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作 者:李秋文[1] 肖文华[1] 萨仁高娃[1] 董伟伟[1] 赵惠霞[1] 段昕妤[1] 朱建华[1] 康欢荣[1] 付艳[1] 郝怡鑫[1] 王如良[1] 宋林萍[1] 叶明[1]
机构地区:[1]解放军总医院第一附属医院肿瘤一科,北京100048
出 处:《中华肝脏病杂志》2012年第3期231-235,共5页Chinese Journal of Hepatology
摘 要:目的探讨组蛋白去乙酰化酶抑制剂MS-275修饰的肝癌细胞exosome所含肿瘤抗原和免疫分子的变化。方法采用蔗糖梯度离心联合超滤技术分离和纯化肝癌HepG2细胞在MS-275处理前后分泌的exosome,并用免疫电镜和Westemblot杂交技术分别观察exosome表达热休克蛋白70、人类白细胞抗原(HLA)-Ⅰ、HLA—Ⅱ、NY-ESO-1和CD80等免疫分子的变化。MS-275处理前后样本均数的比较用配对f检验。结果经MS-275处理后,肝癌HepG2细胞分泌exosome的数量和总蛋白含量均较处理前显著增加,分别为(303.0±3.2)×10^6个/ml比(236.0±2.8)×10^6个/ml和(1.37±0.18)mg比(0.78±0.11)mg,t值分别为27.29和4.85,P值均〈0.01。免疫电镜下,100个exosome所含免疫分子HSP70、HLA—I、HLA-DR和CD80胶体金颗粒数均明显增加,分别为(79.8±5.2)个比(60.2±3.2)个、(65.6±6.3)个比(38.4±2.7)个、(26.9±2.7)个比(3.0±0.16)个和(42.7±5.2)个比(28.9±1.9)个,t值分别为5.56、6.87、15.3和4.32,P值〈0.01或〈0.05;NY—ESO-1所含胶体金颗粒数无明显变化[(40.2±4.6)个比(32.2±4.5)个,t=2.15,P〉0.051。Westernblot结果也证实了上述免疫电镜的发现。结论MS-275修饰肝癌细胞能显著改变exosome各种免疫分子的含量和种类,有利于增强exosome的免疫刺激作用,为体外大规模制备抗肝癌亚细胞瘤苗提供了新思路。Objective To investigate the effects of the histone deacetylase inhibitor, MS-275, on the immune molecule content and categories in hepatocarcinoma exosomes. Methods Exosomes were isolated from the human hepatocarcinoma cell lines, HepG2 and Hep3b, and purified by a combination technique of ultrfiltration centrifugation and sucrose gradient ultracentrifugation. The expressions of heat shock protein (HSP)70, human leukocyte antigen CI-ILA)-I, HLA-DR cluster of differentiation (CD) 80 and NY-ESO-1 on exosomes were analyzed with immunoeleetron microscopy and Western blotting before and after MS-275 treatment. Intergroup differences were statistically analyzed by the Student's paired t-test. Results MS-275 treatment of both HepG2 and Hep3b cell types significantly increased the numbers of exosomes, their total protein content, and expression of liSP70, I-ILA-I and CD80 (per 100 exosomes), as compared to non-treated cells (all, P 〈 0.01). MS-275 was also found to induce de novo expression of HLA-DR, but had no significant effect on NY-ESO-1 expression (p 〉 0.05). The findings from immunoelecton microscopy confirmed those from Western blotting. Conclusion The histone deacetylase inhibitor, MS-275, can significantly alter the immune molecule content and categories in exosomes of hepatocarcinoma cells. The differential expression profile may reflect an anti-cancer immune response and represent molecular targets for novel anti-hepatoma therapeutic or preventative strategies.
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