CAJ | 학술논문

目的：通过检测羧甲基茯苓多糖（ carboxymethytl pachymaram ，CMP）处理后人外周血源性树突状细胞（dendritic cells，DC）中细胞因子信号转导抑制分子-1（SOCS-1）基因甲基化水平及表达情况，探讨 CMP 对 DC 体外成熟的影响。方法通过重组人粒细胞-巨噬细胞集落刺激因子（rhGM-CSF）和IL-4体外诱导人外周血源性单核细胞源DC，并促进成熟，分别加入不同浓度的CMP （终浓度为10、50、100 mg/L），应用甲基化特异性PCR（ methylation specific PCR ，MSP）和real-time PCR法分别分析SOCS-1基因甲基化水平及表达情况；应用流式细胞术、混合淋巴细胞反应（ MLR）和ELISA分别检测DC的表面标志、刺激淋巴细胞增殖能力和IL-12分泌的变化。结果经CMP刺激后DC中SOCS-1基因甲基化水平明显增高，SOCS-1基因表达水平显著降低，表面标志（ CD80、CD83、CD86、HLA-DR）、刺激淋巴细胞增殖能力及IL-12的分泌水平均上调，且呈剂量依赖趋势。50 mg/L和100 mg/L CMP刺激组SOCS-1基因甲基化水平、IL-12分泌量及刺激淋巴细胞增殖指数均显著高于对照组，SOCS-1基因表达水平显著低于对照组。100 mg/L CMP刺激组CD80、CD83和HLA-DR表达水平显著高于对照组。结论 CMP能促进人外周血源性DC中SOCS-1基因甲基化，减少SOCS-1基因表达，进而促进其体外成熟。
목적：통과검측최갑기복령다당（ carboxymethytl pachymaram ，CMP）처리후인외주혈원성수돌상세포（dendritic cells，DC）중세포인자신호전도억제분자-1（SOCS-1）기인갑기화수평급표체정황，탐토 CMP 대 DC 체외성숙적영향。방법통과중조인립세포-거서세포집락자격인자（rhGM-CSF）화IL-4체외유도인외주혈원성단핵세포원DC，병촉진성숙，분별가입불동농도적CMP （종농도위10、50、100 mg/L），응용갑기화특이성PCR（ methylation specific PCR ，MSP）화real-time PCR법분별분석SOCS-1기인갑기화수평급표체정황；응용류식세포술、혼합림파세포반응（ MLR）화ELISA분별검측DC적표면표지、자격림파세포증식능력화IL-12분비적변화。결과경CMP자격후DC중SOCS-1기인갑기화수평명현증고，SOCS-1기인표체수평현저강저，표면표지（ CD80、CD83、CD86、HLA-DR）、자격림파세포증식능력급IL-12적분비수평균상조，차정제량의뢰추세。50 mg/L화100 mg/L CMP자격조SOCS-1기인갑기화수평、IL-12분비량급자격림파세포증식지수균현저고우대조조，SOCS-1기인표체수평현저저우대조조。100 mg/L CMP자격조CD80、CD83화HLA-DR표체수평현저고우대조조。결론 CMP능촉진인외주혈원성DC중SOCS-1기인갑기화，감소SOCS-1기인표체，진이촉진기체외성숙。
Objective To investigate the effects of carboxymethytl pachymaram ( CMP ) on the methylation of SOCS-1 (suppressor of cytokine signaling-1) gene and the in vitro maturation of human mono-cyte-derived dendritic cells (DCs).Methods Human DCs were induced from the peripheral blood mono-cytes in vitro with the treatment of recombined human GM-CSF and interleukin-4 ( IL-4 ) and cultured with different concentrations of CMP (10, 50, and 100 mg/L).The methylation and expression of SOCS-1 gene were analyzed by methylation-specific polymerase chain reaction (MSP) and real-time PCR, respectively. The phenotypic markers of DCs were detected by flow cytometry .Mixed lymphocyte reaction ( MLR) and ELISA were performed to measure the lymphocyte proliferation induced by DCs and IL-12 secretion by DCs . Results CMP promoted the methylation of SOCS-1 gene, but inhibited the expression of SOCS-1 gene in dendritic cells at the concentrations of 50 mg/L and 100 mg/L.The expression of phenotypic markers (CD80, CD83, CD86 and HLA-DR), IL-12 secretion and lymphocyte proliferation induced by DCs were significantly enhanced in a dose dependent manner with the treatment of CMP .Compared with control group , the levels of methylated SOCS-1 gene and IL-12 and the lymphocyte proliferation index were increased upon the stimulation with 50 mg/L and 100 mg/L of CMP (P<0.01), but the expression of SOCS-1 gene was de-creased.The expression of CD80, CD83 and HLA-DR on DCs in the presence of 100 mg/L of CMP were higher than those of control group (P<0.05).Conclusion CMP could induce the methylation of SOCS-1 gene and the maturation of DCs derived from peripheral blood monocytes .