江艷艷 王曉平 李偉利 田雪 姜茜茜 孫乾斌 張倩 馬林 王偉 李春 王勇
摘要 膿毒癥引發(fā)的心肌損傷是重癥監(jiān)護(hù)病房膿毒癥患者死亡的主要原因之一。通過(guò)研究脂多糖(LPS)誘導(dǎo)的膿毒癥致心肌損傷模型,探討保護(hù)心肌損傷的治療方法與策略,能為臨床治療膿毒癥引起的心肌損傷提供實(shí)驗(yàn)依據(jù)。自噬通常被認(rèn)為是降解多余、受損的細(xì)胞成分實(shí)現(xiàn)物質(zhì)循環(huán)再利用的調(diào)節(jié)機(jī)制,在保護(hù)心肌細(xì)胞結(jié)構(gòu)和功能方面發(fā)揮重要的調(diào)控作用。最新研究發(fā)現(xiàn)自噬失衡是引發(fā)心臟毒性損傷的關(guān)鍵機(jī)制,而通過(guò)自噬調(diào)節(jié)可以改善LPS引發(fā)的心肌損傷?,F(xiàn)系統(tǒng)梳理自噬信號(hào)通路在LPS引發(fā)的心肌損傷中的作用機(jī)制及中醫(yī)藥治療的研究進(jìn)展,為膿毒癥引起的心肌損傷的治療提供新的思路和方法。
關(guān)鍵詞 自噬;自噬通量;脂多糖;膿毒癥;心肌損傷;中藥復(fù)方;雙熒光標(biāo)記;中醫(yī)藥
Abstract Myocardial injury caused by sepsis is one of the leading causes of death in patients with sepsis in intensive care units.Studying the model of myocardial injury induced by Lipopolysaccharide(LPS)and exploring the therapeutic methods and strategies for myocardial injury can provide experiment basis for clinical treatment of myocardial injury induced by sepsis.Autophagy is generally considered to be a regulatory mechanism that degrades excess and damaged cell components to realize material recycling and reuse,and plays an important regulatory role in protecting the structure and function of cardiomyocytes.The latest research has found that autophagy imbalance is the key mechanism leading to cardiotoxic damage.The regulation of autophagy can improve myocardial damage caused by LPS.This paper aims to systematically sort out the mechanism of autophagy signaling pathway in myocardial injury caused by LPS and the research progress of traditional Chinese medicine treatment,and provide new ideas and methods for the treatment of myocardial injury caused by sepsis.
