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中华细胞与干细胞杂志(电子版)

中华细胞与干细胞杂志(电子版) ›› 2017, Vol. 07 ›› Issue (05) : 277 -286. doi: 10.3877/cma.j.issn.2095-1221.2017.05.005

所属专题： 文献；

论著

不同注射途径的脂肪间充质干细胞对大鼠肠炎疗效的实验研究
付正伟1, 张振宇1, 葛海燕1,†()   
  1. 1. 201200　上海，同济大学附属东方医院胃肠外科
  • 收稿日期:2017-05-19 出版日期:2017-10-01
  • 通信作者: 葛海燕
  • 基金资助:
    上海市科学技术委员会面上项目(34119b0600；16411970800); 上海市卫生局面上项目(20134194)

Effect of adipose tissue derived mesenchymal stem cells via different routes of injection on rats with colitis

Zhengwei Fu1, Zhenyu Zhang1, Haiyan Ge1,†()   

  1. 1. Department f Gastrointestinal Surgery, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 201200, China
  • Received:2017-05-19 Published:2017-10-01
  • Corresponding author: Haiyan Ge
  • About author:
    Corresponding author: Ge Haiyan, Email:
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付正伟, 张振宇, 葛海燕. 不同注射途径的脂肪间充质干细胞对大鼠肠炎疗效的实验研究[J]. 中华细胞与干细胞杂志(电子版), 2017, 07(05): 277-286.

Zhengwei Fu, Zhenyu Zhang, Haiyan Ge. Effect of adipose tissue derived mesenchymal stem cells via different routes of injection on rats with colitis[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2017, 07(05): 277-286.

目的

探讨脂肪间充质干细胞（ADMSCs）腹腔及静脉注射对三硝基苯磺酸（TNBS）诱导肠炎疗效的影响。

方法

将32只大鼠完全随机分成正常组、腹腔注射组、静脉注射组及模型组，每组8只。用TNBS诱导炎症性肠病（IBD）动物模型，腹腔及静脉注入ADMSCs，记录大鼠的疾病活动指数（DAI）、观察结肠宏观损伤及微观变化、测定结肠的髓过氧化物酶（MPO）活性、检测结肠Ki-67+细胞的表达、比较血液中IL-1β、TNF-α浓度及结肠TGF-β、IL-6、IL-17A、IL-10的基因表达水平。采用单因素方差分析及独立t检验进行统计学分析。

结果

ADMSCs腹腔注射组（98.05±0.63）g高于静脉注射组[（94.32?±?0.48）g，t?= 12.281，P?=?0.000]，同时腹腔注射组DAI评分1.71±0.75低于静脉注射组3.57?±?0.97，（t?=?-3.980，P?=?0.002）。另外发现腹腔注射组结肠组织内髓过氧化物酶MPO浓度（95.75±5.52）U/g低于静脉注射组（74.37±5.12）U/g，（t =-7.513，P = 0.000），腹腔注射组结肠病理评分2.14?±?0.69低于静脉注射组3.57±0.76，（t =-3.612，P = 0.004）。结肠免疫荧光检查发现腹腔注射组比静脉注射组有更多的Ki-67+细胞。酶联免疫吸附测定（ELISA）发现腹腔注射组血浆中IL-1β的浓度（130.71?±?7.08）pg/ml比静脉注射组（163?±?9.09）pg/ml低，（t =-8.518，P?=?0.000），同样腹腔注射组血浆中TNF-α的浓度（201.71±6.75）pg/ml也比静脉注射组（242.28?±8.30）pg/ml低，（t =-10.033，P = 0.000）。此外，结肠组织实时定量聚合酶联反应（RT-qPCR）的结果显示腹腔注射组IL-6 mRNA的表达4.34±0.48比静脉注射低6.15?±?1.05，（t?=?-4.147，P?=?0.001），腹腔注射组IL-17A mRNA的表达2.61±0.53也比静脉注射低3.57?±?0.46，（t?=?-4.301，P = 0.001）。然而，腹腔注射组IL-10 mRNA的表达水平37.75?±?4.46比静脉注射组高27.68±2.25，（t = 5.327，P?= 0.001），腹腔注射组TGF-β mRNA的表达水平15.82?±?0.99也比静脉注射组高11.97±2.25，（t = 3.740，P = 0.003）。

