细胞因子体外诱导慢性髓细胞性白血病树突状细胞分化的研究

In Vitro Cytokines-Induced Differentiation in Mononuclear Cell Derived Dendritic Cells from Chronic Myeloid Leukemia

本研究探讨干扰素(IFN)治疗慢性粒细胞性白血病(CML)的可能新机制,为临床治疗CML提供新的思路。用Ficoll密度离心法分离骨髓单个核细胞(BMMNC),用不同细胞因子组合体外诱导培养树突状细胞(DC),在倒置显微镜及光镜下观察DC形态,应用流式细胞术检测其表面标志(CD1a,CD83,CD86,HLA-ABC,HLA-DR,CD54),MTT法检测其混合淋巴细胞反应(MLR)能力。结果表明:经不同细胞因子组合所诱导出的DC,其特征性表面分子表达率均高于培养前的DC(P<0.01);IFN-α+GM-CSF组DC表达HLA-ABC、HLA-DR明显高于IL-4+GM-CSF培养组(P<0.05);而IFN-α+GM-CSF+IL-4组DC的CD86表达率及MLR水平均高于其它细胞因子组合组(P<0.05)。干扰素耐药组DC表面分子表达率及MLR水平较新诊断组和干扰素治疗有效组均明显减低(P<0.05),但A、B、C各组的CD86表达率及MLR水平在IFN-α+GM-CSF+IL-4因子组合条件下无明显差异。结论:CML骨髓单个核细胞在含有IFN-α的细胞因子组合条件下可分化为具有形态和免疫表型特征的DC,且表达较高的I类和II类分子、共刺激分子、黏附分子及MLR,表明干扰素治疗CML的机理可能与DC有关,这一结果提示通过增强内源性DC功能可能提高CML临床疗效。

The purpose of this study was to investigate the mechanism of effects of interferon-alpha （IFN-α） on chronic myeloid leukemia （CML）. Bone marrow mononuclear cells（BMMNC） were obtained from heparinized blood of CML patients by Ficoll-Paque density gradient centrifugation. The expressions of CD1a, CD83, CD86, HLA-ABC, HLA-DR and CD54 on DC induced by IFN-α ＋ GM-CSF, IFN-α ＋ GM-CSF ＋ IL-4 and IL-4 ＋ GM-CSF for 7 days in vitro were assayed by flow cytometry. The morphologic features were observed by transmission and optical microscopy. The mixed lymphocyte reactions （MLR） with DC were evaluated by MTr assay. The results showed that the DC cultured in different cytokine combinations expressed significantly higher levels of CDI a, HLA-ABC, HLA-DR, CD86, CD54, and CD83 than those in the precultured. The DC growing with IFN-α ＋ GM-CSF expressed significantly higher levels of HLA-ABC, HLA-DR than those in GM-CSF ＋ IL-4. The CD86 expression and MLR levels in IFN-α ＋ GM-CSF ＋ IL-4 increased significantly. The expression rate of DC antigens and MLR in the IFN resistant group significantly lower than those in the newly diagnosed and the effectively treated groups after at least 6 months of IFN-α treatment （P 〈 0.05 ） . The DC from the IFN resistant group did not express significantly CD86 and MLR in IFN-α ＋ GM-CSF ＋ IL-4 groups compared to those in the newly diagnosed and IFN effective treated groups. It is concluded that the BMMNC from CML cultured in combination with IFN-α and other cytokines can be induced into DC with typical morphologic and immunophenotypic characteristics. Addition of IFN-α ＋ GM-CSF ＋ IL-4 to DC cultures can significantly up-regulate the expression of major histocompatibility complex molecules, co-stimulatory molecules and various adhesion molecules. The deficiency of DC differentiation and function may play a role in the development of clinical resistance to IFN-α.