Keywords Autophagy; Autophagy flux; Lipopolysaccharide; Sepsis; Myocardial Injury; Chinese medicine compound; Double fluorescent labeling; Traditional Chinese medicine
中圖分類號(hào):R228文獻(xiàn)標(biāo)識(shí)碼:Adoi:10.3969/j.issn.1673-7202.2021.14.025
膿毒癥是由細(xì)菌、真菌或病毒引起的感染性疾病,嚴(yán)重時(shí)會(huì)導(dǎo)致器官功能障礙[1],其中以心臟功能障礙最為多見(jiàn),但其機(jī)制尚未完全闡明[2]。大量實(shí)驗(yàn)研究采用腹腔注射脂多糖(LPS)建立膿毒型心肌損傷模型,進(jìn)而模擬臨床膿毒癥情況下心肌損傷的病理特征并研究其具體機(jī)制。LPS是革蘭陰性細(xì)菌細(xì)胞壁外壁的組成成分,由3個(gè)通過(guò)共價(jià)鍵連接的結(jié)構(gòu)域組成,包括脂A的糖脂部分、糖鏈以及核心低聚糖[3]。其中脂A被稱為內(nèi)毒素,是LPS的毒性成分,可以顯著促進(jìn)炎癥和免疫應(yīng)答反應(yīng)。作為目前研究最多的細(xì)菌免疫刺激成分之一,LPS很難從細(xì)胞壁脫落,當(dāng)細(xì)菌死亡時(shí)它會(huì)通過(guò)溶解、破壞細(xì)胞來(lái)脫落[4]。研究表明當(dāng)LPS進(jìn)入機(jī)體后,游離的LPS可以與免疫細(xì)胞相互作用,并誘導(dǎo)炎癥介質(zhì)、促凋亡因子和促纖維化因子等的釋放,導(dǎo)致心臟冠狀動(dòng)脈血流減少,心律失常,心臟前負(fù)荷、后負(fù)荷改變等變化,直接損傷心肌細(xì)胞,影響心功能[5],造成心肌纖維化和心肌梗死等心臟疾病[6-7]。盡管現(xiàn)代醫(yī)學(xué)在危重癥的分子生物學(xué)方面已經(jīng)取得了重大進(jìn)展,但心功能障礙依舊是膿毒癥患者死亡的主要原因之一[8]。最新研究表明,LPS刺激會(huì)導(dǎo)致機(jī)體內(nèi)細(xì)胞自噬水平的變化,其中心臟自噬的改變尤為顯著。自噬作為細(xì)胞在應(yīng)激狀態(tài)下將自身受損或衰老的細(xì)胞器和錯(cuò)誤折疊的蛋白降解以維持自身能量需求的一種適應(yīng)性反應(yīng),在其失調(diào)時(shí)同樣會(huì)對(duì)細(xì)胞產(chǎn)生有害作用[9]。因此深入探討自噬在LPS誘導(dǎo)的心臟損傷中的作用機(jī)制以及基于自噬調(diào)控的治療方法具有重要的臨床意義。
1 自噬的分類
在哺乳動(dòng)物細(xì)胞中,自噬有2種不同的分類方法:1)根據(jù)細(xì)胞內(nèi)底物運(yùn)送到溶酶體腔方式的不同分為3種主要類型:大自噬,微自噬和伴侶介導(dǎo)的自噬[10]。2)根據(jù)自噬對(duì)降解底物的選擇性分為兩類:選擇性自噬和非選擇性自噬[10]。盡管每種自噬在形態(tài)上不同,但最終都會(huì)將底物運(yùn)送至溶酶體進(jìn)行降解和再循環(huán)。大自噬依賴于自噬體的形成,包裹底物運(yùn)送至溶酶體;微自噬是通過(guò)溶酶體膜的內(nèi)陷直接吸收底物;分子伴侶介導(dǎo)的自噬直接將未折疊的蛋白質(zhì)轉(zhuǎn)運(yùn)通過(guò)溶酶體膜[11]?,F(xiàn)代研究目前主要集中在大自噬(下文所指自噬),具體過(guò)程分以下幾個(gè)階段進(jìn)行:包括分隔膜的初步形成及延展、吞噬小泡的形成、自噬體成熟和底物隔離、自噬體-溶酶體膜融合以及自噬溶酶體的內(nèi)容物被溶酶體內(nèi)水解酶降解,最終自噬溶酶體的降解物被釋放以進(jìn)行物質(zhì)循環(huán)[12]。