Use of Animal Models in the Study of Colitis Use of Animal Models in the Study of Colitis

Inflammatory bowel diseases (IBDs) relate to chronic inflammations in different parts of the gastrointestinal (GI) tract involving both ulcerative colitis (UC) and Crohn’s dis ease (CD). Ulcerative colitis begins in the rectum and extends continuously up the colon. Notably, CD may affect any area of the GIT, from the mouth to the anus. Various condi tions may influence the genesis of the disease, such as genetics, environment, intestinal microbiota and the presence of agents of enteric infections. Experimental models are therefore suitably used to investigate the various etiological factors; similarly colitis can be induced by genetic modification, cell transfer, spontaneous inflammation and chemi cal agents. The objective of this chapter is to present current concept on animal models of inflammatory bowel diseases. These models are crucial for the understanding of inflam matory bowel diseases, development of alternative treatments and more effective thera - peutic agents thus contributing to the control of the disease.


Introduction
The immune system has a mechanism of response to tissue injury and antigenic stimuli, whose main function is to protect the body. This response is called inflammation. It is characterized by the interaction of several types of cells and signaling molecules, to promote the repair of damaged tissue [1,2]. Despite its protective function, the inflammatory response must not be prolonged, since exacerbated inflammation may cause physical loss of function in the organ [3].
The various manifestations characterized by chronic inflammation include inflammatory bowel diseases (IBDs), such as ulcerative colitis (UC) and Crohn's diseases (CD) [4,5]. Their clinical presentations can be overlapping as well as distinct [6,7]. UC is generally associated to superficial inflammation of the colon mucosa and CD comprises transmural damage that may occur in any segment of the gastrointestinal tract (GI) [8].
The IBDs are common among white people between the ages of 20 and 40 years old and the second peak of the disease occurs around 55 years of age. The distribution of this disease in males and females seems similar, though CD is more frequent in women [9].
The genesis of UC and CD involves multiple factors, such as genetic, environmental, intestinal microbiota and the presence of enteric infections. These factors may affect the immune system, by expanding and perpetuating the inflammatory process [10].
The treatment of IBD is considered unsatisfactory, because among other factors, cure may not be possible, only remission of the disease and/or treatment of symptoms. Several classes of drugs are currently used to treat IBD, such as aminosalicylates, corticosteroids and immunosuppressives [11]. The odds remain that these drugs have many side effects, are expensive and lose their efficacy after a certain period of medication [12].
Therefore, concise experimental studies are needed to unravel the etiology and pathophysiological mechanisms of IBD for development of alternative treatments and/or more effective therapeutic agents to treat patients with ulcerative colitis. Thus, the use of experimental animal models becomes essential to elucidate the mechanism(s) involved and accelerate the development of a specific and effective therapy.

Animal models for inflammatory bowel disease: colitis
The use of animal models is crucial for the investigation of the processes involved in the genesis of IBD. Although the development of the disease in animals does not present all the mechanisms observed in humans, animal models still are the most effective way to study the factors involved in the pathogenesis of BD. It would be difficult to conduct such a study in humans. Additionally, the animals are very useful for the assessment of new therapeutic approaches [13].
Over time, several experimental models of IBD were developed to conduct experimental studies, particularly mice and rats. The animal models of intestinal inflammation can be divided into four groups according to the type of induction namely: genetic modification, cell transfer, spontaneous inflammation and chemical agents [14][15][16].
The ideal experimental model is characterized by morphological disorders and intestinal inflammation, in addition to clinical symptoms, pathophysiology and development of the disease similar or equal to human condition IBD [17]. The chapter thus presents animal models developed for the study of and experimentation on colitis.

