**Animal Inhalation Models to Investigate Modulation of Inflammatory Bowel Diseases Inflammatory Bowel Diseases**

**Animal Inhalation Models to Investigate Modulation of** 

DOI: 10.5772/intechopen.69569

Giuseppe Lo Sasso, Walter K. Schlage, Blaine Phillips, Manuel C. Peitsch and Julia Hoeng Phillips, Manuel C. Peitsch and Julia Hoeng Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

Giuseppe Lo Sasso, Walter K. Schlage, Blaine

http://dx.doi.org/10.5772/intechopen.69569

**Abstract**

Inflammatory bowel diseases (IBDs) comprise primarily two disease manifestations, ulcerative colitis (UC) and Crohn's disease (CD), each with distinctive clinical and pathological features. Environmental and clinical factors strongly affect the development and clinical outcomes of IBDs. Among environmental factors, cigarette smoke (CS) is considered the most important risk factor for CD, while it attenuates the disease course of UC. Various animal models have been used to assess the impact of CS on intestinal pathophysiology. This chapter examines the suitability of animal inhalation/smoke exposure models for assessing the contrary effects of CS on UC and CD. It presents an updated literature review of IBD mouse models and a description of possible mechanisms relevant to relationships between IBD and smoking. In addition, it summarises various technical inhalation approaches, in the context of mouse disease models of IBD.

**Keywords:** inhalation, inflammatory bowel disease, animal models, cigarette smoke, ulcerative colitis, Crohn's disease

#### **1. Introduction**

Inflammatory bowel disease (IBD) is a chronic inflammatory condition of the gastrointestinal tract encompassing two main disease manifestations, Crohn's disease (CD) and ulcerative colitis (UC) [1].

CD and UC have many similarities in symptoms and disease phenotypes, making diagnosis challenging [2]. Currently, criteria for distinguishing these two manifestations are based exclusively on histopathological and endoscopic examinations [3]. Thus, UC is defined as a chronic, non-transmural inflammatory disease characterised by diffuse mucosal inflammation

Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons

involving only the colon. Its primary clinical symptom is bloody diarrhoea [2, 4–7]. As UC is an inflammatory disease, the state of the immune system is a fundamental aspect of the disorder, with an atypical T helper cell (Th)2 response, mediated by natural killer T cells that secrete interleukin (IL)-13 [1, 8, 9]. CD is a relapsing, transmural inflammatory disease that may affect the entire gastrointestinal tract. Its major clinical symptom is abdominal pain or nonspecific abdominal symptoms and bloody diarrhoea is rare. The T cell profile in CD is different from that of UC and, in fact, a Th1 cytokine profile is dominant in patients with CD [4, 7, 10, 11]. Notably, innate immune responses are similarly activated in both CD and UC [12]. Several studies suggested that IBD pathologies result from an inappropriate inflammatory response to intestinal microbes in a genetically susceptible host, with consequent alteration of the intestinal epithelium.

genetic engineering technologies, enabled the development of numerous murine models that, in turn, substantially increased the understanding of intestinal inflammatory processes [23, 24]. Arguably, none of these models can completely recapitulate the complexity of human IBD, but they can provide valuable information about major aspects of the disease, thereby enabling a common set of principles of human IBD pathogenesis to be established. This book chapter reviews key studies conducted in animal inhalation/smoke exposure models aimed at evaluating the different modulation of UC and CD by CS. The application of inhalation technology to rodents, reproducing the clinical effects of smoking on colonic inflammation, will increase the chances of identifying new anti-inflammatory molecular

Animal Inhalation Models to Investigate Modulation of Inflammatory Bowel Diseases

http://dx.doi.org/10.5772/intechopen.69569

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mechanisms and possibly therapeutics, finally increasing the chances of IBDs defeat.

