| Objective 1. Identification and expansion of rat bone marrow mesenchymal stromal cells(BMSCs) in vitro; 2. Development of experimental ulcerative colitis model in rat;3. Comparison of the efficacy of intracardiac or intravenous transplantation of BMSCs in experimental colitic rats.Methods The surface expression of CD34,CD44,CD45 and CD90 on cultured BMSCs from male SD rats were detected by flow cytometry. These BMSCs can be induced to differentiate into osteogenic or adipogenic cells under certain conditions. The rat UC model was induced by 5%TNBS/ethanol enema. The colitic rats were randomly divided into groups A, B and C.Health rats without any treatment were regarded as normal control(Group D). Rats in group A received intracardial injection of 0.5 ml of BMSCs suspension(1×106) in 24 h after the TNBS/ethanol enema. Rats in group B administered 0.5 ml of BMSCs suspension via tail vein(1×106).Rats in group C were injected equal volume of normal saline via tail vein. Disease activity index(DAI) was recorded at day 7 after BMSCs injection. The rats were then sacrificed and the colonic specimen were collected, fixed and sectioned. The colonic expression of Human Sex-determining region Y protein(SRY protein), Leucine-rich repeat containing G protein coupled receptor 5(Lgr5), Factor Associated Suicide(FAS), inducible Nitric Oxide Synthase(i NOS)were investigated immunohistochemically.Results 1. BMSCs were able to attach the wall of the culture dish and displayed spindle-like in shape. In culture, the cells were arranged in gyrate manner with expression of CD34 of 0.78%, CD44 of 99.38%, CD45 of 0.43%, and CD90 of 98.78%.These BMSCs are able to differentiate into osteogenic or adipogenic cells under certain conditions. 2. The rats administered TNBS/ethanol became dull, body weight reduced and stools loose or purulent.These symptoms reach the peak in 3 days after the enema. 3. After BMSCs transplantion, statistical significance of DAI was detected among groups A, B and C(F=90.603,P<0.05) although the difference between group A and B was not significant.Pathology of the colon disclosed formation of giant ulcer with lymphocyte and neutrophilic cell infiltration. The mucosal injury and inflammation in group A was not as severe as in group B. The SRY protein can be detected in rats received BMSCs transplantation, and its expression in group A was stronger in comparison with group B(t=15.224,p<0.05). BMSCs transplantation reduced the i NOS expression in groups A and B compared with saline controls, and the intracardiac administration of BMSCs was more effective than intravenous injection of the cells(F=33.8,p<0.05). The Fas expression in tissue was significantly different among the animals tested(F=38.6,p<0.05). Compared with the normal control group, the expression of Fas in group C administered saline was increased(t=9.494,p<0.05). This expression was reduced in rats received BMSCs transplantation through either intracardiac or intravenous injections compared with the animal underwent intravenous injection of saline. Furthermore, the Fas expression in group A was lower than those in group B. The LGR-5 expression in the colon tissue of those received BMSCs or saline was significantly different(F=24.7, p<0.05). This expression was significantly increase in those underwent BMSCs transplantation in comparison with that in saline control, and the animals received intracardiac transplantation was even higher than those with intravenous administration of BMSCs.Conclusion Intracardiac injection of BMSCs is more effective than intravenous injection in rats with UC. BMSCs play therapeutic roles for rat with UC via promoting intestinal stem cell regenration, inhibiting intestinal cells apoptosis, diminishinginflammtory response and oxidative stress. |
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