诱导性多能干细胞移植治疗小鼠系统性红斑狼疮
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摘要
背景:自体间充质干细胞的生物学特性易受疾病本身影响,寻找不受疾病影响的低免疫原性干细胞成为研究热点。目前,自体来源诱导性多能干细胞移植治疗系统性红斑狼疮尚未见报道。目的:评价自体成纤维细胞来源小鼠诱导性多能干细胞移植治疗系统性红斑狼疮的效果。方法:将54只8周龄MRL/lpr小鼠随机分为3组,模型对照组不给予干细胞注射等处理,试剂对照组尾静脉注射PBS溶液,干细胞治疗组尾静脉注射自体皮肤成纤维细胞来源诱导性多能干细胞。细胞治疗4周,ELISA法检测各组小鼠血清肌酐、尿素氮、抗双链DNA抗体和抗核抗体水平;石蜡切片苏木精-伊红染色观察小鼠肾脏组织形态结构;考马斯亮蓝法检测尿蛋白浓度;流式细胞术检测小鼠外周血调节性T细胞百分率、脾脏树突状细胞百分率、脾脏Th1/Th2比值。结果与结论:(1)与模型对照组和试剂对照组比较,干细胞治疗组血清肌酐、尿素氮、抗双链DNA抗体和抗核抗体水平降低,尿蛋白浓度降低,外周血调节性T细胞百分率、脾脏树突状细胞百分率以及脾脏Th1/Th2比值升高;(2)与模型对照组和试剂对照组比较,干细胞治疗组小鼠肾脏肾小球体积缩小,系膜细胞减少,肾小管上皮细胞变性改善,肾小管腔内管型减少;(3)上述结果说明,自体皮肤成纤维细胞来源诱导性多能干细胞移植可治疗系统性红斑狼疮小鼠,减轻肾组织损伤,改善肾功能并发挥免疫调节作用。
BACKGROUND: The biological characteristics of autologous mesenchymal stem cells for transplantation of systemic lupus erythematosus(SLE) are susceptible to the disease. Cell therapy with low immunogenic stem cells that are not affected by the disease has become a research hotspot. To date, treatment of systemic lupus erythematosus with autologous induced pluripotent stem cell transplantation has not been reported.OBJECTIVE: To evaluate the therapeutic effect of transplantation of induced pluripotent stem cells derived from autologous fibroblasts in mice with systemic lupus erythematosus. METHODS: Fifty-four 8-week-old MRL/lpr mice were randomly divided into three groups. Model control group was not treated, reagent control group was injected with PBS solution through tail vein, and stem cell treatment group was injected with autologous induced pluripotent stem cells through tail vein. Serum creatinine, urea nitrogen, anti-double-stranded DNA antibodies and anti-nuclear antibodies were detected by ELISA at 4 weeks of cell treatment. Kidney tissue morphology was observed by hematoxylin-eosin staining on paraffin sections. Urinary protein concentrations were detected by Coomassie brilliant blue method. Flow cytometry was used to detect the percentage of regulatory T cells in peripheral blood, percentage of splenic dendritic cells, and ratio of splenic helper T1 cells to helper T2 cells. RESULTS AND CONCLUSION: The levels of serum creatinine, urea nitrogen, anti-double-stranded DNA antibody and anti-nuclear antibody in the stem cell treatment group were lower than those in the model control and reagent control groups, while decreased urine protein concentration, increased percentages of regulatory T cells in peripheral blood, increased percentage of splenic dendritic cells, and increased splenic Th1/Th2 ratio were detected in the stem cell treatment group as compared with the other two groups. Compared with the model control and reagent control groups, there was decreased glomerular volume, reduced number of mesangial cells, degeneration of renal tubular epithelial cells and decreased tubular type in the stem cell treatment group. In summary, induced pluripotent stem cell transplantation from autologous skin fibroblasts for treat systemic lupus erythematosus can alleviate renal tissue damage, improve renal function and play an immunoregulatory role in mice.
