脐带间充质干细胞体外调节再障患者T淋巴细胞表达IL-2，IL-6，IL-17，IFN-γ的研究
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摘要
目的：从人脐带中分离、培养间充质干细胞(mesenchymal stem cells MSCs)并予以鉴定,研究其对再生障碍性贫血(aplastic anemia AA)患者外周血单个核细胞的部分细胞因子表达的调节作用,以期为临床治疗再障、移植物抗宿主病及提高异基因造血干细胞移植成功率等提供细胞学方法及实验依据。
方法：采用组织块培养法从脐带分离、培养MSCs；用流式细胞术(FCM)鉴定MSCs的表型；分离再障患者外周血单个核细胞与MSCs共培养；用ELISA法定量测定共培养前后细胞因子IL-2、IL-6、IL-17、IFN-γ的含量。
结果：脐带来源MSCs表型为HLA-I+, CD29+, CD31-, CD44+, CD105+;脐带来源MSCs与再生障碍性贫血患者外周血单个核细胞共培养,共培养前后各个因子浓度的均数及标准差分别为：IL-2：共培养前32.303±8.461pg/ml共培养后36.076±25.339 pg/ml,(P=0.573>0.05),差异无统计学意义；IL-6：共培养前3.372±1.741 pg/ml,共培养后1141.062±347.634 pg/ml,(P=0.001<0.05),差异有统计学意义,共培养后浓度显著上升；IL-17：共培养前6.610±1.261 pg/ml,共培养后12.714±3.500 pg/ml,(P<0.01),差异有统计学意义,共培养后浓度上升；IFN-γ：共培养前42.302±23.851pg/ml,共培养后86.508±59.266 pg/ml,(P=0.031＜0.05),差异有统计学意义,共培养后浓度上升。
结论：研究证明可以从脐带组织中成功分离出脐带MSCs,其具有与骨髓MSCs相似的表型特征；将其与再生障碍性贫血患者外周血单个核细胞共培养,使再生障碍性贫血患者外周血T淋巴细胞表达IL-6,IL-17,IFN-γ增加,而IL-2的浓度无明显改变；共培养使非再障血液病患者外周血T淋巴细胞表达IFN-γ减少,使非再障血液病患者外周血T淋巴细胞表达IL-6增加,对IL-17表达无明显影响。
objective:isolating and culturing mesenchymal stem cells from human umbilical cord, and identifying them, to study the regulation effects to partial cytokines secretion of aplastic anemia patients' Peripheral blood mononuclearcell, it also provide Cytologic method and experimental basis for clinical therapy of aplastic anemia and graft versus host disease.
Methods:mesenchymal stem cells were isolated and cultured from human umbilical cord by the tissue pieces culture method; analising mesenchymal stem cells'markers by flow cytometry; isolating aplastic anemia patients' Peripheral blood mononuclearcells and coculturing with mesenchymal stem cells; detecting the cytokines IL-2, IL-6, IL-17, IFN-γconcentration before and after coculturing by enzyme linked immunosorbent assay;
Results:The human umbilical cord mesenchymal stem cells' markers are HLA-I+, CD29+, CD31-, CD44+, CD 105+; human umbilical cord mesenchymal stem cells cocultured with aplastic anemia patients' Peripheral blood mononuclearcells, before and after coculturing, the cytokines concentration are:IL-2:before coculturing 32.303±8.461pg/ml, after coculturing 36.076±25.339pg/ml(P=0.573＞0.05). difference has no statistical significance; IL-6:before coculturing 3.372±1.741 pg/ml, after coculturing 1141.062±347.634 pg/ml (P=0.001＜0.05), difference has statistical significance, the concentration rised after coculturing; IL-17: before coculturing 6.610±1.261 pg/ml, after coculturing 12.714±3.500 pg/ml (P=0.000＜0.05), difference has statistical significance, the concentration rised after coculturing; IFN-γ:before coculturing 42.302±23.851 pg/ml, after coculturing 86.508±59.266 pg/ml (P=0.031＜0.05), difference has statistical significance, the concentration rised after coculturing.
