張緒超,趙 力,陳 懿,李冬雪,胡 虹,儲(chǔ) 剛,吳 敏,3*

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環(huán)境持久性自由基及其介導(dǎo)的生物學(xué)損傷

張緒超1,趙 力1,陳 懿1,李冬雪2,胡 虹2,儲(chǔ) 剛1,吳 敏1,3*

(1.昆明理工大學(xué)環(huán)境科學(xué)與工程學(xué)院,云南 昆明 650500；2.昆明理工大學(xué)醫(yī)學(xué)院,云南 昆明 650500；3.云南省土壤固碳與污染控制重點(diǎn)實(shí)驗(yàn)室,云南 昆明 650500)

近年來,新型環(huán)境風(fēng)險(xiǎn)物質(zhì)環(huán)境持久性自由基(EPFRs)被發(fā)現(xiàn)廣泛分布于不同來源的環(huán)境介質(zhì)中,如燃燒顆粒物、土壤/沉積物、天然有機(jī)質(zhì)等.因其穩(wěn)定性、持久性,且可以隨著環(huán)境介質(zhì)遷移和轉(zhuǎn)化,EPFRs的生態(tài)環(huán)境風(fēng)險(xiǎn)可能被忽視.基于此,本文系統(tǒng)總結(jié)了存在于環(huán)境介質(zhì)中的EPFRs,并歸納其分布特征;闡述了其介導(dǎo)的組織損傷,包括肺損傷、心血管損傷、神經(jīng)毒性損傷、DNA以及細(xì)胞色素等生物大分子損傷;詳述了主要由氧化應(yīng)激、炎癥、免疫反應(yīng)以及代謝異常引起的損傷機(jī)理;最后,總結(jié)并展望了有關(guān)研究所存在的問題和未來研究方向,以期為EPFRs的生態(tài)健康風(fēng)險(xiǎn)評(píng)價(jià)和政策標(biāo)準(zhǔn)制定提供參考.

環(huán)境持久性自由基；地球表層系統(tǒng)；活性氧；環(huán)境風(fēng)險(xiǎn)；影響機(jī)理

環(huán)境持久性自由基(EPFRs)因其能在環(huán)境中穩(wěn)定存在,且穩(wěn)定時(shí)間長達(dá)數(shù)月甚至更久,被視為一種新型環(huán)境風(fēng)險(xiǎn)物質(zhì)[1-5].目前,EPFRs能夠在燃料燃燒、生物質(zhì)熱解、垃圾焚燒以及金屬冶煉等人為活動(dòng)過程中被釋放,存在于大氣顆粒物(PM)和土壤等環(huán)境介質(zhì)中,對(duì)人體具有不可知的潛在危害以及潛在的環(huán)境毒理效應(yīng).EPFRs能隨著環(huán)境介質(zhì)遷移并進(jìn)入生物體內(nèi),可能誘導(dǎo)機(jī)體產(chǎn)生氧化應(yīng)激反應(yīng),導(dǎo)致機(jī)體出現(xiàn)炎癥和免疫反應(yīng),破壞機(jī)體正常代謝.近年來,關(guān)于EPFRs的相關(guān)研究主要集中在對(duì)其產(chǎn)生的機(jī)理分析[6-10]和對(duì)污染物環(huán)境行為的影響研究[11-13].然而,針對(duì)EPFRs本身環(huán)境風(fēng)險(xiǎn)的系統(tǒng)研究,以及其存在的環(huán)境介質(zhì)風(fēng)險(xiǎn)的全面評(píng)價(jià)仍然缺乏.主要原因:有關(guān)EPFRs的研究還沒有得到足夠的重視,現(xiàn)階段存在EPFRs的環(huán)境介質(zhì)污染物(如焦化地土壤)風(fēng)險(xiǎn)的檢測(cè)、評(píng)價(jià)以及產(chǎn)生風(fēng)險(xiǎn)機(jī)理的解釋,通常只考慮了金屬元素和多環(huán)芳烴類化合物(PAHs)等傳統(tǒng)污染物[14-16];研究者忽視了實(shí)驗(yàn)過程中傳統(tǒng)有機(jī)溶劑萃取以及其他化學(xué)分析方法,可能會(huì)把持久性半醌類自由基轉(zhuǎn)換為苯酚、對(duì)苯醌類分子污染物[17].此外,研究發(fā)現(xiàn)含有EPFRs的顆粒物引起的細(xì)胞毒性比顆粒物本身更強(qiáng),導(dǎo)致細(xì)胞功能缺陷更嚴(yán)重[18-20].現(xiàn)有的國家標(biāo)準(zhǔn)還未將EPFRs納入污染物管控范疇, 這可能致使當(dāng)環(huán)境介質(zhì)污染物(如大氣顆粒物)的水平遠(yuǎn)遠(yuǎn)低于標(biāo)準(zhǔn)時(shí),仍然會(huì)對(duì)公共健康產(chǎn)生威脅[21-23].本文從EPFRs存在的環(huán)境介質(zhì)、介導(dǎo)產(chǎn)生的生物學(xué)損傷和引起損傷的機(jī)理三部分(圖1),論述了EPFRs的研究現(xiàn)狀,總結(jié)了EPFRs 在環(huán)境風(fēng)險(xiǎn)評(píng)估中的不足,為今后EPFRs的研究、風(fēng)險(xiǎn)控制和政策制定提供基礎(chǔ)數(shù)據(jù)和技術(shù)指導(dǎo).