Genetic modification
The development and use of genetically modified animals have contributed to the investigation of various issues related to the IBD, and the most frequent are connected to the triggering of inflammation through the impairment of various functions such as epithelial barrier and detection of bacterial components, signaling of the innate immune response, immune regulation and signaling response to stress [18]. Studies involving the human genome have shown the association of more than 160 genes connected to IBD [19] (Table 1).
When the intestinal epithelial barrier is impaired, the intestinal mucosa has contact with bacteria from the lumen, which generates the inflammatory responses that will lead to the development of IBD. Studies have demonstrated the relationship between CD and certain genes involved in the intestinal epithelial barrier function. Animal models that express defects in this barrier are discussed and are essential in the investigation of IBD pathogenesis [18].
a. C57BL/6 mouse expressing a dominant negative N-cadherin (NCADΔ) N-cadherin is a glycoprotein related to epithelial cell adhesion and is essential for normal development. Mice with a dominant-negative mutation of N-cadherin developed porous regions in the intestinal epithelium and intercellular adhesion disorder, developing transmural inflammation of the jejunum similar to CD at 3 months old. The organization of the small intestinal epithelium makes it possible to use chimeric mice, generated from normal blastocyts and genetically manipulated embryonic stem cells, to study cadherin function. Therefore, embryonic stem cells were transfected with a dominant negative N-cadherin mutant (NCADΔ) under the control of promoters active in small intestinal epithelial cells and then introduced into C57BL/6 mouse blastocysts [20].
b. FVB mice FVB mice originated from Swiss syngeneic strains subjected to processes that triggered intolerance or sensitivity to histamine. FVB mice were derived from the syngeneic Swiss strains, selected from crosses in which progenies received pertussis vaccination and challenged with histamine thus developing resistance or sensitivity to it. From this experimental system, two strains were selected: one resistant to the histamine factor (HSFR/N) and another sensitive (HSFS/N). Within the latter group, in the early 1970s, a group of eight syngeneic generations, HSFS/N, carrying Fv-1b alleles and susceptible to the Friend leukemia virus, lineage B, were selected. They are good breeders with large progenies and the eggs of the FVB/N lineage favor in vitro fertilization procedures since it has a huge pro-nucleus that facilitates DNA microinjection and the production of transgenic [21]. The surface of the intestinal mucosa has many components essential for its protection, such as MUC2 that has a lubricating function and also provides a protective barrier between the contents of the lumen and mucosal layers. Wild-type (Muc2 + / + ), Muc2 + / − and Muc2 − / − littermates are mostly used and scores recorded weekly until the age of 16 weeks to obtain a disease activity index (DAI). The previously mentioned Muc2 −/− mice were backcrossed onto a 129SV (Charles River, Maastricht, The Netherlands) genetic background for nine generations followed by intercrosses to breed animals homozygous for the Muc2 disruption. Mice lacking this component may develop IBD in the distal colon at approximately 5 weeks of age [22]. Lack of Muc2 Development of IBD in the distal colon [22] Specific deletion of C1galt1 in epithelial cells Inflammation in the distal colon [23] MDR1a gene deleted Development of colitis [24] Absence of IRE1 α Spontaneous colitis [26] Nuclear kappa B (NF-kB) deactivated in intestinal epithelial cells Development of colitis with apoptosis of epithelial cells [27,28] Specific deletion of type II receptor for TGB-β in CD4+-specific T cells and dendritic cells Spontaneous inflammation in several organs and colitis [29,30] Absence of SMAD4 Inflammation through the entire gastrointestinal tract [31] Absence of IL-2 and deficiency of JAK 3 (component of IL receptor) Autoimmune disease with colitis and other disorders; spontaneous colitis [32][33][34] Absence of IL-10 and deletion of CRF2-4 (component of IL-10 receptor) Spontaneous development of colitis; chronic colitis [38,39] Absence of TCR α Spontaneous triggering of colitis [40] Absence of STAT3 in macrophages and neutrophils Spontaneous onset of colitis [41] Super expression of STAT4 and immunization with DNP-KLH/CFA Development of transmural colitis [43] Absence of Gαi2 Triggering of colitis and colorectal cancer [44][45][46] Absence of enzyme A20 Inflammation in the intestine and in other organs [47] Absence of T-bet and T and B cells Onset of colitis similar to UC [48] Altered levels of TNF-α Development of inflammation of the ileum and in the distal colon [49][50][51] Super expression of IL-7 Spontaneous onset of colitis [53] Absence of Gpx1 and Gpx2 Triggering of colitis and ileitis [54] Specific deletion of XBP1 factor Development of ileitis [55]  Deletion of C1galt in epithelial cells was also induced in adult animals and, 10 days later, the development of colitis was also observed [23].