结论

ADMSCs腹腔注射优于静脉注射并可能成为ADMSCs治疗IBD的较好选择。

关键词: 炎症性肠病, 脂肪间充质干细胞, 注射途径
Objective

To investigate the effect of adipose tissue derived mesenchymal stem cells (ADMSCs) via intraperitoneal and intravenous injection on colitis induced by trinitrobenzenesulfonic acid (TNBS).

Methods

Totally 32 rats were randomly divided into a normal group, an intraperitoneal injection group, an intravenous injection group and a model group (n = 8). The animal model of inflammatory bowel disease (IBD) was induced by TNBS and ADMSCs were administrated intraperitoneally and intravenously. Disease activity index (DAI) were recorded, the microscopic changes and macroscopic damage of colon were observed, myeloperoxidase (MPO) activity were determined and the expression of Ki-67+cell were measured in colon. The expressions of IL-1βand TNF-αin the peripheral blood were compared and the expression level of gene (TGF-α, IL-6, IL-17A, IL-10) were determined in the colon. Single factor analysis of variance and independentttest were used for statistical analysis.

Results

Weight recovery in the intraperitoneal injection group 98.05±0.63 was higher than that in the intravenous injection group 94.32±0.48, (t= 12.281,P= 0.000) in day 6. At the same time, the DAI score in the intraperitoneal injection group 1.71±0.75 was significantly lower than that in intravenous injection group 3.57±0.97, (t= -3.980,P= 0.002). In addition, the MPO concentration of colon tissues in the intraperitoneal injection group (95.75±5.52) U/g was lower than that in the intravenous injection group (74.37±5.12)U/g, (t=-7.513,P= 0.000). The pathological score of colon in the intraperitoneal injection group 2.14±0.69 was lower than that in the intravenous injection group 3.57±0.76, (t= -3.612,P= 0.004). The immunofluorescence of the colonic tissues revealed more Ki67+cells in the intraperitoneal injection group than in the intravenous injection group. ELISA showed that the intraperitoneal injection group (130.71±7.08) pg/ml had lower concentration of IL-1βin plasma than did the intravenous injection group (163?±9.09) pg/ml, (t =-8.518,P= 0.000). The plasma concentration of TNF-αin the intraperitoneal injection group (201.71±6.75) pg/ml was also lower than that in the intravenous injection group (242.28±8.30) pg/ml, (t=-10.033,P= 0.000). In addition, the results of RT-qPCR in the colon tissues showed that the expression levels of IL-6 mRNA in the intraperitoneal injection group (4.34±0.48) were lower than those in the intravenous injection group (6.15±1.05,t=-4.147,P= 0.001) and the expression levels of IL-17A mRNA in intraperitoneal injection group 2.61±0.53 were also lower than those in the intravenous injection group (3.57±0.46,t= -4.301,P= 0.001). However, the expression levels of IL-10 mRNA in the intraperitoneal injection group (37.75±4.46) were higher than those in the intravenous injection group (27.68±2.25,t= 5.327,P?= 0.001) and the expression levels of TGF-βmRNA in the intraperitoneal injection group (15.82±0.99) were higher than those in the intravenous injection group (11.97±2.25,t= 3.740,P= 0.003).

Conclusion

Intraperitoneal injection of ADMSCs is better than intravenous injection, which may be a better route of administration for ADMSCs treating IBD.

