Exploring Colitis through Dynamic T Cell Adoptive Transfer Models

Abstract

Numerous animal models of colitis have provided important insights into the pathogenesis of inflammatory bowel disease (IBD), contributing to a better understanding of the underlying mechanisms for IBD. As aberrant CD4⁺ T cell responses play a critical role in the pathogenesis and development of IBD, T cell adoptive transfer models of colitis have become a valuable tool in investigating the immunopathogenesis of intestinal inflammation. While the adoptive transfer of CD4⁺ CD45RB^hi T cells into immunedeficient recipient mice was the first discovered and is currently the most widely used model, several variations of the T cell transfer model have also been developed with distinct features. Here, we describe the history, principle, and characteristics of adoptive transfer colitis models and discuss their strengths, limitations, and applications.

Key Messages.

What is already known? Animal models of colitis, including adoptive transfer models, have provided important insights into the pathogenesis of inflammatory bowel disease.

What is new here? We summarize different types of T cell adoptive transfer models of colitis by mainly focusing on the CD45RB^hi T cell adoptive transfer model, and briefly describe some other models.

How can this study help patient care? Understanding the strengths and limitations of each T cell transfer model allows researchers to choose an ideal model or combine different models, which will lead to a more comprehensive understanding of the mechanisms underlying inflammatory bowel disease.

Introduction

Inflammatory bowel disease (IBD), primarily Crohn’s disease (CD) and ulcerative colitis (UC), is a chronic inflammatory disorder occurring in the gastrointestinal tract characterized by diarrhea and abdominal pain with potential complications. In order to uncover the etiology of IBD and develop potential treatments against intestinal inflammation, several animal models of colitis have been established to mimic the features of human IBD.¹ However, no single animal model can fully reflect the complexity of human IBD because of the multifactorial feature of this disorder, in which genetics, environment, and immune responses are all involved.²

CD4⁺ T cells are important members of adaptive immune systems and mediate the pathogenesis of several autoimmune diseases. In IBD, CD4⁺ T cells are overactivated and produce excessive proinflammatory cytokines that contribute to the development of intestinal inflammation.³ The introduction of T cell adoptive transfer models not only dissects the importance of T cells in regulating intestinal homeostasis and inflammation, but also delineates the distinct features and functions of different T cell subsets in colitis. Adoptive transfer models of colitis have helped to find the translational and therapeutic implications of genetically modified T cell subsets in colitis and test drugs in preventing and treating intestinal inflammation. In addition, the interactions among T cells, innate cells, and microbiota could be well investigated using transfer models of colitis. In this review, we summarize different CD4⁺ T cell adoptive transfer models of colitis, with a main focus on CD45RB^hi T cell adoptive transfer model, and briefly describe several other models with their features, applications, strengths, and limitations.

CD4⁺ T Cells in IBD

The intestine is a major interface with the external environment, which harbors a gut microbiota containing huge amounts of bacterial, fungal, and viral species, and continuously encounters various pathogens.⁴ A well-established mucosal immune system associated with the intestinal lining contains a complex and highly specialized network of innate and adaptive immune cells uniquely equipped to manage these diverse challenges.⁵ Multiple regulatory mechanisms maintain intestinal homeostasis. One such mechanism consists of numerous specialized lymphocyte populations in the intestines with highly compartmentalized distribution and functional attributes.⁶ Among those lymphocytes, CD4⁺ T cells are a major population involved in mediating various host protective and homeostatic responses.³ However, the accumulation of CD4⁺ T cells in the intestine is also a key feature of IBD. A variety of mouse models that develop chronic intestinal inflammation resembling IBD have provided strong support for the crucial role of CD4⁺ T cells in mediating the pathogenesis of IBD, presenting a potential therapeutic target for the treatment of IBD as well.