e. MDR1a −/− mice
The multiple drug resistance gene (MDR1) codes the multiple drug carrier (P-glycoprotein) in intestinal epithelial cells. Around 25% of FVB mice with the gene MDR1a "deactivated" developed colitis at 1 year of age [24]. In the Japanese population, the MDR1 gene was related to susceptibility to late onset of UC [25]. To obtain FVB.mdr1a−/− mice, FVB control mice and mdr1a −/− mice were bred and maintained in the SPF facility. Mdr 1a −/− mice had been backcrossed onto the FVB strain for at least seven generations for models obtained at Taconic Farms [24].

g. NEMO IEC−KO mice
Animals with deactivation of nuclear factor-kappa B (NF-ĸβ) binding in intestinal epithelial cells, through conditional ablation of NEMO or both IKK1 (IKKa) and IKK2 (IKKb)-IKK subunits essential for NF-ĸβ activation, have an exacerbated response to luminal microbiota, which leads to the development of colitis with apoptosis of epithelial cells at the age of 6 weeks. To investigate the function of NF-ĸβ signaling in the gut epithelium in vivo, this was generated in mice lacking NEMO specifically in intestinal epithelial cells (NEMO IEC−KO mice) by crossing mice carrying loxP-flanked (floxed, FL) NEMO (Ikbkg) alleles with Villin-Cre transgenics. These experiments were performed using mice backcrossed into the C57BL/6 genetic background for at least five generations. They were used as wild-type controls. The mice used were housed in individually ventilated cage systems, in either specific pathogen-free or conventional animal facilities [27,28]. Therefore, regardless of the immune system, intestinal cells clearly provide protection against bacterial pathogens of the microbiota [18].

Transgenic animal models expressing colitis through modulation of immune regulatory agents
Mice with several genetic disorders associated with immune regulation were examined, revealing the development of colitis that was related to different substances [18]. These models with genetic modifications that affect immune regulation and trigger IBD are discussed below.
a. Type II receptor for TGF-β or SMAD4 absence The transforming growth factor-beta 1 (TGF-β1) acts both in immune regulation and in the differentiation of regulatory T cells. Transgenic animals with dominant-negative mutant TGF-β type II receptor in specific CD4+ T cells showed spontaneous inflammation at around 3-4 months of age, and there was increased expression of Th1 and Th2 cytokines in several body organs such as colon, liver, stomach, duodenum, pancreas and kidney. These animals were created by using DNA microinjection into (C57BL/6xC3H) F1 fertilized oocytes, which results in six transgene-positive mice. All pups in the experimental cycle appeared normal at 2 weeks of age, but three died between 2 and 4 weeks of age. Three remaining transgenepositive mice appeared healthy and were bred; their progeny was further used to identify transgenic lines expressing a functional dominant-negative TGF-β receptor type II [29].
SMAD4 acts as a mediator in intracellular signaling of TGF-β. To inactivate specifically Smad4 in T cells, it was necessary to cross mice with a transgene encoding a Cre-recombinase driven by the Lck promoter (Smad4co/co; Lck-Cre) or the CD4 promoter (Smad4co/co; CD4-Cre) with the strain carrying the Smad4 conditional allele (Smad4co/co). Mice without SMAD4 in specific T cells and those without SMAD4 in CD4+ T cells had inflammation through the entire gastrointestinal tract at 3 months of age [31].

b. Mutations affecting the interleukin receptor (IL-2)
Interleukin-2 (IL-2) is related to activated-induced cell death and to the triggering of regulatory T cells function. Deficient IL-2 mice develop spontaneous autoimmune colitis, gastritis, hepatitis, pneumonia, pancreatitis, nephritis and hemolytic anemia [32]. Janus tyrosine kinase 3 (JAK 3) acts as a transducer of interleukin-2 receptor common gamma chain. Deficiency of JAK 3 led to the spontaneous development of colitis in mice [33]. The animal model that uses IL-2 is characterized by association of colitis with the autoimmune mechanism, which was demonstrated in the spontaneous onset of colitis without the presence of microorganisms [34].
Similarly, IL-10 is a regulatory cytokine with IBD susceptibility genes not only in adults but also in children [35][36][37]. IL-10 deficient mice develop spontaneous colitis [38]. Mice with deleted CRF2-4, a component of the IL-10 receptor, develop splenomegaly and disorders such as chronic colitis [39].

