外周血單核細(xì)胞亞群及其趨化因子在急性冠脈綜合征早期的表達(dá)特點(diǎn)

許苗苗 沈 偉 施海明 莊心宇 劉湘緒 歐 洋 孫晟甲 吳幫衛(wèi) 朱志棟 陳羽斐

(復(fù)旦大學(xué)附屬華山醫(yī)院心內(nèi)科 上海 200040)

外周血單核細(xì)胞亞群及其趨化因子在急性冠脈綜合征早期的表達(dá)特點(diǎn)

許苗苗 沈 偉△施海明 莊心宇 劉湘緒 歐 洋 孫晟甲 吳幫衛(wèi) 朱志棟 陳羽斐

(復(fù)旦大學(xué)附屬華山醫(yī)院心內(nèi)科 上海 200040)

目的 探討急性冠脈綜合征(acute coronary syndrome,ACS)早期單核細(xì)胞亞群及其趨化因子即單核細(xì)胞趨化蛋白-1 (monocyte chemoattractant protein,MCP-1)和不規(guī)則趨化因子(fractalkine,FKN)的表達(dá)特點(diǎn),并分析其相關(guān)性。方法 選取我院2016年9月至12月以胸痛癥狀入院擬行冠脈造影術(shù)(coronary angiography,CAG)的患者。手術(shù)當(dāng)天術(shù)前抽取靜脈血,采用流式細(xì)胞術(shù)檢測(cè)外周血單核細(xì)胞(monocyte,Mon) 3個(gè)亞型的含量及其比例,依據(jù)分化抗原-14 (cluster differentiation-14,CD-14)和CD16表達(dá)分為3個(gè)亞型即CD14+CD16-Mon (Mon1)、CD14+CD16+Mon (Mon2)和CD14-CD16+Mon (Mon3);手術(shù)當(dāng)天術(shù)前及術(shù)后一天抽取靜脈血,ELISA檢測(cè)Mon1的趨化因子MCP-1和Mon3的趨化因子FKN水平,比較不同組MCP-1-Mon1和FKN-Mon3水平變化,并分析其相關(guān)性。結(jié)果 共入選70例患者,結(jié)合其臨床癥狀、心肌標(biāo)志物、心電圖、CAG檢查結(jié)果進(jìn)行診斷分組:急性心肌梗死(acute myocardium infarction,AMI)組患者30例、不穩(wěn)定性心絞痛(unstable angina pectoris,UAP)組患者25例、CAG完全正常者(對(duì)照組) 15例。流式細(xì)胞術(shù)結(jié)果顯示AMI組Mon1所占比例高于UAP組和正常對(duì)照組(P<0.05),Mon3在各組間尚無(wú)差異。AMI組患者外周血Mon3/Mon1比值低于對(duì)照組(P<0.05)。AMI組和UAP組患者FKN、MCP-1和紅細(xì)胞分布寬度均高于對(duì)照組,并且FKN和Mon3具有強(qiáng)相關(guān)性(P<0.05;R=0.650 2)。 結(jié)論 單核細(xì)胞亞群(Mon1和Mon3)在ACS早期水平增高,并伴有其負(fù)責(zé)招募的趨化因子(MCP-1和FKN)增加,且FKN和Mon3具有強(qiáng)相關(guān)性,提示MCP-1-Mon1和FKN-Mon3兩條通路可能參與患者ACS早期病理生理過(guò)程。

急性心肌梗死; 急性冠脈綜合征; 單核細(xì)胞亞群; 單核細(xì)胞趨化蛋白-1; 不規(guī)則趨化因子

急性冠狀動(dòng)脈綜合征(acute coronary syndromes,ACS)是以冠狀動(dòng)脈粥樣硬化(coronary arteriosclerosis,AS)斑塊破裂或糜爛、潰瘍,繼發(fā)血管完全或不完全閉塞性血栓形成為病理基礎(chǔ)的一組臨床綜合征,包括急性心肌梗死(acute myocardium infarction,AMI)和不穩(wěn)定性心絞痛(unstable angina pectoris,UAP)。AS以單核細(xì)胞(monocyte,Mon)黏附到內(nèi)皮細(xì)胞并遷移至動(dòng)脈壁下和脂質(zhì)積累為特征[1]。

單核細(xì)胞分為三個(gè)亞型:CD14+CD16-、CD14+CD16+、CD14-CD16+,分別簡(jiǎn)稱為Mon1 (炎癥型)、Mon2 (中間型)、Mon3 (定居型)[2]。單核細(xì)胞在骨髓(骨髓造血系統(tǒng))和脾臟(髓外造血系統(tǒng))產(chǎn)生。Mon1首先在損傷部位積累并分化為巨噬細(xì)胞,然后吞噬脂質(zhì)和膽固醇結(jié)晶變?yōu)楦缓|(zhì)的泡沫細(xì)胞。Mon2功能尚不清楚。Mon3在脈管系統(tǒng)“巡邏”,然后進(jìn)入損傷部位(數(shù)量少于Mon1)。ACS發(fā)生時(shí)的多重病理生理和交感反應(yīng)最終介導(dǎo)了骨髓造血系統(tǒng)祖細(xì)胞的釋放瀑布,也是心梗后激活骨髓的過(guò)程[3]。動(dòng)物實(shí)驗(yàn)已證實(shí),Mon1依賴單核細(xì)胞趨化蛋白-1 (monocyte chemoattractant protein,MCP-1)與其表面受體結(jié)合,募集并黏附到血管內(nèi)皮細(xì)胞,引起炎性反應(yīng)并促進(jìn)粥樣斑塊形成,同時(shí)Mon1可在急性心肌梗死發(fā)生后發(fā)揮清除壞死心肌殘骸的作用[3];Mon3依賴不規(guī)則趨化因子(fractalkine,FKN)募集到血管損傷部位,可促進(jìn)斑塊內(nèi)血管新生從而促進(jìn)斑塊的破裂、糜爛和血栓形成,導(dǎo)致冠狀動(dòng)脈完全或不完全閉塞[2],同時(shí)Mon3可在AMI后的增殖修復(fù)階段促進(jìn)缺血心肌血管新生和重建。

然而,國(guó)內(nèi)外關(guān)于人的單核細(xì)胞亞群在ACS患者外周血中水平的研究很少,尚未有以單核細(xì)胞在外周血的趨化及招募為切入點(diǎn)的研究。臨床試驗(yàn)已證實(shí)ACS患者血漿中FKN表達(dá)上調(diào)會(huì)促使AS斑塊不穩(wěn)定的風(fēng)險(xiǎn)指標(biāo)升高[4]。動(dòng)物實(shí)驗(yàn)表明,依賴單核細(xì)胞不同亞型分化水平及其趨化因子FKN和MCP-1介導(dǎo)的信號(hào)招募機(jī)制,單核細(xì)胞亞群比例和分布在ACS中發(fā)揮重要作用。為進(jìn)一步研究在人體內(nèi),ACS早期階段MCP-1-Mon1和FKN-Mon3的水平及相關(guān)性,本臨床試驗(yàn)首次同步檢測(cè)MCP-1-Mon1和FKN-Mon3的表達(dá)特點(diǎn),對(duì)單核細(xì)胞亞群及其趨化因子在ACS早期的診斷具有很強(qiáng)的臨床意義。

資 料 和 方 法

研究對(duì)象 連續(xù)收集2016年9月至12月在復(fù)旦大學(xué)附屬華山醫(yī)院心內(nèi)科以胸痛癥狀就診,并擬行冠脈造影術(shù)(coronary angiography,CAG)的住院患者。按照入選標(biāo)準(zhