CAJ | 학술논문

背景：间充质干细胞条件培养基被认为是干细胞移植的良好替代方案。既往研究表明，炎症因子的诱导激活能增强间充质干细胞多种生物学潜能，而正常状态的间充质干细胞可能由于免疫活性和迁徙能力不足，无法有效修复损伤组织。目的：探讨炎症预激活前后骨髓间充质干细胞条件培养基对急性辐射损伤小肠上皮治疗作用的差异。方法：分离、培养、鉴定SD幼鼠骨髓间充质干细胞，分别与正常对照和辐射损伤的小肠隐窝细胞株IEC-6在Transwel培养板中共培养24 h，预刺激的骨髓间充质干细胞继续单独培养48 h，收集到的上清液分别为正常状态间充质干细胞条件培养基(MSC-CMNOR)和辐射炎症预激活间充质干细胞条件培养基(MSC-CMIR)。将成年SD大鼠随机分为4组，正常对照组、辐射损伤组、MSC-CMNOR治疗组和MSC-CMIR治疗组，以14 Gy剂量一次性腹部局部照射制备急性小肠辐射损伤大鼠模型，模型制备后尾静脉注射和植入式胶囊渗透压泵腹腔植入联合给药。于治疗后第1，3，7天取小肠组织检测短回路电流和血清木糖水平观察小肠分泌、吸收功能变化。治疗后第3天取小肠组织行电镜观察超微结构改变，治疗后第1，3，5，7，14天取小肠组织行苏木精-伊红染色观察小肠黏膜组织学改变。记录各组大鼠生存状态及生存时间。结果与结论：输注MSC-CM IR后，大鼠小肠短回路电流差值、血清木糖水平较辐射损伤组升高，差异有显著性意义(P <0.05)。苏木精-伊红染色和电镜观察显示从治疗后第3天起MSC-CMIR组小肠黏膜病理改变明显减轻，上皮厚度、紧密连接间隙明显优于辐射损伤组。生存分析显示MSC-CMIR组生存率和平均生存时间明显改善(P <0.05)，而MSC-CMNOR组的各项指标与辐射损伤组比较差异无显著性意义(P >0.05)。结果表明炎症激活状态下的骨髓间充质干细胞条件培养基可促进辐射损伤小肠结构和功能的修复。揹景：間充質榦細胞條件培養基被認為是榦細胞移植的良好替代方案。既往研究錶明，炎癥因子的誘導激活能增彊間充質榦細胞多種生物學潛能，而正常狀態的間充質榦細胞可能由于免疫活性和遷徙能力不足，無法有效脩複損傷組織。目的：探討炎癥預激活前後骨髓間充質榦細胞條件培養基對急性輻射損傷小腸上皮治療作用的差異。方法：分離、培養、鑒定SD幼鼠骨髓間充質榦細胞，分彆與正常對照和輻射損傷的小腸隱窩細胞株IEC-6在Transwel培養闆中共培養24 h，預刺激的骨髓間充質榦細胞繼續單獨培養48 h，收集到的上清液分彆為正常狀態間充質榦細胞條件培養基(MSC-CMNOR)和輻射炎癥預激活間充質榦細胞條件培養基(MSC-CMIR)。將成年SD大鼠隨機分為4組，正常對照組、輻射損傷組、MSC-CMNOR治療組和MSC-CMIR治療組，以14 Gy劑量一次性腹部跼部照射製備急性小腸輻射損傷大鼠模型，模型製備後尾靜脈註射和植入式膠囊滲透壓泵腹腔植入聯閤給藥。于治療後第1，3，7天取小腸組織檢測短迴路電流和血清木糖水平觀察小腸分泌、吸收功能變化。治療後第3天取小腸組織行電鏡觀察超微結構改變，治療後第1，3，5，7，14天取小腸組織行囌木精-伊紅染色觀察小腸黏膜組織學改變。記錄各組大鼠生存狀態及生存時間。結果與結論：輸註MSC-CM IR後，大鼠小腸短迴路電流差值、血清木糖水平較輻射損傷組升高，差異有顯著性意義(P <0.05)。囌木精-伊紅染色和電鏡觀察顯示從治療後第3天起MSC-CMIR組小腸黏膜病理改變明顯減輕，上皮厚度、緊密連接間隙明顯優于輻射損傷組。生存分析顯示MSC-CMIR組生存率和平均生存時間明顯改善(P <0.05)，而MSC-CMNOR組的各項指標與輻射損傷組比較差異無顯著性意義(P >0.05)。結果錶明炎癥激活狀態下的骨髓間充質榦細胞條件培養基可促進輻射損傷小腸結構和功能的脩複。
배경：간충질간세포조건배양기피인위시간세포이식적량호체대방안。기왕연구표명，염증인자적유도격활능증강간충질간세포다충생물학잠능，이정상상태적간충질간세포가능유우면역활성화천사능력불족，무법유효수복손상조직。목적：탐토염증예격활전후골수간충질간세포조건배양기대급성복사손상소장상피치료작용적차이。방법：분리、배양、감정SD유서골수간충질간세포，분별여정상대조화복사손상적소장은와세포주IEC-6재Transwel배양판중공배양24 h，예자격적골수간충질간세포계속단독배양48 h，수집도적상청액분별위정상상태간충질간세포조건배양기(MSC-CMNOR)화복사염증예격활간충질간세포조건배양기(MSC-CMIR)。장성년SD대서수궤분위4조，정상대조조、복사손상조、MSC-CMNOR치료조화MSC-CMIR치료조，이14 Gy제량일차성복부국부조사제비급성소장복사손상대서모형，모형제비후미정맥주사화식입식효낭삼투압빙복강식입연합급약。우치료후제1，3，7천취소장조직검측단회로전류화혈청목당수평관찰소장분비、흡수공능변화。치료후제3천취소장조직행전경관찰초미결구개변，치료후제1，3，5，7，14천취소장조직행소목정-이홍염색관찰소장점막조직학개변。기록각조대서생존상태급생존시간。결과여결론：수주MSC-CM IR후，대서소장단회로전류차치、혈청목당수평교복사손상조승고，차이유현저성의의(P <0.05)。소목정-이홍염색화전경관찰현시종치료후제3천기MSC-CMIR조소장점막병리개변명현감경，상피후도、긴밀련접간극명현우우복사손상조。생존분석현시MSC-CMIR조생존솔화평균생존시간명현개선(P <0.05)，이MSC-CMNOR조적각항지표여복사손상조비교차이무현저성의의(P >0.05)。결과표명염증격활상태하적골수간충질간세포조건배양기가촉진복사손상소장결구화공능적수복。
