我们发现纳秒脉电场作用于肝癌细胞后，导致共培养的单核细胞对其吞噬发生率显著提高；用于消融局部肿瘤能抑制肺转移发生。这种新的物理消融方法诱导的肿瘤细胞免疫原性改变，能否引起持续稳定的免疫识别和抗原递呈，诱导肿瘤细胞特异性免疫反应发生，对其临床转化至关重要。本项目研究纳秒脉冲电场引起肝癌细胞膜磷脂酰丝氨酸外翻与引起的其它肿瘤细胞免疫原性相关的分子如胞内尿酸、ATP、Calreticulin、HMGB1及MHC I类等空间位置变化的相关性；在诱导肝癌细胞被单核细胞吞噬的同时，对巨噬细胞M1/M2分布状态变化的影响，以及未成熟DC细胞对经电场诱导的肿瘤细胞的反应；动物实验中，治疗导致的肿瘤及微环境中巨噬细胞和CD8+ T 细胞表型变化，及其对血管新生和趋化等相关因子水平的影响。阐明基于这种新疗法诱导肿瘤细胞特异性T细胞毒性的可行性，为优化该技术的临床转化研究提供理论依据。

We recently found nanosecond pulsed electric field (nsPEF) can trigger engulfment of hepatocellular carcinoma cells by co-culturing macrophages in vitro; and the incidence of pulmonary metastases reduced significantly while the primary tumor in subcutaneous xenoplanted model had been ablated by nsPEF with fractionated low dose remedy. It is crucial to the clinical translation of this modality that the possibility of synergistic tumor cell specific cytotoxic T cell response stimulated by this modality alone or combined with other current therapies. This proposal is designed to illustrate the relation of phosphatidylserine externalization induced by nsPEF with dispelling of other sealed molecules possessing immunogenicity in living tumor cells, and responses of macrophages and antigen presenting cells to the immuno-recognizing signals revealed by nsPEF treatment in vivo and in vitro. Researches focused on three subjects: 1. the relations of membrane phosphatidylserine translocation with other immune recognition or ectopic immuno-molecules (uric acid, ATP, Calreticulin, High mobility Group Box1, CD154 and MHC I molecules) in tumor cells treated by nsPEF. 2. M1/2 state shifts of monocytes in co-culture with treated tumor cells; responses of naive dendritic cells to stimulated cancer cells. 3. The phenotype changes of local tumor associated macrophage (TAM), antigen presenting dendritic cells and CD8+ T cells infiltration to treated tumor tissue and their environment after tumor ablated with nsPEF; as well as the serum concentration changes concerning angiogenesis and chemotaxis. To provide theoretical basis for the clinical translation of pulsed power related technology by clarify the efficiency and internal mechanism of tumor specific T cell cytotoxicity induced by nsPEF treatments with cytology, immunology and tumor biology methods.