圖1 EPFRs的環(huán)境介質(zhì)、損傷機(jī)理及其影響

1 EPFRs存在的環(huán)境介質(zhì)

1.1 大氣顆粒物中的EPFRs

1.1.1 室外大氣顆粒物 大氣顆粒物是EPFRs存在的環(huán)境介質(zhì)之一[24-25].有研究報(bào)道,可通過使用電子順磁共振儀(EPR)檢測(cè)PM中的持久性半醌類自由基的信號(hào),并且隨顆粒物粒徑變小,EPFRs濃度變高[28].通常,粒徑越小的PM因其傾向于沉積在下呼吸道和肺泡,更容易引起呼吸道毒性和功能障礙[29]; EPFRs也會(huì)隨其遷移到相應(yīng)的組織部位,引起更廣泛的組織損傷[30].在從美國各城市收集的PM樣品中已經(jīng)發(fā)現(xiàn)了類似半醌類自由基的存在,體外細(xì)胞實(shí)驗(yàn)表明EPFRs可以造成細(xì)胞的DNA損傷[18].近期研究表明,含有EPFRs的PM會(huì)引起并加重人體免疫、呼吸和心血管系統(tǒng)方面的疾病[30-32].

以往研究PM對(duì)呼吸系統(tǒng)的損傷時(shí),一般考慮常規(guī)燃燒污染物(如PM、有機(jī)污染物和重金屬)的貢獻(xiàn)[33-34],卻忽視了EPFRs的存在.Franklin等[35]和Wang等[36]先后注意到急性心血管疾病和癌癥的發(fā)生,無法使用傳統(tǒng)環(huán)境污染物的毒性風(fēng)險(xiǎn)進(jìn)行解釋, EPFRs卻有可能誘發(fā)這些疾病.Dellinger等[1,3,37]鑒定了幾乎所有燃燒源的產(chǎn)物都會(huì)存在EPFRs,同時(shí)開發(fā)了一套使用顆粒物-自由基的模型化合物進(jìn)行實(shí)驗(yàn)的方法.利用模型化合物,證明EPFRs在環(huán)境暴露引起的疾病中的主導(dǎo)作用,以及EPFRs能引起比環(huán)境介質(zhì)本身更強(qiáng)的生物活性和毒性[20,30,32,38].此外,在塑料燃燒后產(chǎn)生的顆粒煙霧排放物和殘留固體灰分中都檢測(cè)到以碳和氧為中心的EPFRs,進(jìn)入空氣后,能不斷進(jìn)行遷移從而產(chǎn)生更大范圍的風(fēng)險(xiǎn)[39].

綜上所述,各種燃料(如油、煤、木材、木炭、煙草)和塑料垃圾燃燒時(shí),不僅會(huì)產(chǎn)生大量傳統(tǒng)的燃燒副產(chǎn)物,也會(huì)產(chǎn)生穩(wěn)定的EPFRs.這些EPFRs隨著環(huán)境介質(zhì)進(jìn)行遷移轉(zhuǎn)化,使風(fēng)險(xiǎn)提高.目前,對(duì)于垃圾焚燒等過程中納米級(jí)顆粒物的歸趨及其形成的EPFRs的風(fēng)險(xiǎn)研究相對(duì)較少.因此,PM中的EPFRs潛在的環(huán)境風(fēng)險(xiǎn)應(yīng)得到足夠的重視和關(guān)注.

1.1.2 室內(nèi)空氣顆粒物 室內(nèi)PM主要來源于烹飪爐具產(chǎn)生的油煙煙霧、生物質(zhì)燃燒過程中的混合物及香煙煙霧等介質(zhì),在這些介質(zhì)中檢測(cè)到與室外PM相似的EPFRs信號(hào)[40-41].研究表明,香煙煙霧的煙焦油中的 EPFRs主要是醌/氫醌類復(fù)合體[42-43];此類EPFRs在體外容易與氧氣反應(yīng)生成羥基自由基等活性氧自由基(ROS),ROS可能引起機(jī)體的氧化損傷.研究結(jié)果顯示電子煙同樣存在健康風(fēng)險(xiǎn),其煙霧中的EPFRs信號(hào)強(qiáng)度高達(dá)7×1011radicals/puff; 將小鼠暴露在電子煙煙霧2周后,小鼠肺部出現(xiàn)了明顯的氧化應(yīng)激反應(yīng)和巨噬細(xì)胞引起的炎癥反應(yīng)[44].含有EPFRs的香煙煙霧會(huì)削弱病人針對(duì)細(xì)菌和病毒的先天及適應(yīng)性免疫應(yīng)答反應(yīng),增加慢性阻塞性肺疾病發(fā)病率和死亡率[45-46].Wang等[36]從居民住宅區(qū)收集到人們?nèi)粘Ｊ褂玫牡碗A煤中檢測(cè)到EPR信號(hào)高達(dá)2.49× 1019spins/g,而且煤燃燒爐煙囪出口的煙塵EPR信號(hào)強(qiáng)度可達(dá)3.10×1019spins/g,這些EPFRs在18個(gè)月后依然保持在一個(gè)數(shù)量級(jí)上;研究者注意到當(dāng)?shù)胤伟┑母甙l(fā)生率無法完全用傳統(tǒng)的化學(xué)物質(zhì)分析,據(jù)此推測(cè)這可能與長期暴露在EPFRs的環(huán)境中有關(guān).因此,嬰幼兒、老人及病人等在室內(nèi)時(shí)間長的人群需要格外注意家庭煙囪煙氣和煙霧的影響,這些室內(nèi)空氣顆粒物是EPFRs存在的重要環(huán)境介質(zhì).