Conclusion:Mesenchymal stem cells can be successfully isolated from human umbilical cord tissue; coculturing mesenchymal stem cells with aplastic anemia patients'Peripheral blood mononuclearcells, the IL-6, IL-17, IFN-γsecretion of aplastic anemia patients'Peripheral blood mononuclearcells rised, IL-2 has no statistical significance; IL-2,IL-6 secretion of control group rised after coculturing; IFN-γsecretion of control group decreased after coculturing; IL-17 of control group has no statistical significance.
方法：采用组织块培养法从脐带分离、培养MSCs；用流式细胞术(FCM)鉴定MSCs的表型；分离再障患者外周血单个核细胞与MSCs共培养；用ELISA法定量测定共培养前后细胞因子IL-2、IL-6、IL-17、IFN-γ的含量。
结果：脐带来源MSCs表型为HLA-I+, CD29+, CD31-, CD44+, CD105+;脐带来源MSCs与再生障碍性贫血患者外周血单个核细胞共培养,共培养前后各个因子浓度的均数及标准差分别为：IL-2：共培养前32.303±8.461pg/ml共培养后36.076±25.339 pg/ml,(P=0.573>0.05),差异无统计学意义；IL-6：共培养前3.372±1.741 pg/ml,共培养后1141.062±347.634 pg/ml,(P=0.001<0.05),差异有统计学意义,共培养后浓度显著上升；IL-17：共培养前6.610±1.261 pg/ml,共培养后12.714±3.500 pg/ml,(P<0.01),差异有统计学意义,共培养后浓度上升；IFN-γ：共培养前42.302±23.851pg/ml,共培养后86.508±59.266 pg/ml,(P=0.031＜0.05),差异有统计学意义,共培养后浓度上升。
结论：研究证明可以从脐带组织中成功分离出脐带MSCs,其具有与骨髓MSCs相似的表型特征；将其与再生障碍性贫血患者外周血单个核细胞共培养,使再生障碍性贫血患者外周血T淋巴细胞表达IL-6,IL-17,IFN-γ增加,而IL-2的浓度无明显改变；共培养使非再障血液病患者外周血T淋巴细胞表达IFN-γ减少,使非再障血液病患者外周血T淋巴细胞表达IL-6增加,对IL-17表达无明显影响。
objective:isolating and culturing mesenchymal stem cells from human umbilical cord, and identifying them, to study the regulation effects to partial cytokines secretion of aplastic anemia patients' Peripheral blood mononuclearcell, it also provide Cytologic method and experimental basis for clinical therapy of aplastic anemia and graft versus host disease.
Methods:mesenchymal stem cells were isolated and cultured from human umbilical cord by the tissue pieces culture method; analising mesenchymal stem cells'markers by flow cytometry; isolating aplastic anemia patients' Peripheral blood mononuclearcells and coculturing with mesenchymal stem cells; detecting the cytokines IL-2, IL-6, IL-17, IFN-γconcentration before and after coculturing by enzyme linked immunosorbent assay;
Results:The human umbilical cord mesenchymal stem cells' markers are HLA-I+, CD29+, CD31-, CD44+, CD 105+; human umbilical cord mesenchymal stem cells cocultured with aplastic anemia patients' Peripheral blood mononuclearcells, before and after coculturing, the cytokines concentration are:IL-2:before coculturing 32.303±8.461pg/ml, after coculturing 36.076±25.339pg/ml(P=0.573＞0.05). difference has no statistical significance; IL-6:before coculturing 3.372±1.741 pg/ml, after coculturing 1141.062±347.634 pg/ml (P=0.001＜0.05), difference has statistical significance, the concentration rised after coculturing; IL-17: before coculturing 6.610±1.261 pg/ml, after coculturing 12.714±3.500 pg/ml (P=0.000＜0.05), difference has statistical significance, the concentration rised after coculturing; IFN-γ:before coculturing 42.302±23.851 pg/ml, after coculturing 86.508±59.266 pg/ml (P=0.031＜0.05), difference has statistical significance, the concentration rised after coculturing.
