胶原诱导关节炎模型中髓系来源抑制细胞通过白细胞介素-1β促进Th17细胞分化的作用
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摘要
目的通过胶原诱导关节炎模型探讨MDSCs对Th17细胞分化的作用及可能的机制,为阐明自身免疫性关节炎发病机制和寻找新的治疗靶点提供研究依据。方法 SPF级雄性DBA1/J小鼠20只随机分为胶原诱导关节炎(CIA)组和正常对照组,每组10只。采用CIA法制备自身免疫性关节炎小鼠模型。收集各组小鼠脾脏中骨髓来源抑制细胞(MDSCs)和Th17细胞的频率,分别采用流式细胞术和ELISA法测定小鼠血浆中炎性因子白细胞介素(IL)-β、IL-17及抑炎因子IL-10水平,采用实时荧光定量PCR技术测定细胞中Th17分化相关因子STAT3 mRNA、视黄酸相关孤儿受体(ROR)αmRNA、RORγt mRNA相对表达量。采用在Th17细胞分化培养条件下[添加IL-6+转化生长因子β(TGF-β)] MDSCs与小鼠幼稚CD4+T细胞共培养评估CIA小鼠中MDSCs对Th17细胞分化的影响,在培养过程中采用IL-1β中和抗体(IL-1βm Ab)和拮抗剂(IL-1ra)阻断IL-1β信号的方法以观察MDSCs对Th17细胞分化影响。结果与正常对照组小鼠比较,CIA组小鼠MDSCs比例明显升高[(20. 1±2. 6)%vs.(2. 6±1. 7)%],血浆炎症因子IL-17水平明显升高[(1. 3±0. 2)%vs.(0. 3±0. 1)%], CIA组血浆IL-β水平明显升高,抑炎因子IL-10水平明显降低,RORγtmRNA相对表达量明显增加,差异均有统计学意义(均P <0. 01)。CIA组小鼠MDSCs比例及Th17细胞比例变化呈明显正相关。CIA小鼠中MDSCs抑制CD4+T细胞增生并减少细胞分泌的γ干扰素(IFN-γ),但体外MDSCs和CD4+T细胞共培养结果显示MDSCs促进T17细胞分化,且使细胞上清液中IL-17A质量浓度升高,STAT3和RORγtmRNA相对表达量上调,培养液中IL-1β质量浓度升高。阻断IL-1β信号可以减少MDSCs对Th17细胞分化的作用。结论自身免疫性关节炎小鼠MDSCs具有促炎症反应的作用,MDSCs依靠IL-1β上调RORγt的作用而促进Th17细胞分化,这可能是自身免疫性关节炎发病过程中的一个关键因素。
Objective To investigate the role and mechanism of myeloid-derived suppressor cells( MDSCs) on Th17 cell differentiation by using collagen induced arthritis( CIA) model. To elucidate the pathogenesis of autoimmune arthritis and find new therapeutic targets. Methods SPF male DBA1/J mice were randomly divided into CIA group( n = 10) and control group( n = 10). An autoimmune arthritis mouse model was prepared by CIA method. The proportion of MDSCs and Th17 cells were examined in each group by flow cytometry.The levels of plasma IL-1β,IL-10,and IL-17 A were detected by ELISA. The relative expression levels of factor-related to Th17 differentiation,such as STAT3 mRNA,retinoic acid associated orphine receptor( ROR) αmRNA and RORγt mRNA,were detected by real-time fluorescence quantitative PCR. The effects of MDSCs on the differentiation of Th17 cells in CIA mice were assessed by co-culture of MDSCs with mouse naive CD4+T cells under the condition of Th17 cell differentiation( IL-6+TGF-β). It was also observed by adding IL-1βm Ab or IL-1 ra when co-culturing MDSCs with CD4+T cells. Results Compared with the control group,the proportion of MDSCs and plasma IL-17 level were significantly elevated [MDSCs:( 20. 1 ± 2. 6) % vs.( 2. 6 ±1. 17) %; IL-17:( 1. 3±0. 2) % vs.( 0. 3±0. 1) %],the levels of IL-1β and the relative expression of RORγt mRNA were significantly increased and IL-10 was lowed( P< 0. 01). The proportion of MDSCs was positively correlated to the proportion of Th17 cells in CIA mice. MDSCs inhibited the proliferation of CD4+T cells and the secretion of gamma interferon( IFN-γ) in CIA. In contrast,MDSCs promoted Th17 cells differentiation in vitro,followed by significant increase of IL-17 A,IL-1β,and upregulation of STAT3 and RORγt. The effect of MDSCs in differentiation of Th17 cells was down-regulated when IL-1β pathway was blocked. Conclusions MDSCs play a role in promoting inflammation by driving Th17 cell differentiation in autoimmune arthritis,which depends on IL-1β pathway. MDSCs may be a potential target in treatment of autoimmune arthritis.
