that DSS has been shown to inhibit the activities of both polymerase and reverse transcriptase. Viennois et al⁴⁹ recommended that regardless of the duration or percentage of DSS treatment, RNA purification should be conducted to ensure the reliability of gene expression analysis in the colonic mucosa of DSS-treated mice. Ho et al⁵⁰ harnessed scRNA-Seq to unravel the transcriptome of cells within the colons of mice afflicted with DSS-induced colitis. Intriguingly, their findings revealed that Serpina3n, a serine protease inhibitor that is distinctly expressed in stromal cell clusters during inflammation resolution, engages with an elastase as a potential target. Genetic ablation of the Serpina3n gene postponed the resolution of DSS-induced inflammation, while systemic administration of Serpina3n fostered inflammation resolution to mitigate colitis symptoms.

Parigi et al⁵¹ used scRNA-Seq analysis to illuminate the spatially organized transcriptional programs that define distinct mucosal healing compartments and reveal regions characterized by dominant interconnected pathways. Notably, this study demonstrated that p53 activation is decreased in well-defined areas with upregulation of proliferating epithelial stem cells in the colons of mice recovering from DSS-induced colitis. Additionally, Zhang et al⁵² utilized scRNA-Seq to unveil the altered immune cell landscape resulting from IL-17B deficiency in DSS-induced colitic mice. Their findings revealed that IL-17B plays a protective role in colitis by curbing neutrophil infiltration and suppressing pro-inflammatory cytokine production in intestinal macrophages within the colon. Administration of IL-17B was found to ameliorate colitis in mice, suggesting its potential as a novel inhibitory cytokine that could serve as both a biomarker and a protective target for colitis therapy.

Strains Used for DSS-induced Colitis

Dextran sodium sulfate-induced colitis is typically established in laboratory mice or house mice due to their small size, short lifespan, and high reproductive rate. Several inbred mouse species are commonly used for DSS-induced models. The BALB/c and C57BL/6 mice, which are the most widely used inbred strains, differ in their susceptibility and responsiveness to DSS. Melgar et al demonstrated that after the removal of DSS applied for 1 cycle, BALB/c mice recovered from inflammation, but C57BL/6 mice developed chronic-phase colitis.⁵³ Studies also showed that BALB/c and C57BL/6J mice secreted different sets of cytokines in the chronic phase of DSS colitis. These differences were detected by immunohistochemical (IHC) staining of colonic tissues with infiltration of inflammatory cells,⁵³ and the results indicated that genetic factors are involved in the progression of the inflammatory response. After applying a standardized protocol of 3.5% DSS (36 to 45-kDa) for 5 days to 9 mouse strains, Mähler et al performed quantitative histological analysis to identify major differences in DSS responsiveness among strains.⁵⁴ The authors discovered that the C3H/HeJ, NOD/Ltj, and NOD-SCID inbred strains are highly susceptible to DSS and develop severe inflammation in the cecum, while 129/SvPas and DBA/2J inbred mice are less susceptible to DSS. Moreover, they found that the degree of susceptibility depends on the anatomical site: the severity of DSS-induced lesions increased from the proximal to the distal colon in most inbred strains, and male mice presented greater susceptibility to DSS in the colon than the cecum.⁵⁴

In addition to the previously mentioned inbred strains, CD-1 is a commonly used outbred mouse strain that is also susceptible to developing inflammation and can be used in establishing the DSS-induced colitis model.⁵⁵ The choice of strain is important, given that different strains have different genetic backgrounds and varied susceptibilities to colitis, which may affect the study results. The differences in susceptibility to DSS-induced lesions reflect a combination of differences in the ability of the mucosa to withstand inflammatory damage and the ability to recover from inflammatory injury.

Research has highlighted the pivotal role of enteric bacteria in shaping DSS-induced colitis. Gut microbiota and associated components are reportedly involved in the pathogenesis of colitis, and their absence considerably reduces the colonic inflammation triggered by DSS.⁵⁶ Hans et al reported that antibiotic treatment could reduce the infiltration of granulocytes to the mucosa and improve the histological index of acute-phase DSS colitis.⁵⁷ Nell et al showed that gnotobiotic mice were not prone to developing DSS-induced colitis until they were reconstituted with bacteria representing normal constituents of the luminal microbiota.⁵⁸ Lobionda et al further demonstrated that commensal bacteria and innate immunity together play an important role in the development of DSS-induced intestinal inflammation.⁵⁹ When BALB/c (wild-type) and immunodeficient SCID (lacking T and B lymphocytes) mice were maintained under conventional conditions, DSS treatment induced more substantial changes in the colonic mucosa of BALB/c mice compared with SCID mice. Conventionally reared BALB/c mice showed complete epithelium loss and severe inflammatory cell infiltration. In contrast, BALB/c and SCID mice reared in germ-free conditions developed minor signs of mucosal inflammation after DSS treatment.^59-61 These studies offered direct evidence that intestinal bacteria contribute to the pathogenic mechanisms underlying DSS-induced colitis.

