Ileal Motility pattern In a Mouse Disease Model of Ulcerative Colitis pattern In a

Background: Ulcerative colitis (UC), is an immune-mediated illness characterized by chronic intestinal inflammation. It is described as a multifactorial disease that involves both genetic and environmental components. UC might result in malabsorption, anemia, and colorectal cancers. There is no definitive treatment for ulcerative colitis, but treatment can help to suppress the inflammatory response and manage symptoms. Objective: The aim of this research project is to study the ileal motility pattern in a disease model of ulcerative colitis in mice. Methods: Ulcerative colitis was induced in mice following specific protocol by adding DSS substance in the drinking water of mice, then small strip of distal part of ileal tissue was cut and threaded to force transducer in pan lab setup which is connected to a computer screen to record muscle longitudinal activity. Acetylcholine and atropine were added and change in muscle activity was observed. Data was analyzed using Graph pad Prism Program (p value ˂0.05 was considered significant). Results: The results showed regular and fast contractions with sharp peaks in both normal and DSS treated mice. It has been found that there is no significant difference in ileal contractions in both groups. Conclusion: The overall findings showed that ulcerative colitis does not affect ileal motility as ileal tissue showed normal response to different chemicals that were added in the experiment. Model of Ulcerative Colitis.


Literature Review
The small intestine is the longest part of the gastrointestinal tract and is about 20 feet long. The journey of food that passes from the stomach into the small intestine passes through three parts: first it enters the duodenum, then the jejunum, and finally the ileum. The ileum is the final section of the small intestine. The ileum follows the duodenum and jejunum and is separated from the cecum by the ileocecal valve. In humans, the ileum is about 2-4 m long, and the pH is usually between 7 and 8 (neutral or alkaline).
The function of the ileum is mainly to absorb vitamin B 12 and bile salts and whatever products of digestion that were not absorbed by the jejunum. The wall itself is made up of folds, each of which has many tiny finger-like projections known as villi on its surface.
It has a large surface area for adsorption of digestion end products.
Layers of circular and longitudinal smooth muscle enable the chyme, which is the digested food, to be pushed along the ileum by waves of muscle contractions called peristalsis. The remaining chyme is then passed to the colon. It is very important as it can be affected in a few diseases such as ulcerative colitis.

Inflammatory bowel disease (IBD) encompasses a range of
intestinal pathologies, the most common of which are ulcerative colitis (UC) and Crohn's Disease (CD). Both UC and CD, when present in the ileum, generate a similar symptom profile which can include diarrhea, rectal bleeding, abdominal pain, and weight loss.
Although the pathogenesis of IBD remains unknown, it is described as a multifactorial disease that involves both genetic and environmental components. UC might result in malabsorption, anemia, colorectal cancers and other complications outside the intestine. There is no specific diagnostic test for ulcerative colitis.
However, some findings on biopsy and barium x-rays, as well as appearance during endoscopy give a clear diagnosis in most cases.
Diagnosis of ulcerative colitis is based on medical history and physical examination, and the results of laboratory and endoscopic tests. There is no definitive treatment for ulcerative colitis, but treatment can help to suppress the inflammatory response and manage symptoms.
There are numerous and variable animal models of colonic inflammation that resemble several features of IBD. Animal models of colitis range from those arising spontaneously in susceptible strains of certain species to those requiring administration of specific concentrations of colitis-inducing chemicals, such as dextran sulphate sodium (DSS). Chemically induced models of gut inflammation are the most used and best described models of IBD [1]. An ideal experimental model of IBD should mimic human IBD, either ulcerative colitis or Crohn's disease (CD), quite closely. In addition, it should be simple to induce, not expensive and highly reproducible (Elson et al. 1995). Unfortunately, there is no single model available that meets these criteria. The dextran sulphate sodium (DSS) model of colitis is widely perceived as a good model of experimental colitis because it has similarities to human IBD in aetiology, pathology, pathogenesis and therapeutic response. In addition, induction is cheap and simple, and may be acute or chronic. (Elson et al. 1995). There has been suggestion that ulcerative colitis might affect the motility pattern of ileum which consequently leads to abnormal function. Therefore, the aim of this is experiment is to study the ileal motility pattern in a disease model of ulcerative colitis in mice. This model represents certain pathophysiological features of human's ulcerative colitis involving weight loss and diarrhea.

