机构地区:[1]解放军第四军医大学西京医院心脏外科,陕西省西安市710032 [2]解放军第四军医大学免疫教研室,陕西省西安市710032 [3]解放军第四军医大学西京医院骨科,陕西省西安市710032 [4]沈阳军区总医院心脏外科,辽宁省沈阳市110016
出 处:《中国临床康复》2005年第15期120-122,F003,共4页Chinese Journal of Clinical Rehabilitation
摘 要:目的:观察在体外混合培养条件下,抗原能否诱导外周淋巴细胞反应抑制,建立体外淋巴细胞反应抑制模型。方法:实验于2003-10/2004-04在解放军第四军医大学西京医院心脏外科研究所实验室进行。取健康自愿者新鲜血液用于分离外周淋巴细胞。在体外条件下,以同种异体外周淋巴细胞分作供体和受体进行混合培养,采用四甲基偶氮唑盐比色法、电镜、姬姆萨-瑞氏染色、流式细胞仪检测对照组D组:首次混合培养淋巴细胞(PMLC),即受体淋巴细胞(R)+用丝裂霉素处理过的供体淋巴细胞(DBm)混合培养72h所得,A组(PMLC+DBm)、B组(PMLC+白细胞介素2中和单抗)、C组(PMLC+DBm+白细胞介素2中和单抗)等4组淋巴细胞的活性变化、活性变化的原因及其数量的变化。结果:①4组混合培养淋巴细胞在电镜、显微镜下变化:均可见细胞质染色浓集、核固缩裂解、凋亡小体形成。②四甲基偶氮唑盐比色法检测4组细胞活性即刺激指数:B组和C组比A、D组明显减弱(3.152±0.322,1.802±0.115,4.293±0.269,3.709±0.278,P<0.05);C组细胞活性比B组也明显减弱(P<0.05)。③流式细胞仪检测淋巴细胞凋亡率:单向混合淋巴细胞培养后的T淋巴细胞的凋亡率Bf组和Cf组高于Df对照组和Af组(11.25±0.78)%,(31.65±1.33)%,(5.95±0.24)%,(2.15±0.04)%,P<0.01;AIM:To observe whether antigens can induce the restrained peripheral lymphocytes response under mixed culture in vitro,and establish a model of restrained lymphocytes response in vitro. METHODS:The experiment was carried out in the laboratory of Institute of Cardiosurgery,Xijing Hospital,Fourth Military Medical University of Chinese PLA from October 2003 to April 2004.Blood samples obtained from healthy volunteers were prepared for isolation of peripheral lymphocytes.In vitro,homologuous peripheral lymphocytes were mixed cultured as donator and receptor respectively,methyl-thiazol-tetrazolium(MTT),electron microscopy,Giemsa-Wright's staining, and flow cytometry were used to detect the changes of the lymphocytes' activity and the reasons, and the change of the number in the four groups:control group(group D:primarily mixed cultured lymphocytic cells,which were obtained from the 72-hour mixed culture of lymphocytes of receptor and lymphocytes of donator treated with mitomycin),primarily mixed-cultured lymphocytic cells+lymphocytes of donator treated with mitomycin group(group A),primarily mixed-cultured lymphocytic cells+anti-interleukin-2 neutralizing monoclonal antibody group(group B) and primarily mixed-cultured lymphocytic cells+lymphocytes of donator treated with mitomycin+anti-interleukin-2 neutralizing monoclonal antibody group(group C). RESULTS:①Changes of mixed cultured lymphocytes under electronic microscopy and microscopy:chromatin condensation,shrinkage and splitting of nucleus and formation of poptotic body were observed in all the four groups.②Activity of lymphocytes(stimulation index) in the four groups:It was obviously weakened in group B(3.152±0.322) and group C(1.802±0.115) as compared with group A(4.293±0.269) and group D (3.709±0.278) (P< 0.05);The stimulation index in group C was also markedly decreased as compared with that in group B(P< 0.05).③Apoptosis rate of lymphocytes detected with flow cytometry:After unidirectional mixed culture of lymphocytes, the apoptosis rates of lymp
关 键 词:淋巴细胞反应 抑制模型 培养条件 抗原诱导 四甲基偶氮唑盐比色法 单向混合淋巴细胞培养 流式细胞仪检测 外周淋巴细胞凋亡 培养淋巴细胞 白细胞介素2 淋巴细胞凋亡率 第四军医大学 混合培养 活性变化 健康自愿者 T淋巴细胞
分 类 号:R512.620.3[医药卫生—内科学] R151.3[医药卫生—临床医学]
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