BACKGROUND: The biological characteristics of autologous mesenchymal stem cells for transplantation of systemic lupus erythematosus(SLE) are susceptible to the disease. Cell therapy with low immunogenic stem cells that are not affected by the disease has become a research hotspot. To date, treatment of systemic lupus erythematosus with autologous induced pluripotent stem cell transplantation has not been reported.OBJECTIVE: To evaluate the therapeutic effect of transplantation of induced pluripotent stem cells derived from autologous fibroblasts in mice with systemic lupus erythematosus. METHODS: Fifty-four 8-week-old MRL/lpr mice were randomly divided into three groups. Model control group was not treated, reagent control group was injected with PBS solution through tail vein, and stem cell treatment group was injected with autologous induced pluripotent stem cells through tail vein. Serum creatinine, urea nitrogen, anti-double-stranded DNA antibodies and anti-nuclear antibodies were detected by ELISA at 4 weeks of cell treatment. Kidney tissue morphology was observed by hematoxylin-eosin staining on paraffin sections. Urinary protein concentrations were detected by Coomassie brilliant blue method. Flow cytometry was used to detect the percentage of regulatory T cells in peripheral blood, percentage of splenic dendritic cells, and ratio of splenic helper T1 cells to helper T2 cells. RESULTS AND CONCLUSION: The levels of serum creatinine, urea nitrogen, anti-double-stranded DNA antibody and anti-nuclear antibody in the stem cell treatment group were lower than those in the model control and reagent control groups, while decreased urine protein concentration, increased percentages of regulatory T cells in peripheral blood, increased percentage of splenic dendritic cells, and increased splenic Th1/Th2 ratio were detected in the stem cell treatment group as compared with the other two groups. Compared with the model control and reagent control groups, there was decreased glomerular volume, reduced number of mesangial cells, degeneration of renal tubular epithelial cells and decreased tubular type in the stem cell treatment group. In summary, induced pluripotent stem cell transplantation from autologous skin fibroblasts for treat systemic lupus erythematosus can alleviate renal tissue damage, improve renal function and play an immunoregulatory role in mice.
引文
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[22]Fliefel R,Ehrenfeld M,Otto S.Induced pluripotent stem cells(iPSCs)as a new source of bone in reconstructive surgery:Asystematic review and meta-analysis of preclinical studies.JTissue Eng Regen Med.2018;12(7):1780-1797.
[23]Celhar T,Fairhurst AM.Modelling clinical systemic lupus erythematosus:similarities,differences and success stories.Rheumatology(Oxford).2017;56(suppl_1):i88-i99.
[24]De Groof A,Hémon P,Mignen O,et al.Dysregulated Lymphoid Cell Populations in Mouse Models of Systemic Lupus Erythematosus.Clin Rev Allergy Immunol.2017;53(2):181-197.
[25]Shin MS,Lee N,Kang I.Effector T-cell subsets in systemic lupus erythematosus:update focusing on Th17 cells.Curr Opin Rheumatol.2011;23(5):444-448.
[26]Smirnova EV,Krasnova TN,Proskurnina EV,et al.Role of neutrophil dysfunction in the pathogenesis of systemic lupus erythematosus.Ter Arkh.2017;89(12):110-113.
[27]Dammacco R.Systemic lupus erythematosus and ocular involvement:an overview.Clin Exp Med.2018;18(2):135-149.
[28]álvarez K,Vasquez G.Damage-associated molecular patterns and their role as initiators of inflammatory and auto-immune signals in systemic lupus erythematosus.Int Rev Immunol.2017;36(5):259-270.
[29]Mok CC.Calcineurin inhibitors in systemic lupus erythematosus.Best Pract Res Clin Rheumatol.2017;31(3):429-438.
[30]Floris A,Piga M,Cauli A,et al.Predictors of flares in Systemic Lupus Erythematosus:Preventive therapeutic intervention based on serial anti-dsDNA antibodies assessment.Analysis of a monocentric cohort and literature review.Autoimmun Rev.2016;15(7):656-663.