In response to gut microbiota stimulation, CD4⁺ T cells can develop into different subsets of T cells based on the expression of distinct transcription factors and production of cytokines, namely T helper 1 (Th1), Th2, Th17, and regulatory T (Treg) cells.^3,7 Th1 cells produce signature cytokine interferon γ (IFNγ), promoted by interleukin (IL)-12 via induction of T-bet and Stat4; Th2 cells produce signature cytokines IL-4, IL-5, and IL-13, via induction of Gata3 and Stat6 by IL-4; Th17 cells produce signature cytokines IL-17, IL-21, and IL-22, through induction of RORγt and Stat3 by transforming growth factor β (TGFβ) and IL-6; and Treg cells produce TGFβ and IL-10 via induction of Foxp3. While Th1 and Th17 cells have been implicated in the pathogenesis of different types of CD, Th2 cells mediate the pathogenesis of certain types of UC.⁷ In contrast, Treg cells suppress the pathogenesis of both CD and UC.⁸ T cell development in the intestines is tightly controlled by gut microbiota. Various gut bacteria have been shown to regulate the development of different subsets of T cells differentially. While segmented filamentous bacteria induce mucosal Th17 cells,⁹Klebsiella bacteria induce Th1 cells¹⁰ and Clostridia strains promote Treg cells.¹¹ Earlier evidence showed a decline in peripheral Treg cells in IBD patients,^12,13 which had been attractively hypothesized to attribute to the development of IBD. However, emerging data have demonstrated an increase in Treg cells in the inflamed intestinal mucosa of IBD patients,^14-16 the presence of which is crucial in preventing inflammation from becoming too severe, and thereby detrimental. Thus, both homeostatic and inflammatory mechanisms mediated by CD4⁺ T cells coexist under inflammatory conditions in IBD.

CD4⁺ CD45RB^hi T Cell Adoptive Transfer Model of Colitis

History

The CD4⁺ CD45RB^hi T cell adoptive transfer model of colitis was established in the early 1990s.^17,18 CD45, a member of the protein tyrosine phosphatase family, is expressed on most hematopoietic cells.¹⁹ Due to the differential splice of exons A, B, and C, CD45 exists in various isoforms, which define different subpopulations of immune cells, including CD4⁺ T cells.¹⁹ Based on the expression of CD45RB, murine CD4⁺ T cells can be divided into CD45RB^high and CD45RB^low subtypes, which produce distinct cytokines in response to various stimuli.^20,21 To determine the function of CD45RB^hi and CD45RB^lo CD4⁺ T Cells, Powrie et al¹⁷ and Morrissey et al¹⁸ isolated CD4⁺ T cells with the brightest CD45RB staining (CD45RB^hi) and CD4⁺ T cells with the lowest CD45RB staining (CD45RB^lo) and transferred them into syngeneic severe combined immunodeficient (SCID) recipient mice via intravenous injection. The recipient SCID mice that received CD4⁺ CD45RB^hi T cells developed severe colitis, which was prevented by the cotransfer of CD45RB^lo CD4⁺ T cells,¹⁷ thus establishing the CD4⁺ CD45RB^hi T cell adoptive transfer model of colitis. Those studies not only established an elegant experimental colitis model resembling IBD but also revealed a crucial regulatory interaction between these 2 CD4⁺ T cell subsets. In addition, they provided a conceptual breakthrough that both pathogenic and regulatory CD4⁺ T cells coexist under steady conditions.^17,22 Although this intestinal inflammation is predominantly located in the large intestine, which extends from the cecum to the rectum,²³ CD45RB^hi T cells also induce inflammation in the small intestine.²⁴ The characteristics of the colitis induced by CD45RB^hi T cells include leukocyte infiltration of the lamina propria, extending to the depth of entire intestine; epithelial change (eg, hyperplasia, crypt disorganization, goblet cell loss); and mucosal erosion and ulceration.²³ Later on, other immunodeficient mice were also used as the recipients of CD45RB^hi T cells for this model (ie, Rag1^−/− mice, Rag2^−/− mice, and TCRβxδ^−/− mice). Although both Rag^−/− mice and SCID mice do not have mature T cells and B cells, SCID mice can become leaky,²⁵ meaning that some functional T and B cells can develop, while Rag mutations do not. Thus, Rag^−/− mice offer numerous advantages over SCID mice. TCRβxδ^−/− mice lack T cells but have functional B cells, making them more representative of physiological conditions. This offers advantages over Rag^−/− mice and SCID mice in the study of the role of B cells in the pathogenesis and regulation of colitis. It is worth noting that high IFNγ-producing natural killer cells occupy tissues (eg, intestine) of these immunedeficient recipient mice, which could affect the severity of colitis induced by CD45RB^hi T cells.²⁶