Objective To investigate the role and mechanism of myeloid-derived suppressor cells( MDSCs) on Th17 cell differentiation by using collagen induced arthritis( CIA) model. To elucidate the pathogenesis of autoimmune arthritis and find new therapeutic targets. Methods SPF male DBA1/J mice were randomly divided into CIA group( n = 10) and control group( n = 10). An autoimmune arthritis mouse model was prepared by CIA method. The proportion of MDSCs and Th17 cells were examined in each group by flow cytometry.The levels of plasma IL-1β,IL-10,and IL-17 A were detected by ELISA. The relative expression levels of factor-related to Th17 differentiation,such as STAT3 mRNA,retinoic acid associated orphine receptor( ROR) αmRNA and RORγt mRNA,were detected by real-time fluorescence quantitative PCR. The effects of MDSCs on the differentiation of Th17 cells in CIA mice were assessed by co-culture of MDSCs with mouse naive CD4+T cells under the condition of Th17 cell differentiation( IL-6+TGF-β). It was also observed by adding IL-1βm Ab or IL-1 ra when co-culturing MDSCs with CD4+T cells. Results Compared with the control group,the proportion of MDSCs and plasma IL-17 level were significantly elevated [MDSCs:( 20. 1 ± 2. 6) % vs.( 2. 6 ±1. 17) %; IL-17:( 1. 3±0. 2) % vs.( 0. 3±0. 1) %],the levels of IL-1β and the relative expression of RORγt mRNA were significantly increased and IL-10 was lowed( P< 0. 01). The proportion of MDSCs was positively correlated to the proportion of Th17 cells in CIA mice. MDSCs inhibited the proliferation of CD4+T cells and the secretion of gamma interferon( IFN-γ) in CIA. In contrast,MDSCs promoted Th17 cells differentiation in vitro,followed by significant increase of IL-17 A,IL-1β,and upregulation of STAT3 and RORγt. The effect of MDSCs in differentiation of Th17 cells was down-regulated when IL-1β pathway was blocked. Conclusions MDSCs play a role in promoting inflammation by driving Th17 cell differentiation in autoimmune arthritis,which depends on IL-1β pathway. MDSCs may be a potential target in treatment of autoimmune arthritis.
引文
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[18] Wang W,Jiao Z,Duan T,et al. Functional characterization of myeloid-derived suppressor cell subpopulations during the development of experimental arthritis[J]. Eur J Immunol,2015,45:464-473.
[19] Zhu B,Bando Y,Xiao S,et al. CD11b+Ly-6C(hi)suppressive monocytes in experimental autoimmune encephalomyelitis[J]. J Immunol,2007,179:5228-5237.
[20] Zhang H,Wang S,Huang Y,et al. Myeloid-derived suppressor cells are proinflammatory and regulate collagen-induced arthritis through manipulating Th17 cell differentiation[J]. Clin Immunol,2015,157:175-186.
[21] King IL, Dickendesher TL, Segal BM. Circulating Ly-6C+myeloid precursors migrate to the CNS and play a pathogenic role during autoimmune demyelinating disease[J]. Blood,2009,113:3190-3197.
[22] Azawa T,Shibata M,Gonda K,et al. Increased IL-17 production correlates with immunosuppression involving myeloid-derived suppressor cells and nutritional impairment in patients with various gastrointestinal cancers[J]. Mol Clin Oncol,2013,1:675-679.
[23] Chatterjee S,Das S,Chakraborty P,et al. Myeloid derived suppressor cells(MDSCs)can induce the generation of Th17 response from na6ve CD4+T cells[J]. Immunobiology,2013,218:718-724.
[24] Chung Y,Chang SH,Martinez GJ,et al. Critical regulation of early Th17 cell differentiation by interleukin-1 signaling[J].Immunity,2009,30:576-587.
[25] Evans HG,Gullick NJ,Kelly S,et al. In vivo activated monocytes from the site of inflammation in humans specifically promote Th17responses[J]. Proc Natl Acad Sci U S A,2009,106:6232-6237.
[26] Zhang L,Zhang Z,Zhang H,et al. Myeloid-derived suppressor cells protect mouse models from autoimmune arthritis via controlling inflammatory response[J]. Inflammation,2014,37:670-677.
[27] Jiao Z,Hua S,Wang W,et al. Increased circulating myeloid-derived suppressor cellscorrelated negatively with Th17 cells in patients with reumatoid arthritis[J]. Scand J Rheumatol,2013,42:85-90.
[2] Lubberts E,van den Bersselaar L,Oppers-Walgreen B,et al. IL-17 promotes bone erosion in murine collagen-induced arthritis through loss of the receptor activator of NF-kappa Bligand/osteoprotegerin balance[J].J Immunol,2003,170:2655-2662.
[3] Diaz-Montero CM,Salem ML,Nishimura MI,et al. Increased circulating myeloid-derived suppressor cells correlate with clinical cancer stage,metastatic tumor burden,and doxorubicin-cyclophosphamide chemotherapy[J]. Cancer Immunol Immunother,2009,58:49-59.
[4] Solito S,Falisi E,Diaz-Montero CM,et al. A human promyelocytic-like population is responsible for the immune suppression mediated by myeloid-derived suppressor cells[J]. Blood,2011,118:2254-2265.
