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目的研究慢性乙肝合并肝细胞癌(HCC)患者外周血髓样树突状细胞(mDC)负载乙型肝炎病毒(HBV)抗原后表型和功能的变化。方法用成分血分离机采集21例肝细胞癌患者和4名健康人的外周血单核细胞(PBMC),用无血清AIMV培养基加入粒细胞巨噬细胞集落刺激因子(GMCSF)和白细胞介素(IL)4等因子诱导mDC分化、发育和成熟,同时加入HBcAg或HBsAg致敏mDC,通过流式细胞仪分析mDC表面分子的表达,用酶联免疫吸附试验(ELISA)检测mDC培养上清中IL12、IL10的含量,通过自体混合淋巴细胞反应(AMLR)比较抗原致敏后mDC刺激淋巴细胞增殖的能力。结果在肝细胞癌患者中,(1)经HBcAg致敏的mDC表面分子CD80、CD86、CD40、HLADR的表达率(55%±26%、80%±13%、70%±13%、73%±24%)明显高于无抗原致敏组mDC(29%±25%、35%±18%、44%±26%、45%±23%,P<005);经HBsAg致敏的mDC仅HLADR的表达率(63%±15%)高于无抗原致敏组(45%±23%,P<005),其他表面分子差异无统计学意义。(2)在AMLR中经HBV抗原致敏的mDC刺激淋巴细胞增殖的能力明显高于无抗原致敏组(P<005),而且HBcAg的致敏效果优于HBsAg的致敏效果(P<005)。(3)经HBcAg致敏的mDC分泌IL10和IL12的量(236pg/ml±95pg/ml,733pg/ml±212pg/ml)明显高于经HBsAg致敏的mDC(35pg/ml±9pg/ml,135pg/ml±63p
Objective To study the phenotypic and functional changes of peripheral blood mononuclear cells (mDC) loaded with hepatitis B virus (HBV) antigen in patients with chronic hepatitis B and hepatocellular carcinoma (HCC). Methods Peripheral blood mononuclear cells (PBMCs) from 21 patients with hepatocellular carcinoma and 4 healthy individuals were collected by using a blood separator. The serum free AIMV was added into the granulocyte macrophage colony stimulating factor (GMCSF) and interleukin (IL) 4 and other factors to induce the differentiation, development and maturation of mDC. Meanwhile, HBcAg or HBsAg-sensitized mDCs were added to analyze the expression of mDC surface molecules by flow cytometry. The expression of mDC was detected by enzyme-linked immunosorbent assay (ELISA) IL12, IL10 content by autologous mixed lymphocyte reaction (AMLR) compared antigen sensitized mDC stimulation of lymphocyte proliferation. Results In hepatocellular carcinoma patients, (1) the expression rates of CD80, CD86, CD40 and HLADR on HBcAg-sensitized mDC were 55% ± 26%, 80% ± 13%, 70% ± 13%, 73% ± 24%) were significantly higher than that of mDC without antigen-sensitized group (29% ± 25%, 35% ± 18%, 44% ± 26%, 45% ± 23%, P <005) The expression rate of HLADR (63% ± 15%) was higher than that without antigen sensitization (45% ± 23%, P <005). There was no significant difference in other surface molecules. (2) The ability of mDC stimulated by HBV antigen in AMLR to stimulate the proliferation of lymphocytes was significantly higher than that without antigen sensitization (P <005), and the sensitization effect of HBcAg was better than that of HBsAg (P <0.05 ). (3) The amount of IL10 and IL12 secreted by HBcAg-sensitized mDC was significantly higher (236pg / ml ± 95pg / ml, 733pg / ml ± 212pg / ml) than HBsAg-sensitized mDCs 135 pg / ml ± 63 p