Brief Description of the Procedure

After sacrificing the mice following the Institutional Animal Care and Use Committee protocol, spleens are taken to make a single-cell suspension. After removing red blood cells using cell lysing buffer, the cells are incubated in a tube with magnetic anti-mouse CD4 beads or a negative selection cocktail in the proper buffer for a time suggested by the manufacturer, to separate CD4⁺ T cells using a magnet. After washing with phosphate-buffered saline, the cells are stained with surface antibodies, CD4 antibody, and CD45RB antibody in an optimal combination of fluorescence following a standard flow cytometry staining protocol. Live CD4⁺ CD45RB^hi T cells can be sorted using a cell sorter. The procedure and the representative histopathology are shown in Figure 1A, B. In the original report, 40% to 50% of the CD4⁺ T cells with the brightest CD45RB staining were sorted and used for intravenous injection into SCID mice. However, the percentages of CD45RB^hi T cells and the cell number used for inducing colitis in recipient mice are not identical among different research groups. Based on our experience, sorting 10% to 15% of the brightest CD45RB staining in live CD4⁺ T cells for transfer (1 × 10⁵ cells/mouse) generates reproducible intestinal inflammation in the Rag1^−/− and TCRβxδ^−/− recipient mice. There are some tips for the techniques on cell staining and sorting, which are critical to purify the correct population for inducing colitis. First, it is important to eliminate the dead cells by viability staining (eg, DAPI, propidium iodide, and 7-aminoactinomycin D) before sorting. Second, setting up the template for sorting (eg, gating, sorting speed and pressure, and yield model) is helpful to the consistency of inducing colitis with fewer errors. In addition, the routes of cell transfer (eg, intraperitoneal, intravenous) affect the time of colitis induction. The recipient mice develop intestinal inflammation from 3 weeks and can be sacrificed at 5 to 8 weeks post–T cell transfer, but it may vary within different animal facilities. Cecal and colonic tissues are rolled, fixed in 10% neutral buffered formalin, dehydrated with a series of alcohol, and embedded in paraffin. After staining with hematoxylin and eosin, the histopathological analysis is assessed by the scores of the following parameters: the levels of lamina propria inflammation (score 0-3), goblet cell loss (score 0-3), abnormal crypt (score 0-3), the presence or absence of crypt abscesses (score 0-1), the severity of mucosal erosion and ulceration (score 0-3), and the presence or absence of submucosal change (score 0-2).²⁷

Figure 1.

The procedure of T cell transfer and histopathology of recipient mice. (A) Splenic CD4⁺ T cells were isolated from the donor mice, and CD4⁺ CD45RB^hi T cells or Foxp3⁺ regulatory T (Treg) cells were purified by cell sorting. Subsequently, CD4⁺ CD45RB^hi T cells with or without Treg cells were transferred into Rag1^−/− mice by intravenous injection. (B) The recipients were sacrificed 6 weeks post–cell transfer. The representative hematoxylin and eosin image of colonic changes in recipients was shown. PBS, phosphate-buffered saline; WT, wild-type.

Derivations of CD4⁺ CD45RB^hi T Cell Transfer Model

CD45RB^hi T cells are primarily naïve T cells, which have several other surface markers. Thus, there are several variations of naïve T cell adoptive transfer models of colitis using different markers, such as CD62L and CD25.^28,29 CD62L, also called L-selectin, is a homing molecule and a naïve T cell marker shed from the cell membrane following T cell activation.³⁰ Therefore, CD62L^−/low could serve as a marker for activated T cells. CD25, an IL-2 receptor alpha, is a regulatory T cell marker.³¹ It is also highly expressed in activated T cells.³² Thus, CD25⁻ serves as a naïve T cell marker. Transfer of CD4⁺ CD62L⁺ T cells or CD4⁺ CD25⁻ T cells or CD4⁺ CD62L⁺ CD25⁻ T cells into SCID mice/Rag^−/− mice/TCRβxδ^−/− mice induces chronic colitis, similar to the transfer of CD4⁺ CD45RB^hi T cells. Practically, the transfer of CD4⁺ CD62L⁺ CD25⁻ T cells induces colitis more consistently. Compared with CD45RB^hi T cells, it is relatively easier to sort CD62L⁺ or CD25⁻ CD4⁺ naïve T cells with a higher yield. The time of T cell induction of colitis is different using different markers, which also depends on the animal housing facilities. Thus, it is hard to compare which one is better. It should be noted that the severity of colitis might be different in SCID mice, Rag^−/− mice, and TCRβxδ^−/− mice.²⁴