1.2 土壤/沉積物中的EPFRs

土壤/沉積物是環(huán)境中有機(jī)污染物主要的匯,其組成主要有有機(jī)質(zhì)、金屬氧化物、粘土礦物等,構(gòu)成了EPFRs形成的必要條件[3,36].Nwos等[47]證明不需要經(jīng)過生物途徑,土壤即可形成持續(xù)時(shí)間足夠長的自由基.這說明土壤有機(jī)質(zhì)和金屬氧化物可能產(chǎn)生EPFRs.

研究發(fā)現(xiàn),五氯苯酚污染的土壤和沉積物中有很強(qiáng)的EPFRs信號(hào),其強(qiáng)度最強(qiáng)可高出空白對(duì)照土壤30倍,并且在超過10a時(shí)間里EPFRs仍不斷產(chǎn)生[48-49].另一研究發(fā)現(xiàn),通過低溫?zé)崽幚硇迯?fù)被污染的土壤反而可以增強(qiáng)EPFRs的強(qiáng)度[4].最近,在燃煤區(qū)域的土樣中檢測(cè)到類似的EPFRs信號(hào).在云南宣威地區(qū),發(fā)現(xiàn)土壤中含有大量EPFRs,并推測(cè)該發(fā)現(xiàn)可能是這一地區(qū)癌癥高發(fā)的原因之一[36].Jia等[26]在焦化地周邊的土壤中檢測(cè)到EPFRs的強(qiáng)度達(dá)到3×1017radicals/g,同時(shí)說明了PAHs對(duì)EPFRs的形成起著主要作用.在自然條件下,采煤和焦化廠附近的土地會(huì)形成EPFRs,改變濕度和光照會(huì)形成其他類型的EPFRs,這些新的EPFRs可能具有不同的性質(zhì)和毒性[50].研究者從土壤里提取了腐殖質(zhì)(土壤有機(jī)質(zhì)主要成分),從中檢測(cè)到半醌類自由基和以碳為中心的芳香類的EPR信號(hào),其強(qiáng)度也會(huì)因?yàn)闈穸群凸庹盏母淖兌兓痆50-52],在土壤中, EPFRs參與氧化還原過程,而在水體環(huán)境中,它可以誘導(dǎo)產(chǎn)生ROS[25,53-54],使土壤生物處在氧化脅迫的環(huán)境下.Liao等[55]和趙力等[56]檢測(cè)到生物炭中有以氧和碳為中心的EPR信號(hào),使用羥基捕獲劑5,5-二甲基-1-吡咯啉-N-氧化物(DMPO),成功捕獲到生物炭誘導(dǎo)水體產(chǎn)生的羥基自由基.因此,土壤/沉積物中的EPFRs是確實(shí)存在的,但其對(duì)周邊地區(qū)的人群和動(dòng)植物的健康風(fēng)險(xiǎn)還需要進(jìn)一步研究.

1.3 水體中的EPFRs

從德國和馮諾斯坎底亞采集的23個(gè)地表水樣中提取到的天然有機(jī)質(zhì)檢測(cè)到有機(jī)自由基濃度最高可達(dá)到1.84′1017spins/g[48].研究發(fā)現(xiàn)球形納米顆粒和纖維會(huì)自發(fā)形成一種新的穩(wěn)定的三苯甲基自由基,這個(gè)自發(fā)反應(yīng)可以發(fā)生在室溫水溶液中,甚至低溫冷凍的極端環(huán)境中[57],這也使得這類EPFRs的風(fēng)險(xiǎn)難以控制.鑒于納米材料的廣泛生產(chǎn)和使用,包括水體在內(nèi)的各種環(huán)境介質(zhì)中,納米材料所形成的EPFRs引起的生態(tài)風(fēng)險(xiǎn)和人體健康問題值得關(guān)注.另外,含有EPFRs的生物炭也會(huì)以各種途徑進(jìn)入水體,從而對(duì)水中的生物產(chǎn)生威脅.

綜上所述,EPFRs普遍存在于環(huán)境介質(zhì)中(大氣圈、巖石圈和水圈等).環(huán)境介質(zhì)的組成、性質(zhì)以及環(huán)境行為會(huì)影響EPFRs的類型以及其環(huán)境行為,EPFRs也可以隨著環(huán)境介質(zhì)在環(huán)境中遷移轉(zhuǎn)化,導(dǎo)致EPFRs風(fēng)險(xiǎn)增加.此外,目前評(píng)估環(huán)境介質(zhì)中污染物風(fēng)險(xiǎn)時(shí),沒有考慮到EPFRs的存在,導(dǎo)致風(fēng)險(xiǎn)評(píng)估的誤差.

2 EPFRs對(duì)生物體的環(huán)境健康風(fēng)險(xiǎn)

本文主要以人和動(dòng)物為例,介紹EPFRs介導(dǎo)的組織損傷.EPFRs可以通過呼吸直接進(jìn)入呼吸系統(tǒng)或直接暴露的方式,引發(fā)肺部、心血管、免疫和神經(jīng)系統(tǒng)等在內(nèi)多種組織的損傷(表1).