Key words: Inflammatory bowel disease, Adipose tissue derived mesenchymal stem cells, Routes of injection
表1 疾病活动指数(DAI)评分
分值 相对体重下降（﹪） 大便隐血 大便性状
0 < 1 阴性 正常
1 1 ~ 5 隐血阳性 软便
2 5 ~ 10 ? 大便松散
3 10 ~ 15 ? 稀糊样便
4 > 20 肉眼血便 水样腹泻
表2 RT-qPCR中的引物序列
基因 正向引物（5'-3'） 反向引物（5'-3'）
IL-17A AAGCACAGAAAGCATGATCCG GAGTCCAGGGTGAAGTGGA
IL-10 CCTTCCTCCGTGTGGTTTGA TTCTTGAGGGCCACTTCGTA
IL-6 GCCAGATAGAGTCGTTGCCC AGCTAGTTGCCGTGTGTCTG
TGF-β CCCACATGAGATCAGCCTCC GGAGTTGGGTGGCAAGAGTT
GAPDH TCAATGAAGGGGTCGTTGAT CGTCCCGTAGACAAAATGGT
图1 倒置荧光显微镜下观察脂肪间充质干细胞（×100）
图2 流式细胞术对脂肪间充质干细胞特征性标记物的检测
表3 各组大鼠每日的相对体重变化(±s)
分组 动物数 第1天 第2天 第3天 第4天 第5天 第6天
正常组 8 101.00±0.84 102.07±0.77 102.74±0.64 103.08±0.40 103.37±0.44 103.74±0.33
腹腔注射组 8 96.80±0.90 94.81±0.80 93.27±0.46 5.22±0.51 96.24±0.56 98.05±0.63
静脉注射组 8 97.55±1.33 93.18±0.53 92.14±0.62 92.70±0.53a 93.47±0.48a 94.32±0.48a
模型组 8 97.95±1.00 92.10±0.67 89.88±0.78 90.67±0.71a 91.41±0.51a 92.37±0.45a
F值 ? 22.145 283.773 548.216 680.569 751.259 727.073
P值 ? 0.186 < 0.001 < 0.001 < 0.001 < 0.001 < 0.001
表4 各组大鼠每日疾病活动指数(DAI)评分(±s)
分组 动物数 第1天 第2天 第3天 第4天 第5天 第6天
正常组 8 0 0 0 0 0 0
腹腔注射组 8 1.42±0.53 6.57±0.97 7.87±1.06 5.87±1.06 3.00±0.81 1.71±0.75
静脉注射组 8 2.14±0.69 6.00±1.15 9.42±0.97 8.14±1.34a 5.42±0.97a 3.57±0.97a
模型组 8 2.14±0.69 7.00±0.81 9.71±0.95 9.14±1.06b 7.28±1.11b 6.00±0.81b
F值 ? 23.077 102.500 195.222 114.651 97.400 84.587
P值 ? 0.024 < 0.001 < 0.001 < 0.001 < 0.001 < 0.001
表5 大鼠结肠髓过氧化物酶(MPO)浓度、结肠长度、结肠宏观损伤评分及组织学评分(±s)
分组 动物数 MPO（U/g） 结肠长度（cm） 宏观损伤评分 组织学评分
正常组 8 56.41±5.57 19.92±1.59 0.18±0.06 0.17±0.07
腹腔注射组 8 74.37±5.12 18.01±0.99 1.57±0.53 2.14±0.69
静脉注射组 8 95.75±5.52a 15.38±0.78a 2.42±0.53a 3.57±0.76a
模型组 8 172.61±11.27a 14.45±1.06a 4.28±0.75a 7.87±0.69a
F值 ? 340.746 32.841 71.795 188.748
P值 ? < 0.001 < 0.001 < 0.001 < 0.001
图3 脂肪间充质干细胞不同注射途径对肠炎宏观损伤的影响
图4 正置荧光显微镜下观察脂肪间充质干细胞不同注射途径对肠炎的病理影响（HE染色，×100）
图5 正置荧光显微镜下观察脂肪间充质干细胞不同注射途径对肠炎结肠黏膜修复的影响（免疫荧光染色，×100）
表6 大鼠血清炎性因子的浓度及结肠局部炎性因子的mRNA的表达(±s)
分组 动物数 IL-1β（pg/ml） TNF-α（pg/ml） IL-6（2-ΔΔCt） IL-17A（2-ΔΔCt） IL-10（2-ΔΔCt） TGF-β（2-ΔΔCt）
正常组 8 81.71±6.67 140.57±7.65 1.08±0.13 1.07±0.22 1.10±0.14 1.34±0.31
腹腔注射组 8 130.71±7.08 201.71±6.75 4.34±0.48 2.61±0.53 37.75±4.46 15.82±0.99
静脉注射组 8 163.00±9.09a 242.28±8.30a 6.15±1.05a 3.57±0.46a 27.68±2.25a 11.97±2.25a
模型组 8 200.42±7.97a 295.00±6.78a 8.60±0.67a 8.78±0.22a 8.25±2.25a 5.41±0.49a
F值 ? 339.835 543.776 137.346 258.390 311.892 132.632
P值 ? < 0.001 < 0.001 < 0.001 < 0.001 < 0.001 < 0.001
1
Wang Y, Ouyang Q. APDW 2004 Chinese IBD working group.ulcerative colitis in China:retrospective analysis of 3100 hospitalized patients[J].J Gastroenterol Hepatol, 2007, 22(9):1450-1455.
2