Major Discoveries From Using CD4⁺ CD45RB^hi T Cell Adoptive Transfer Model

As an important and reproducible chronic colitis model, the CD4⁺ CD45RB^hi T cell adoptive transfer model has contributed significantly to understanding IBD pathogenesis. Numerous important discoveries have been made using this model. Here, we only describe a few related to the role of T cells in regulating colitis to serve as examples.

Gut microbiota is required for colitis development

Because the intestinal epithelial barrier is compromised in immunedeficient mice, gut microbiota could advance into intestinal lamina propria to drive inflammation. Transfer of CD4⁺ CD45RB^hi T cells induces less severe colitis in immunedeficient mice with reduced numbers of enteric flora, and does not induce colitis in germ-free mice.^33-35 This suggests that gut microbiota is indispensable in the induction of colitis. The composition of intestinal microbiota could vary based on facility-to-facility, room-to-room, parent-to-parent, and cage-to-cage variability. We suggest using recipient mice from breeding pairs that are weaned into the same cage to minimize parent-to-parent and cage-to-cage variations. Alternatively, adding fecal pellets or bedding from other colitic mice in the colony to the cages of recipients can equalize the microbiota.

Certain bacteria or cocktails have been identified in inducing colitis and determining the effect of specific bacteria on T cell functions using the CD4⁺ CD45RB^hi T cell transfer model. For example, monoassociation of SCID mice with Helicobacter muridarum leads to accelerated colitis,³⁵ while segmented filamentous bacteria induces colitis in germ-free SCID mice only when co-colonization with a defined cocktail of specific pathogen-free bacteria.³⁴ In addition, the prominent human symbiont Bacteroides fragilis is beneficial for protection against colitis in immunedeficient mice colonized with H. hepaticus through the molecule polysaccharide A.³⁶B. fragilis was also shown to facilitate Treg cell differentiation in the gut to induce mucosal tolerance.³⁷ More recently, several studies utilizing the CD4⁺ CD45RB^hi T cell transfer model have revealed that gut microbiota–derived metabolites play a role in mediating intestinal inflammation.³⁸ These studies highlight the complex interplay among the gut microbiota, T cell responses, and intestinal inflammation, suggesting that manipulation of the gut microbiota or their metabolites might be a potential therapeutic strategy for the treatment of colitis.

Th1 and Th17 cells are pathogenic in inducing colitis

Intestinal CD4⁺ T cells in recipient immunodeficient mice reconstituted with CD4⁺ CD45RB^hi T cells produce a high level of IFNγ, a major cytokine of Th1 cells, and treatment with anti-IFNγ almost completely abrogates the colitis.²² Additionally, blocking IL-12, critical in Th1 differentiation, suppresses CD4⁺ CD45RB^hi T cell–induced colitis.⁹ CD4⁺ CD45RB^hi T cells lacking T-bet, a transcription factor of Th1 lineage, fail to induce colitis in reconstituted SCID mice.³⁹ Therefore, CD4⁺ CD45RB^hi T cell transfer–induced colitis was considered a Th1-dominant intestinal inflammation model, which contributed to the establishment of the commonly held view that Th1 cells are fundamental in the pathogenesis of IBD, before the discovery of Th17 cells.⁴⁰