c. Mice with T cell-related mutations
The T cell receptor (TCR) is essential for the recognition of antigens by T cells. Approximately 60% of mice without TCR α, TCR component, developed Th2-mediated spontaneous colitis.
The animals used in this study were produced by crossing TCR β × mutant mice with TCR mutant mice, and generated TCR β × gamma double mutant mice, which are deficient both in αβ and γ cells. By mutating, the recombination-activating gene RAG-1 mice totally deficient in mature T and B lymphocytes were produced [40].
The transcription factor, signal transducer and activator of transcription 3 (STAT3) acts in the control of various innate immune responses such as differentiation of Th17 cells and epithelial renovation. Its gene is susceptible to UC and CD [35,36]. Animals with intestinal epithelial cells deficient in STAT3 developed spontaneous colitis. In response to inflammatory stimuli, LysMcre/Stat flox/− mice develop enterocolitis even though they produce more IL-10 than normal animals in the same condition. To disrupt the Stat3 gene specifically in macrophages and neutrophils, two types of animals were crossed, mice in which the Stat3 gene is flanked by two loxP sites (Stat3 flox/flox ) and the other mouse in which the Cre cDNA is inserted into the lysozyme M gene by a knock-in approach (LysMcre mice) [41].
STAT4 is a transcription signal transducer and activator factor that acts in the development of Th1 cells. Its gene was considered susceptible to UC in individuals with white skin [42]. d. Gαi2 −/− mice Gαi2 signal transduction proteins participate in a wide signaling system. Mice that lack Gαi2 proteins showed dysfunctions related to T cells and developed colitis when they were 3-4 weeks old and colorectal cancer at approximately 3-4 months of age. Gαi2-deficient (Gαi2 −/− ) mice on a 129SvEv X C57BL/6 background (backcrossed four or five generations into 129SvEv and then intercrossed) and on a pure 129SvEv background were used [44][45][46].
e. A20 −/− mice A20 is an enzyme related to ubiquitin, which in turn is associated to NF-ĸβ1 activation. It acts in the prevention of tissue destruction and inflammation mediated by innate immune cells, in addition to regulating skin differentiation. To study the functions of A20 in vivo, A20-deficient (A20 −/− ) was generated by gene targeting. Rodents that lack this enzyme developed inflammation in various organs, including bowel, when they were 3-6 weeks olds [47].
f. T-bet −/− mice T-bet is a transcription factor that regulates the differentiation of T cells into Th1 cells. To investigate whether the T-bet's protective function is linked to innate immunity, the T-bet −/− mouse developed on the RAG2 −/− background was examined. The mice experimental model that lacked the T-bet factor showed no colitis. On the other hand, a new mouse model through which animals without T-bet were crossed with RAG2 mice lacking T and B cells showed the development of colitis similar to what occurs in UC at 4 weeks of age, therefore, T-bet expression protects against colitis and T-bet −/− × RAG2 −/− mice develop spontaneous colitis [48].

g. TNF ΔARE mice
TNF-α is related to the onset of CD, because of the development of spontaneous inflammation in the ileum of mice with increased TNF-α activity [49]. Animals with intrinsic defect related to post-transcriptional regulation of TNF mRNA (TNF ΔARE rat model) developed inflammation, especially in the terminal ileum and subsequently in the proximal colon [50,51]. To develop TNFΔARE mice, a 69bp deletion encompassing TNF ARE was conducted in embryonic stem cells in conjunction with the neomycin resistance marker (neo) flanked by loxP sequences. The excision of the neon gene and generation of TNFΔARE rat lines was due to the transient expression of the Cre recombinase in fertilized oocytes from the previously mentioned mice. The TNFΔARE homozygous mice developed early pathologies and also had a short shelf life. To reverse this problem, the effects of the TNFΔARE mutation on TNF biosynthesis were investigated in TNFΔARE/− hemizygous mice generated by the crossing of TNFΔARE/+ to TNFΔARE/− mice. The use of hemizygous TNFΔARE/− mice made it possible to investigate the regulatory role of the ARE element in a monoallelic system without the interference of a wild type TNF allele in the generation of TNF. So, a defective function of ARE may be etiopathogenic for the development of analogous human pathologies, in this case, Crohn's-like inflammatory bowel disease.
h. SRα/IL-7 mice IL-7 has an important role in the homeostasis of T cells, and the gene linked to the IL-7 receptor was related to UC [35,36]. To study the IL-7 role, it was critical to develop the model SRα/ IL-7 transgenic mice using 2.1 kb SRα/lL-7 DNA fragment that was linearized by AatH and ApaU digestion. Microinjection into the pronuclei of fertilized eggs of C57BL/6 J mice was performed with ~10 3 copies of the SRα/lL-7 DNA fragment according to the standard procedure [52]. Rodents with super expression of IL-7 developed spontaneous colitis at about 4-12 weeks of age [53].