APDW2004 Chinese IBD Working Group. Retrospective analysis of 515 cases of Crohn's disease hospitalization in China: nationwide study from 1990 to 2003[J]. J Gastroenterol Hepatol, 2006, 21(6):1009-1015.
3
Danese S. New therapies for inflammatory bowel disease: from the bench to the bedside[J].Gut, 2012, 61(6):918-932.
4
樊利芳，董卫国.肠干细胞与结直肠肿瘤干细胞研究进展[J].世界华人消化杂志, 2008, 16(36):4075-4080.
5
Gao X, Yang RP, Chen MH, et al. Risk factors for surgery and postoperative recurrence: analysis of a South China cohort with Crohn's disease[J]. Scand J Gastroenterol, 2012, 47(10):1181-1191.
6
Danese S, Rutella S, Vetrano S. Mesenchymal stromal cells in inflammatory bowel disease:conspirators within the 'colitogenic niche'?[J]. Gut, 2013, 62(8):1098-1099.
7
Liang J, Zhang H, Wang D, et al. Allogeneic mesenchymal stem cell transplantation in seven patients with refractory inflammatory bowel disease[J]. Gut, 2012, 61(3):468-469.
8
Bortolotti F, Ukovich L, Razban V, et al.In vivotherapeutic potential of mesenchymal stromal cells depends on the source and the isolation procedure[J]. Stem Cell Reports, 2015, 4(3):332-339.
9
Ding DC, Chang YH, Shyu WC, et al. Human umbilical cord mesenchymal stem cells: a new era for stem cell therapy[J]. Cell Transplant, 2015, 24(3):339-347.
10
Pacini S, Petrini I. Are MSCs angiogenic cells? New insights on human nestin-positive bone marrow-derived multipotent cells[J]. Front cell Dev Biol, 2014, 2:20.
11
Freeman FE, Haugh MG, Mcnamara LM. Anin vitrobone tissue regeneration strategy combining chondrogenic and vascular priming enhances the mineralization potential of mesenchymal stem cells in vitro while also allowing for vessel formation[J]. Tissue Eng Part A, 2015, 21(7/8):1320-1332.
12
Drakos PE, Nagler A, Or R. Case of crohn's disease in bone marrow transplantation[J]. Am J Hematol, 1993, 43(2):157-158.
13
Griffin MD, Elliman SJ, Cahill E, et al. Concise review:adult mesenchymal stromal cell therapy for inflammatory diseases: how well are we joining the dots?[J]. Stem Cells, 2013, 31(10):2033-2041.
14
Castelo-Branco MT, Soares ID, Lopes DV, et al. Intraperitoneal but not intravenous cryopreserved mesenchymal stromal cells home to the inflamed colon and ameliorate experimental colitis[J]. PLoS One, 2012, 7(3):e33360.
15
Gonçalves Fda C, Schneider N, Pinto FO, et al. Intravenous vs intraperitoneal mesenchymal stem cells administration: what is the best route for treating experimental colitis?[J]. World J Gastroenterol, 2014, 20(48):18228-18239.
16