A series of studies have supported the importance of Th17 cells in regulating intestinal inflammation.^41,42 IL-23, comprising IL-12p40 and IL-23p19 subunits, is a key cytokine driving Th17 cells,⁴³ which mainly produce IL-17A, IL-17F, and IL-22. RORγt is the master transcription factor of Th17 cells.⁴⁴ Th17 cells exist in the intestine in the CD4⁺ CD45RB^hi T cell transfer model,⁴⁵ and deficiency of IL-23 receptor or RORγt in CD4⁺ CD45RB^hi T cells reduces the severity of colitis in Rag1^−/− mice.^46,47 In addition, the colitis is highly attenuated in Rag1^−/− recipients with a deficiency in the IL-23p19 subunit (lacking IL-23) but not the IL-12p35 subunit (lacking IL-12).⁴⁸ Furthermore, genome-wide association studies have identified that IL-23 receptor variants are a risk factor for human IBD.⁴⁹ All the evidence support that Th17 cells mediate intestinal inflammation.

Treg cells prevent and treat colitis

Treg cells are a subset of CD4⁺ T cells that play an important role in controlling excessive immune responses and maintaining intestinal inflammation. Dysfunction of Treg cells in the intestine leads to uncontrolled inflammation in the gut and contributes to the development of colitis.³

In the original reports of the CD4⁺ CD45RB^hi T cell transfer model, while CD4⁺ CD45RB^hi T cells are pathogenic in inducing intestinal inflammation, the CD4⁺ CD45RB^lo T cells, a subpopulation of T cells with low expression of the CD45RB protein, are not able to induce colitis, and even prevent the intestinal inflammation induced by CD4⁺ CD45RB^hi T cells in SCID mice.^17,18 These elegant studies highlight that the imbalance of T cell subpopulations is involved in the development of intestinal inflammation. CD4⁺ CD45RB^lo T cells have been shown to have regulatory properties and suppress inflammation by producing TGF-β and IL-10.^50,51 Specifically, CD4⁺ CD25⁺ CD45RB^lo T cells (ie, Treg cells), other than CD4⁺ CD25⁻ CD45RB^lo T cells, are responsible for the suppression of colitis.³¹ Further, Foxp3 has been recognized as the transcription factor of Treg cells.^52,53 Transfer of Foxp3⁺ Treg cells inhibits colitis induced by CD4⁺ CD45RB^hi T cells,^54,55 and the procedure and the representative histopathology are shown in Figure 1A, B. Interestingly, in vitro differentiated Treg cells also have the capacity to suppress colitis.⁵⁶

The transfer of Treg cells in animal models of colitis with promising results suggests that this approach might be a potential therapeutic strategy for human colitis. Many registered trials of Treg therapy for IBD are either completed or ongoing. Several groups have reported the success of using Treg transfer to treat IBD patients.⁵⁷ However, clinical use of Treg cells for IBD patients has been limited by small sample sizes and is still in the early stages of investigation. More efforts are needed to determine the safety and efficacy of this approach. In addition, it is important to investigate whether gene-modified Treg cells are more potent for suppressing colitis and if diet, microbiota, or other factors affect the efficiency of Treg cell inhibition of colitis.

Regulatory B cells inhibit colitis

In addition to producing antibodies upon immunization and infection, B cells have been found to play multiple functions in various conditions. Regulatory B cells maintain intestinal homeostasis and inhibit colitis. Early studies by Mizoguchi and Bhan’s group demonstrated that although B cells were not required for the initiation of colitis in TCRα^−/− mice, B cells and autoantibodies suppressed colitis, mediated by the clearance of apoptotic cells and the related self-antigens.⁵⁸ While the recipient Rag1^−/− mice developed a more rapid and severe form of colitis upon transfer of CD4⁺ CD45RB^hi T cells, TCRβxδ^−/− mice, which lack B cells, developed a later and more moderate form of colitis,²⁴ supporting the concept that B cells function as regulatory cells in the regulation of colitis. More recent works further demonstrate that IL-10–producing regulatory B cells inhibit colitis by inducing T regulatory-1 cells when cotransferred with naïve CD4⁺ T cells into Rag2^−/− mice.⁵⁹ Furthermore, intestinal CD11b⁺ B cells suppress colitis development by secreting IgA.⁶⁰

