血红素加氧酶-1对小鼠树突状细胞免疫表型和功能的影响

Effects of Heme Oxygenase-1 on Dendritic Cells Immunophenotype and Function in Mouse

目的通过体外分离培养骨髓来源的树突状细胞(DCs),经血红素和锡-原卟啉(SnPP)干预DCs中血红素加氧酶-1(HO-1)表达,探讨HO-1对DCs免疫表型和功能的影响。方法采用重组(rm)小鼠粒细胞-巨噬细胞集落刺激因子(rmGM-CSF)和rmIL-4体外诱导小鼠骨髓来源的DCs,至第4天,随机分为血红素组(H组)、对照组(C组)和SnPP组(S组),给予5×104μmol/L的血红素和SnPP共同培养,诱导HO-1表达,第6天用50μg/L的TNF-α刺激成熟。通过流式细胞分选仪测定DCs表面分子CD11c/CD40,CD11c/CD80,CD11c/CD86和CD11c/I-Ad的表达,分析DCs免疫表型变化;经同种异体混合淋巴细胞培养,采用CCK-8测定各组A值变化,计算刺激指数,分析DC刺激T淋巴细胞增殖能力的变化。结果体外成功诱导出骨髓源性DCs;用血红素诱导HO-1高表达后,与C组和S组比较,H组的DCs表面分子表达呈下降趋势,但其差异无统计学意义(P>0.05);同种异体混合淋巴细胞培养后,刺激细胞与效应细胞按110混合时,刺激指数C组(0.379±0.006)、H组(0.104±0.124)、S组(1.021±0.082)间差异有统计学意义(P<0.05);按120混合时,C组(1.590±0.137)、H组(1.153±0.072)、S组(1.665±0.139)间差异有统计学意义(P<0.05);按140混合时,C组(1.239±0.068)、H组(0.893±0.002)、S组(1.631±0.156)间差异有统计学意义(P<0.05)。结论HO-1对DCs表型成熟无明显影响,但可抑制DCs刺激T淋巴细胞的增殖能力。

Objective To explore the effects of heine oxygenase - 1 （ HO - 1 ） on dendritic cells （DCs） immunophenotype and function by treating with hemin and Sn - protoporphyrin （SnPP） in vitro. Methods Mice bone marrow DCs （BMDCs） were generated by culture with recombination of granulocyte - macrophage colony - stimulating （ rmGM - CSF） and rmIL - 4, and on day - 4 they were divided into 3 groups randomly： group C, group H and group S. 5 × 10^4 μmol/L heroin or SnPP was administrated to the culture to induce HO - 1, and 50 μg/L TNF - α was used to stimulate DCs maturation on day -6. The expressions of CD11c/CD40,CD11c/CD80, CD11c/CD86 and CD11c/I - A^d on DCs were detected by fluorescence activated cell storting（FACS） to analyze the immunophenotype of DCs. The function of DCs to stimulate T cells proliferation was analyzed by allogeneic mixed lymphocyte culture using CCK - 8 to detect the difference of A value difference among groups and stimulation index （SI） was calculated. Results BMDCs were obtained successfully. Up - expression of HO - 1 was induced by heroin, and the surface molecules on DCs were down - regulated but there was no statistically significance （ P 〉 0.05 ）. After allogeneic mixed lymphocyte culture,when the ratio of stimulatory cells and effector cells was 1 ： 10, the SI of group C（0. 379 ±0. 006） ,group H（0. 104 ±0. 124） and group S（ 1. 021 ±0. 082 ） had significant difference （ P 〈 0.05 ） ; when 1 ： 20, group C （ 1. 590 ± 0. 137 ）, group H （ 1. 153 ± 0.072） and group S（ 1. 665 ± 0. 139 ）had significant difference （P 〈 0.05 ） ;when 1 ： 40, group C （1. 239 ± 0. 068 ）, group H（0. 893 ± 0. 002） and group S（ 1. 631 ±0. 156 ）had significant difference （P 〈 0.05 ）. Conclusions This study suggests that HO - 1 may reduce phenotypic maturation of DCs but there was no statistical significance. However, HO - 1 inhibits the ability of DCs to stimulate T lymphocyte proliferation.