i. Double-KO mice deficient in both Gpx1 and Gpx2 gene expression
Activation of endoplasmic reticulum (ER) stress responses of intestinal epithelial cells to the metabolic by products of bacteria in the intestinal lumen is necessary for intestinal homeostasis and prevention of chronic inflammation [18]. The glutathione peroxidase (GPX) enzyme is selenium-dependent and can reduce a broad range of hydroperoxides. The inflammatory process is associated with increased lipid peroxidation, and hydroperoxide appears to mediate IBD-related symptoms. To clarify the role of GPX in the intestinal epithelial cells, a homozygous double-KO mice with interruption in the expression of Gpx1 and Gpx2 genes was generated. These animals have signs and symptoms related to inflammatory bowel disease. The Ye-Shih Ho (Wayne State University, Detroit, MI) generated Gpx1-KO mice, as C57BL/6 J and 129Sv/J hybrids. Likewise, the generation of Gpx2-KO mice as B6 and 129S3 hybrids occurred at the City of Hope (COH; Duarte, CA) Transgenic Mouse Core. These mice were housed at the COH Animal Resources Center in ventilated cage racks. So, it was possible to verify that mice deficient in Gpx1 and GPX2 developed spontaneous colitis and ileitis GPX2 at about 3 weeks of age [54].
The transcription factor XBP1 is involved in the response to ER stress. XBP1 was related to CD and UC. To understand the function of this factor, Xbp1 exon 2 was almost completely disrupted in the intestinal tissue (small intestine and colon). Mice with specific deletion of this factor in intestinal epithelial cells developed spontaneous ileitis due to the lack of Paneth cells that secrete protective cells [55].

Adoptive transfer model of colitis
The referred methodology induces intestinal inflammation through selective transfer of certain types of immune cells in animals with impaired immune systems (RAG-1 knockout and RAG-2 knockout and SCID pigs). The study based on these models allows the investigation of immune factors, particularly the role of pathogenic T cells and T cell-mediated colitis on mucosal inflammation [18] ( Table 2).

a. CD4+ CD45RB hi model
The transfer of T CD4+ cells with high levels of protein CD45RB to mice deficient in T cells (RAG −/− or SCID) is the transfer methodology most commonly used to induce colitis, and it increases intestinal inflammation approximately 5-10 weeks after the transfer [56][57][58]. This method causes an inflammation similar to that of models that use chemical agents and is characterized by hyperplasia and inflammation in the colon, generally causing the death of the animal [59].

CD4 + CD45RB hi
Intestinal inflammation with Th1 responses to antigens of bacteria in the lumen [56][57][58][59][60][61] SheLAg-specific CD4+ T cells Colitis caused by one single bacterial antigen [62] Hsp60-specific CD8+ T cells Severe inflammation in the mucosa of the small intestine, regardless of the animal's microbiota [63]  This model of cell transfer triggers Th1 responses to bacteria in the lumen [60]. After cell transfer, naive T cells react to antigens of the luminal bacteria and expand rapidly in the lymph nodes and colon mucosa, becoming oligoclonal. The presence of pathogenic effector Th1 cells was possible due to the absence of regulatory T cells [18,61].
b. SheLAg-specific CD4+ T cells H. hepaticus Ag (SHelAg)-specific CD4+ Th1 clones transfer the disease to RAG1 knockout mice deficient in T cells and infected with a pathogenic bacterium (H. hepaticus) causing colitis between 7 and 8 weeks after the transfer, while the non-infected receptors did not develop colitis. It is inferred, therefore, that pathogenic T cells that respond to a single bacterial antigen may lead to the development of colitis [62].

c. Hsp60-specific CD8+ T cells
In general, heat shock proteins (HSP) are antigens considered potential targets to autoimmune disease. Then, the transfer of CD8+ T cells responsive to HSP60 for immunodeficient animals can trigger severe inflammation in the mucosa of the small intestine regardless of the animal's microbiota [63].
The severity of inflammation by adoptive transfer depends on the donor animal, as well as to the recipient [64,65]. A study aimed to protocol standardization found that Balb/SCID mice develop a more severe and faster type of colitis C57BL/6 RAG-1 −/− mice [66].