Other T Cell Transfer Models of Colitis

CBir1 Transgenic T Cell Transfer Model

As accumulating evidence highlights a crucial role of gut bacterial antigen–specific T cells in the pathogenesis of IBD, utilizing T cells specific for a defined gut bacterial antigen will provide insights into how T cells respond to the gut bacterial antigens to regulate colitis.⁶¹ Gut bacterial flagellin, the component of flagella, has been shown to contribute to the pathogenesis of IBD.^62,63 In multiple animal models, flagellin induces or exacerbates intestinal inflammation.⁶² CBir1 flagellin, one of the gut bacterial flagellins produced by Lachnospiraceae bacteria, has been identified as a dominant flagellin antigen with a role in both innate and adaptive immunity.⁶⁴ About 50% of CD patients demonstrate antibodies to CBir1 flagellin.⁶³ CBir1 reactive CD4⁺ T cells and CD154⁺ effector memory CD4⁺ T cells are present in CD patients.^65-67 CBir1 T cell receptor (TCR) transgenic mice have been generated, which use TCRVβ8.3 and Vα6.⁶⁸ Transfer of CD4⁺ T cells from CBir1 Tg mice, which are reactive to CBir1 flagellin, into Rag^−/− mice or TCRβxδ^−/− mice induces colitis, thus setting up the CBir1 T cell adoptive transfer model of colitis.⁶¹ In this model, the colitis develops within 2 to 3 weeks after cell transfer and reaches its peak in 4 to 6 weeks in various animal facilities. All T cell subsets (ie, Th1, Th17, and Treg cells) are present in the lamina propria of the inflamed intestines, thus providing a unique colitis model to study subsets of T cells in the pathogenesis and regulation of chronic colitis. While the colonization of CBir1-expressing bacteria in the recipient mice is crucial for the induction of colitis in this model, a recent study indicates that the proliferating T cells in the recipient mouse respond not only to CBir1 flagellin, but also to other antigens recognized by the second TCR via endogenous rearrangement carried by CBir1 TCRα chains.⁶⁹ In some animal facilities, while wild-type CBir1 T cells induce colitis in Rag^−/− recipients, Rag^−/− CBir1 transgenic T cells do not induce colitis, suggesting that induction of colitis by the gut T cells responding to a specific gut bacterial antigen may require other interrelated commensal bacteria, for example Helicobacter spp, to function as an adjuvant in this model.

Th1 and Th17 Effector T Cell Transfer Model

As the roles of Th1 and Th17 cells in the induction of colitis are well established, the transfer of in vitro polarized polyclonal Th1 effector cells can induce colitis in immune-deficient mice.^45,70 The severity of colitis induced by the in vitro differentiated Th1 effector cells in immunedeficient mice may vary from the colitis induced by CD4⁺ CD45RB^hi T cells, which several possibilities might explain. First, the way T cells are activated and polarized in vitro and in vivo is not identical, which might make the in vitro polarized Th1 cells less or more pathogenic in inducing colitis. Second, the ability of proliferation and migration of in vitro polarized Th1 cells might be relatively low. The Th1 effector cell transfer model provides an excellent platform to investigate the effect of different genes and drugs on Th1 induction of colitis.

Not surprisingly, the transfer of polarized Th17 cells into immunedeficient mice leads to severe colitis comparable to that induced by CD45RB^hi cells.⁷¹ This model provides an excellent tool to investigate the role of Th17 cells in regulating colitis and the plasticity of Th17 cells in the intestine, and to manipulate Th17 cells to affect colitis. This model has demonstrated that IL-17A-producing Th17 cells can transition to IFNγ-producing Th1-like cells, and Th17 cells deficient in IFNγ fail to induce intestinal inflammation.⁷² These results have nicely resolved the previous contradiction that naϊve T cells deficient in IL-17A lead to excessive Th1 responses and accelerate wasting disease and intestinal inflammation.⁷³ The Th17 cell transfer model provides a valuable tool for understanding the Th17 responses and plasticity in the intestines.