Obermeier F, Kojouharoff G, Hans W, et al. Interferon-gamma (IFN-gamma)- and tumour necrosis factor (TNF)-induced nitric oxide as toxic effector molecule in chronic dextran sulphate Sodium (DSS)-induced colitis in mice[J]. Clin Exp Immunol, 1999, 116(2):238-245.
17
Rivera DG, Hernández I, Merino N, et al. Mangifera indica L. extract (Vimang) and mangiferin reduce the airway inflammation and Th2 cytokines in murine model of allergic asthma[J]. J Pharm Pharmacol, 2011, 63(10):1336-1345.
18
Strober W, Fuss IJ, Blumberg RS. The immunology of mucosal models of inflammation[J]. Annu Rev Immunol, 2002, 20(1):495-549.
19
Dominici M, Le Blanc K, Mueller I, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement[J]. Cytotherapy, 2006, 8(4):315-317.
20
Bradley PP, Priebat DA, Christensen RD, et al. Measurement of cutaneous inflammation:estimation of neutrophil content with an enzyme marker[J]. J Invest Dermatol, 1982, 78(3):206-209.
21
Tozaki H, Fujita T, Odoriba T, et al. Validation of a pharmacokinetic model of colon-specific drug delivery and the therapeutic effects of chitosan capsules containing 5-aminosalicylic acid on 2,4,6-trinitrobenzenesulphonic acid-induced colitis in rats[J]. J Pharm Pharmacol, 1999, 51(10):1107-1112.
22
Bouma G, Strober W. The immunological and genetic basis of inflammatory bowel disease[J]. Nat Rev Immunol, 2003, 3(7):521-533.
23
Neurath MF, Pettersson S, Meyer Zum Büschenfelde KH, et al. Local administration of antisense phosphorothioate oligonucleotides to the p65 subunit of NF-kappa B abrogates established experimental colitis in mice[J]. Nat Med, 1996, 2(9):998-1004.
24
Sands BE, Anderson FH, Bernstein CN, et al. Infliximab maintenance therapy for fistulizing Crohn's disease[J]. N Engl J Med, 2004, 350(9):876-885.
25
Li LL, Zhang S, Zhang Y, et al. Paracrine action mediate the antifibrotic effect of transplanted mesenchymal stem cells in a rat model of global heart failure[J]. Mol Biol Rep, 2009, 36(4):725-731.
26
Nasef A, Mathieu N, Chapel A, et al. Immunosuppressive effects of mesenchymal stem cells: involvement of HLA-G[J]. Transplantation, 2007, 84(2):231-237.
27
Begue B, Wajant H, Bambou JC, et al. Implication of TNF-related apoptosis-inducing ligand in inflammatory intestinal epithelial lesions[J]. Gastroenterology, 2006, 130(7):1962-1974.
28
Liu Z, Colpaert S, Broeck CD, et al. Expression of interleukin-15 in inflammatory bowel disease[J]. Gastroenterology, 1998, 114(4):A1024.
29
Zhang Z, Zheng M, Bindas J, et al. Critical role of IL-17 receptor signaling in acute TNBS-induced colitis[J]. Inflamm Bowel Dis, 2006, 12(5):382-388.
30
Mangan PR, Harrington LE, O'quinn DB, et al. Transforming growth factor-beta induces development of the T(H)17 lineage[J]. Nature, 2006, 441(790):231-234.