Models of spontaneous colitis
These models concern both the animals that develop spontaneous inflammation in the mucosa and those with inadequate mucosal immune response due to defect or genetic modification [56]. Spontaneous models are of great interest for the study of intestinal inflammation since, as in disease in humans, there is no external manipulation. The murine model C3H/HeJBir, for example, is characterized by chronic inflammation in the colon and in the cecal region [67] (Table 3).

a. C3H/HeJBir mouse model
Because of the occurrence of soft, light-colored feces and/or secondary perianal ulceration, and histological evidence of colitis, without pathogens, mice of the C3H/HeJ strain are likely to have a genetic predisposition to develop a form of IBD, with perianal ulceration, it has the pedigree to generate a "high-susceptibility" substrain, C3H/HeJBir, with a high incidence of spontaneous colitis [68].

Models
Main results References

C3H/HeJBir
Chronic inflammation in the colon and cecal region similar to CD [67, 69]

SAMP1/YitFc
Severe inflammation in the terminal ileum, similar to CD [70,71]  The wounds observed in this mouse model occur in the ileocecal region and the colon. Clinical manifestations begin at 3-4 weeks of age and last up to 10-12 weeks of age, approximately. The characteristics of the colitis developed by this methodology are similar to those of CD [17,69].
b. SAMP1/YitFc model Investigation of the mechanisms of CD can be based on the model that uses SAMP1/YitFc mice, where there is spontaneous and/or severe inflammation of the terminal segment of the ileum, which is the primary site of the wounds in CD, this disease model in animals also resembles human CD with regard to histologic features [15,70,71]. For the purposes of these experiments, a colony of SAMP1/Yit mice in a barrier facility at the University of Virginia was established from animals provided by Yakult Central Institute for Microbiological Research (Tokyo, Japan). The SAMP1/Yit (H-2 k ) mouse strain was originally derived from AKR mice (original parents purchased from The Jackson Laboratories, Bar Harbor, Maine, USA) [70]. The findings from the adoptive transfer research have made it possible to infer that CD4+ T cells that generate a Th1-like cytokine profile, for example, IFN-γ and TNF, act as mediators of intestinal inflammation observed in SAMP1/ Yit mice.

Chemically induced models of intestinal inflammation
Experimental models of colitis induced by chemical agents are used in the analysis of pathological mechanisms of IBD as well as the development of therapeutic agents. The referred method facilitates the induction of colitis. Some chemicals, such as dextran sodium sulfate (DSS) and trinitrobenzene sulfonic acid (TNBS) are the most commonly used substances. Models with such agents have been used for more than 20 years for the investigation and circumstances that affect the development of IBD [16] ( Table 4).
Induction of inflammation by chemical agents can cause disorders in the barrier of the intestinal mucosa and/or cause hapten-induced hypersensitivity reactions [18]. Several factors are related to the magnitude of the onset of colitis in models induced by chemical substances. Although these models do not have the complexity of human IBD, they can contribute to the study of the processes involved in the control of the disease [72].

DNBS
Extensive tissue damage and acute inflammatory process similar characteristics to CD [80] Oxazolone Inflammation of the distal colon with clinical similarities to UC [74]  Use of Animal Models in the Study of Colitis http://dx.doi.org/10.5772/intechopen.75608 a. DSS approach DSS methodology has been used to induce colitis since 1985 because the disease induced is very similar to the human disease [73]. Its mechanism of action is related to direct epithelial toxicity, which impacts on the epithelial barrier of the intestinal mucosa [18]. In the referred methodology, the substance is diluted in water for animal drinking, for a period ranging from 5 to 7 days. The inflammation induced by DSS has characteristics similar to diarrhea with bleeding, ulcerations and granulocyte infiltration. Dysplasia often occurs during the chronic stage of the inflammation, which is very similar to ulcerative colitis in humans [16,74].
The inflammatory immune response occurs immediately after disturbance of the intestinal epithelium, due to contact of the antigens in the lumen with the immune cells of the mucosa and submucosa areas [72].
Acute and chronic forms of the disease can be induced, according to the doses and treatment cycles. DSS was also used to induce IBD in transgenic and immunodeficient animals, and most of them are prone to intestinal inflammation. Also, the symptoms and the pathology are more severe with the association of this chemical agent [18]. DSS-induced colitis is frequently used in studies on innate immune processes involved in the onset of IBD and also in restoring the intestinal barrier integrity [72].

