癌细胞表面电荷特征及粒细胞识别机制研究

Granulocytes are the most abundant and most functionally vigorous part of the human immune system. The known function of the granulocytes is to rapidly eradicate the invading bacteria and fungi. The most important research result of my lab in recent years since 1999 is the fact that the granulocytes have the naturally existing activities of killing a variety of cancer cells. In last several decades, there have been numerous speculations and theories on how granulocytes recognize pathogenic cells. However, even until now, there still has not been a theory that can completely and logically explain the mechanism of granulocyte recognition of target cells. In an expected way, the pilot study of our group may have provided an interesting clue. We discovered that all cancer cells, bacteria and fungi had had the unique negative charges on their surfaces whereas the normal cells don’t, as they are either charge-neutral or slightly positively charged. Therefore we hypothesize that the mechanism of granulocyte recognition is not based on any kind of specific surface molecules, but rather on the negative electrical charges of the cell surfaces. The current study project is to utilize the novel nanoprobes we recently developed and the additional ones we will construct to neutralize the surface charges of cells in order to evaluate whether such neutralizations could affect the ability of granulocytes to bind and to kill target cancer cells.

粒细胞是人体免疫系统中功能最强数量最大的组成部分。粒细胞的已知功能是快速消灭感染机体的细菌及真菌。本团队近十几年的独创研究更发现健康人体内粒细胞还有广谱杀伤癌细胞的固有活性。长年来科学界对粒细胞识别病原细胞的机制已有众多猜测，但至今仍旧没有一种合理完整的解释来说明粒细胞的靶向机制即如何识别区分正常细胞与病原细胞。本课题组前导实验结果提供了一条意外线索。我们发现作为粒细胞的靶向细胞的各种癌细胞，细菌和真菌的表面都均带有大量表面负电荷，而且正常细胞则持电中性或带微量正电荷。我们提出的待验证假设是粒细胞识别以上列举的病原细胞的机制很可能不是任何细胞表面特殊分子，而是细胞表面电荷。我们计划在本科研课题中使用现有的带电荷纳米材料和将利用类似方法合成的带电纳米材料来干扰和中和细胞表面电荷，从而观察这种细胞表面电荷干扰是否能够影响粒细胞又癌细胞的表面结合，是否能够影响粒细胞杀伤癌细胞的活性。

“抗癌免疫监控系统”是机体消除潜在癌变细胞、保持健康的重要防御体系。长期以来，“抗癌免疫监控系统”相关研究工作的重点一直是单个核细胞（包括T细胞、B细胞、NK细胞等），而非粒细胞。1999年，本课题组崔征教授团队偶然发现了抗癌小鼠（Spontaneous regression/complete resistance mice, SR/CR）的存在。相关研究工作显示，SR/CR小鼠的粒细胞具有极强的杀癌活性，其在SR/CR小鼠对抗恶性肿瘤细胞侵袭时，起着极为重要的防御作用。此后几年间，本课题组也陆续在许多健康志愿者血液中，检测出粒细胞的杀癌活性。以上种种现象提示我们，粒细胞在机体抗癌免疫监控系统中，起着不容忽视的重要作用。基于本课题组的前期工作，我们详细探讨了不同种类细胞表面电荷性质，并进一步详细探讨了人类粒细胞对癌细胞的识别机制。此部分研究中，我们利用本课题组前期自主设计合成的多功能纳米颗粒（Nanoparticles, NPs），探讨不同细胞表面电荷性质。NPs+及NPs-捕获细胞效率对比试验、迪夫快速染色试验、及激光共聚焦显微镜观察结果显示，癌细胞表面呈强负电性、粒细胞表面呈弱正电性、单个核细胞表面呈电中性。NPs屏蔽细胞表面电荷实验表明，细胞表面正负电荷相互吸引，是粒细胞识别癌细胞机制之一，但与单个核细胞识别癌细胞机制无关。3-溴丙酮酸抑制糖酵解通路实验进一步显示，癌细胞糖酵解通路是粒细胞识别癌细胞的基础。癌细胞糖酵解通路异常旺盛，导致癌细胞表面携带大量负电荷，粒细胞可利用其细胞表面的相反电荷识别癌细胞，进而完成杀伤过程。本项研究，阐明了粒细胞识别癌细胞的特有机制，为后续癌症的临床检验及治疗，提供了全新的研发思路。