線粒體自噬(Mitophagy)是選擇性自噬的一種,其對(duì)受損或衰老線粒體的選擇性降解在維持能量穩(wěn)態(tài)和減少由功能障礙線粒體產(chǎn)生的活性氧和線粒體DNA方面起著重要作用[13]。線粒體作為細(xì)胞中的能量供應(yīng)器對(duì)心肌細(xì)胞至關(guān)重要。一些研究發(fā)現(xiàn)LPS會(huì)損害心肌細(xì)胞線粒體,通過(guò)調(diào)節(jié)線粒體自噬水平能減緩LPS誘導(dǎo)的心肌損傷[14-15]。然而自噬在LPS誘導(dǎo)的心肌病中的具體機(jī)制還需進(jìn)一步研究。
2 自噬的過(guò)程及調(diào)節(jié)因子
自噬的誘因復(fù)雜,主要分為胞內(nèi)和胞外,胞內(nèi)信號(hào)來(lái)源于損傷、衰老的細(xì)胞器及長(zhǎng)壽命蛋白等,胞外信號(hào)包括缺氧、饑餓、生長(zhǎng)因子(如胰島素、表皮生長(zhǎng)因子、成纖維細(xì)胞生長(zhǎng)因子、血小板來(lái)源增殖因子以及生長(zhǎng)激素釋放抑制因子等)等[12]。自噬的調(diào)節(jié)因子也十分多樣,在特定條件下,自噬對(duì)調(diào)節(jié)因子傳導(dǎo)的環(huán)境信號(hào)做出反應(yīng)[16]。1)饑餓是一種有效的自噬生理調(diào)節(jié)因子,可以通過(guò)哺乳動(dòng)物雷帕霉素靶點(diǎn)蛋白(Mammalian Target of Rapamycin,mTOR)調(diào)控自噬,mTOR主要包括大分子復(fù)合物mTORC1[17]。2)另外生長(zhǎng)因子刺激Ⅰ類磷脂酰肌醇3-激酶(Phosphoinositide 3-kinase,PI3K)-蛋白激酶B(Protein Kinase B,PKB)途徑或其他營(yíng)養(yǎng)相關(guān)信號(hào)(如亮氨酸),激活mTORC1負(fù)調(diào)控UNC-51激酶1復(fù)合物(ULK1、ATG13、ATG101和FIP200);相反,能量耗竭則抑制mTORC1,允許激活ULK1介導(dǎo)的自噬關(guān)鍵起始步驟[18]。程序性死亡受體-1(Beclin1)與膜泡分揀蛋白34(Vacuolar Protein Sorting 34,VPS34)及VPS15形成復(fù)合物啟動(dòng)自噬體的形成[19];隨后其他自噬相關(guān)蛋白(Autophagy Related Proteins,ATG)被募集以促進(jìn)膜的延伸,許多ATG蛋白在酵母和哺乳動(dòng)物之間是保守的,并參與泛素樣結(jié)合系統(tǒng)(該系統(tǒng)包括泛素激酶E1的Atg7,2個(gè)泛素結(jié)合酶E2的Atg10和Atg3,以及2個(gè)泛素類蛋白Atg8和Atg12,對(duì)自噬泡的成熟和包裹物的招募具有重要作用)形成自噬體,自噬體延伸需要2個(gè)泛素樣結(jié)合系統(tǒng):ATG5-ATG12結(jié)合系統(tǒng)和微管結(jié)合蛋白輕鏈3(Microtubule-associated Protein 1A/1B-light Chain 3-ATG8)結(jié)合系統(tǒng);當(dāng)膜結(jié)構(gòu)進(jìn)一步延展時(shí),自噬體與底物上的受體或銜接蛋白結(jié)合,以促進(jìn)底物進(jìn)入自噬體;隨后包裹底物的自噬體在相關(guān)分子Rab(Rab5、Rab7)、UVRAG、SNARE等的作用下與溶酶體融合,在溶酶體水解酶作用下降解底物實(shí)現(xiàn)物質(zhì)再循環(huán)。溶酶體的生物活性和功能取決于溶酶體內(nèi)環(huán)境酸堿度、溶酶體水解酶活性及溶酶體相關(guān)蛋白LAMP1、LAMP2的調(diào)節(jié)[20]。