表1 EPFRs存在的環(huán)境介質(zhì)及其影響

續(xù)表1

注: 表中列出了EPFRs對(duì)動(dòng)物、植物和人的影響. ①飛灰:由工業(yè)廢料焚化產(chǎn)生[61];② MCP230:由5%的氧化銅和95%的二氧化硅(直徑小于0.2μm)顆粒組成,并在大于230°C的溫度下負(fù)載鄰氯苯酚或鄰二氯苯,分別稱之為MCP230或DCB230[20];③Slican/MCP:合成方法同MCP230[62];④DCB230:合成方法同MCP230[3,63];⑤生物炭:氮?dú)猸h(huán)境中,將水稻、玉米和小麥秸稈分別在不同溫度下熱解(200~500°C)冷卻到室溫得到[55];⑥puff與g(克)相對(duì)應(yīng),此單位暫無對(duì)應(yīng)中文翻譯,每puff在原文中對(duì)應(yīng)35mL體積氣體.*:值和強(qiáng)度分別是電子順磁共振儀器(EPR)中用來分析自由基類型和電子自旋數(shù)的值,標(biāo)*表示原文中未出現(xiàn),這里從其參考文獻(xiàn)中摘錄;EPFRs強(qiáng)度的絕對(duì)定量還沒有統(tǒng)一的標(biāo)準(zhǔn),而其強(qiáng)度與EPR信號(hào)吸收峰的面積成正比,因此一般使用吸收峰的積分面積與標(biāo)準(zhǔn)樣品1,1-二苯基-2-苦基肼基(DPPH)做對(duì)比,并用質(zhì)量或體積標(biāo)準(zhǔn)化后計(jì)算出相對(duì)強(qiáng)度,文獻(xiàn)中的單位常使用spins/g或radicals/g或radicals/puff.通常,g-Factor<2.0030時(shí),EPFRs是以碳為中心的自由基;2.00302.0040時(shí),EPFRs是以氧為中心的自由基[10,55];“-”表示文章中沒有相應(yīng)的數(shù)據(jù).

2.1 肺損傷

外源化學(xué)物可經(jīng)呼吸道和其他途徑到達(dá)肺組織,引起肺的損傷.流行病學(xué)研究表明,環(huán)境空氣中PM數(shù)量增加與死亡率升高、哮喘惡化、呼吸道感染等疾病有一定的關(guān)系[76-77].如前文所說,EPFRs會(huì)隨著顆粒物遷移并進(jìn)入呼吸系統(tǒng),從而增加疾病患病率或加重病情.

起初,研究人員在香煙煙霧中發(fā)現(xiàn)醌類/氫醌類復(fù)合物可產(chǎn)生超氧化物損傷細(xì)胞,最終導(dǎo)致肺氣腫和癌癥的發(fā)生[43].使用EPR檢測(cè)了長時(shí)間吸煙者、癌癥患者和煤炭礦工工人的肺部組織存在EPFRs,并觀察到相對(duì)嚴(yán)重的病情[58-59,78-79].機(jī)動(dòng)車輛和垃圾燃燒產(chǎn)生的顆粒物會(huì)對(duì)肺上皮細(xì)胞[80]和肺部[76,81]造成損害,這些PM中含有與香煙煙霧相似的半醌類自由基信號(hào),在肺部沉積位置參與氧化還原過程不斷產(chǎn)生ROS[76],甚至與過渡金屬等協(xié)同作用產(chǎn)生氧化應(yīng)激反應(yīng)[81],引發(fā)肺功能異常[80,82].

Dellinger等[3,37,83]合成了一種與PM有相似EPR信號(hào)的模型化合物.利用模型化合物,發(fā)現(xiàn)沉積在呼吸道中的EPFRs通過生成ROS使肺部產(chǎn)生氧化應(yīng)激和炎癥反應(yīng)對(duì)肺部造成危害,與沒有EPFRs的對(duì)照相比,實(shí)驗(yàn)組產(chǎn)生的ROS引發(fā)連鎖脂質(zhì)過氧化反應(yīng)破壞了支氣管上皮細(xì)胞(BEAS-2B)的細(xì)胞膜完整性[18],降低了細(xì)胞內(nèi)的谷胱甘肽和抗氧化酶的水平,表現(xiàn)出更強(qiáng)的細(xì)胞毒性[20,62].最近的試驗(yàn)說明EPFRs有改變免疫反應(yīng)的可能,暴露在EPFRs的環(huán)境后,肺部便處于氧化應(yīng)激狀態(tài),這種狀態(tài)抑制了免疫應(yīng)答反應(yīng)[69].用收集的車輛尾氣中的顆粒物進(jìn)行試驗(yàn),肺部的免疫應(yīng)答反應(yīng)被延遲,病毒感染的可能性增加,通過捕獲液相中的超氧陰離子和羥基自由基以及毒性實(shí)驗(yàn),證實(shí)了含有EPFRs的PM引起的氧化應(yīng)激是造成細(xì)胞毒性和肺組織損傷的主要可能機(jī)制[84-85].