b. TBNS
In hapten-induced colitis, the TNBS substances dinitrobenzene sulfonic acid (DNBS) or oxazolone dissolved in ethanol can be administered rectally. The type of response triggered (Th1/ Th2) will depend on the hapten selected, as well as on the susceptibility of the animal, and route of administration of the chemical agent. Ethanol is used to disrupt the barrier of the intestinal mucosa, while the agent (TNBS, DNBS or oxazolone) is associated with an autologous protein or microbiota byproducts, becoming immunogenic, and triggering the interaction of specific antigens and cells of the immune system [18,75,76]. The IBD models induced by hapten, similar to those that use DSS, can also be applied in transgenic and immunodeficient animals, as well as in those susceptible to immune-mediated colitis [77][78][79][80].
In the beginning of the experiment with TNBS-induced model, one single dose can be administered, resulting in acute inflammation, with Th1 response occurring in 2-3 days. Initial sensitization with TNBS can be done by rectal application or via skin, and another administration must be performed 6 days later for the development of delayed hypersensitivity response to TNBS-haptenized colonic proteins [81].
TNBS is administered rectally with ethanol, amidst haptenization of the proteins from the microbiota and the emergence of T CD4+ cells. The severity and the extent of the inflammation depend on genetic factors inherent in the animals and the presence or absence of bacteria that activate T cells [71]. T CD4 + cells are strongly related to Th1-mediated immune responses through IL-12 cytokines, culminating in transmural colitis similar to CD [80,82].
The IBD model that uses DNBS is similar to the one induced by TNBS, because there is extensive tissue damage and acute inflammatory process. Also, as it occurs in CD, it depends on T CD4+ cells [80].
Oxazolone is generally administered after subcutaneous pre-sensitization (abdomen), which stimulates greater formation of Th2 cytokines and inflammation of the distal colon resembling ulcerative colitis [74].
Like TBNS, oxazolone is also administered rectally. Acute inflammation of the distal mucosa and submucosa generally involves ulcers and infiltrations of neutrophils, macrophages and lymphocytes. The type of response triggered depends on the dose of the substance administered. Overall, this model is useful for the investigation of specific characteristics of the inflammatory process in chronic IBD [72]. Use of Animal Models in the Study of Colitis http://dx.doi.org/10.5772/intechopen.75608 Table 5 shows a comparative presentation of the various ways colitis disease is induced as discussed herein. Each of the several models discussed here have similarities with human IBD, and offer advantages for the study of certain aspects involved in the pathogenesis of the disease. Induction by chemical agents is often used widely by researchers mainly because it is easier, although it does not contribute to the elucidation of inflammatory and immunological processes involved in IBD [14,18]. Genetic modification allows the study of specific aspects present in IBD; however, such form of induction does not allow to analyze the disease in a global way, and contributes less to the test of efficacy of new drugs [14,18,19]. Cellular transfer helps to study specific issues, contributing to the investigation of the role of regulatory T cells and mucosal immune regulation, but adaptive immunity alone does not act in the onset of IBD [14,18]. The spine development of colitis reproduces the disease as in humans (since there is no intervention), allowing the investigation of complex and multifactor situations that are present in the disease, nevertheless, there is a low availability of models reported in the literature [14,18,66].

Final considerations
The animal models described here can contribute to the understanding of various mechanisms involved in IBD, as well as to the development of new drugs to improve the treatment. It would be extremely difficult to carry out these investigations and experimentation in humans. However, due to a good number of animal models available for this disease, care should be taken in the selection of animal/model choice. Investigators should take into consideration the purpose of their study, for example, the type of compound to be tested and/or pathophysiological mechanisms to be investigated.