[5] Ochoa AC,Zea AH,Hernandez C,et al. Arginase,prosta-glandins,and myeloid-derived suppressor cells in renal cell carcinoma[J]. Clin Cancer Res,2007,13(2 Pt 2):721s-726s.
[6] Gabrilovich DI,Nagaraj S. Myeloid-derived suppressor cells as regulators of the immune system[J]. Nat Rev Immunol,2009,9:162-74.
[7] Strand-Rosenberg S,Sinha P. Myloid-derived suppressor cells:linking inflammation and cancer[J]. J Immunol,2009,182:4499-4506.
[8] Ioannou M,Alissafi T,Lazaridis I,et al. Crucial role of granulocytic myeloid-derived suppressor cells in the regulation of central nervous system autoimmunedisease[J]. J Immunol,2012,188:136-1146.
[9] Yi H,Guo C,Yu X,et al. Mouse CD11b+Gr-1+myeloid cells can promote Th17 cell differentiation and experimental autoimmune encephalomyelitis[J].J Immunol,2012,189:4295-4304.
[10] Haile LA,von Wasielewski R,Gamrekelashvili J,et al. Myeloidderived suppressor cells in inflammatory bowel disease:a new immunoregulatory pathway[J]. Gastroenterology,2008,135:871-881.
[11] Yin B,Ma G,Yen CY,et al. Myeloid-derived suppressor cellsprevent type 1 diabetes in murine models[J]. J Immunol,2010,185:5828-5834.
[12] Fujii W,Ashihara E,Hirai H,et al. Myeloid-derived suppressor cells play crucial roles in the regulation of mouse collagen-induced arthritis[J].J Immunol,2013,191:1073-1081.
[13] Inglis JJ,Simelyte E, Mc Cann FE, et al. Protocol for the induction of arthritis in C57BL/6 mice[J]. Nat Protoc,2008,3:612-618.
[14] Gonzalez-Rey E,Chorny A,Varela N,et al. Therapeutic effect of urocortin on collagen-induced arthritis by down-regulation of inflammatory and Th1 responses and induction of regulatory T cells[J]. Arthritis Rheum,2007,56:531-543.
[15] Gabrilovich DI,Nagaraj S. Myeloid-derived suppressor cells as regulators of the immune system[J].Nat Rev Immunol,2009,9:162-174.
[16] Pastua A,Marcinkiewicz J. Myeloid-derived suppressor cells:a double-edged sword?[J].Int J Exp Pathol,2011,92:73-78.
[17] le Blanc K,Jitschin R,Mougiakakos D. Myeloid-derived suppressor cells in allogeneic hematopoietic stem cell transplantation:A double-edged sword?[J/OL]. Oncoimmunology, 2013, 2:e25009[2018-03-20].https://www.tandfonline.com/doi/full/10.4161/onci.25009.
[18] Wang W,Jiao Z,Duan T,et al. Functional characterization of myeloid-derived suppressor cell subpopulations during the development of experimental arthritis[J]. Eur J Immunol,2015,45:464-473.
[19] Zhu B,Bando Y,Xiao S,et al. CD11b+Ly-6C(hi)suppressive monocytes in experimental autoimmune encephalomyelitis[J]. J Immunol,2007,179:5228-5237.
[20] Zhang H,Wang S,Huang Y,et al. Myeloid-derived suppressor cells are proinflammatory and regulate collagen-induced arthritis through manipulating Th17 cell differentiation[J]. Clin Immunol,2015,157:175-186.
[21] King IL, Dickendesher TL, Segal BM. Circulating Ly-6C+myeloid precursors migrate to the CNS and play a pathogenic role during autoimmune demyelinating disease[J]. Blood,2009,113:3190-3197.
[22] Azawa T,Shibata M,Gonda K,et al. Increased IL-17 production correlates with immunosuppression involving myeloid-derived suppressor cells and nutritional impairment in patients with various gastrointestinal cancers[J]. Mol Clin Oncol,2013,1:675-679.
[23] Chatterjee S,Das S,Chakraborty P,et al. Myeloid derived suppressor cells(MDSCs)can induce the generation of Th17 response from na6ve CD4+T cells[J]. Immunobiology,2013,218:718-724.
[24] Chung Y,Chang SH,Martinez GJ,et al. Critical regulation of early Th17 cell differentiation by interleukin-1 signaling[J].Immunity,2009,30:576-587.
[25] Evans HG,Gullick NJ,Kelly S,et al. In vivo activated monocytes from the site of inflammation in humans specifically promote Th17responses[J]. Proc Natl Acad Sci U S A,2009,106:6232-6237.
[26] Zhang L,Zhang Z,Zhang H,et al. Myeloid-derived suppressor cells protect mouse models from autoimmune arthritis via controlling inflammatory response[J]. Inflammation,2014,37:670-677.
[27] Jiao Z,Hua S,Wang W,et al. Increased circulating myeloid-derived suppressor cellscorrelated negatively with Th17 cells in patients with reumatoid arthritis[J]. Scand J Rheumatol,2013,42:85-90.