同時(shí),EPFRs會(huì)引起氣道中的淋巴細(xì)胞、嗜中性粒細(xì)胞、細(xì)胞因子的改變,導(dǎo)致氣道高反應(yīng)性,增加肺部炎癥[65,67],這種癥狀甚至?xí)茐淖哟姆尾棵庖甙l(fā)育[30,68].哮喘的發(fā)展和惡化可能與EPFRs引發(fā)的氧化應(yīng)激有關(guān),EPFRs誘導(dǎo)抗原呈遞細(xì)胞成熟,使肺部發(fā)生炎癥而加重哮喘[67].此外,EPFRs可誘導(dǎo)上皮間質(zhì)轉(zhuǎn)化,不可逆的改變氣道結(jié)構(gòu)和功能,顯著影響肺部的發(fā)育[38].感染流感后,肺部巨噬細(xì)胞減少,抗細(xì)菌/病毒和宿主防御體系受損,疾病嚴(yán)重程度增加,進(jìn)而引起發(fā)病率和死亡率升高[44,69,75].

綜上,氧化應(yīng)激和炎癥反應(yīng)以及免疫抑制是造成呼吸系統(tǒng)功能降低的重要機(jī)制.而認(rèn)識(shí)到EPFRs的存在可以幫助人們重新理解致病機(jī)制,如哮喘加重的原因.

2.2 心血管損傷

有毒污染物會(huì)增加心率和心律失常的發(fā)生率,加劇動(dòng)脈粥樣硬化、冠心病和外周動(dòng)脈疾病,增加心絞痛、心肌缺血、中風(fēng)等疾病的發(fā)病率,研究表明, EPFRs與這些疾病的發(fā)生存在一定相關(guān)關(guān)系[86-87].

盡管超細(xì)顆粒物可以進(jìn)入細(xì)胞并引起主動(dòng)脈粥樣硬化[88-89],但細(xì)顆粒物中的EPFRs風(fēng)險(xiǎn)往往被忽視了.利用DCB230,發(fā)現(xiàn)EPFRs使心臟處于促炎狀態(tài),增加肺動(dòng)脈壓,降低心臟左心室基線功能,并在短暫的缺血再灌注后加重左心室功能障礙,表現(xiàn)出明顯的心臟毒性[19,66,71].此外,EPFRs刺激產(chǎn)生的氧化應(yīng)激能引起左心室肌細(xì)胞中的線粒體去極化,啟動(dòng)細(xì)胞凋亡程序,造成心肌細(xì)胞的細(xì)胞毒性[74].

EPFRs介導(dǎo)的心血管損傷的潛在機(jī)制可能是氧化應(yīng)激和炎癥反應(yīng)改變了免疫應(yīng)答反應(yīng),導(dǎo)致局部乃至全身的炎癥,從而觸發(fā)疾病的發(fā)生.然而,目前的研究只是通過氧化應(yīng)激的標(biāo)志物和已經(jīng)發(fā)生的炎癥反應(yīng)來解釋疾病的發(fā)生過程,沒有給出具體的致病機(jī)理以及EPFRs直接引起損傷的觀測(cè).

2.3 神經(jīng)毒性作用

EPFRs能夠引起機(jī)體氧化應(yīng)激反應(yīng),而氧化應(yīng)激在神經(jīng)退行性病變中起到重要作用.以中樞神經(jīng)系統(tǒng)為例,通常該部位存在較低濃度的抗氧化酶,因此生物大分子最容易受到氧化.在阿爾茲海默癥、帕金森氏病、運(yùn)動(dòng)神經(jīng)元病等疾病的臨床研究中,氧化應(yīng)激可以引起神經(jīng)元的缺失和神經(jīng)變性[90].研究者利用模式生物秀麗隱桿線蟲()首次探討了土壤改良劑生物炭中的EPFRs的神經(jīng)毒性,結(jié)果表明生物炭中的EPFRs在高濃度的時(shí)候會(huì)顯著抑制的身體彎曲頻率和相對(duì)運(yùn)動(dòng)長度,并有抑制化學(xué)感知功能的可能性,這對(duì)于生物炭在應(yīng)用過程中被忽視的風(fēng)險(xiǎn)進(jìn)行了補(bǔ)充和校正[91].

2.4 DNA和細(xì)胞色素等生物大分子損傷

研究發(fā)現(xiàn)香煙煙焦油水提取物中的半醌類自由基會(huì)使DNA產(chǎn)生切口[60],存在于PM、TiO2、石棉、陶瓷/玻璃纖維等介質(zhì)中的EPFRs也能引起這樣的損傷[92],通過檢測(cè)DNA羥基化產(chǎn)物說明EPFRs能促進(jìn)ROS的產(chǎn)生從而引起氧化損傷[61,93-94].為研究氧化應(yīng)激的調(diào)控機(jī)制,研究者使用電子顯微鏡觀察到較小尺寸(<2.5μm)的PM出現(xiàn)在線粒體上,表明EPFRs可隨之遷移進(jìn)入細(xì)胞內(nèi)并引起線粒體嚴(yán)重的結(jié)構(gòu)損傷[88,94].此外,EPFRs還會(huì)抑制肝微粒體中的細(xì)胞色素P450的正常功能,使P450參與的代謝不能正常進(jìn)行,從而影響生物體代謝和消除異物的能力[70,72-73].

綜上,EPFRs通過損傷細(xì)胞結(jié)構(gòu)和正常細(xì)胞內(nèi)蛋白質(zhì)的表達(dá),致使代謝異常,最終導(dǎo)致機(jī)體的發(fā)育異常和疾病發(fā)生.EPFRs引起的氧化應(yīng)激和代謝異常具有潛在的危害作用.

