Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/5087
標題: 人類乳癌細胞株中多氯聯苯對雌性激素(17β-estradiol)所誘發細胞毒性、核酸損害與基因表現失衡之影響
Effects of polychlorinated biphenyls (PCBs) on the induction of cell toxicity, DNA damage and altered gene expression by 17β-estradiol in human breast carcinoma cell lines
作者: 周雅炘
Chou, Ya-Hsin
關鍵字: 多氯聯苯
PCB
雌性激素
estrogen
出版社: 環境工程學系
摘要: 本研究主旨在於探討多氯聯苯(PCB126與PCB153)對雌性激素(17β-estrodiol,E2)於人類乳癌細胞株ERα(+)/MCF-7與ERα(-)/MDA-MB-231,誘發基因表現失衡、氧化壓力、核酸氧化損害、及細胞毒性之影響。研究結果顯示,E2、PCB126、PCB153於MCF-7和MDA-MB-231細胞中所誘發之細胞毒性皆具有時間及濃度效應,且在此兩株人類乳癌細胞中,PCB153所誘發之細胞毒性明顯大於PCB126。研究結果顯示,在不引起細胞毒性濃度下,E2於MDA-MB-231細胞中會誘發細胞內ROS增加,且E2濃度愈高,ROS濃度增加愈明顯,若預處理PCB126則不會影響由E2所誘發之ROS生成,但細胞若預處理PCB153,則會抑制由E2所誘發之ROS生成,而α-naphthoflavone (a CYP1A1/1B1 inhibitor)能有效地抑制E2於MDA-MB-231細胞中所造成之ROS生成;此外,在MCF-7細胞中,E2不能誘發細胞內ROS的生成,細胞若預處理PCB126,亦不能促進E2誘發細胞內ROS量增加,但細胞若預處理PCB153後,則能促進E2誘發細胞內ROS量增加。更進一步研究指出,E2於正常生理濃度(1-10 nM)即可誘發MDA-MB-231細胞DNA strand breaks之形成,進而活化PARP-1 [poly(ADP-ribose) polymerase-1],誘發細胞內NAD+及NAD(P)H濃度下降,細胞若預處理PCB126後,對於E2所誘發之PARP-1之活化並無影響,但細胞若預處理PCB153,則會抑制由E2所誘發之PARP-1之活化。E2 (0.1 nM-10 nM)於MCF-7細胞中並無法誘發NAD(P)H下降,而MCF-7細胞預處理PCB153後,能促進E2誘發細胞內NAD(P)H濃度下降。由半定量之RT-PCR結果顯示,E2可誘發MDA-MB-231細胞中CYP1A1、CYP1B1、hOGG1和XRCC1之基因表現量,預處理PCB153可抑制由E2所誘發CYP1A1、CYP1B1、hOGG及XRCC1 mRNA之基因表現量;而單獨的PCB153與E2可誘發MCF-7細胞中CYP1A1和CYP1B1之基因表現,預處理PCB153可促進由E2所誘發CYP1A1之基因表現量,單獨之PCB153可抑制MCF-7細胞中hOGG1及XRCC1之基因表現量。由RT-PCR結果顯示,PCB153之暴露會影響E2代謝與核酸修補相關基因之表現。綜合以上結果顯示,共平面和非共平面多氯聯苯同屬物對於雌性激素於人類乳癌細胞株所誘發不同程度的ROS形成、DNA損害、和細胞毒性皆能產生干擾效應,且雌性激素受體存在與否能影響PCB同屬物調控E2誘發DNA損害作用和細胞毒性。
The objective of this research is to investigate the effects of polychlorinated biphenyls (PCBs) (i.e. PCB126 and PCB153), on the induction of imbalances in gene expression, formation of reactive oxygen species (ROS), oxidative DNA damage and cell toxicity by 17β-estradiol (E2) in human breast cancer cell lines, including estrogen receptor α (ERα)(+)/MCF-7 and ERα(-)/MDA-MB-231 cells. Results indicated that E2, PCB126, and PCB153 induced concentration- and time-dependent increases in cytotoxic response in both MCF-7 and MDA-MB-231 cells. The extent of cytotoxic response in human breast cancer cells was greater for PCB153 than for PCB126 in these two cell lines. The data also showed that at non-cytotoxic concentrations, E2 induces concentrations-dependent increases in intracellular levels of ROS in MDA-MB-231 cells. Pretreatment with PCB126 did not alter the induction of ROS by E2 in MDA-MB-231 cells. α-Naphthoflavone (a CYP1A1/1B1 inhibitor) completely blocked the E2-induced increases in the amount of intracellular ROS in MDA-MB-231 cells pretreatment of PCB126 (20 μM) and PCB mixture [PCB126 (20μM)+PCB153 (1μM)]. In contrast, induction of ROS formation by E2 in MDA-MB-231 cells was inhibited with pretreatment of PCB153. Additionally, E2 did not induce increases in intracellular ROS in MCF-7 cells. However, when cells were pretreated with PCB153, increases in intracellular ROS was detected in E2-treated MCF-7 cells. Further investigation indicated that E2 induces decreases in intracellular NAD(P)H and NAD+ by PARP-1 [poly(ADP-ribose) polymerase-1] activation through formation of DNA strand breaks in MDA-MB-231 cells at physiologically relevant concentration (1-10 nM). Pretreatment with PCB126 did not alter the E2-induced PARP-1 activation in MDA-MB-231 cells. In contrast, pretreatment of PCB153 inhibits the E2-induced PARP-1 activation in MDA-MB-231 cells. When MCF-7 cells were exposed to E2 (0.1 nM-10 nM) alone, we did not observe decreases in intracellular NAD(P)H, whereas pretreatment of cells with PCB153 significantly reduces intracellular NAD(P)H in MCF-7 cells exposed to E2. Results from semi-quantitative reverse-transcription polymerase chain reaction (RT-PCR) assay indicated that E2 induced increases in the expression of CYP1A1, CYP1B1, and hOGG1, XRCC1 genes in MDA-MB-231 cells. Pretreatment of PCB153 inhibits the E2-induced CYP1A1, CYP1B1, hOGG1 and XRCC1 gene expression in MDA-MB-231 cells. PCB153 and E2 alone induced increases in the expression of CYP1A1 and CYP1B1 in MCF-7 cells. Pretreatment of PCB153 increases the E2-induced CYP1A1 gene expression in MCF-7 cells. PCB153 alone can inhibits the hOGG1 and XRCC1 gene expression in MCF-7 cels. This result suggests that exposure to PCB153 alters gene expression in the disposition of estrogen and DNA repair. In conclusions, this evidence suggests that both planar and non-planar PCB congeners modulate different profile of estrogen-induced formation of ROS, DNA lesions, and cell toxicity in human breast cancer cells. The data also suggests that the status of estrogen receptor a may play a role in modulating the E2-mediated induction of oxidative DNA lesions and cell toxicity in human breast cancer cells.
URI: http://hdl.handle.net/11455/5087
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