慢性特发性血小板减少性紫癜患者外周血树突细胞的变化及意义

Alteration and biological significance of peripheral dendritic cells in patients with chronic idiopathic thrombocytopenic purpura

目的探讨慢性特发性血小板减少性紫癜（ITP）患者体内树突细胞（DC）数量和功能的变化。方法慢性ITP患者予以大剂量地塞米松（HD—DXM）冲击治疗，剂量40mg／d，口服，连续4d，观察短期疗效。用流式细胞术检测患者外周血中髓系DC（mDC）和浆细胞样DC（pDC）在治疗前后绝对数量的变化及与CD4^＋CD25^＋调节性T（Treg）细胞的相关性；流式细胞术检测外周血总DC表面协同刺激分子的表达。将慢性ITP患者外周血单核细胞来源的DC、CD4^＋T淋巴细胞与自身血小板或健康人同源异基因血小板混合培养，观察CD4^＋T淋巴细胞增殖情况，检测DC血小板相关抗原的呈递功能。结果与正常对照组比较，慢性ITP患者外周血pDC和mDC绝对数量均无明显变化（P值均〉0．05）；而外周血CD4^＋FOXP3^＋T细胞数明显降低（P〈0．01），pDC和mDC与CD4^＋FOXP3^＋T细胞数之间均存在负相关性（r=-0．396，P=0．045和r=-0．410，P=0．037）。HD—DXM治疗初始反应率达92．3％，pDC绝对数量较治疗前减少达75．5％（P〈0．01）；而mDC较治疗前虽增加了24．3％（P〈0．05），但mDC上CD11c表达的平均荧光强度（MFI）从治疗前340±30降至199±21（P〈0．01）；CD4^＋FOXP3^＋T细胞数较治疗前显著增加（P〈0．01），治疗后pDC与CD4^＋FOXP3^＋T细胞存在负相关（r=-0．524，P=0．006），而mDC与CD4^＋FOXP3^＋T细胞间无相关性（r=-0．360，P=0．071）。慢性ITP患者外周血DC表面协同刺激分子CD86表达的MFI明显高于正常对照组（P〈0．05）；CD86、CD40、CD80表达阳性率及CD40、CD80表达的MFI与正常对照组比较差异均无统计学意义（P值均〉0．05）。慢性ITP患者CD4^＋T淋巴细胞增殖反应强度高于对照组（P〈0．05），DC对血小板相关抗原呈递功能亢进。结论DC可能参与了慢性ITP的免疫发病机制，并与CD4^＋CD25^＋Treg细胞有一定的关联。

Objective To investigate the quantity and function of circulating dendritic cells （DC） in patients with chronic idiopathic thrombocytopenic purpura（ITP）. Methods High dose dexamethasone（ HD-DXM ） at a dose of 40 mg orally per day for four consecutive days Was the initial treatment for chronic ITP patients. Flow cytometry was used to analyze the number of myeloid DC （mDC）, plasma cytoid DC（pDC） and CD4^＋ FOXP3^＋ T cells in patients before and after the treatment, meanwhile the co-stimulatory molecules on circulating DCs were assayed as well. Monocyte-derived DCs and CD4^＋T cells were co-cultured with autologous or allogeneic normal fresh platelets and after 6 days of incubation ^3H-TdR was used to assay the proliferation of CD4^＋ T cells. Results The absolute numbers of circulating mDC and pDC were not significantly different between pre-treatment patients and healthy controls （P 〉 0.05 and P 〉 0.05 ）. However, percentage of CD4^＋ FOXP3^＋ T cells was decreased （ P 〈 0.01 ）, and their percentage was inversely correlated with the number of pDC and mDC （ r = - 0. 396, P = 0. 045 and r = - 0. 410, P = 0. 037）. The initial response rate to HD-DXM was 92.3%. After 4-days treatment, CD4^＋ FOXP3^＋ Treg cells increased （ P 〈 0.01 ） while pDCs decreased （P 〈 0.01 ）. Although mDCs increased after HD-DXM （P 〈 0.05 ）, their CD11c expression level was decreased （ P 〈 0.01 ）, the mean fluorescence intensity （MFI） decreased from 340 ± 30 before treatment to 199 ± 21 after treatment. The inverse correlation between pDCs and CD4^＋ FOXP3^＋ Treg cells remained （r = -0. 524, P = 0. 006） while that between mDCs and Treg cells disappeared （ r = - 0. 360,P = 0.071 ）. The MFI of CD86 on DCs was higher in ITP patients than in healthy controls （ P 〈 0.05 ）, while the proportions of CD86, CD40, CD80 and the MFI of CD40, CD80 in ITP patients were normal （P 〉 0.05 ）. DCs from chronic ITP patients co-cultured with autologous or allogeneic platelets were highly efficient in stimulating autologous CD4^＋ T cells proliferaton as compared to those derived from healthy donors （P 〈 0.05 and P 〈 0.05 ）. Conclusion DCs may play a role in the pathogenesis of chronic ITP in relation with CD4^＋ CD25^＋ Treg cells.