3 EPFRs介導(dǎo)的損傷機(jī)制

EPFRs與機(jī)體接觸之后,其損傷的風(fēng)險(xiǎn)來源主要有2部分:EPFRs在體外產(chǎn)生ROS,讓機(jī)體處于氧化脅迫狀態(tài);EPFRs進(jìn)入體內(nèi)產(chǎn)生ROS,損傷生物體內(nèi)的生物大分子,從而造成損傷.EPFRs對(duì)機(jī)體是否有直接作用和損傷,現(xiàn)有的研究還很少提及到.本文總結(jié)了EPFRs所能引起的損傷機(jī)理和途徑(圖2),并將其主要?dú)w為氧化應(yīng)激、炎癥和免疫反應(yīng),以及代謝異常.

圖2 EPFRs損傷機(jī)理

1.EPFRs誘導(dǎo)細(xì)胞體內(nèi)ROS產(chǎn)生并與細(xì)胞膜作用;2.ROS與線粒體膜作用;3.ROS與溶酶體膜作用;4.ROS與蛋白質(zhì)作用;5.ROS與多聚核苷酸作用;6.EPFRs與內(nèi)質(zhì)網(wǎng)上的細(xì)胞色素P450作用;7.EPFRs與線粒體直接作用;8.EPFRs誘導(dǎo)細(xì)胞因子(如IL-6)釋放;9.EPFRs在細(xì)胞體外誘導(dǎo)ROS產(chǎn)生

3.1 氧化應(yīng)激

ROS引起的氧化應(yīng)激是EPFRs介導(dǎo)生物損傷的主要原因.ROS的氧化能力一般比較強(qiáng),化學(xué)性質(zhì)非?；顫?如羥基自由基(×OH)、超氧陰離子(O- 2×)、單線態(tài)氧(1O2)等[90],其存在時(shí)間往往較短,因此稱之為瞬時(shí)自由基.EPFRs通過氧化還原循環(huán)不僅可以在體外刺激產(chǎn)生ROS[58],也可以誘導(dǎo)體內(nèi)產(chǎn)生ROS[30,38,90],ROS大量聚集后可損害細(xì)胞中幾乎所有的生物大分子(如脂質(zhì)、糖、蛋白質(zhì)和多聚核苷酸),這種損害引起的次生產(chǎn)物還可繼續(xù)產(chǎn)生與ROS同樣的損害作用,并有可能破壞抗氧化防御體系引起細(xì)胞和組織的生理及病理的損傷,造成機(jī)體的廣泛損傷,即氧化損傷[90].

在體外實(shí)驗(yàn)中,香煙煙霧、PM和DCB230中的醌類/氫醌類自由基參與氧化還原循環(huán),減少了分子氧,產(chǎn)生超氧化物,形成H2O2和×OH,通過共價(jià)結(jié)合的方式損傷DNA[58],且添加抗氧化物質(zhì)如超氧化物歧化酶(SOD)等能夠清除瞬時(shí)自由基,減輕其損傷程度[15,38,58,94].ROS還可以通過氧化膜磷脂(即脂質(zhì)過氧化)在細(xì)胞膜上引發(fā)連鎖反應(yīng),并在膜內(nèi)產(chǎn)生H2O2,脂質(zhì)過氧化導(dǎo)致丙二醛和8-異構(gòu)前列腺素等氧化產(chǎn)物的產(chǎn)生,現(xiàn)階段主要是通過檢測(cè)這些產(chǎn)物確定氧化應(yīng)激的程度[95],同時(shí)丙二醛能與蛋白質(zhì)和酶發(fā)生交聯(lián)反應(yīng)而擴(kuò)大損傷,同樣地這些損傷可以用白藜蘆醇來減緩[20,30,62].使用來自骨髓的抗原呈遞樹突狀細(xì)胞(DCs)也發(fā)現(xiàn)谷胱甘肽(GSH)的明顯變化,說明EPFRs有可能誘導(dǎo)DCs的成熟,從而引起Th17細(xì)胞炎癥[67].使用HL-1心肌細(xì)胞檢測(cè)EPFRs對(duì)細(xì)胞的毒性作用,發(fā)現(xiàn)線粒體膜電位在早期開始下降(線粒體去極化),伴隨著胱天蛋白酶增加,細(xì)胞凋亡通路被激活,后期釋放出乳氨酸脫氫酶(細(xì)胞死亡標(biāo)志物)證明細(xì)胞已經(jīng)死亡[32,74].

類似的,當(dāng)EPFRs通過各種方式進(jìn)入體內(nèi)也會(huì)對(duì)機(jī)體產(chǎn)生損傷.EPFRs使細(xì)胞產(chǎn)生氧化應(yīng)激反應(yīng)時(shí),過量的ROS會(huì)通過自由基鏈?zhǔn)椒磻?yīng),迅速擴(kuò)散并改變酶、脂質(zhì)、蛋白質(zhì)、多聚核甘酸的結(jié)構(gòu)和功能,最終誘發(fā)機(jī)體各種肺部、心血管等疾病[30,90].EPFRs增強(qiáng)DCs抗原攝取和共刺激受體表達(dá),導(dǎo)致白細(xì)胞介素IL-13增加和干擾素IFN-γ減少,輔助性T細(xì)胞數(shù)量增加[96],這將可能導(dǎo)致哮喘的發(fā)生[67,82]. MCP230/DCP230能夠在細(xì)胞中產(chǎn)生ROS,伴隨著氧化應(yīng)激反應(yīng)和炎癥反應(yīng),體內(nèi)基線心臟功能改變,在短暫的缺血再灌注后加重左心室功能障礙[62,66].此外,EPFRs會(huì)引起溶酶體和線粒體膜改變,氧化應(yīng)激驅(qū)使BEAS-2B經(jīng)歷上皮-間質(zhì)轉(zhuǎn)化(EMT),導(dǎo)致氣道結(jié)構(gòu)和功能的不可逆損傷[38].心肌細(xì)胞暴露在DCB230后會(huì)死亡,死亡的內(nèi)在凋亡途徑關(guān)鍵是線粒體膜去極化,其次是細(xì)胞色素從線粒體轉(zhuǎn)移到細(xì)胞質(zhì),最終導(dǎo)致胱天蛋白酶激活聚合酶,結(jié)果就是細(xì)胞凋亡導(dǎo)致疾病發(fā)生和加重[70,73-74].