Dextran sodium sulfate is often used to compare colitis susceptibility in genetically engineered mice (GEM) vs WT mice. Genetically engineered mice are particularly valuable for investigating the fundamental immunologic mechanisms of UC, as they allow researchers to manipulate specific genes involved in colitis pathogenesis through knockout or knock-in techniques.^62,63 For instance, deletion of suppressor of cytokine signaling-1 (SOCS-1), a negative regulator of interferon-gamma/signal transducer and activator of transcription 1 (IFN-γ/STAT1) signaling, increases susceptibility to DSS. This suggests that SOCS-1 may protect against DSS-induced colitis by inhibiting IFN-γ/STAT1 signaling.⁶⁴

In addition to mice, many protocols have been established and validated to generate the DSS-induced acute colitis in the Sprague-Dawley (SPD) rat,⁶⁵ and DSS-induced intestinal inflammation has been examined in large animals, such as swine. Young et al observed increased expression of TNF-α, IL-6, IFN-γ, and IL-17A in swine administered with DSS.⁶⁶ These signs are similar to those observed in patients suffering from active IBD. Other studies showed increased crypt destruction, mucosal erosion, and lymphocyte infiltration in the mucous tissue of swine following DSS treatment.⁶⁷ Recently, nonhuman primate models, including rhesus macaques (Macaca mulatta) and African green monkeys (Chlorocebus sabaeus), were compared under DSS-induced chronic colitis. Macaques were treated with 3 cycles of water with DSS followed by water without DSS, and the researchers reported finding “few-to-no overt clinical signs.”⁶⁸ However, when African green monkeys with longstanding simian immunodeficiency virus infection were treated with DSS for 10 days, colonoscopy revealed multifocal mucosal thickening, redness, and ulcerations.⁶⁸

Clinical Signs and Histopathology of DSS-induced Colitis

Oral administration of DSS in mice causes human UC-like pathologies because DSS damages colonic epithelial cells and compromises colonic mucosal barrier function. Therefore, clinical observations in mice with DSS-induced colitis are similar to signs of human colitis.⁶⁹ Weight loss, diarrhea, and occult blood in feces are commonly seen in the DSS-induced acute colitis mouse model. The clinical manifestations of DSS colitis differ between the acute and chronic phases.⁷⁰ Acute colitis is usually induced by treating the animal with 2% to 5% DSS for a short period (4-9 days); it mostly involves the activation of macrophages and neutrophils. Mice with acute colitis show signs of piloerection and anemia,^71,72 and histologic analysis often reveals mucin depletion, necrosis, and epithelial damage in colon tissues.³⁰ However, typical histological observations in human acute colitis, such as cryptitis and crypt abscesses, are seen in only approximately 50% of acute colitis C57BL/6 mice and are rare in acute colitis BALB/c mice.⁵³ Neutrophil infiltration into the lamina propria and submucosa is commonly found in acute colitis mice; these neutrophils can be detected by immunofluorescent staining and used to characterize the severity of colitis in the acute stage.⁷⁰

The induction of chronic colitis requires long-term continuous administration of a low dose of DSS or repeated cyclic administration of DSS. In chronic colitis, lymphocytes are activated, but the clinical signs may not reflect the severity of the inflammation or histological features found in the colon.⁷⁰ The histological changes of chronic inflammation often consist of mononuclear leucocyte infiltration, crypt architectural disarray, deep mucosal lymphocytosis, and an increase in the distance between crypt bases and muscular mucosa.²⁰ Some studies also observed transmural inflammation with lymphoid follicles in C57BL/6 mice with chronic colitis, and moderate epithelial regenerative atypia-simulating dysplasia was reportedly found at the edge of chronic erosions.^64,72 Given the patchy nature of the colonic lesions observed in DSS-induced chronic inflammation, Kitajima et al suggested that analysis of longitudinal sections and a few sections at an intermediate distance (>0.1 mm) could better reflect the actual damage of DSS colitis compared with analysis of cross sections.²²

Crucially, the chronic DSS colitis model is the prevailing approach for gaining insights into intestinal fibrosis. The repetitive administration of DSS has the capability to induce fibrosis in mice by inciting inflammation and causing damage to the intestinal lining. This damage, in turn, triggers the activation of fibroblasts—cells responsible for generating collagen and other proteins that constitute scar tissue. The accumulation of such scar tissue within the intestines can lead to the narrowing of the intestinal lumen, potentially obstructing the passage of food.

This model of chronic DSS-induced fibrosis stands as a dependable avenue for investigating the roles of various factors in the development of intestinal fibrosis, encompassing genetics, immunity, and microbiota. Moreover, it serves as a platform for evaluating the effectiveness of novel treatments for intestinal fibrosis. However, it is noteworthy that this model does not encompass the formation of strictures or fistulas, which are common complications associated with intestinal fibrosis in humans.^73,74 It is vital to acknowledge certain limitations tied to the utilization of DSS for inducing fibrosis. These include the potential time and resource intensity involved in setting up and maintaining the model, as well as its possible impact on the well-being of the mice. Thus, adhering to ethical guidelines is of utmost importance when working with this model.