3 自噬信號(hào)通路
自噬信號(hào)通路有很多報(bào)道,目前對(duì)自噬信號(hào)通路的研究主要集中在:1)mTOR信號(hào)通路;2)腺苷酸活化蛋白激酶(AMP-activated Protein Kinase,AMPK)信號(hào)通路;3)Beclin1-Bcl-2(Bcl2,B細(xì)胞淋巴瘤-2,抑制凋亡的線粒體內(nèi)膜蛋白)信號(hào)通路;4)轉(zhuǎn)錄因子EB(Transcription Factor EB,TFEB)信號(hào)通路。
3.1 mTOR信號(hào)通路 作為自噬負(fù)性調(diào)控因子中研究的熱點(diǎn),mTOR是進(jìn)化上保守的絲氨酸/蘇氨酸蛋白激酶,能與多種結(jié)合蛋白相互作用,其中研究較多的mTORC1由mTOR、RAPTOR、PRAS40、DEPTOR、MLST8等組成[21]。mTORC1調(diào)控自噬的機(jī)制包括:1)直接磷酸化ULK1的Ser757位點(diǎn)進(jìn)而抑制ULK1的活性,抑制自噬體的形成[11];2)磷酸化AMBRA1(自噬/Beclin1調(diào)控因子)進(jìn)而抑制ULK1穩(wěn)定性,抑制自噬體的形成[22];3)磷酸化PI3K復(fù)合物中的Atg14進(jìn)而抑制自噬體的形成[23];4)抑制TFEB的核定位進(jìn)而抑制溶酶體相關(guān)基因的轉(zhuǎn)錄,抑制自噬溶酶體的形成及降解[24]。mTORC1的上游信號(hào)包括:1)胰島素或生長(zhǎng)因子信號(hào)主要通過(guò)PI3K-AKT-TSC復(fù)合物-RHEB激活mTORC1[23];2)活化的AKT直接磷酸化抑制PRAS40(mTORC1的抑制蛋白激酶),從而激活mTORC1抑制自噬[25]。在LPS誘導(dǎo)的心肌損傷中,通過(guò)激活mTOR通路,調(diào)節(jié)自噬可以保護(hù)心肌細(xì)胞。如Han等[26]研究發(fā)現(xiàn)阿托伐他汀(Atorvastatin)通過(guò)抑制mTOR上游蛋白AKT,mTOR和mTOR下游核糖體蛋白S6激酶(p70S6K)的磷酸化促進(jìn)自噬,改善LPS誘導(dǎo)的炎癥反應(yīng),保護(hù)心臟。Zhou等[27]發(fā)現(xiàn)在LPS誘導(dǎo)的腸道上皮炎癥模型中,通過(guò)抑制mTOR的活性激活A(yù)tg5促進(jìn)自噬小體的形成,會(huì)減輕LPS誘導(dǎo)的腸道上皮細(xì)胞炎癥反應(yīng)。
3.2 AMPK信號(hào)通路 AMPK是一個(gè)高度保守的激酶,通過(guò)協(xié)調(diào)多種代謝通路來(lái)維持全身能量代謝的穩(wěn)態(tài)[28]。以α,β,和γ異源三聚體的形式存在于多種組織中,其亞基的多個(gè)位點(diǎn)可通過(guò)被磷酸化和去磷酸化而發(fā)揮自噬調(diào)節(jié)作用[29]。AMPK調(diào)控自噬的機(jī)制包括:1)磷酸化TSC2和RAPTOR抑制mTORC1,從而促進(jìn)自噬[30-31];2)直接磷酸化UKL1的Ser555和Ser317位點(diǎn),激活ULK1,進(jìn)而啟動(dòng)自噬體的形成;3)激活FOXO3,TFEB和BRD4等轉(zhuǎn)錄因子的表達(dá)間接促進(jìn)自噬[32];4)調(diào)節(jié)細(xì)胞周期負(fù)性調(diào)控因子p27Kip1的磷酸化而促進(jìn)自噬[33]。