Conclusion:Mesenchymal stem cells can be successfully isolated from human umbilical cord tissue; coculturing mesenchymal stem cells with aplastic anemia patients'Peripheral blood mononuclearcells, the IL-6, IL-17, IFN-γsecretion of aplastic anemia patients'Peripheral blood mononuclearcells rised, IL-2 has no statistical significance; IL-2,IL-6 secretion of control group rised after coculturing; IFN-γsecretion of control group decreased after coculturing; IL-17 of control group has no statistical significance.
引文
1 Young NS,. Calado RT, Scheinberg P. Current concepts in the pathophysiology and treatment of aplastic anemia.Blood.2006,108(8),2509-2519.
2 Bacigalupo A, Oneto R, Bruno B,etal. Current Results of Bone Marrow Transplantation in Patients with Acquired Severe Aplastic Anemia. Acta Haematol 2000; 103:19-25
3 Corcione A, Benvenuto F, Ferretti E,etal. Human mesenchymal stem cells modulate B-cell functions. Blood,2006,107 (1):367-372.
4 Nauta AJ, Kruisselbrink AB, Lurvink E,etal. Mesenchymal Stem Cells Inhibit Generation and Function of Both CD34+-Derived and Monocyte-Derived Dendritic CellsJ. Immunol,2006,177(4): 2080-2087.
5 Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood,2005; 105(4):1815-1822
6 Le Blanc K, Rasmusson I, Sundberg B,etal. Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells. The Lancet,2004; 363(9419),1439-1441
7 Zappia E, Casazza S, Pedemonte E,etal. Mesenchymal stem cells ameliorate experimental autoimmune encephalomyelitis inducing T-cell anergy. Blood,2005; 106 (5):1755-1761
8 Gerdoni E, Gallo B, Casazza S,etal. Mesenchymal Stem Cells Effectively Modulate Pathogenic Immune Response inExperimental AutoimmuneEncephalomyelitis. Ann Neurol 2007;61(3):219-227
9 Le Blanc K, Tammika C, Rosendahlc K,etal. HLA expression and immunologic propertiesof differentiated and undifferentiated mesenchymal stem cells. Exp Hematol.2003; 31 (10): 890-896
10 叶任高.陆再英.内科学。人民卫生出版社,2006：572-576。
11 张之南。血液病学。北京：人民卫生出版社.2003：618-633.
12 Young NS,. Calado RT, Scheinberg P. Current concepts in the pathophysiology and treatment of aplastic anemia.Blood.2006,108(8),2509-2519.
13 司徒镇强,吴军正。细胞培养。世界图书出版公司,2006：58-64。
14 Abdallah BM, Boissy P, Tan Q, etal. dlkl/FA1 Regulates the Function of Human Bone Marrow Mesenchymal Stem Cells by Modulating Gene Expression of Pro-inflammatory Cytokines and Immune Response-related Factors. [J]. THE JOURNAL OF BIOLOGICAL CHEMISTRY 2007;282 (10).7339-7351.
15 Tang KC, Trzaska KA, Smirnov SV,etal. Down-Regulation of MHC Ⅱ in Mesenchymal Stem Cells at High IFN-{gamma} Can Be Partly Explained by Cytoplasmic Retention of CIITA.[J]. The Journal of Immunology.2008;180(3);1826-1833.
16 Romieu-Mourez R, Francois M, Boivin MN.etal. Cytokine Modulation of TLR Expression and Activation in Mesenchymal Stromal Cells Leads to a Proinflammatory Phenotype.[J]. The Journal of Immunology,2009,182(12),7963-7973
17 Stagg J, Pommey S, Eliopoulos N.Interferon-γ-stimulated marrow stromal cells:a new type of nonhematopoietic antigen-presenting cell [J].Blood,2006,107(6):2570-2577.
18 Francois M, Romieu-Mourez R, Stock-Martineau S, etal. Mesenchymal stromal cells cross-present soluble exogenous antigens as part of their antigen-presenting cell properties. Blood, 2009,114 (13):2632-2638.