**2.1. Methods of acute and chronic pulmonary delivery of aerosols to rodents**

The technical means for pulmonary delivery of aerosols (either small molecules, proteins or mixtures) may employ either direct intratracheal administration or, alternatively, inhalation

For acute pulmonary delivery of an agent, intratracheal administration may be ideal. Its main advantages are that it requires little infrastructure or equipment and can be performed in a basic *in vivo* lab environment [25]. In addition, dose delivery can be accurately and reproducibly estimated [26]. However, this method also has several shortcomings, such as need for anaesthesia, inability to administer volatile agents or gases and unequal distribution in the lungs, resulting in minimal exposure to the alveoli. Overall, such concerns make intratracheal administration a less suitable method for subchronic or chronic pulmonary

For subchronic or chronic administration of aerosols to rodents, repeated inhalation exposure systems are preferred. Thus, animals are exposed to aerosols within a confined environment for a fixed daily duration. In the field of toxicology, testing guidelines for repeated dose exposure for toxicological assessments, such as the OECD TG413 guideline, recommend up to 6 h per day exposure for a 90 day exposure period. However, for therapeutic or disease modelling purposes, the exposure period must be determined empirically, based on the effective dose and the time needed for the target biological effect to occur. Importantly, exposure systems must enable consistent delivery of aerosols, at concentrations that are stable during the exposure period, and with appropriate aerosol properties to enable efficient inhalation

Principally, two types of exposure chambers are routinely used to administer aerosols to rodents, whole body or nose-only exposure chambers, each with its own advantages and disadvantages [27]. Whole body exposure systems are restraint free, as the animals are placed into an exposure chamber, either in a cage or on a mesh or grid surface, depending on the specific system. Both chambers are technically simple, assuming sufficient infrastructure (aerosol

**2. Technical aspects of inhalation**

delivery.

and uptake [27].

exposure, the latter often requiring restraint of animals.

During IBD development, the paracellular space in the intestinal epithelium becomes more permeable, impacting defensive strategies naturally activated by specialized epithelial cells, including goblet and Paneth cells [13–16]. This process primes a positive feedback loop, with increased exposure to the intestinal microbiota, leading to amplification of the inflammatory response. Observations in patients or animal models show that host-microbiome interactions and microbiome fluctuations play prominent roles in such inflammatory processes [17, 18]. However, whether these alterations contribute to the disease, or simply reflect secondary changes caused by the inflammation, is still under debate.

Indeed, the basic aetiology of IBD is still unclear and the potential factors contributing to the pathogenesis of the disease, such as dysbiosis, epithelial and/or immune system dysfunctions and oxidative stress, represent the major research topics in the IBD field. Moreover, new area of interest arose from the necessity of understanding the potential environmental causes behind the disease onset.

Among the environmental factors associated with IBDs, the most significant causes are cigarette smoke (CS) and nicotine, and these inversely affect the risk and course of UC and CD. The relationship between smoking and IBD has been known for many years, with the first report of a negative correlation between IBD and smoking, in a cohort of UC patients, published 40 years ago [19]. Since then, there have been numerous epidemiological, clinical and pre-clinical studies describing the dual effects of active smoking in the two forms of IBD [20, 21]. CS is associated with a higher risk for developing CD and a worse outcome in CD patients. In contrast, UC is considered a non-smokers' disease, with a significantly lower risk of disease development in current smokers. Despite the considerable research on smoking and IBD, the molecular mechanisms for CS-induced impacts on IBD development, as well as the specific CS components responsible, are not well understood [22].

To better understand the different aetiological factors in the onset of IBD, a variety of disease models were developed. Human and *in vitro* studies have historical limitations because of design complexity, duration and cost or, for *in vitro* studies, the lack of translational applicability. Therefore, animal models are advantageous by allowing *in vivo* experiments to be conducted under more easily controlled conditions than those in human studies, while providing the organism complexity lacking in *in vitro* systems. Increased knowledge of mucosal immunity and host-microbiome interactions and dynamic, as well as the availability of new genetic engineering technologies, enabled the development of numerous murine models that, in turn, substantially increased the understanding of intestinal inflammatory processes [23, 24]. Arguably, none of these models can completely recapitulate the complexity of human IBD, but they can provide valuable information about major aspects of the disease, thereby enabling a common set of principles of human IBD pathogenesis to be established.

This book chapter reviews key studies conducted in animal inhalation/smoke exposure models aimed at evaluating the different modulation of UC and CD by CS. The application of inhalation technology to rodents, reproducing the clinical effects of smoking on colonic inflammation, will increase the chances of identifying new anti-inflammatory molecular mechanisms and possibly therapeutics, finally increasing the chances of IBDs defeat.
