| Objective: To assess the therapeutic effects of adipose-derivedmesenchymal stem cells (Ad-MSC) on radiation-induced intestinalinjury.Method: A total of152male Sprague-Dawley rats, weighting400~450g, were enrolled in this study. Firstly,36rats were selected forevaluating the relationship between the ionizing doses and tissue damage.The rats were classified into four groups. Each group contained9rats.They were respectively irradiated with0,8.5,12and15grays (Gy) totheir whole abdomens. During a period of14days after irradiation, allrats were weighted every other day, and the incidences of diarrhea andoccult blood were recorded. Besides, the histological changes wereanalyzed using H&E staining. The data from the four groups werecompared using One-way ANOVA method. After that, we selected40rats to evaluate the effect of Ad-MSC on maintaining the integrity ofintestinal epithelium after irradiation. Herein,34rats, receiving15Gyionizing irradiation, were used for establishing models ofradiation-induced intestinal injury. Thereafter,17rats were respectivelyadministrated with a dose of5×106cells intrapersonally (Ad-MSCgroup). Another17rats were respectively injected with1.5ml phosphate buffered saline (PBS group). The remaining6rats were used as thecontrols (Normal group). At10,20and30days post-irradiation, theinjured tissue was freshly isolated for analyzing the integrity of intestinalepithelium, including their morphology using H&E staining,proliferative cells and intestinal stem cells within crypts using eitherKi67or Bmi1IHC-staining. TUNEL for apoptotic cells and thegene-profiles containing EGF, KGF, E-Cadherin, Bcl-2and Bax usingReal-time PCR. Besides, the survival of rats and their body-weightvariation were also recored. Next, another40rats were used in the studyof Ad-MSC promoting micro-vascular reconstruction within injuredintestine. The approaches of grouping and modeling were as same asthose in previous study. A t10,20and30days post-irradiation, each3rats from either the PBS group or the Ad-MSC group were sacrificed.The samples were used for identifying the CD31and vWFdouble-positive cells in the injured tissues by IF-staining. In addition,the integrity of vasculature was analyzed by VE-Cadherin IHC-staining,and the density of micro-blood vessels was tested using Isolectin-B4.Markers including CD31, CD34, CD45, CD105, CD133and Ki67wereused for analyzing the mobilized HSC or HPC within injured tissue.Besides, the expressions of VEGF, HGF and SDF-1in irradiatedintestine were also tested using real-time PCR. Lastly, the remaining36rats were used for analyzing the anti-inflammatory effect of Ad-MSC. There were6,15and15rats in the normal, PBS and Ad-MSC groups,respectively. At3.5,7,10,20and30days post-irradiation, each3ratsfrom either the PBS group or the Ad-MSC group were sacrificed. Theperipheral blood and thymus were harvested for computing thepercentages of CD4/CD25/Foxp(3) triple-positive regulatory T cells.Besides, levels of IL-10in serum were tested using ELISA method. Thedeposition of collagen was detected by using Masson staining.IHC-staining for MPO was used for identifying the infiltration ofinflammatory cells within injured tissues. The data from the PBS andAd-MSC groups were compared using Paired t-test.Result: We observed that the degrees of tissue damage depended on theionizing doses, and there were the typical lesions of radiation-inducedintestinal injury in the rats receiving15Gy irradiation. As comparedwith the PBS group, Ad-MSC could promote the proliferation of cellswithin crypts and increase the the number of Bmi1-positve intestinalstem cells through the high expressions of EGF and KGF. Besides, theup-regulated expression of Bcl-2and down-regulated expression of Baxin the Ad-MSC group inhibited the apoptosis of epithelial cells. Theresults also showed that Ad-MSC could promote the neovascularizationin the damaged intestine by triggering mobilization of HSC and HPCtogether with up-regulated expressions of VEGF, HGF and SDF-1.Additionally, after delivery of Ad-MSC, the percentages of CD4/CD25/Foxp(3) triple-positive regulatory T cells increased in boththymus and peripheral blood, which resulted in a high serum level ofIL-10facilitating inflammation in injured tissues.Conclusion: Ad-MSC had the healing roles on radiation-inducedintestinal injury, representing by the restoring the integrity of epithelium,promoting neovascularization within injured tissues and mitigating theinflammation. Consequently, Ad-MSC had the potentials formanagement of radiation-induced intestinal injury. |
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