19 Eliopoulos N, Stagg J, Lejeune L,etal. Allogeneic marrow stromal cells areimmune rejected by MHC class I and II mismatchedrecipient mice. Blood.2005;106:4057-4065.
20 Glennie S, Soeiro I, Dyson P, etal. Bone marrow mesenchymal stem cells induce division arrest anergy of activated T cells. [J].Blood,2005 105 (7).2821-2827
21 Hu KX, Zhao SF, Sun QY,etal. Effect of bone marrow mesenchymal stem cells on immunoregulation in H-2 haploidentical bone marrow transplantation mice. Zhonghua Xue Ye Xue Za Zhi,2007; 28(8):505-9.
22 Beyth S, Borovsky Z, Mevorach D,etal. Human mesenchymal stem cells alter antigen-presenting cell maturationand induce T-cell unresponsiveness. Blood,2005; 105(5): 2214-2219.
23 Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood,2005; 105(4):1815-1822
24 Guo Z, Zheng C, Chen Z, etal. Fetal BM-derived mesenchymal stem cells promote the expansion of human Th17 cells, but inhibit the production of Thl cells. Eur J Immunol,2009; 39(10):2840-9
25 Gu Y, Hu X, Liu C, etal. Interleukin (IL)-17 promotes macrophages to produce IL-8, IL-6 and tumour necrosis factor-alpha in aplastic anaemia. Br J Haematol,2008; 142(1):109-14.
26 Zoumbos NC, Gascon P, Dieu JY,et al. Circulating activated suppressor Tlymphocytes in aplastic anemia. N Engl J Med,1985;312:257-265
27 Nistico A, Young NS. γ——interferon gene expression in the bone marrow of patients with acquired aplastic anemia. Ann Inter Med,1994; 120:463-469
28 Morris TC, Vincent PC, Young GA,et al. CFU C inhibitors in aplastic anemia. Blut,1984;48:61-65
29 Kawano Y, Takaue Y, Hirao A. Synergistic effect of recombinant interferon- gamma and interleukin- 3 on the growth of immature human hematopoietic progenitors. Blood,1991;77:2118-2121
30 Tichelli A, Gratwohl A, Wursch A,et al. Late haematological complications in severe aplastic anemia. Br J Haematol,1988; 69:413-418
31 于文强,孙秉中。免疫功能紊乱与再生障碍性贫血。细胞与分子免疫学杂志1999年第1期第11卷：1365-1370
32 Rasmusson I, Le Blanc K, Sundberg B,etal. Mesenchymal stem cells stimulate antibody secretion in human B cells. Scand J Immunol,2007; 65(4):336-43.
33 Horwitz EM. Culture conditions shape mesenchymal stromal cell phenotype and function. Cytotherapy,2009; 11(2):101-2
34 Zhu GR, Zhou XY, Lu H,etal. Human bone marrow mesenchymal stem cells express multiple hematopoietic growth factors. Zhongguo Shi Yan Xue Ye Xue Za Zhi,2003; 11(2):115-9.
35 Chen B, Hu J, Liao H, etal. Flk-1+ mesenchymal stem cells aggravate collagen-induced arthritis by up-regulating interleukin-6. Clin Exp Immunol.2010; 159(3):292-302.
36 Qiao Z, Zhao XJ. Study on Mechanism of Human Marrow Mesenchymal Stem Cell in Treating Patients with Aplastic Anemia. Blood (ASH Annual Meeting Abstracts) 2007 110: Abstract 4099
37 Jin JG, Hu JW, Ning HM, The effect of MSC on cytokine production by naive T cell differentiated in vitro. Zhonghua Xue Ye Xue Za Zhi.2005; 26(6):339-41.
1 Abdallah BM, Boissy P, Tan Q, etal. dlkl/FA1 Regulates the Function of Human Bone Marrow Mesenchymal Stem Cells by Modulating Gene Expression of Pro-inflammatory Cytokines and Immune Response-related Factors. [J]. THE JOURNAL OF BIOLOGICAL CHEMISTRY 2007; 282(10):7339-7351.
