脂质体包裹RNA和裸RNA在肿瘤RNA疫苗中作用的比较

Comparison of RNA/liposome complexes with naked RNA

目的:比较有脂质体包裹的肿瘤RNA与无脂质体包裹的裸RNA转染未成熟树突状细胞(DC)后DC的成熟情况及其诱导的CTL对靶细胞的杀伤作用,以确定裸RNA能否有效转染DC。方法:从肺癌细胞系CALU-6中提取总RNA。采集肺癌患者PBMC富集血分离单个核细胞培养DC,第5天时将DC分成3组,A组用阳离子脂质体DMRIE-C与RNA共同转染DC,B组只用RNA转染DC,C组为DC对照组,继续培养5d后测定转染后3组DC的表面标志。转染后的A、B两组DC与自体淋巴细胞混合,诱导特异性CTL,LDH法测定诱导出的CTL的特异性杀伤活性。结果:与未进行转染的DC相比,A、B两组肿瘤细胞RNA转染DC后,CD1a、CD83及CD86、HLA-ABC都出现大幅度上调表达,说明两种方法转染RNA都可诱导DC的成熟。上述DC与自体淋巴细胞混合培养均可诱导出CTL,LDH释放实验显示,在效靶比40:1、20:1及10:1时两组DC诱导的CTL对同种靶细胞的杀伤活性比较无统计学差异,提示脂质体包裹RNA和裸RNA转染DC均可诱导特异性抗肿瘤免疫反应。结论:不需脂质体包裹的裸RNA与用脂质体包裹的RNA转染未成熟DC都能实现肿瘤细胞RNA信息进入DC,并一过性表达相应抗原,诱导DC成熟。二者都具有在体外诱导出自体CTL的特异性杀伤靶细胞的能力。

Objective： To compare the function of naked RNA and RNA/liposome complexes transfecting DC in inducing DCs maturity and priming CTL response. Methods： Total RNA was isolated from the lung cancer cell line CALU-6 and DCs were generated from PBMCs of lung cancer patients. Immature DCs were divided into 3 groups： DCs transfected with RNA/liposome complexes （Group A）, DCs transfected with naked RNA （Group B） and DCs control （Group C）. The phenotypes of 3 groups of DCs were characterized after 5 days culture. In group A or B, DCs were mixed with lymphocytes to induce CTLs. And the cytotoxicity was measured using LDH method. Results： Comparing with untranstected DCs, both A and B group of DCs expressed high levels of CD1a, CD83, CD86, ther naked RNA or RNA/liposome complexes can and HLA-ABC, which suggested that transfection of eiinduce maturation of DCs. CTLs were induced from the lymphocytes cocultured with DCs of each group. The result of LDH assay suggested there was no difference of cytotoxicity of CTLs between naked RNA and RNA/liposome complexes transfected DCs. Conclusion： Naked RNA is transfected into DCs to express tumor antigens and induce DCs maturation. Comparing with RNA/liposome complexes, naked RNA pulsed DCs can directly prime CTL response in vitro.