本文選題：多環(huán)芳烴 + 遺傳易感性�。� 參考：《中國疾病預防控制中心》2010年博士論文

【摘要】： 焦爐逸散物已被國際癌癥研究署定義為1類致癌物,即確定的人類致癌物。焦爐逸散物(Coke oven emission, COE)中的致癌性多環(huán)芳烴(Polycyclic aromatic hydrocarbons, PAH)是導致焦爐工肺癌的主要原因,在我國焦爐工肺癌被列為八種法定職業(yè)腫瘤之一。研究表明,基因組不穩(wěn)定性是多環(huán)芳烴暴露到肺癌發(fā)生的重要生物學事件。多環(huán)芳烴暴露工人染色體損傷水平較非暴露組明顯升高,但在相同暴露條件下,其損傷水平不盡相同,提示人群易感性的存在。 本研究探討共濟失調(diào)毛細血管擴張癥突變基因(ATM)多態(tài)性與多環(huán)芳烴暴露致染色體損傷遺傳易感性及相關分子機制。結(jié)果如下： 一、對140名多環(huán)芳烴暴露工人和66名非暴露工人外周血淋巴細胞微核率與共濟失調(diào)毛細血管擴張癥突變基因多態(tài)性關系的分析顯示,在校正性別、年齡、吸煙和自然對數(shù)轉(zhuǎn)換的尿1-OHP后,PAH暴露組中ATM基因rs600931 GA和GA+AA基因型攜帶者外周血淋巴細胞微核率(11.14±6.92‰和10.57±6.82‰)明顯高于GG基因型攜帶者(7.66±5.69‰,P=0.015和0.038)；rs189037 GA和GA+AA基因型個體外周血淋巴細胞微核率(10.99±6.90‰和10.51±6.77‰)明顯高于GG基因型攜帶者(7.72±5.83‰,P=0.018和0.035)；rs624366 GC和GC+CC基因型個體外周血淋巴細胞微核率(11.34±6.74‰和10.73±6.63‰)明顯高于GG基因型攜帶者(7.61±6.07‰,P=0.001和0.003)；rs227092 GT基因型個體外周血淋巴細胞微核率(10.78±6.60‰)明顯高于GG基因型攜帶者(7.91±6.30‰,P=0.025)。單體型對GGGGTGC/AAACATT攜帶者外周血淋巴細胞微核率(12.05±7.40‰)明顯高于GGGGTGC/GGGGTGC攜帶者(7.51±6.20‰,P=0.007)。 二、利用胞質(zhì)阻滯細胞微核(Cytokinesis-block micronucleus, CBMN)實驗探討ATM干擾細胞和空載質(zhì)粒細胞對B(a)P誘導所致遺傳損傷的差異。結(jié)果顯示,隨著B(a)P作用濃度的增加,ATM干擾細胞和空載質(zhì)粒細胞CBMN率均逐漸升高,呈明顯的劑量反應關系。在4μM和8μM作用濃度時,ATM干擾細胞CBMN率明顯高于空載質(zhì)粒細胞CBMN率,差異具有統(tǒng)計學意義(P0.05)。 采用單細胞凝膠電泳技術探討ATM干擾細胞和空載質(zhì)粒細胞對博萊霉素誘導的DNA修復能力(DNA repair capacity, DRC)差異。結(jié)果顯示,在10μg/ml博萊霉素誘導下16HBE-siATM細胞的DRC(56.664±0.78%)明顯低于16HBE-siGFP細胞(70.13±3.82%),差異具有統(tǒng)計學意義(P=0.039)；與16HBE細胞的表現(xiàn)相似,HEK-siATM細胞DRC(63.86±2.17%)亦明顯低于HEK-siGFP細胞(73.904±1.67%),差異具有統(tǒng)計學意義(P=0.044)。 三、采用流式細胞技術探討ATM干擾細胞和空載質(zhì)粒細胞對B(a)P誘導的細胞周期變化。結(jié)果顯示,與16HBE-siGFP細胞相比,4μM B(a)P使16HBE-siATM細胞G1期明顯縮短,G2期明顯延長；與HEK-siGFP相比,4μM B(a)P使HEK-siATM細胞G1期和G2期均明顯縮短,S期明顯延長。 采用Western-blotting技術探討B(tài)(a)P作用下ATN、P53、NBS1和rH2AX蛋白表達情況。結(jié)果顯示,與對應的溶劑對照組比較,8μM B(a)P作用24h后,16HBE-siGFP和HEK-siGFP細胞ATM總蛋白表達無明顯變化；與對應的溶劑對照組比較,8μMB(a)P可使ATM干擾細胞及空載質(zhì)粒細胞P53、NBS1和rH2AX表達水平均升高。 四、利用雙熒光素酶報告基因?qū)嶒炋接慉TM基因5'UTR (rs189037GA)和3'UTR (rs227092 GT)變異體外驅(qū)動報告基因表達的差異。結(jié)果顯示,在三種肺組織來源的細胞系(16HBE細胞、CCC-HPF-1細胞和MRC-5細胞)中,含rs189037G等位基因的5'UTR和含rs189037A等位基因的5'UTR調(diào)控報告基因表達的差異均沒有統(tǒng)計學意義(P0.05)；含rs227092G等位基因的3'UTR和含rs227092T等位基因的3'UTR調(diào)控報告基因表達的差異亦均沒有統(tǒng)計學意義(P0.05)。 總之,本研究探討了ATM基因多態(tài)性與多環(huán)芳烴暴露致染色體損傷遺傳易感性關系及易感性差異的分子機制,揭示了ATM基因變異可能是多環(huán)芳烴暴露致染色體損傷的易感因素之一,證實了ATM在B(a)P致染色體損傷修復及細胞周期調(diào)控中起重要作用。
[Abstract]:Coke oven escapes have been defined by the international cancer research agency as 1 types of carcinogens, known as human carcinogens. Carcinogenic polycyclic aromatic hydrocarbons (Polycyclic aromatic hydrocarbons, PAH) in Coke oven emission (COE) are the main causes of lung cancer in coke oven workers. Lung cancer in coke oven workers in China is listed as eight legal occupational tumors. The study shows that genomic instability is an important biological event of polycyclic aromatic hydrocarbons (PAHs) exposure to lung cancer. The levels of chromosomal damage in workers exposed to polycyclic aromatic hydrocarbons are significantly higher than those in non exposed groups, but under the same exposure conditions, the level of damage is not the same, suggesting the presence of susceptibility to the population.
In this study, the genetic susceptibility and molecular mechanisms of chromosomal damage induced by ataxia telangiectasia (ATM) and polycyclic aromatic hydrocarbons (PAH) exposure were investigated. The results were as follows:
The analysis of the relationship between the micronucleus rate of peripheral lymphocyte micronucleus and ataxia telangiectasia in 140 polycyclic aromatic exposed workers and 66 non exposed workers showed that the ATM gene rs600931 GA and GA+AA genotype carriers in the PAH exposure group were after the correction of sex, age, smoking and natural logarithmic conversion of urine 1-OHP. The micronucleus rate of lymphocyte in peripheral blood (11.14 + 6.92% and 10.57 + 6.82 per 1000) was significantly higher than that of GG genotype (7.66 + 5.69%, P=0.015 and 0.038). The micronucleus rate of lymphocytes in rs189037 GA and GA+AA genotype (10.99 + 6.90% and 10.51 +%) was significantly higher than that of GG based carriers (7.72 + 5.83 per thousand, P=0.018 and 0.035); rs624366 G The micronucleus rate of lymphocytes in C and GC+CC genotypes (11.34 + 6.74% and 10.73 + 6.63%) was significantly higher than that of GG genotype carriers (7.61 + 6.07 per 1000, P=0.001 and 0.003), and the micronucleus rate (10.78 + 6.60%) in rs227092 GT genotype was significantly higher than that of GG genotype (7.91 + 6.30%, P=0.025). Monopal to GGGGTG The rate of micronucleus in peripheral blood lymphocytes of C/AAACATT carriers (12.05 + 7.40 per thousand) was significantly higher than that of GGGGTGC/GGGGTGC carriers (7.51 + 6.20 per thousand, P=0.007).
Two, Cytokinesis-block micronucleus (CBMN) was used to investigate the difference between B (a) P induced genetic damage induced by ATM interference cells and empty plasmid cells. The results showed that the CBMN rates of ATM and no-load granulocytes increased with the increase of B (a) P concentration, and showed a significant dose response relationship. At the concentration of 4 M and 8 M, the CBMN rate of ATM interference cells was significantly higher than the CBMN rate of empty plasmid cells, the difference was statistically significant (P0.05).
The difference between the DNA repair ability (DNA repair capacity, DRC) induced by bleomycin induced by ATM interference cells and empty plasmid cells was investigated by single cell gel electrophoresis. The results showed that the DRC (56.664 + 0.78%) of 16HBE-siATM cells was significantly lower than that of 16HBE-siGFP cells (70.13 + 3.82%) under the induction of bleomycin in 10 mu g/ml, and the difference was statistically significant. Meaning (P=0.039); similar to the expression of 16HBE cells, DRC (63.86 + 2.17%) of HEK-siATM cells was also significantly lower than that of HEK-siGFP cells (73.904 + 1.67%), and the difference was statistically significant (P=0.044).
Three, the cell cycle of B (a) P induced by ATM and unloaded plasmid cells was investigated by flow cytometry. The results showed that compared with 16HBE-siGFP cells, 4 M B (a) P significantly shortened the G1 phase of 16HBE-siATM cells and prolonged the G2 phase obviously. Lengthening.
The expression of ATN, P53, NBS1 and rH2AX protein under the action of B (a) P was investigated by Western-blotting technique. The results showed that there was no obvious change in the total protein expression of 8 u M B (a) P as compared with that of the corresponding solvent control group. The expression level of P53, NBS1 and rH2AX in granulocyte increased.
Four, using the double luciferase reporter gene experiment to investigate the differences in the expression of the ATM gene 5'UTR (rs189037GA) and 3'UTR (rs227092 GT) variation in vitro driven reporter gene. The results showed that the 5'UTR and rs189037A alleles containing the rs189037G allele were found in the cell lines of the three lung tissue sources (16HBE, CCC-HPF-1, and MRC-5). There was no significant difference in the expression of 5'UTR regulatory report gene (P0.05), and there was no statistical difference between the 3'UTR and the rs227092T alleles containing the rs227092G allele and the 3'UTR regulation of the rs227092T allele. (P0.05).
In conclusion, this study explored the genetic polymorphisms of ATM gene and the molecular mechanism of susceptibility to chromosomal damage induced by polycyclic aromatic hydrocarbons (PAH) exposure, and revealed that the ATM gene mutation may be one of the susceptible factors of chromosomal damage induced by polycyclic aromatic hydrocarbons (PAH) exposure, which confirms that ATM plays a role in the repair of chromosome damage induced by B (a) P and the regulation of cell cycle. Important role.