2 Tang KC, Trzaska KA, Smirnov SV,etal. Down-Regulation of MHC II in Mesenchymal Stem Cells at High IFN-{gamma} Can Be Partly Explained by Cytoplasmic Retention of CIITA.[J]. The Journal of Immunology.2008;180(3):1826-1833.
3 Romieu-Mourez R, Francois M, Boivin MN,etal. Cytokine Modulation of TLR Expression and Activation in Mesenchymal Stromal Cells Leads to a Proinflammatory Phenotype.[J]. The Journal of Immunology.2009;182(12):7963-7973
4 Tse WT, Pendleton JD, Beyer WM,etal. Suppression of allogeneic T-cell proliferationby human marrow stromal cells:implicationsin transplantation. Transplantation.2003;75:389-397.
5 Koc ON, Day J, Nieder M, et al. Allogeneic mesenchymalstem cell infusion for treatment ofmetachromatic leukodystrophy (MLD) and Hurlersyndrome (MPS-IH). Bone Marrow Transplant.2002;30:215-222.
6 Stagg J, Pommey S, Eliopoulos N.Interferon-y-stimulated marrow stromal cells:a new type of nonhematopoietic antigen-presenting cell [J].Blood; 2006,107(6):2570-2577.
7 Francois M, Romieu-Mourez R, Stock-Martineau S. etal. Mesenchymal stromal cells cross-present soluble exogenous antigens as part of their antigen-presenting cell properties. Blood.2009;114 (13):2632-2638.
8.Eliopoulos N, Stagg J, Lejeune L, Pommey S, Galipeau J. Allogeneic marrow stromal cells areimmune rejected by MHC class I and II mismatchedrecipient mice. Blood.2005; 106:4057-4065.
9 Grinnemo KH, Mansson A. Deligren G, et al. Xenoreactivityand engraftment of human mesenchymalstem cells transplanted into infarcted ratmyocardium. [J]. Thorac Cardiovasc Surg. 2004; 127:1293-1300
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11 Phinney DG Biochemical heterogeneity of mesenchymal stem cell populations:clues to their therapeutic efficacy. Cell Cycle.2007; 6(23):2884-9.
12 Hu KX, Zhao SF, Sun QY.Effect of bone marrow mesenchymal stem cells on immunoregulation in H-2 haploidentical bone marrow transplantation mice. Zhonghua Xue Ye Xue Za Zhi 2007; 28(8):505-9.
13 Glennie S, Soeiro I, Dyson PJ, etal. Bone marrow mesenchymal stem cells induce division arrest anergy of activated T cells. [J].Blood,2005; 105(7):2821-2827
14. Selmani Z, Naji A, Zidi I, etal. Human leukocyte antigen-G5 secretion by human mesenchymal stem cells is required to suppress T lymphocyte and natural killer function and to induce CD4+CD25highFOXP3+ regulatory T cells.[J]. Stem Cells,2008,26(1):212-222.
15. Zheng ZH, Li XY, Ding J,etal. Allogeneic mesenchymal stem cell and mesenchymal stem cell-differentiated chondrocyte suppress the responses of type II collagen-reactive T cells in rheumatoid arthritis.[J]. Rheumatology,2008;47(1):22-30
16 Phinney DG, Hill K, Michelson C,etal. Biological activities encoded by the murine mesenchymal stem cell transcriptome provide a basis for their developmental potential and broad therapeutic efficacy. Stem Cells,2006; 24(1):186-98.
17. Karlsson H, Samarasinghe S, Ball LM,etal. Mesenchymal stem cells exert differential effects on alloantigen and virus-specific T-cell responses.[J]. Blood,2008,112(3):532-541.
18. Corcione A, Benvenuto F, Ferretti E,etal. Human mesenchymal stem cells modulate B-cell functions.[J]. Blood,2006,107(1):367-372.