AMPK的上游信號(hào)包括:1)缺氧時(shí),細(xì)胞內(nèi)AMP或ADP水平升高,AMP或ADP可以與AMPK的γ亞基結(jié)合激活A(yù)MPK[34];2)肝激酶B1(Liver Kinase B1,LKB1),鈣調(diào)蛋白依賴性蛋白激酶酶β(CaMKKβ)以及轉(zhuǎn)化生長(zhǎng)因子激活酶(TGFβ-activated Kinase,TbAK1)也可激活A(yù)MPK[35-37]。研究發(fā)現(xiàn)通過(guò)AMPK通路調(diào)節(jié)心肌自噬水平可以對(duì)抗LPS誘導(dǎo)的心肌損傷。如Zhang等[38]發(fā)現(xiàn)LPS可以明顯促進(jìn)心肌細(xì)胞中AMPK上游調(diào)節(jié)因子蛋白磷酸酶2(PP2A)和蛋白磷酸酶2C(PP2C)的表達(dá)抑制AMPK,激活mTOR,從而改變心肌細(xì)胞自噬水平。
3.3 Beclin1-Bcl-2信號(hào)通路 Beclin 1是酵母自噬相關(guān)蛋白Atg6與Vps30在哺乳動(dòng)物中的同源物,與PI3K-Ⅲ相互作用產(chǎn)生磷脂酰肌醇3-磷酸(Phosphatidylinositol 3-phosphate,PI3P),PI3P是自噬小體形成所必需的效應(yīng)蛋白[39]。抗凋亡蛋白Bcl-2能與Beclin1相互作用,調(diào)節(jié)細(xì)胞內(nèi)的自噬水平[40]。Beclin1-Bcl-2調(diào)節(jié)自噬的機(jī)制為:正常情況下,Beclin1與液泡蛋白分選蛋白(Vacuolar Protein Sorting,Vps34),p150,Atg14三者結(jié)合形成復(fù)合物,Bcl-2通過(guò)BH3結(jié)構(gòu)域與Beclin1相互作用,抑制Beclin1活性,從而抑制Beclin1介導(dǎo)的自噬;在缺氧或饑餓等應(yīng)激條件下,Bcl-2會(huì)與Beclin1復(fù)合物解離,誘導(dǎo)Beclin1引起的自噬[41]。研究發(fā)現(xiàn)在受到外界刺激時(shí),負(fù)調(diào)控自噬活性的Bcl-2家族蛋白(Bcl-2/XL)可以被各種激酶(包括DAPK、ROCK1、Mst1和JNK1)調(diào)控,促進(jìn)其與Beclin1的解離誘導(dǎo)自噬[42]。Sun等[43]的研究發(fā)現(xiàn)在LPS誘導(dǎo)的心肌損傷模型中,通過(guò)調(diào)節(jié)Beclin 1的表達(dá)改善細(xì)胞自噬水平會(huì)減輕LPS誘導(dǎo)的心肌損傷。
3.4 轉(zhuǎn)錄因子EB(Transcription Factor EB,TFEB)信號(hào)通路 TFEB是自噬的主要轉(zhuǎn)錄調(diào)控因子之一,它促進(jìn)自噬體形成、溶酶體生物發(fā)生,是溶酶體功能的關(guān)鍵調(diào)控因子[44]。在饑餓或溶酶體功能障礙的情況下,TFEB被磷酸化,進(jìn)而迅速轉(zhuǎn)移到細(xì)胞核并激活其靶基因的轉(zhuǎn)錄[45]。TFEB調(diào)控自噬的機(jī)制包括:1)直接與協(xié)調(diào)溶酶體表達(dá)和調(diào)控的基序結(jié)合,促進(jìn)包含其啟動(dòng)子調(diào)控基序的整個(gè)基因網(wǎng)絡(luò)的表達(dá),增加溶酶體數(shù)量和溶酶體酶的釋放[46-47];2)與眾多自噬基因的啟動(dòng)子區(qū)域結(jié)合,并誘導(dǎo)自噬體生物發(fā)生和自噬體溶酶體融合;3)TFEB過(guò)表達(dá)還可以導(dǎo)致大體積自噬底物(如長(zhǎng)壽蛋白)的降解增強(qiáng)、脂滴清除和線粒體損傷;4)參與調(diào)節(jié)特異性細(xì)胞器自噬,如脂噬和線粒體自噬[45]。