Generally, both acute and chronic colitis can be monitored with noninvasive methods, such as endoscopy, stool consistency check, hemoccult positivity assay, and testing of biomarkers, such as fecal lipocalin 2.⁷⁵ Clinical scores and the Disease Activity Index (DAI) can be generated based on these observations. Calculating DAI scores normally involves summing 3 individual scores (ie, weight loss, stool consistency, and hematochezia/bleeding), with a higher score indicating more pronounced inflammation (Table 1).⁷⁶ The myeloperoxidase (MPO) assay can be performed in both noninvasive (live animal imaging) and invasive (dissected colon tissues) contexts.⁷⁵ Other histopathological parameters, including colon length, spleen weight, histology, immunohistochemistry, and epithelial cell proliferation and migration, could be collected after the terminal process.⁷⁷

Limitations

Colitis induced by DSS reproduces some of the key features of human IBD, including inflammation, diarrhea, and abnormal feces. This model is easily accessible because it is typically less expensive to use than other genetically modified animal models (such as IL-10 KO) and is relatively easy to maintain.⁷⁸ While variations in cages or facilities could potentially influence the model, it is important to note that the DSS-induced acute colitis model remains robust in its demonstration.²⁹ Its reliability is predominantly linked to the quality of the DSS, encompassing a range of molecular weights, rather than being solely facility-dependent. It is also a very flexible model that allows researchers to study how various interventions, such as medications or dietary changes, affect the disease course.⁷⁹

While the DSS-induced colitis model has been put to good use in many preclinical studies, culminating in the successful development of several drugs (as outlined in Table 2), it is important to recognize several limitations that should be considered when applying this model to studying human IBD. One major limitation is that DSS-induced colitis does not fully replicate the complex pathogenesis of human IBD,²⁹ which is influenced by various environmental, genetic, and immunological factors.⁹² The inflammation induced by DSS is more severe and occurs more rapidly than that in human IBD. Colitis induced by DSS presents indiscriminate epithelial cell damage and extensive microbial invasion into the lamina propria, which are unlikely to occur in human IBD.⁹³ The difference between DSS colitis and human IBD is also reflected in the observation that DSS-induced chronic colitis is associated with mixed T-cell pro-inflammatory cytokine responses, whereas chronic human IBD is associated with polarized T-cell responses.^15,36

Notably, DSS-induced colitis primarily reproduces the symptoms and pathology of UC, a type of IBD that affects the colon and rectum. The model is less effective at recapitulating the symptoms and pathology of Crohn’s disease, which is another type of IBD that can affect any part of the gastrointestinal tract.⁹⁴ In addition, the DSS-induced colitis model is highly dependent on the utilized mouse strain: different strains may differ in their susceptibilities to the development of inflammation and/or their responses to various interventions. These discrepancies can complicate efforts to compare results across studies and generalize the findings to other mouse strains or humans.

Conclusion

Since its introduction, the DSS-induced colitis model has been widely used in research to study and evaluate potential therapies for IBD. This model is simple to induce and inexpensive, and it is among the models most commonly used to investigate different aspects of IBD, including its pathogenesis, genetic predisposition, and immune mechanisms, as well as the effect of diet and the role of microbiota in the pathogenesis of IBD. The polydispersity of the DSS largely shapes its in vivo behavior: the size of the DSS (expressed as a MW range) critically influences its colitis-inducing effects. Thus, researchers must exercise prudence when selecting the appropriate molecular weight range for DSS, specifically within the range of 30 to 50k.

While the DSS-induced colitis model does not exactly replicate human IBD, it has advanced our understanding of the disease and exhibited some translational value. Many of the mechanisms and pathways that have been identified using this model have also been found to be relevant in human IBD. Some model-validated interventions have been tested in clinical trials in humans and revealed efficacy in reducing inflammation and improving symptoms in people with IBD. Several pharmacological therapeutic strategies against IBD were developed based on DSS-induced colitis. Nonpharmacological therapies have also been developed based on the DSS-induced colitis model; these include dietary interventions, such as the use of probiotics or prebiotics to modulate the microbiome and reduce inflammation. The efficacy of interventions has also been studied using the DSS-induced colitis model. Overall, while the DSS-induced mouse model of colitis has limitations and is not a perfect replica of human IBD, it has been and will continue to be a valuable tool for advancing our understanding of the disease and identifying potential therapeutics for use in humans.

Author Contributions

Conceptualization, CH.Y.; resources, CH.Y.; writing—original draft preparation, CH.Y.; writing—review and editing, D.M.; funding acquisition, D.M.. All authors have read and agreed to the published version of the article.

Funding

This research was funded by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), grant number RO1-DK-116306, Department of Veterans Affairs, grant number BX002526. D.M. is a recipient of a Senior Research Career Scientist Award (BX004476) from the Department of Veterans Affairs.

Conflicts of Interest

The authors declare no conflict of interest.

References