Unfractionated IL-10–Deficient CD4⁺ T Cell Transfer Model

IL-10 is an anti-inflammatory cytokine that plays a crucial role in regulating the immune response in colitis. The Il10 locus has been linked with susceptibility to IBD.⁷⁴ In addition, therapies aimed at increasing IL-10 levels have shown promise in treating colitis in animal experiments and clinical studies.⁷⁵

Unfractionated IL-10–deficient CD4⁺ T cells can induce colitis upon transfer into immunedeficient mice to similar levels induced by CD4⁺ CD45RB^hi T cells.⁷⁶ Using unfractionated IL-10–deficient CD4⁺ T cells can simplify the preparation and purification of a specific subpopulation of T cells for transfer, making it a more accessible tool for investigators with few resources and less experience in the transfer model. In addition, using unfractionated IL-10–deficient T cells may reflect the complexity of different T cell subpopulations in the gut. While it may provide a more comprehensive understanding of the role of CD4⁺ T cells in the development of colitis, there are also some disadvantages to this model. First, IL-10^−/− T cells are unsuitable for analyzing factors regulating IL-10 production in CD4⁺ T cells. Furthermore, using this model to translate the findings to the human condition is challenging, as IL-10–deficient T cells cannot accurately reflect the natural characteristics of T cells in IBD patients. It is also not ideal to evaluate the potential therapeutic interventions for IBD if IL-10 might be the target.

Strengths, Limitations, and Strategies

Each of the adoptive T cell transfer models of colitis has advantages and disadvantages (Table 1). Overall, the colitis in these T cell transfer models is chronic, mimicking human IBD. In addition, based on the goals of the investigations, it is flexible for the investigators to choose the type of T cells and the recipient mice to be used. This allows investigators to study the role of specific T cell subsets in intestinal homeostasis and inflammation. Besides, researchers have a high degree of experimental control over factors, including the timing of treatment and other factors that may influence the development of colitis. All these strengths make the T cell adoptive transfer models an ideal tool for investigating the mechanisms involved in the pathogenesis of IBD and testing potential drugs or other therapies for treating colitis.

Table 1.

Pros and cons of different T cell transfer–induced colitis models.

Model	Pros	Cons
CD4+ CD45RBhi T cells	Widely recognized and used; Genetically modified cells can be used	Complicated cell preparation; Time-consuming; Beginner-unfriendly
CBir1 transgenic T cells	Gut microbiota antigen-specific T cells; Relatively simpler cell preparation	Not commercially available; CBir1-expressing bacteria required
Th1 and Th17 effector T cells	Providing platforms to investigate the function and plasticity of effector cells in colitis	In vitro differentiation required
IL-10–deficient unfractionated CD4+ T cells	Simple cell preparation; Reflecting the complexity of different T cell subsets; Beginner-friendly	Unsuitable for analyzing factors regulating IL-10 production in CD4+ T cells

Abbreviations: IL, interleukin; Th, T helper.

Despite these strengths, it is essential to understand that the T cell transfer models have flaws and limitations, which should be taken into account when interpreting the findings. First, transferring T cells into immunedeficient mice does not accurately reflect the human immune system and the development of colitis in humans. Besides, these models only mimic certain aspects of human colitis, thus limiting their ability to reflect the intricacy of human IBD. The intestinal microbiota in mice differ from those in humans, which could further restrict the relevance of the T cell transfer models to human IBD.

Therefore, it is important to use complementary models (eg, chemical-induced colitis and genetic mice) to validate the findings obtained from T cell transfer models. Using the combinations of a variety of models can help to gain more accurate and reliable conclusions, leading to a more comprehensive understanding of the mechanisms underlying IBD. In addition, it is necessary to increase the relevance of colitis models to human IBD by developing improved techniques, such as incorporating the human gut microbiome and using T cells from human donors (healthy control individuals or IBD patients) transferred into genetically engineered mice that better mimic human IBD.

Conclusions

Adoptive T cell transfer models of colitis are an informative and valuable tool in studying intestinal inflammation, providing insights into the complex interplay of genetic, environmental, and immunological factors that drive the development of IBD. As each T cell transfer model has unique advantages and limitations, careful considerations are needed to choose an ideal model or to combine different models, which depends on the specific questions being investigated and the availability of appropriate resources.

Author Contribution

Conceptualization, draft preparation, review and editing: W.Y. and Y.C.; funding acquisition: Y.C.

Funding

This work was supported by National Institutes of Health grants DK112436, DK125011, and DK135193.

Conflicts of Interest

The authors declare no competing interests.