許多研究發(fā)現(xiàn)通過(guò)調(diào)控TFEB的磷酸化水平來(lái)控制細(xì)胞自噬水平,對(duì)疾病的發(fā)生發(fā)展有良好的調(diào)節(jié)作用。如Kim等[48]研究發(fā)現(xiàn)通過(guò)CO激活蛋白激酶R(PKR)樣內(nèi)質(zhì)網(wǎng)激酶(PERK)和Ca2+水平和鈣調(diào)磷酸酶活性,增加了TFEB的核轉(zhuǎn)運(yùn),提高了鼠肝細(xì)胞和肝組織中線粒體自噬的水平和溶酶體基因Lamp1、CathB、TPP1等的表達(dá)。
雖然LPS調(diào)節(jié)自噬的機(jī)制尚未完全闡明,但許多研究證實(shí)傳統(tǒng)中藥通過(guò)多途徑調(diào)節(jié)心肌細(xì)胞mTOR、AMPK以及自噬體形成關(guān)鍵蛋白的水平,有望保護(hù)心肌細(xì)胞對(duì)抗LPS誘導(dǎo)的心肌損傷[38,49]。
4 自噬與LPS誘導(dǎo)心肌損傷的關(guān)系
LPS引起的膿毒癥造成的急性休克和心肌損傷,一直是臨床醫(yī)生和科學(xué)家高度關(guān)注的問(wèn)題,然而其致病機(jī)制尚未明確。許多研究已經(jīng)探討了LPS誘導(dǎo)的心肌損傷的分子機(jī)制,先前的研究表明炎癥反應(yīng),微循環(huán)受損和細(xì)胞病變性缺氧是LPS誘導(dǎo)心臟損傷的重要病因[50-51]。而隨著自噬在應(yīng)激狀態(tài)下維持細(xì)胞內(nèi)環(huán)境穩(wěn)態(tài)和保護(hù)心肌細(xì)胞的作用被重視,自噬調(diào)控逐漸成為了治療膿毒癥中心肌損傷的關(guān)鍵機(jī)制[17,52]。
最近的研究發(fā)現(xiàn)LPS刺激產(chǎn)生的過(guò)量炎癥介質(zhì)會(huì)引起小鼠心肌自噬水平發(fā)生顯著變化[53]。然而不同研究者對(duì)小鼠心肌自噬水平的變化有不同的觀點(diǎn):一些研究者認(rèn)為自噬是細(xì)胞在應(yīng)激狀態(tài)下的適應(yīng)性反應(yīng),通過(guò)提高心肌細(xì)胞自噬通量可以對(duì)抗LPS誘導(dǎo)的心肌損傷,如Pan等[54]研究發(fā)現(xiàn)褪黑素可以調(diào)節(jié)線粒體解偶聯(lián)蛋白(UCP),提高LPS誘導(dǎo)的心肌細(xì)胞自噬水平,保護(hù)線粒體的結(jié)構(gòu)和功能,緩解LPS誘導(dǎo)的心肌損傷;Unuma等[55]發(fā)現(xiàn)鈷原卟啉可以激活心肌細(xì)胞TFEB和LAMP蛋白促進(jìn)自噬體形成,保護(hù)心肌細(xì)胞,緩解LPS誘導(dǎo)的心肌損傷。也有研究者認(rèn)為L(zhǎng)PS誘導(dǎo)的心肌細(xì)胞自噬是過(guò)度自噬會(huì)損害心肌細(xì)胞,如Li等[56]認(rèn)為L(zhǎng)PS誘導(dǎo)的心肌細(xì)胞自噬是過(guò)量的,他們發(fā)現(xiàn)一種重要的生物活性肽阿拉曼定(Alamandine)可以在一定程度上抑制LPS處理小鼠心臟中Atg3和Atg5 mRNA水平的升高,減少LPS誘導(dǎo)的微管結(jié)合蛋白輕鏈3(Microtubule-associated Protein 1A/1B-light Chain 3,MAP-LC3)的增加,降低LPS處理鼠的自噬水平的強(qiáng)烈增加,減弱LPS對(duì)小鼠心臟結(jié)構(gòu)和功能的破壞。LPS刺激劑量、刺激方式及刺激時(shí)間以及判斷自噬水平的監(jiān)測(cè)方法的差異,均會(huì)導(dǎo)致自噬調(diào)控結(jié)果在同一膿毒癥模型中的表現(xiàn)不一致。