19. Comoli P, Ginevri F, Maccario R,etal. Human mesenchymal stem cells inhibit antibody production inducedin vitro by allostimulation.[J]. Nephrol Dial Transplant,2008,23:1196-1202
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21. Meisel R, Zibert A, Laryea M, et al. Human bonemarrow stromal cells inhibit allogeneic T-cell responsesby indoleamine 2,3-dioxygenasemediatedtryptophan degradation. Blood.2004; 103:4619-4621.
22. Barry FP, Murphy JM, English K, Mahon BP. Immunogenicity of adult mesenchymal stem cells:lessons from the fetal allograft. Stem Cells.2005; 14:252-265.
23 Spaggiari GM, Abdelrazik H, Becchetti F.etal.MSCs inhibit monocyte-derived DC maturation and function by selectively interfering with the generation of immature DCs:central role of MSC-derived prostaglandin E2 [J].Blood,2009,113(26):6576-6583.
24 Tasso R, Augello A, Carida M,etal. Development of sarcomas in mice implanted with mesenchymal stem cells seeded onto bioscaffolds.[J]. Carcinogenesis 2009; 30(1):150-157
25 Bacigalupo A, Valle M, Podesta M, et al. T-cellsuppression mediated by mesenchymal stemcells is deficient in patients with severe aplasticanemia.[J].Exp Hematol. 2005;33:819-827.
26. Li JP, Zheng CL, Han ZC. Abnormal immunity and stem/progenitor cells in acquired aplastic anemia.[J]. Crit Rev Oncol Hematol,2009; 12(1):1105-1107
27. Del Papa N, Quirici N, Soligo D, et al. Bone marrow endothelial progenitors are defective in systemicsclerosis.[J]. Arthritis Rheum.2006;54:2605-2615.
28. Nauta AJ, Fibbe WE. Immunomodulatory properties of mesenchymal stromal cells.[J]. Blood, 2007;110(10):3499-3506
2 Bacigalupo A, Oneto R, Bruno B,etal. Current Results of Bone Marrow Transplantation in Patients with Acquired Severe Aplastic Anemia. Acta Haematol 2000; 103:19-25
3 Corcione A, Benvenuto F, Ferretti E,etal. Human mesenchymal stem cells modulate B-cell functions. Blood,2006,107 (1):367-372.
4 Nauta AJ, Kruisselbrink AB, Lurvink E,etal. Mesenchymal Stem Cells Inhibit Generation and Function of Both CD34+-Derived and Monocyte-Derived Dendritic CellsJ. Immunol,2006,177(4): 2080-2087.
5 Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood,2005; 105(4):1815-1822
6 Le Blanc K, Rasmusson I, Sundberg B,etal. Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells. The Lancet,2004; 363(9419),1439-1441
7 Zappia E, Casazza S, Pedemonte E,etal. Mesenchymal stem cells ameliorate experimental autoimmune encephalomyelitis inducing T-cell anergy. Blood,2005; 106 (5):1755-1761
8 Gerdoni E, Gallo B, Casazza S,etal. Mesenchymal Stem Cells Effectively Modulate Pathogenic Immune Response inExperimental AutoimmuneEncephalomyelitis. Ann Neurol 2007;61(3):219-227
9 Le Blanc K, Tammika C, Rosendahlc K,etal. HLA expression and immunologic propertiesof differentiated and undifferentiated mesenchymal stem cells. Exp Hematol.2003; 31 (10): 890-896
10 叶任高.陆再英.内科学。人民卫生出版社,2006：572-576。
11 张之南。血液病学。北京：人民卫生出版社.2003：618-633.
12 Young NS,. Calado RT, Scheinberg P. Current concepts in the pathophysiology and treatment of aplastic anemia.Blood.2006,108(8),2509-2519.
13 司徒镇强,吴军正。细胞培养。世界图书出版公司,2006：58-64。
14 Abdallah BM, Boissy P, Tan Q, etal. dlkl/FA1 Regulates the Function of Human Bone Marrow Mesenchymal Stem Cells by Modulating Gene Expression of Pro-inflammatory Cytokines and Immune Response-related Factors. [J]. THE JOURNAL OF BIOLOGICAL CHEMISTRY 2007;282 (10).7339-7351.
