以BALB/c小鼠为模型, 探讨H7N9流感病毒灭活疫苗免疫小鼠后所诱导的长效体液免疫应答的动态变化。不同剂量的流感H7N9全病毒灭活疫苗单独或辅以MF59佐剂肌肉注射免疫小鼠一次。连续采集免疫后小鼠15个月的血清, 用ELISA方法检测特异性IgG抗体水平, 血凝抑制(hemagglutination inhibition, HI)试验和微量中和(microneutralization, MN)试验检测第15个月时的HI抗体和中和抗体效价。实验结果发现, 小鼠血清中的特异性IgG抗体水平随时间变化持续缓慢上升, 第5个月时达到顶峰, 随后略有下降但一直持续平稳状态; IgG抗体滴度与疫苗剂量成正相关, 且添加佐剂能提高抗体滴度。HI及MN抗体检测表明, 免疫后第15个月产生的抗体能有效中和病毒, 且抗体跟疫苗剂量成正比。以上研究表明, H7N9流感病毒灭活疫苗免疫小鼠一次诱导产生的特异性抗体能在较长期内保持比较平稳的抗体滴度, 为小鼠提供免疫保护; 增加抗原剂量和添加MF59佐剂能增加疫苗特异性抗体水平。该研究为H7N9流感疫苗产生的长期保护效应提供了一定的数据积累和参考。

Photonics immunotherapy is a novel cancer treatment strategy that combines local phototherapy and immunotherapy. Phototherapy is a noninvasive or minimally invasive therapeutic strategy for local treatment of cancer, which can destroy tumor cells and release tumor antigens, inducing an in situ antitumor immune response. Immunotherapy, including the use of antibodies, vaccines, immunoadjuvants and cytokines, when combined with phototherapy, could bring a synergistic effect to stimulate a host immune response that effectuates a long-term antitumor immunity. This review will focus on the development of photonics immunotherapy and its systemic antitumor immunological effects.

目的: 通过靶向CTLA4 siRNA探讨CTLA4在T调节细胞诱导异种抗原免疫耐受中是否发挥功能及其作用机制。方法: 体外扩增培养利用磁珠分选出的T调节细胞, AO/PI染色计算活率并通过细胞计数作出生长曲线, 流式细胞仪检测扩增后细胞表型; 采用Alex488染料标记siRNA, 通过流式细胞仪检测siRNA的转染效率; 实时定量 PCR检测靶向CTLA4 siRNA的沉默效率; 流式细胞术检测CTLA4 在蛋白水平的变化; 混合淋巴实验检测CTLA4表达下调后, T调节细胞的功能变化。结果: 经过4周体外扩增, T调节细胞能够增长约1 200倍, 并具有高活率和高纯度; siRNA的转染效率为61.8%± 4.5%; Realtime PCR和流式细胞术检测CTLA4在mRNA水平和蛋白水平均有不同程度下降, 其结果与对照组相比差异显著 （P<0.05）; 混合淋巴实验结果显示T效应细胞在受到异种抗原刺激时会发生增殖, Treg细胞能够抑制这种增殖, 但是CTLA4表达下调后明显减弱了T调节细胞的抑制能力。进一步, 在DC细胞参与的混合淋巴实验中发现Treg能够通过DC抑制T效应细胞对异种抗原的应答, 而siCTLA4-Treg不能通过DC抑制T效应细胞增殖。结论: CTLA4在Treg介导的异种免疫应答中发挥着重要作用, 这种作用方式可能是通过直接作用于T效应细胞或者间接通过DC细胞作用于T效应细胞发挥作用。靶向CTLA4 siRNA能够下调Treg 细胞CTLA4的表达, 影响Treg细胞抑制异种抗原引起T效应细胞应答的能力。

The ideal treatment modality for metastatic cancer would be a local treatment that can destroy primary tumors while inducing an effective systemic anti-tumor response. To this end, we developed laser immunotherapy, combining photothermal laser application with an immunoadjuvant for the treatment of metastatic cancer. Additionally, to enhance the selective photothermal effect, we integrated light-absorbing nanomaterials into this innovative treatment. Specifically, we developed an immunologically modified carbon nanotube combining single-walled carbon nanotubes (SWNTs) with the immunoadjuvant glycated chitosan (GC). To determine the effectiveness of laser irradiation, a series of experiments were performed using two different irradiation durations — 5 and 10 min. Rats were inoculated with DMBA-4 cancer cells, a metastatic cancer cell line. The treatment group of rats receiving laser irradiation for 10 min had a 50% longterm survival rate without residual primary or metastatic tumors. The treatment group of rats receiving laser irradiation for 5 min had no long-term survivors; all rats died with multiple metastases at several distant sites. Therefore, LasertSWNT–GC treatment with 10 min of laser irradiation proved to be effective at reducing tumor size and inducing long-term anti-tumor immunity.