| Aim:1. To establish mouse model of colon slow transit motility and study the pathogenesis of slow-transit constipation.2. To study the change of ultrastructure characters of interstitial cells of Cajal (ICC) and the relationship between ICC and enteric nerves system in mouse colon slow-transit motility.3. To investigate the change of ICC and nitric oxide synthase(NOS) in mouse colon slow-transit motility, and relationship between them.Methods:1. The mouse model was established by subcutaneous injection of morphine, the group I of test (1.25mg/kg/d × 45d, n=10), the group II of test (2.5mg/kg/d × 45d, n=10), and one control group(the same dosage of 9 g/L sodium chloride solution/d × 45d,n=10). Fecal weight and character were recorded daily, and transit function of intestine was measured by activated charcoal suspension pushing test in order to compare with test groups and control group.2. Tissue of adult ICR mouse colon in two test groups and control group were observed with conventional electron microscopy, including their ultrastructure, distribution of ICC and the relationship between ICCand the enteric nerves.3. The mouse model have been established and obtain proximal and distal colon tissue. All tissues was embedded by paraffin wax. The change of ICC and NOS were observed by immunohistochemistry, and the relationship between ICC and nNOS were observed by double immunofluorescence.Results:1. Fecal weight daily was no difference in three groups, but fecal character in the I and II groups of test was more dry and hard(Bristol classes: l-2grade) than in control group(Bristol classes: 4-5grade). Intestinal transit rate in the group I of test was 51.1 ± 14.1 % , the group II of test was 49.7 ± 13.6 % , and control group was 66.9 ±13.1 % , the two test groups were lower than control group (P<0.05 ), but there were no difference in the two test groups (P>0.05 ) .2. ICC were characterized by long processes, large oval nuclei and scant perinuclear cytoplasm. The cytoplasm contained numerous mitochondria, abundant smooth and rough endoplasmic reticulum. ICC in the circular muscle layer and longitudinal muscle layer of mice colon formed synaptic-like contacts and their processes were often associated with neighboring nerve trunks. The group I of test, ICC were characterized by short and small processes, swelling mitochondria, vague mitochondrial crista, unintegrity nucleimembrane. The group II of test,ICC characterized by decreased cytoplasm, more shorter and smaller processes, expanding endoplasmic reticulum, unintegrity nuclei membrane.3. The area of c-kit~+ cells in mouse proximal colon tissue of the group I of test was 2.00 ± 0.44 ten thousand μm2, the group II of test was 1.27 ±0.34 ten thousand μm2, and control group was 4.28 ±0.38 ten thousand μm2, two test groups were more obviously decreased than control group (P<0.01) , and there was a significant difference in the group II of test vs the group I of test (P<0.05 ) . The area of nNOS~+ cells in mouse proximal colon tissue of the group I of test was 1.69 ± 0.27 ten thousand μm2, the group II of test was 1.25 ±0.23 ten thousand μm2, and control group was 2.38 ± 0.22 ten thousand μm2, two test groups were more obviously decreased than control group (P<0.01) , and there was a significant decrease in thegroup II of test vs the group I of test (P<0.05 ). There was no difference in the area of c-kit~+ cells and nNOS~+ cells of mouse distal colon tissue of two test groups and control group (P>0.05) . c-kit~+ cells and nNOS~+ were expressed in identical sites of mouse colon tissue of two test groups and control group by double immunofluorescence.Conclusions:1. There are dry and hard fecal character and decrease of intestinal transit rate in mouse of morphine induced colon slow-transit motility.Basically, this mouse model conform to clinic and pathophysiology of slow-transit constipation.2. ICC in colon tissue have unique ultrastructure characters, ICC ultrastructure characters have changed in morphine induced colon slow...
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