[31]Bai Y,Tong Y,Liu Y,et al.Self-dsDNA in the pathogenesis of systemic lupus erythematosus.Clin Exp Immunol.2018;191(1):1-10.
[32]Momtazi-Borojeni AA,Haftcheshmeh SM,Esmaeili SA,et al.Curcumin:A natural modulator of immune cells in systemic lupus erythematosus.Autoimmun Rev.2018;17(2):125-135.
[33]王颍翠,潘兴华.间充质干细胞对系统性红斑狼疮T、B细胞的免疫调节机制[J].中国组织工程研究,2017,21(29):4734-4741.
[34]Anolik JH.B cell biology:implications for treatment of systemic lupus erythematosus.Lupus.2013;22(4):342-349.
[35]Wang D,Huang S,Yuan X,et al.The regulation of the Treg/Th17 balance by mesenchymal stem cells in human systemic lupus erythematosus.Cell Mol Immunol.2017;14(5):423-431.
[2]Di Battista M,Marcucci E,Elefante E,et al.One year in review 2018:systemic lupus erythematosus.Clin Exp Rheumatol.2018;36(5):763-777.
[3]Linge P,Fortin PR,Lood C,et al.The non-haemostatic role of platelets in systemic lupus erythematosus.Nat Rev Rheumatol.2018;14(4):195-213.
[4]Oon S,Huq M,Godfrey T,et al.Systematic review,and meta-analysis of steroid-sparing effect,of biologic agents in randomized,placebo-controlled phase 3 trials for systemic lupus erythematosus.Semin Arthritis Rheum.2018;48(2):221-239.
[5]Zhu Y,Feng X.Genetic contribution to mesenchymal stem cell dysfunction in systemic lupus erythematosus.Stem Cell Res Ther.2018;9(1):149.
[6]Wang D,Sun L.Stem cell therapies for systemic lupus erythematosus:current progress and established evidence.Expert Rev Clin Immunol.2015;11(6):763-769.
[7]Chalayeré,Ffrench M,Cathébras P.Aplastic anemia as a feature of systemic lupus erythematosus:a case report and literature review.Rheumatol Int.2015;35(6):1073-1082.
[8]Liang J,Sun L.Mesenchymal stem cells transplantation for systemic lupus erythematosus.Int J Rheum Dis.2015;18(2):164-171.
[9]王治国,刘学明,佟胜全,等.骨髓间充质干细胞移植治疗系统性红斑狼疮[J].中国组织工程研究,2016,20(10):1461-1467.
[10]阮光萍,王金祥,杨建勇,等.系统性红斑狼疮小鼠骨髓间充质干细胞的成骨、成脂分化能力下降[J].中国组织工程研究,2014,18(1):1-6.
[11]Liao J,Chang C,Wu H,et al.Cell-based therapies for systemic lupus erythematosus.Autoimmun Rev.2015;14(1):43-48.
[12]Martin PE,O'Shaughnessy EM,Wright CS,et al.The potential of human induced pluripotent stem cells for modelling diabetic wound healing in vitro.Clin Sci(Lond).2018;132(15):1629-1643.
[13]Zhou R,Jiang G,Tian X,et al.Progress in the molecular mechanisms of genetic epilepsies using patient-induced pluripotent stem cells.Epilepsia Open.2018;3(3):331-339.
[14]Rowland HA,Hooper NM,Kellett KAB.Modelling Sporadic Alzheimer's Disease Using Induced Pluripotent Stem Cells.Neurochem Res.2018;43(12):2179-2198.
[15]Pocock JM,Piers TM.Modelling microglial function with induced pluripotent stem cells:an update.Nat Rev Neurosci.2018;19(8):445-452.
[16]Hwang Y,Broxmeyer HE,Lee MR.Generating autologous hematopoietic cells from human-induced pluripotent stem cells through ectopic expression of transcription factors.Curr Opin Hematol.2017;24(4):283-288.
