【作者】 姚成灿；

【导师】 黄耀熊； 李校坤；

【作者基本信息】 暨南大学 ， 生物医学工程， 2004， 博士

【摘要】 随着移动电话等的普及，人们日常生活中接受到越来越多的射频电磁辐射，因此，射频电磁场的生物学效应已经成为引起人们广泛关注的重要研究领域。电磁场的生物效应包括热效应和非热效应，过去20多年里，对电磁辐射的生物效应，尤其是非热效应进行了大量研究，也提出了各种理论和模型。目前较为一致的看法是，电磁辐射作用的形式和强度大小与对人体及实验动物所引起的各种生物效应有相关性，但是，到目前为止，仍然没有得出一个权威性的结论，尚未阐明其确切效应和内在机制。综合分析以往这些研究，可发现存在多方面的问题，其中一个非常值得注意的问题是，除个别实验报道进行的单项检测外，基本上大部分的实验都没有对在电磁场作用过程当中对细胞产生的各种作用及有关效应进行实时监测。 为此，本文研究移动电话所用900 MHz射频电磁波，在分别对应于手机在通话状态下(15μW／cm~2～31μW／cm~2)，和手机基站、微波通讯机房、微波炉生产线、机场雷达等周围泄露的电磁波(1 mW／cm~2～5 mW／cm~2)功率密度水平辐照下，对分离人红细胞的有关即时效应。之所以选择红细胞，是因为红细胞是生理学、生物物理学和生物电化学研究的理想模型，是细胞生物学基础研究的重要实验对象。本文的主要目的在于，通过多维显微成像技术、快速显微多道分光光度技术、显微动态图像分析技术、相分析电泳激光散射技术等多种手段，研究射频电磁波以及射频电磁波与其他理化因素对红细胞的协同作用。在完整细胞、细胞膜、细胞内分子三个层次水平上，对单个活态人红细胞的形态、大小、膜弹性模量、胞内Hb吸收光谱、膜表面电荷等多个参数的测量和分析。揭示射频电磁波和有关理化因素对单个活态人红细胞结构与功能各种参数即时影响的规律和机制。研究所得结论如下： (1)对应于移动电话手机辐射水平的极低功率密度(15μW／cm~3和31μW／cm~2)900 MHz射频电磁波≤30分钟照射对离体单个活态人红细胞的形态、大小、膜弹性模量、胞内Hb的结构与浓度、膜表面电荷(照射时间为60 min)等都不会产生统计意义的显著影响。而低功率密度(1 mW／cm~2和5 mW／cm~2)900 MHz射频电磁波对单个活态人红细胞的形态、大小、膜弹性模量、胞内Hb吸收光谱、膜表面电荷等有显著影响。 (2)低功率密度900 MHz射频电磁波辐射，通过对红细胞膜的作用可使其表面电暨南大学理学博士学位论文 荷密度与Zeta电位降低，从而影响膜的通透性，引起Hb和离子外泄，同时 也通过对氢键的激励等途径，改变蛋白质的结构和活性直接引起胞内Hb损 失，而使胞内Hb浓度减少。这些变化导致红细胞形态从双凹形向球形或棘形 过渡，接触面积与直径变小，弹性模量增大。 (3)低功率密度900 MHz射频电磁波对红细胞的影响并非单纯取决于辐射的剂量 (即功率密度与时间之乘积)，还与电磁波的照射时间有关。也就是说，作用 除有一功率密度闭值外，还有一作用时间闭值，需超过一定的照射时间才会 引起或出现有关效应。超过这两个闭值后，呈现一定的剂量累积效应。 (4)低功率密度射频电磁波照射对红细胞的作用具有时效效应:电磁波仅照射10 min便可使膜弹性模量显著增加，而停止照射30 min后膜弹性模量会下降至 照前水平;红细胞内Hb浓度与分子构型在照射30 min后才会发生变化，但 停止照射后会维持在停止照射时的水平，不能恢复到照射前水平。说明在各 种结构与功能参数中，红细胞膜的变形能力受射频电磁波辐射作用的反应最 快、最敏感，并有一定的自我恢复能力。而细胞的形态大小和胞内Hb的变性 与流失则不可恢复。这提示有关射频电磁波辐射对细胞作用的研究应在即时 (实时)进行的必要性和重要性。 (5) pH值、温度、胰蛋白酶、射频电磁波等理化因素作用都能引起红细胞形状、 大小、膜力学特性、胞内Hb和表面电荷的变化。在本实验范围内，对红细胞 形态大小的影响作用依强弱次序为pH值>温度>射频电磁波>胰蛋白酶;对膜 弯曲弹性模量影响作用依强弱次序为pH值>温度>胰蛋白酶>射频电磁波;对 膜剪切弹性模量影响作用依大小次序为温度>pH值>胰蛋白酶>射频电磁波; 对胞内Hb浓度影响作用依大小次序为pH值>温度>射频电磁波>胰蛋白酶; 对表面电荷影响作用依大小次序为pH值>温度>胰蛋白酶>射频电磁波。 (6)外加电磁波照射引起的效应与红细胞所处状态密切相关，如果红细胞所处状 态偏离正常生理状态，则更容易在电磁辐射作用下，产生更严重的影响。pH 值、温度、表面电荷等理化因素与极低功率密度900 MHz射频电磁场之间没 有协同作用。pH值、温度、胰蛋白酶等理化因素对900 MHz低功率密度射 频电磁场照射导致的红细胞结构与功能变化具有正协同作用，对于红细胞膜 力学性质的协同作用效果各因素中pH值>温度>表面电荷，对于胞内Hb的协 同作用效果为pH值>表面电荷>温度。 (7)红细胞的表面电荷、大小形态、膜弹性模量及胞内Hb的结构与功能等参数 之间的变化相互紧密相关，互为影响。各种理化因素依其作用方式不同可能更多还原