31
Manel N, Unutmaz D, Littman DR. The differentiation of human T(H)-17 cells requires transforming growth factor-beta and induction of the nuclear receptor RORgammat[J]. Nat Immunol, 2008, 9(6):641-649.
32
Sakaguchi S. Naturally arising CD4+regulatory t cells for immunologic self-tolerance and negative control of immune responses[J]. Annu Rev Immunol, 2004, 22(22):531-562.
33
Izcue A, Coombes JL, Powrie F. Regulatory T cells suppress systemic and mucosal immune activation to control intestinal inflammation[J]. Immunol Rev, 2006, 212(1):256-271.
34
Prevosto C, Zancolli M, Canevali P, et al. Generation of CD4+or CD8+regulatory T cells upon mesenchymal stem cell-lymphocyte interaction[J]. Haematologica, 2007, 92(7):881-888.
35
Parekkadan B, Upadhyay R, Dunham J, et al. Bone marrow stromal cell transplants prevent experimental enterocolitis and require host CD11b+splenocytes[J]. Gastroenterology, 2011, 140(3):966-975.
36
Antunes MA, Abreu SC, Cruz FF, et al. Effects of different mesenchymal stromal cell sources and delivery routes in experimental emphysema[J]. Respir Res, 2014, 15:118.
37
Fischer UM, Harting MT, Jimenez F, et al. Pulmonary passage is a major obstacle for intravenous stem cell delivery: the pulmonary first-pass effect[J]. Stem Cells Dev, 2009, 18(5):683-692.
38
Schrepfer S, Deuse T, Reichenspurner H, et al. Stem cell transplantation: the lung barrier[J].Transplant Proc, 2007, 39(2):573-576.
39
Von Bahr L, Batsis I, Moll G, et al. Analysis of tissues following mesenchymal stromal cell therapy in humans indicates limited long-term engraftment and no ectopic tissue formation[J]. Stem Cells, 2012, 30(7):1575-1578.
40
Wang M, Liang C, Hu H, et al. Intraperitoneal injection (IP), Intravenous injection (IV) or anal injection (AI)? Best way for mesenchymal stem cells transplantation for colitis[J]. Sci Rep, 2016, 6: 30696.
41
Lm MM. The problem is obvious, the solution is not:numbers do matter in cardiac cell therapy!EXPERT'S PERSPECTIVE[J].Cardiovasc Res, 2012, 96(2):210-213.
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[11] 王梦, 徐东燕, 张晓雨, 赵海剑. 伴有肛周疾病的炎症性肠病患者肛门功能及生活质量分析[J]. 中华消化病与影像杂志(电子版), 2022, 12(04): 224-227.
[12] 杨翠萍, 全旭, 孙顺昌, 张梦茵, 张金叶, 贾颖, 俞骁珺, 谢玲, 蔡波尔, 吴云林, 陈平. 上海市嘉定区炎症性肠病患者的粪便钙卫蛋白检测研究[J]. 中华消化病与影像杂志(电子版), 2021, 11(05): 193-195.
[13] 江学良. 关于制订我国炎症性肠病中西医结合质量控制标准的建议[J]. 中华消化病与影像杂志(电子版), 2020, 10(06): 241-243.
[14] 费诗茵, 陈洁, 刘军, 王婷婷, 周盟, 何家俊. 炎症性肠病相关药物性胰腺炎的研究进展[J]. 中华临床医师杂志(电子版), 2021, 15(03): 213-217.
[15] 杨全龙, 范崇熙, 石学汇, 徐梦楠, 孙涛, 宁守斌. 炎症性肠病发病机制及与肠道菌群关系的研究进展[J]. 中华胃肠内镜电子杂志, 2022, 09(03): 148-151.
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