[17]Nikitina TV,Menzorov AG,Kashevarova AA,et al.Induced pluripotent stem cell line,IMGTi003-A,derived from skin fibroblasts of an intellectually disabled patient with ring chromosome 13.Stem Cell Res.2018;33:260-264.
[18]丁桃,李阳,唐睿珠,等.急性肺损伤患者诱导性多能干细胞系的建立[J].南方医科大学学报,2014,34(10):1414-1419.
[19]陈旭东,范文娟,袁科理,等.不同培养条件对小鼠诱导性多能干细胞分化为神经元样细胞的影响[J].细胞与分子免疫学杂志,2015,31(9):1216-1219,1223.
[20]Schwarz N,Uysal B,Rosa F,et al.Generation of an induced pluripotent stem cell(iPSC)line from a patient with developmental and epileptic encephalopathy carrying a KCNA2(p.Leu328Val)mutation.Stem Cell Res.2018;33:6-9.
[21]Sui W,Hou X,Che W,et al.Hematopoietic and mesenchymal stem cell transplantation for severe and refractory systemic lupus erythematosus.Clin Immunol.2013;148(2):186-197.
[22]Fliefel R,Ehrenfeld M,Otto S.Induced pluripotent stem cells(iPSCs)as a new source of bone in reconstructive surgery:Asystematic review and meta-analysis of preclinical studies.JTissue Eng Regen Med.2018;12(7):1780-1797.
[23]Celhar T,Fairhurst AM.Modelling clinical systemic lupus erythematosus:similarities,differences and success stories.Rheumatology(Oxford).2017;56(suppl_1):i88-i99.
[24]De Groof A,Hémon P,Mignen O,et al.Dysregulated Lymphoid Cell Populations in Mouse Models of Systemic Lupus Erythematosus.Clin Rev Allergy Immunol.2017;53(2):181-197.
[25]Shin MS,Lee N,Kang I.Effector T-cell subsets in systemic lupus erythematosus:update focusing on Th17 cells.Curr Opin Rheumatol.2011;23(5):444-448.
[26]Smirnova EV,Krasnova TN,Proskurnina EV,et al.Role of neutrophil dysfunction in the pathogenesis of systemic lupus erythematosus.Ter Arkh.2017;89(12):110-113.
[27]Dammacco R.Systemic lupus erythematosus and ocular involvement:an overview.Clin Exp Med.2018;18(2):135-149.
[28]álvarez K,Vasquez G.Damage-associated molecular patterns and their role as initiators of inflammatory and auto-immune signals in systemic lupus erythematosus.Int Rev Immunol.2017;36(5):259-270.
[29]Mok CC.Calcineurin inhibitors in systemic lupus erythematosus.Best Pract Res Clin Rheumatol.2017;31(3):429-438.
[30]Floris A,Piga M,Cauli A,et al.Predictors of flares in Systemic Lupus Erythematosus:Preventive therapeutic intervention based on serial anti-dsDNA antibodies assessment.Analysis of a monocentric cohort and literature review.Autoimmun Rev.2016;15(7):656-663.
[31]Bai Y,Tong Y,Liu Y,et al.Self-dsDNA in the pathogenesis of systemic lupus erythematosus.Clin Exp Immunol.2018;191(1):1-10.
[32]Momtazi-Borojeni AA,Haftcheshmeh SM,Esmaeili SA,et al.Curcumin:A natural modulator of immune cells in systemic lupus erythematosus.Autoimmun Rev.2018;17(2):125-135.
[33]王颍翠,潘兴华.间充质干细胞对系统性红斑狼疮T、B细胞的免疫调节机制[J].中国组织工程研究,2017,21(29):4734-4741.
[34]Anolik JH.B cell biology:implications for treatment of systemic lupus erythematosus.Lupus.2013;22(4):342-349.
[35]Wang D,Huang S,Yuan X,et al.The regulation of the Treg/Th17 balance by mesenchymal stem cells in human systemic lupus erythematosus.Cell Mol Immunol.2017;14(5):423-431.