【Abstract】 With the population of mobile phone, people daily have received more and more radiation of radio frequency electromagnetic wave (RF-EMW). Therefore, the biological effects of RF-EMW have become an important research topics. The biological effects of electromagnetic wave include thermal effects and non-thermal effects. In the past 20 years, many researches have been performed and various theories and models have been proposed on RF-EMW biological effects, especially the non-thermal effects. The unanimous opinion at present is that the form and intensity of electromagnetic radiation is correlated to various biological effects caused by it. Up to now, however, there has been neither authoritative conclusion nor exact effects and intrinsic mechanism for this issue. Many limitations occurred in previous work. One distinct problem is that, except for some reports on single examinations, most experiments did not perform real-time measurements/monitoring on the instant effects caused by electromagnetic field to the cells.Therefore, in this thesis, the instant effects of 900 MHz RF-EMW on isolated human red blood cell (RBC) were studied. Two levels of power density were used in the investigation, the first group is an ultra-low level group with power densities of 15 μW/cm2and 31 μW/cm2, corresponding to the radiation level from two kinds of mobile phones respectively. The second group is a low level group with power densities of 1 mW/cm2and 5 mW/cm2 respectively, corresponding to the electromagnetic waves leaked from the mobile phone base station, microwave communication station, microwave production lines, and the aerodrome control rada. The RBC is chosen as the target cells because it is an ideal model for physiological, biophysical and bioelectrochemical and cell biological study. The main target of this work is to study the effects of RF-EMW itself on RBC, and the cooperative effects of the RF-EMW and other physical and chemical factors. Some advanced techniques like multi-dimensional micro-imaging technique, fast multi-channel micro- spectrophotomery, dynamic micro-imaging analysis technique, and phase analysis electrophoretic laser-scattering technique were employed for the study. Various parameters on both the structures and functions of single living intact human RBC, such as the morphology, size, elastic modulus of membrane, absorption spectrum of intracellular Hb, and surface negative charges of RBCs were determined and analyzed, simultaneously on the intact cell level, membrane level, and molecular level of intracellular protein. According to the experimental results, we have the following remarks:1. The exposure of 900 MHz RF-EMW at ultra-low power density (15 W/cm2 and 31 uW/cm2) has no significant influence on the morphology, size, elastic modulus of membrane, the structure and concentration of intracellular Hb, surface negative charges (exposed for 60min) of isolated single living intact human RBC; while the exposure of 900 MHz RF-EMW at low power density (1 mW/cm2 and 5 mW/cm2) has significant influence on the above issues.2. The exposure of 900 MHz RF-EMW at low power density will interact with the membrane, decrease its surface negative charge density and Zeta potential, influence the permeability of the membrane, and result in the leakage of intracellular Hb and ions. It will also alter the structure and activity of protein through the stimulation of hydrogen bond to cause the leakage of intracellular Hb directly and the decrease of the concentration of intracellular Hb. The changes result in alteration of RBCs from discocytes to sphericytes or echinocytes, the diminution of contact area and diameter, and the increase of elastic modulus of membrane.3. Under the same dose of exposure of the 900 MHz RF-EMW at low power density, two kinds with different power density have different effects on RBCs, which indicates that the influence of RF-EMWs on RBCs is not only determined by the dose of exposure (i.e., product of power density and time), but determined by both the power density and the更多还原