A Research On Correlation Between AQP-1,AQP-4 And Slow Transit Constipation

Objective:Oweing to the changing of dietetic habit and accelerating of pace of living, constipation has already become a common disease which affects people's health and daily life and work seriously in current society. In order to solve this problem, WHO(World Health Organization) especially formulated Rome-Ⅲdiagnostic criteria to guide the diagnosis and treatment of constipation. Among all types of constipation, slow transit constipation is the most common and serious one with the following clinical symptoms: the volume of stool is too small, hard feces, having difficulty during defecation, and sometimes with some special symptoms, such as: defecating forcibly prolongedly,having sensation of rectum fullness or sensation of incomplete bowel evacuation , even defecting with hand's help. Or defecting spontaneously less than twice in one week, or has no awareness of defection for long time without the help of cathartics. Constipation patients especially those patients with slow transit constipation usually suffer a lot, and it always causes other diseases doing harm to people's health. Therefore, curing constipation thoroughly becomes a responsibility of medical workers. Medical worker can cure the disease only if the etiology is found, but the research on etiology of constipation is still at a starting stage. Given this fact, this research attempts to investigate the etiology of slow transit constipation from the perspective of molecular biology. Techniques such as immunohistochemistry and image analysis are used to investigate the difference in AQP-1, AQP-4 expression on the colonic mucosa of patients with slow transit constipation and normal subject. At last, the role that AQP-1 and AQP-4 play in the occurrence and development of slow transit constipation will be deduced from the difference.Method: Take the cases from rectum surgical department of the Third Hospital of Hebei Medical University(cases from Sep.,2008-Nov.,2009 ) who conform to Rome-Ⅲcriteria and without organic diseases such as intestinal obstruction and megacolon after first visit, inquiring disease history in detail, physical examination and laboratory examination. Give each patient of the 25 cases with slow transit constipation 3 markers for 120 hours, and the results showed that there are more than 20% of markers still remain in the colon of the patients ranging from 72-120 hour. After treating all the patients with subtotal colectomy surgery, take two samples respectively from proximal ascending colon and distal descending colon of each patient. Take 20 samples of ascending and descending colon from normal subject(without history of constipation or taking cathartics assisting defecation in long-term and diarrhoea and analogous diseases history ) as control group. Group Method: Samples are divided into four groups:①experimental group1: 25 ascending colon samples taking from 25 slow transit constipation patients(2 males, 23 females, with age from 36-71, averaging 48.60 years old; Constipation history of 1-23 years, averaging 5.25 years);②Experimental group2: descending colon samples from the above 25 patients;③Control group1:20 cases of ascending colon samples from patient without constipation history, 1 female, 19 females, with age from 33-68 years old, averaging 43.70 years;④control group2:taking samples of descending colon from the patients of control group 1. The difference of gender, age, disease history between experimental group and control group are not considered to be statistically significant.All the above samples are perfusion fixed by 4% paraformaldehyde for 24 hours, then the fixed explants will get through conventional gradient dehydration of alcohol, Xylene Transparence, paraffin-embedded in consequence. Then, the embedded paraffin will be serial sectioned by slicer, with a thickness of 5μm; next, four pieces of them are mounted on slide glass coated with protein glycerol ( used in ordinary HE staining ) and polylysine(used in Immunohistochemical staining ); at last, stain the chosen slice with HE and Immunohistochemistry. Finally, observe the expression of AQP-1 and AQP-4 through high-powered microscope and detect the difference of AQP-1 and AQP-4 expression in the colon of normal subject and patient with slow transit constipation using manual counting and automatic image analysis.Results:1. AQP-1 has expressions in colonic mucosa epithelial cell top,basolateral and cavity surface of 25 cases of slow transmit constipation and 20 cases of normal people's, and mainly expresses in absorptive cell with small amount of expression in goblet cells, with the positive expression rate of 100%. Its expression in ascending colon and descending colon has no significant difference(p>0.05).However, AQP-1 expression in colonic mucosal cell of patients with slow transit constipation(experimental group) is obviously higher than that in normal subject(control group). Statistical analysis showed the AQP-1 expression in colonic mucosa of patients with slow transit constipation expression(experimental group) and normal subject(control group) has significant difference, P.<0.05 was considered to be statistically significant.2. There is no AQP-4 expression in the ascending and descending colonic mucosal epithelial cell of normal subject(control group), but small amount of expression in ascending colon of slow transmit constipation patient(experimental group) and no expression in descending colon. Among 25 cases of patients with slow transit constipation, only 12 cases are found to have a small amount of AQP-4 expression in the substrate parts of ascending colonic mucosal epithelial cell with positive expression rate less than 50%. Statistical analysis showed the expression of AQP-4 on colonic mucosa in patients with slow transit constipation expression(experimental group) and normal subject(control group) has no significant difference, P.>0.05 was not considered to be statistically significant.Conclusion:1. AQP-1 has expression in total colon and the expression in ascending and descending colon is same, but its expression in the colon mucous of patient with slow transit constipation is obviously higher than in the colon mucous membrane of normal subject. This phenomenon indicates AQP-1 plays an important role in the process of colon reabsorbing water and AQP-1 may influence the occurrence and development of slow transit constipation.2. AQP-4 has no expression in normal subject's total colon, but has small amount of expression in slow transmit constipation patient's ascending colon and no expression in their descending colon. This indicates AQP-4 doesn't play an important role in colon's re-absorption of water, only participating a little. Therefore, AQP-4 impacts the occurrence and development of slow transmit constipation little.