尤其是針對(duì)自噬水平的判斷,多數(shù)研究集中于自噬相關(guān)蛋白表達(dá)量的研究,而自噬是一個(gè)動(dòng)態(tài)且復(fù)雜的過(guò)程,需要對(duì)其整體通量進(jìn)行綜合評(píng)估。如在研究自噬水平時(shí),通過(guò)自噬指示蛋白的表達(dá)量來(lái)反映細(xì)胞的自噬水平是片面的,應(yīng)在自噬指示蛋白表達(dá)水平研究的基礎(chǔ)上,進(jìn)一步綜合采用GFP-mRFP-LC3雙熒光標(biāo)記、電鏡等方法,對(duì)自噬通量即包括自噬體、自噬溶酶體以及溶酶體活性等多個(gè)環(huán)節(jié)與角度進(jìn)行綜合評(píng)估進(jìn)而判斷自噬通量[57]。同時(shí),研究自噬通量的時(shí)間點(diǎn)也因研究目的不同而異。因此,應(yīng)建立LPS刺激的劑量梯度和時(shí)間梯度,以檢測(cè)并繪制心肌細(xì)胞在體自噬水平變化。綜合以上的研究將為通過(guò)干預(yù)自噬治療LPS引起的心肌損傷的最佳給藥時(shí)間及給藥劑量提供指導(dǎo)。
5 中藥在LPS引發(fā)心肌損傷中的研究進(jìn)展
根據(jù)《國(guó)際膿毒癥和感染性休克管理指南》2016版[58],針對(duì)LPS引發(fā)的心肌損傷強(qiáng)烈建議盡快開(kāi)始抗菌治療,這是由于在治療膿毒癥或膿毒癥休克時(shí),每延遲1 h使用抗生素,都可能導(dǎo)致死亡率增加。然而抗生素作為現(xiàn)代醫(yī)學(xué)普遍認(rèn)可的治療膿毒癥的藥物,其對(duì)膿毒癥引起的心肌損傷未見(jiàn)有效[51]。而且臨床對(duì)抗生素的使用有嚴(yán)格要求,如具有安全一次性或快速輸液優(yōu)勢(shì)的β-內(nèi)酰胺類抗生素,雖可迅速進(jìn)入血液發(fā)揮抗炎、抗菌作用,減輕患者全身及各器官膿毒癥反應(yīng),但大量、長(zhǎng)時(shí)間服用會(huì)產(chǎn)生耐藥性和不良反應(yīng)(如雙硫侖樣作用),因此其使用劑量和時(shí)間有一定限制。除了盡快開(kāi)始抗生素治療外,機(jī)械通氣、糾正低氧和酸中毒、維持血壓和心輸出量等均是治療膿毒癥引起心肌損傷的重要治療方式[59],使用相應(yīng)的治療心血管損傷的藥物,可減輕心肌損傷,但此類藥物只是單純的對(duì)癥治療,對(duì)膿毒癥引發(fā)的心肌損傷仍有局限作用。近來(lái)有研究顯示二甲雙胍(Metformin)、生物活性肽(Alamandine)和右美托咪定(Dexmedetomidine)等藥物均可通過(guò)調(diào)控心肌細(xì)胞自噬,進(jìn)而減輕LPS誘導(dǎo)的心肌損傷[56,60-61]。但這些藥物的具體作用機(jī)制尚未完全闡明,且未應(yīng)用于臨床。因此需要我們繼續(xù)尋找和探索既能保護(hù)LPS誘導(dǎo)心肌損傷同時(shí)不良反應(yīng)小的新藥。
中醫(yī)藥在預(yù)防和治療LPS誘導(dǎo)的心肌損傷中有巨大潛力,研究證實(shí)部分中藥復(fù)方、單味藥及活性成分可通過(guò)抗炎、抗心肌纖維化、抗氧化、抗凋亡等不同途徑減輕LPS誘導(dǎo)的心肌損傷。中藥復(fù)方如:1)強(qiáng)心一號(hào)復(fù)方由水紅花子、黃芪、茯苓、丹參和五味子組成[62],通過(guò)抑制膿毒癥小鼠心肌細(xì)胞凋亡改善病理性心肌肥大;2)傳統(tǒng)中藥復(fù)方四逆湯由附子(制)、干姜、炙甘草組成,可降低膿毒癥中晚期大鼠心肌細(xì)胞中TNF-α、IL-10等炎癥介質(zhì)的表達(dá)水平,改變大鼠體內(nèi)炎癥介質(zhì)大量釋放的狀態(tài),糾正大鼠心肌的免疫紊亂,從而減輕膿毒癥大鼠心肌損傷[63-64]。