極低頻電磁場對於不同人類角質細胞之生物效應研究

在現代生活中，由於家用電器地使用越來越普及，人類已不知不覺暴露於家用電器產生的極低頻電磁場(extremely low-frequency electromagnetic fields；ELF-EMFs) 中。因此，近三十年來，極低頻電磁場是否會危害人類的健康也受到大眾媒體與科學家廣泛關注與討論。對於極低頻電磁場所造成的生物效應研究常有不同的結論，然而這些不一致的結果可能由於不同的實驗條件，例如：實驗對象、照射條件(連續或間歇性)、照射時間及沒有嚴謹的電磁波磁場照射系統(例如：溫度控制及照射環境控制)所導致。在本研究中，我們透過基因表現、蛋白質表現及細胞層級等不同實驗方法，探討極低頻電磁場對於人類角質細胞之生物效應，證實人類皮膚角質細胞Immortalized nontumorigenic human keratinocytes (HaCaT; p53-mutated type)在1.5 mT，60 Hz 極低頻電磁場照射144 小時，會觸發ATM-Chk2-p21 pathway抑制細胞生長，引起細胞G1 phase arrest而降低細胞增生能力。透過西方墨點法，得知極低頻電磁場引發ATM/Chk2 signaling cascades，增加p21蛋白質表現量。相對地，利用siRNA將CHK2 基因 knockdown後，HaCaT 細胞之G1 phase arrest現象即消失。因此，本研究認為極低頻電磁場影響ATM-Chk2-p21 pathway造成HaCaT 細胞發生G1 phase arrest。進一步，利用相同的照射系統與實驗設計，評估極低頻電磁場是否會在不同的人類皮膚角質細胞primary normal human epidermal keratinocytes (NHEK; primary type)造成相似於HaCaT 細胞之效應。實驗結果證實，極低頻電磁場不會影響NHEK 細胞之細胞生長、細胞增生、細胞週期分布及ATM signaling pathway。本研究發現兩種不同的人類皮膚角質細胞對於極低頻電磁場有不同的反應。進一步以極低頻電磁場同時照射HaCaT細胞與 NHEK細胞168 小時，觀察細胞之生長曲線，其結果呈現相同結論。因此，本研究經嚴謹測試之磁場照射控制系統，以基因表現、蛋白質表現及細胞層級等方法證實不同的細胞株對於極低頻電磁場有不同的反應(cell type specific response)，這可能是目前極低頻電磁場呈現出不一致生物效應現象的原因之一。

關鍵字

極低頻電磁場；人類角質細胞；基因微陣列晶片；基因表現；細胞生長；細胞週期

並列摘要

In daily life, humans are exposed to the extremely low-frequency electromagnetic fields (ELF-EMFs) generated by electric appliances, and public concern is increasing regarding the biological effects of such exposure. Numerous studies have yielded inconsistent results regarding the biological effects of ELF-EMF exposure. This study showed that ELF-EMFs activate the ATM-Chk2-p21 pathway in HaCaT cells to decrease cell proliferation. To present well-founded results, we comprehensively evaluated the biological effects of ELF-EMFs at the transcriptional, protein, and cellular levels. Human HaCaT cells from an immortalized epidermal keratinocyte cell line were exposed to a 1.5 mT, 60 Hz ELF-EMF for 144 h. The ELF-EMF caused G1 arrest and decreased colony formation. Protein expression experiments revealed that ELF-EMFs induced the activation of the ATM/Chk2 signaling cascades. In addition, the p21 protein, a regulator of cell cycle progression at G1 and G2/M, exhibited a higher level of expression in exposed HaCaT cells compared with the expression of sham-exposed cells. The ELF-EMF-induced G1 arrest was diminished when the CHK2 gene expression (which encodes checkpoint kinase 2; Chk2) was suppressed by specific small interfering RNA (siRNA). These findings indicated that ELF-EMFs activated the ATM-Chk2-p21 pathway in HaCaT cells resulting in cell cycle arrest at the G1 phase. According to the results of HaCaT cells, it is natural to suspect whether ELF-EMFs cause similar effects in a distinct epidermal keratinocyte, primary normal human epidermal keratinocytes (NHEK), by using the same ELF-EMF exposure system and experimental design. In NHEK cells, ELF-EMFs exerted no effects on cell growth, cell proliferation, cell cycle distribution, and the activation of ATM signaling pathway. This study elucidated that the two types of epidermal keratinocytes differently responded to ELF-EMFs. To further validate this finding, the NHEK and HaCaT cells were simultaneously exposed to ELF-EMFs in the same incubator for 168 h and observed the cell growths. The results of simultaneous exposure in the two cell types showed that the NHEK and HaCaT cells exhibited distinct responses to ELF-EMFs. Thus, the biological effects of ELF-EMFs in epidermal keratinocytes are cell type specific. The findings may partially explain the inconsistent results of previous studies when comparing results across various experimental models. Based on the precise control of the ELF-EMF exposure and rigorous sham-exposure experiments in this study, the experiments at the transcriptional, protein and cellular levels all consistently supported the conclusion.

並列關鍵字

Extremely Low-Frequency Electromagnetic Fields ； ELF-EMFs ； 60 Hz ； HaCaT ； NHEK ； cDNA microarray ； gene expression ； ATM ； Chk2 ； p21 ； cell cycle ； cell growth

參考文獻

1. Humans IWGotEoCRt (2002) Non-ionizing radiation, Part 1: static and extremely low-frequency (ELF) electric and magnetic fields. IARC Monogr Eval Carcinog Risks Hum 80: 1-395.

2. International Commission on Non-Ionizing Radiation P (2009) Guidelines on limits of exposure to static magnetic fields. Health Phys 96: 504-514.

3. (2002) Non-ionizing radiation, Part 1: static and extremely low-frequency (ELF) electric and magnetic fields. IARC Monogr Eval Carcinog Risks Hum 80: 1-395.

4. International Commission on Non-Ionizing Radiation P (2010) Guidelines for limiting exposure to time-varying electric and magnetic fields (1 Hz to 100 kHz). Health Phys 99: 818-836.

5. Wertheimer N, Leeper E (1979) Electrical wiring configurations and childhood cancer. Am J Epidemiol 109: 273-284.

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極低頻電磁場對於不同人類角質細胞之生物效應研究

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