| 摘要: | 香菸為一種含有超過4000多種化學物質的複雜混合物。吸菸會促使癌症的發生，同時損害吸菸者及附近被動吸二手菸者的健康。在最近的研究中證明pyrazine, 2-ethylpyridine, 3-ethylpyridine存在於香菸之中，並且對細胞造成影響。肺癌死亡率為全球所有癌症死亡人數的15％。此外，肺癌發展也伴隨著腫瘤的缺氧，這一點在疾病的進展中已被證明其重要性。另一方面，當藥物在治療疾病的效用降低時我們稱之為抗藥性。而在癌症治療上，患者對治療藥物所產生的抗藥性已成為現今所面臨的一大課題。通常參與這種現象的蛋白質為P-glycoprotein，其可利用主動運輸的方式將各種物質運輸至細胞膜內外。本研究的主要目的是探討已知菸草煙霧成分對人類正常及惡性腫瘤肺細胞之影響。在所設計一連串的實驗之中，我們將已知菸草煙霧成分與肺部細胞共同培養，並置於低氧(5%O2)及正常氧(20%O2)濃度的環境中，觀察細胞生長情形。此外，利用WST與AnnexinV-FITC檢測細胞之存活率與死亡率，接著利用Transwell assay來探討菸草煙霧成分對細胞轉移能力之影響以及 DCFH試劑檢測菸草煙霧成分對細胞內部氧化壓力之影響。另一方面，由於ATP-dependent drug efflux pump P-glycoprotein的過度表現而引起惡性腫瘤細胞的多重抗藥性，在臨床上會影響化學治療的療效。故在實驗的最後我們利用P-glycoprotein的受質Rhodamine123能夠被運輸至細胞內及細胞外的特性，探討菸草煙霧成分對惡性腫瘤細胞抗藥性之影響。結果顯示，菸草煙霧成分使得正常肺細胞增生受到抑制；而在惡性腫瘤肺細胞中則促進細胞增生，且於低氧環境下增生較為迅速。根據我們研究的結果，確定菸草煙霧成分對細胞之損壞作用最低劑量(LOAEL)為10-10M。當培養正常和惡性腫瘤肺細胞於LOAEL之菸草煙霧成分會減少正常和惡性肺細胞存活的細胞百分比，但MRC5細胞在統計上並無顯著之差異。而在細胞凋亡方面，菸草煙霧成分對正常和惡性腫瘤肺細胞皆無明顯影響。細胞內氧化壓力檢測檢測方面，發現菸草煙霧成分誘導細胞內的過氧化氫產生，而使得正常肺細胞之氧化壓力提升。在轉移方面，無論在正常氧或缺氧的環境中，惡性腫瘤肺細胞之轉移並無受到菸草煙霧成分之影響。在研究的最後，這些多環芳族菸草煙霧成分會使得A549細胞中P-glycoprotein數目或活性的增加，證明菸草煙霧成分對細胞抗藥性的發展是顯而易見的。綜合以上研究結果，顯示菸草煙霧成分對人體呼吸系統之不利影響。
Cigarette smoke (CS) is a complex mixture of the gas and particulate phases, in which contain more than 4000 chemical compound. CS will lead to the occurrence of cancer, and damage to smokers, as well as passive smoker’s health. In a recent study of cigarette smoke, pyrazine, 2-ethylpyridine, 3-ethylpyridine were identified in cigarette smoke and shown to be the most potent chemicals in their respective groups to have the strongest effects on cells. Lung cancer is the leading cause of cancer-related mortalities in both genders and this malignancy accounts for 15% of all cancer deaths globally and this malignancy is associated with cigarette smoking. Furthermore, development of lung cancer is also accompanied with tumoral hypoxia, which has been shown to play important role in disease progression. On the other hand, drug resistance, the reduction in effectiveness of a therapeutic agent in curing a disease, is a major problem in cancer management. The purpose of this study is to investigate the effects of known cigarette smoke toxicants components on human normal and malignant tumor lung cells. Our data showed that cigarette smoke components decreased cells proliferation in normal lung cells MRC5 while induced cell proliferation in malignant tumor lung cells. When adenocarcinomic A549 cells were maintained in hypoxia, the stimulatory effect on cell proliferation was enhanced. Based on our results, the lowest observed adverse effect level (LOAEL) was determined to be at 10-10M in normaxia. When cells were administrated with cigarette smoke components at LOAEL, the percentage of viable cells was decreased in both MRC5 and A549 cells; however, the difference is only statistically significant in A549 cells. In addition, the percent of apoptotic cells detected in cultures was comparable to that of control group in both normal and malignant, suggesting that these smoke constituents impair normal cell proliferation via mechanism(s) other than cell program death. Our oxidative stress analysis data showed that cigarette smoke components caused a significant induce in the generation of intracellular H2O2, evidencing oxidative stress induction in both MRC5 and A549 cells. Furthermore, cigarette smoke components did not significant affect invasion of A549 cells in low or normal oxygen environment. Finally, these heterocyclic aromatic smoke components caused a significant decrease in the rhodamine mean intensity of A549 cells, suggesting the development of drug resistance. Taken together, the present finding reveal adverse effects of cigarette smoke components on human respiratory system. |