15 Tang KC, Trzaska KA, Smirnov SV,etal. Down-Regulation of MHC Ⅱ in Mesenchymal Stem Cells at High IFN-{gamma} Can Be Partly Explained by Cytoplasmic Retention of CIITA.[J]. The Journal of Immunology.2008;180(3);1826-1833.
16 Romieu-Mourez R, Francois M, Boivin MN.etal. Cytokine Modulation of TLR Expression and Activation in Mesenchymal Stromal Cells Leads to a Proinflammatory Phenotype.[J]. The Journal of Immunology,2009,182(12),7963-7973
17 Stagg J, Pommey S, Eliopoulos N.Interferon-γ-stimulated marrow stromal cells:a new type of nonhematopoietic antigen-presenting cell [J].Blood,2006,107(6):2570-2577.
18 Francois M, Romieu-Mourez R, Stock-Martineau S, etal. Mesenchymal stromal cells cross-present soluble exogenous antigens as part of their antigen-presenting cell properties. Blood, 2009,114 (13):2632-2638.
19 Eliopoulos N, Stagg J, Lejeune L,etal. Allogeneic marrow stromal cells areimmune rejected by MHC class I and II mismatchedrecipient mice. Blood.2005;106:4057-4065.
20 Glennie S, Soeiro I, Dyson P, etal. Bone marrow mesenchymal stem cells induce division arrest anergy of activated T cells. [J].Blood,2005 105 (7).2821-2827
21 Hu KX, Zhao SF, Sun QY,etal. Effect of bone marrow mesenchymal stem cells on immunoregulation in H-2 haploidentical bone marrow transplantation mice. Zhonghua Xue Ye Xue Za Zhi,2007; 28(8):505-9.
22 Beyth S, Borovsky Z, Mevorach D,etal. Human mesenchymal stem cells alter antigen-presenting cell maturationand induce T-cell unresponsiveness. Blood,2005; 105(5): 2214-2219.
23 Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood,2005; 105(4):1815-1822
24 Guo Z, Zheng C, Chen Z, etal. Fetal BM-derived mesenchymal stem cells promote the expansion of human Th17 cells, but inhibit the production of Thl cells. Eur J Immunol,2009; 39(10):2840-9
25 Gu Y, Hu X, Liu C, etal. Interleukin (IL)-17 promotes macrophages to produce IL-8, IL-6 and tumour necrosis factor-alpha in aplastic anaemia. Br J Haematol,2008; 142(1):109-14.
26 Zoumbos NC, Gascon P, Dieu JY,et al. Circulating activated suppressor Tlymphocytes in aplastic anemia. N Engl J Med,1985;312:257-265
27 Nistico A, Young NS. γ——interferon gene expression in the bone marrow of patients with acquired aplastic anemia. Ann Inter Med,1994; 120:463-469
28 Morris TC, Vincent PC, Young GA,et al. CFU C inhibitors in aplastic anemia. Blut,1984;48:61-65
29 Kawano Y, Takaue Y, Hirao A. Synergistic effect of recombinant interferon- gamma and interleukin- 3 on the growth of immature human hematopoietic progenitors. Blood,1991;77:2118-2121
30 Tichelli A, Gratwohl A, Wursch A,et al. Late haematological complications in severe aplastic anemia. Br J Haematol,1988; 69:413-418
31 于文强,孙秉中。免疫功能紊乱与再生障碍性贫血。细胞与分子免疫学杂志1999年第1期第11卷：1365-1370
32 Rasmusson I, Le Blanc K, Sundberg B,etal. Mesenchymal stem cells stimulate antibody secretion in human B cells. Scand J Immunol,2007; 65(4):336-43.
33 Horwitz EM. Culture conditions shape mesenchymal stromal cell phenotype and function. Cytotherapy,2009; 11(2):101-2
34 Zhu GR, Zhou XY, Lu H,etal. Human bone marrow mesenchymal stem cells express multiple hematopoietic growth factors. Zhongguo Shi Yan Xue Ye Xue Za Zhi,2003; 11(2):115-9.
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