單味藥如:1)人參和西洋參,分別為五加科人參屬植物人參和西洋參的根,可補(bǔ)氣滋陰,提高機(jī)體免疫力,降低小鼠心肌細(xì)胞NADPH氧化酶2(NOX2)表達(dá)水平,從而使心肌細(xì)胞炎癥介質(zhì)超氧陰離子(O2-)以及腫瘤壞死因子-α(TNF-α)的生成減少,減輕LPS引發(fā)的心肌損傷[65];2)紅花,為菊科植物紅花的干燥花,常用來(lái)治療心臟病和炎癥,可以激活抗凋亡蛋白Bcl-2和p-Bad,下調(diào)Bax、細(xì)胞色素C、凋亡蛋白酶(Caspase-9、Caspase-3)和凋亡誘導(dǎo)因子表達(dá),激活胰島素樣生長(zhǎng)因子受體信號(hào)通路,抑制LPS誘導(dǎo)的H9C2心肌細(xì)胞凋亡[7]。中藥活性成分如:1)麝香酮來(lái)源于麝科動(dòng)物林麝或原麝成熟雄體香囊中的干燥分泌物麝香,可以顯著下調(diào)膿毒癥小鼠IL-1β、TNF-α和IL-6等炎癥介質(zhì)的水平,抑制核因子κB,NLRP3炎性小體的活性,減輕心肌梗死后心臟組織巨噬細(xì)胞介導(dǎo)的炎癥反應(yīng),保護(hù)心肌梗死后心臟組織[66];2)楊梅素(Myricetin)為楊梅科植物楊梅的樹(shù)皮中提取的一種黃酮醇類化合物,能抑制一氧化氮合酶(iNOS)的過(guò)表達(dá)和氧化還原酶(SOD和GPx)活性的降低,通過(guò)抗炎,抗氧化減輕LPS誘導(dǎo)的心肌損傷[67];3)蘆丁又稱蕓香苷,廣泛存在于植物的花和果實(shí)中,具有抗氧化、抗炎的作用,可以減少LPS誘導(dǎo)的心肌纖維化[68];4)丹參酮ⅡA磺酸鈉(Sodium TanshinoneⅡA Sulfonate)為唇形科植物丹參的根提取分離得到的二萜醌類化合物,可以減緩LPS誘導(dǎo)的家兔血流動(dòng)力學(xué)惡化,調(diào)節(jié)炎癥介質(zhì)分泌,減輕LPS造成的心肌損傷[69]。
近年來(lái)隨著自噬研究的深入,自噬在維持心臟結(jié)構(gòu)和功能中關(guān)鍵作用已被確認(rèn),以上藥物能否通過(guò)調(diào)節(jié)自噬保護(hù)LPS誘導(dǎo)的心肌損傷具有極大的研究?jī)r(jià)值。另外其他具有心肌保護(hù)作用的中藥復(fù)方及其提取物,如:丹參酮ⅡA、黃芪甲苷、木犀草素、紫荊黃酮、葒草苷、木酚素、丹酚酸B、加味丹參飲、芪丹理心丸、芪藶強(qiáng)心等,在不同心肌損傷模型中通過(guò)影響細(xì)胞自噬水平進(jìn)而治療心肌損傷的效果也被證實(shí)[57,70-74],但能否通過(guò)調(diào)節(jié)自噬水平治療LPS誘導(dǎo)的心肌損傷尚未得到證實(shí)。
6 討論
自噬水平的改變是LPS誘導(dǎo)心肌損傷的重要病理改變,調(diào)節(jié)心臟自噬水平的變化能緩解LPS誘導(dǎo)的心肌損傷。最新研究提示,體內(nèi)注射LPS導(dǎo)致心肌細(xì)胞內(nèi)自噬流的迅速變化,自噬水平的變化與心肌的結(jié)構(gòu)和功能密切相關(guān)。而通過(guò)多渠道調(diào)節(jié)心肌自噬水平顯著減輕了LPS誘導(dǎo)的心肌損傷,這為臨床治療LPS引起的膿毒癥提供了理論基礎(chǔ)。傳統(tǒng)中藥能通過(guò)不同機(jī)制調(diào)節(jié)自噬,保護(hù)多種原因引起的心肌損傷。然而無(wú)論是從機(jī)制還是從藥物篩選上,應(yīng)用中藥調(diào)節(jié)心肌自噬水平治療膿毒癥引起的心肌損傷的研究都十分缺乏,仍需進(jìn)一步探索。
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(2020-09-01收稿 責(zé)任編輯:楊覺(jué)雄)