BACKGROUND:Conditioned medium from mesenchymal stem cels (MSC-CM) may represent a promising alternative to MSCs transplantation. Previous studies have shown that inflammatory activation can strengthen the multiple biological potencies of MSCs; however, normal MSCs with insufficiency of immunocompetence and migration ability are not effective for tissue damage repair. OBJECTIVE:To investigate differential effects of MSC-CM with and without inflammatory activation on radiation-induced intestinal injury.METHODS:MSCs from the bone marrow of SD rats were separated, cultured and identified, and then co-cultured with non-irradiated IEC-6 or irradiated IEC-6 in a transwel system for 24 hours. Then, MSCs with inflammatory activation were cultured alone for another 48 hours. After that, the supernatant was colected as non-activated MSC-CM (MSC-CMNOR) and MSC-CM under radiation-induced inflammatory condition (MSC-CMIR). Rats were exposed to 14 Gy whole abdominal irradiation and randomly divided into four groups: control group, radiation injury group (DMEM/F12), MSC-CMNOR group and MSC-CMIR group. Continuous administration was givenvia tail vein and intraperitoneal implantation of Alzet microosmotic pumps. Intestinal samples were colected at 1, 3, 7 days after radiation for analysis of short circuit variation, at 3 days after radiation for analysis of intestinal epithelium ultrastructure, and at 1, 3, 5, 7, 14 days after radiation for histological observation of the intestinal epithelium using hematoxylin-eosin staining. Blood samples were colected at 1, 3, 7 days after radiation for analysis of serum xylose levels. In addition, the survival state and survival time of rats were observed and recorded. RESULTS AND CONCLUSION: The short circuit variation responding to electrical field stimulation was significantly reduced at al frequencies, but it was significantly improved in the MSC-CMIR group. Similarly, the intestinal absorption (serum xylose levels) was also significantly impaired by irradiation, but improved by delivery of MSC-CMIR (P < 0.05). At 3 days after MSC-CMIR infusion, the intestinal epithelium exhibited an increase in crypt size and vilous length (P < 0.05). Under the electron microscope, a reduction in intestinal microvili and open tight junctions in irradiated intestinal epithelium was found, and the intestine from rats treated with MSC-CMIR had more obvious tight junctions. In addition, treatment with MSC-CMIR dramaticaly improved the survival rate and mean survival time of irradiated rats as compared to those treated with DMEM/F12 or MSC-CMNOR (P < 0.05). Taken together, the present study demonstrated that MSC-CMIR , but not non-activated MSC-CM, improves the structural and functional restoration of the smal intestine after radiation-induced intestinal injury.