總之, EPFRs可以誘導(dǎo)產(chǎn)生ROS,且ROS與各個(gè)生物大分子相互作用,引起細(xì)胞膜、溶酶體膜和線粒體膜結(jié)構(gòu)和功能改變,激活細(xì)胞凋亡通路,引起細(xì)胞和組織凋亡,最終導(dǎo)致機(jī)體正常功能障礙.但是,這些EPFRs影響細(xì)胞的分子機(jī)理以及信號(hào)通路還需要深入研究.

3.2 炎癥和免疫反應(yīng)

解釋EPFRs不利的健康影響時(shí),炎癥的產(chǎn)生也是關(guān)鍵的損傷機(jī)制,這是EPFRS誘導(dǎo)ROS和氧化應(yīng)激能力的直接后果[43].未被清除的ROS導(dǎo)致細(xì)胞因子和趨化因子的基因表達(dá),引起細(xì)胞內(nèi)信號(hào)傳導(dǎo)級(jí)聯(lián)反應(yīng)的激活,炎癥便開始在目標(biāo)組織中局部以及全身產(chǎn)生,并導(dǎo)致遠(yuǎn)離損傷部位的廣泛促炎作用[97-98].因此,EPFRs可能是加重人類疾病惡化的重要風(fēng)險(xiǎn)因素.

半醌類自由基經(jīng)氧化還原過程產(chǎn)生ROS,而ROS能引起炎癥反應(yīng)[80].EPFRs導(dǎo)致肺的炎癥反應(yīng),伴隨著粘膜狀態(tài)和嗜酸性粒細(xì)胞的形態(tài)改變,上皮修復(fù)機(jī)制喪失,并造成呼吸功能障礙和疾病惡化.研究表明,城市中重要的空氣污染物是PM,PM中的EPFRs可能會(huì)通過調(diào)節(jié)促炎細(xì)胞因子的釋放來調(diào)節(jié)上皮細(xì)胞的功能[82].MCP230能夠在肺部支氣管上皮細(xì)胞中誘導(dǎo)細(xì)胞毒性,而且MCP230在較長的暴露時(shí)間和等效劑量下誘發(fā)的毒性更大[14,20,65].除了損傷肺部外,吸入DCB230能產(chǎn)生全身性炎癥,誘導(dǎo)炎性細(xì)胞因子IL-6的增加[19,38].此外,暴露于EPFRs后的心臟會(huì)產(chǎn)生促炎反應(yīng),使心臟更容易受到缺血性損傷[71].所以,EPFRs介導(dǎo)的損傷機(jī)理可能是炎癥和免疫反應(yīng)所導(dǎo)致的結(jié)果.

3.3 代謝異常

代謝異常時(shí)也會(huì)誘發(fā)疾病的產(chǎn)生.一方面,正常細(xì)胞代謝本身會(huì)產(chǎn)生傳統(tǒng)的瞬時(shí)的自由基;另一方面,來自外部各種介質(zhì)中(如PM、香煙煙霧、生物炭等)的EPFRs也可以誘導(dǎo)產(chǎn)生不穩(wěn)定的自由基[90],而后者往往是影響正常代謝過程的主要原因.

在代謝的過程中細(xì)胞色素P450(酶)起到了關(guān)鍵性作用,最近發(fā)現(xiàn)其與許多疾病的發(fā)生有著密切的關(guān)系.P450可能參與ROS的生成,進(jìn)而引發(fā)上述包括氧化應(yīng)激等反應(yīng)過程的發(fā)生.PM中的EPFRs可能會(huì)被P450代謝酶活化成反應(yīng)性親電代謝物,這些物質(zhì)會(huì)對(duì)靶細(xì)胞產(chǎn)生各種毒性作用[15].此外,EPFRs還可以導(dǎo)致不同形式的P450酶活性的不可逆抑制[70,72],使接觸EPFRs的個(gè)體更加容易受到環(huán)境中存在的毒素和致癌物質(zhì)的有害影響[73].因此,EPFRs可以通過影響細(xì)胞色素,引發(fā)細(xì)胞凋亡,導(dǎo)致機(jī)體功能受損.EPFRs導(dǎo)致機(jī)體損傷不是單一機(jī)理的作用,往往是綜合作用過程.深入研究EPFRs損傷機(jī)理,可以有效控制環(huán)境介質(zhì)中的EPFRs風(fēng)險(xiǎn).

綜上所述,EPFRs對(duì)生物的影響,一般需要經(jīng)歷氧化還原的過程,引發(fā)ROS的產(chǎn)生,通過產(chǎn)生氧化應(yīng)激反應(yīng),造成炎癥和免疫反應(yīng),破壞正常的代謝等方式,使機(jī)體的正常功能受損,從而引發(fā)風(fēng)險(xiǎn).