【學位授予單位】：中國疾病預防控制中心
【學位級別】：博士
【學位授予年份】：2010
【分類號】：R394

【參考文獻】

相關期刊論文 前10條

1 王智琴;齊以濤;楊迪;陳倩;周宗燦;王捷;梁震;肖希龍;;苯并芘對HELF細胞周期及相關基因表達的影響[J];癌變.畸變.突變;2009年01期

2 魏莉,辛曉燕,王健,雷迎鋒,薛小平;ATM基因siRNA真核表達載體的構(gòu)建及其對轉(zhuǎn)染宮頸癌細胞株ATM表達的抑制作用[J];第四軍醫(yī)大學學報;2005年05期

3 羅加林,曹建平,朱巍,劉芬菊,王明明,周獻鋒,朱財英,鄭斯英;用HPRT基因突變及CBMN研究AT細胞高輻射敏感性[J];輻射研究與輻射工藝學報;2004年06期

4 俞英欣,萬琪;ATM介導的蛋白磷酸化與DNA損傷的信號轉(zhuǎn)導機制[J];國外醫(yī)學(分子生物學分冊);2003年02期

5 李平,周元國;ATM和ATR:在G_2-M期阻滯與凋亡中的作用及相互關系[J];國外醫(yī)學(分子生物學分冊);2003年03期

6 安社娟,陳家X,陳學敏;多環(huán)芳烴致癌的分子毒理學研究進展[J];國外醫(yī)學(衛(wèi)生學分冊);2005年01期

7 盛方軍,曹建平;ATM參與細胞周期調(diào)控分子機制的研究進展[J];國外醫(yī)學.遺傳學分冊;2005年02期

8 任軍,何偉,彭程,Martin Lavin;ATM對DNA損傷后細胞反應信號傳導通路的調(diào)控作用[J];山東大學學報(醫(yī)學版);2002年06期

9 宋宜;孫志賢;;DNA雙鏈斷裂損傷反應及它的醫(yī)學意義[J];生物化學與生物物理進展;2007年09期

10 余光創(chuàng);秦宜德;伯曉晨;王升啟;;依賴于5′端非編碼區(qū)高級結(jié)構(gòu)的真核生物mRNA翻譯調(diào)控[J];中國生物化學與分子生物學報;2007年11期

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