Main Objective
The aim of this project is to study the ileal motility pattern in a disease model of ulcerative colitis in mice.

a.
To measure spontaneous ileal motility in normal and diseased mice. b.
To study the effects of different concentration of acetylcholine on ileal motility in normal and diseased mice.

Collection of Ileal Tissue
A mixture of Ketamine (140mg/kg) and Xylazine (40mg/ kg) were injected in mice. Then, the abdomen was cut open in the midline and intestine was transected and transferred to cold oxygenated Ringer's solution. The distal part of the ileum was used to assess motility.

Assessment of Ileal Damage Severity
Macroscopic examinations were done to assess the degree of ileal damage, which included stool consistency and blood in the stool. Bleeding was checked using fecal blood occult test (FOBT).
For this test, a small sample of mouse stool is placed on a special card containing guaiac resin as indicator. The developing reagent is a stabilized alcoholic solution of hydrogen peroxide, due to it is peroxidase like activity hemoglobin catalyzes the oxidation of the indicator by hydrogen peroxide causing the test paper to turn to blue in presence of blood (positive blood occult test). If no change in color occurs the test is considered negative. While stool consistency was assessed using a pair of forceps to press down on the feces to determine its consistency (Table 1).

Data Analysis
Data were compared and analyzed using Microsoft Excel version (7) and Graph pad prism software (San Diego, CA,

USA) version (6) using t-test and analysis of variance (ANOVA).
Significance association between different groups, a p value ˂0.05 was statistically significant.

Assessment of Ileal Damage Severity
Macroscopic disease score was used to determine the severity of ulcerative colitis. Disease group showed stool consistency of different degrees some were soft, very soft and others watery stool.
To determine a score for blood in stool, the color of the stool was noted and further validated using a FOBT and determine the degree using the scoring system Table 1. Most control mice showed score of zero which resembles normal stool and negative blood occult test while disease mice showed bloody soft stool with mild blood occult test (score 1), however some were negative blood occult test.

Response to Different Concentrations of ACh in the Presence and Absence of Atropine
In the distal ileum of wild-type mice, acetylcholine caused concentration-dependent contractions (10nM-10 μM) ( Figure   6). Acetylcholine-induced contractions showed no significant difference in both groups. However, the contractions were slightly higher in the control group. Increasing acetylcholine concentrations in both groups result in increased ileal contractility amplitude These ACh-mediated contractions were inhibited by atropine (2µL) in both groups (Figure 7).

Discussion
The current study showed significant loss in body weight in DSS-treated mice compared to control ones. This finding agrees with other previous study which stated that 20 to 30% body weight loss is a sign of colitis in mice models [2]. Generally, loss in body weight is attributed to diarrhea and loss of appetite (Marchione, 2016). However, the current study found lower percentage of body weight loss compared to previous findings which was (20- This experimental study showed also that there were no

Conclusion
This experiment aimed to study the effects of ulcerative colitis in ileal motility in a mice model which resembles ulcerative colitis in human beings because a better understanding of the pathophysiological grounds of altered ileal motility in ulcerative colitis may lead to a more in-depth knowledge of the accompanying symptoms and to better and more targeted therapeutic approaches.
The overall findings suggest that ulcerative colitis has nothing to do with ileal motility, it does not affect the activity of ileum muscles.
Ileal tissue shows normal response to different acetylcholine concentrations in the presence and absence of atropine. However, our findings lead to additional questions, how ulcerative colitis affects the gut mucosal permeability and studying the response to electrical field stimulation.
Limitations a) Limited number of available mice in animal house.

b)
Mice ileal tissue is very soft so easily can be damaged.