4 結(jié)語

新型環(huán)境污染物EPFRs的環(huán)境介質(zhì)幾乎分布于大氣圈(如顆粒物)、巖石圈(如土壤)和水圈(如天然有機(jī)質(zhì))各個(gè)圈層,通過環(huán)境介質(zhì)在圈層間遷移轉(zhuǎn)化,不斷對(duì)生物圈內(nèi)的生物產(chǎn)生影響并有可能造成威脅.影響現(xiàn)有風(fēng)險(xiǎn)研究無法繼續(xù)深入的主要原因如下:一,EPFRs存在的環(huán)境介質(zhì)的成分非常復(fù)雜,造成風(fēng)險(xiǎn)量化困難;二,EPFRs與傳統(tǒng)污染物的相互作用會(huì)導(dǎo)致風(fēng)險(xiǎn)的不確定性,如傳統(tǒng)污染物與EPFRs是否存在協(xié)同作用加重疾病程度;三,大多數(shù)研究中使用的環(huán)境介質(zhì)都存在顆粒團(tuán)聚和沉降等問題[4].進(jìn)行毒性實(shí)驗(yàn)時(shí),人為地使環(huán)境介質(zhì)分散在溶液中可能會(huì)導(dǎo)致EPFRs性質(zhì)發(fā)生變化,從而不能很好的模擬EPFRs的實(shí)際情況,造成風(fēng)險(xiǎn)評(píng)估偏差.為了更好的研究和控制其風(fēng)險(xiǎn),未來還需要解決以下存在的問題.

4.1 加強(qiáng)存在EPFRs環(huán)境介質(zhì)風(fēng)險(xiǎn)的全面評(píng)估.以往研究PM、土壤和納米材料等環(huán)境介質(zhì)風(fēng)險(xiǎn)時(shí)未關(guān)注介質(zhì)中的EPFRs,因此針對(duì)這些環(huán)境介質(zhì)的風(fēng)險(xiǎn)評(píng)估應(yīng)考慮EPFRs的潛在作用,從而更有效地指導(dǎo)和控制環(huán)境介質(zhì)的不利影響.

4.2 EPFRs影響細(xì)胞的分子機(jī)理和信號(hào)通路.未來可以借鑒分子生物學(xué)的手段來探討EPFRs對(duì)細(xì)胞膜、脂質(zhì)和蛋白質(zhì)等的作用,這將有助于了解控制相應(yīng)疾病的發(fā)生和治療.此外,EPFRs對(duì)生物的直接影響仍需深入研究,如EPFRs是否可以直接損傷肺部.

4.3 EPFRs風(fēng)險(xiǎn)量化和模型化合物優(yōu)化問題.建立標(biāo)準(zhǔn)化的模型和分析方法量化風(fēng)險(xiǎn),使得毒性結(jié)果具有可比性.

4.4 EPFRs風(fēng)險(xiǎn)控制相關(guān)政策問題.利用可比較的模型和方法,增加有關(guān)EPFRs對(duì)生物影響的基礎(chǔ)研究,為政策制定提供依據(jù).

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Overlooked risks and influences of environmentally persistent free radicals in the ambient media.

ZHANG Xu-chao1, ZHAO Li1, CHEN Yi1, LI Dong-xue2, HU Hong2, CHU Gang1, WU Min1,3*

(1.Faculty of Environmental Science and Engineering, Kunming University of Science and Technology, Kunming 650500, China；2.Medical School, Kunming University of Science and Technology, Kunming 650032, China；3.Yunnan Provincial Key Lab of Carbon Sequestration and Pollution Control in Soils, Kunming 650500, China)., 2019,39(5)：2180~2189

Environmentally persistent free radicals (EPFRs) are a class of pollutants with emerging concern. They are detected in various environmental media, such as combustion products, soil/sediment, and natural organic matter, and have been attracted a great deal of research interest because of their potential toxic impacts to organisms. In this paper, the detection of EPFRs in ambient media was firstly been summarized. The negative effect or toxicity mediated of EPFRs was been described, including pulmonary injuries, cardiovascular disorders, neurotoxicities and biomacromolecule damages (such as protein, enzyme, and DNA). The mechanisms of these adverse effects (inducing oxidative stress, inflammatory and immune response and metabolic disorder) of EPFRs were discussed. We also discussed the urgently needed future research direction on EPFRs. This paper aims to provide reference for the potential risk assessment, health assessment and policy formulation of EPFRs in the environment.

environmental persistent free radicals；earth surface system；reactive oxygen species；environmental risks；mechanism

X826,R994.6

A

1000-6923(2019)05-2180-10

張緒超(1994-),男,安徽六安人,昆明理工大學(xué)碩士研究生.主要研究方向?yàn)槌志眯宰杂苫沫h(huán)境風(fēng)險(xiǎn),現(xiàn)階段主要利用秀麗隱桿線蟲來探討生物炭的毒性風(fēng)險(xiǎn)來源.發(fā)表論文2篇.

2018-09-10

云南省重點(diǎn)研發(fā)計(jì)劃資助(2018BC004);國家自然科學(xué)基金地區(qū)項(xiàng)目(41663013);國家自然科學(xué)基金重點(diǎn)項(xiàng)目(U1602231)

*責(zé)任作者, 教授, minwup@hotmail.com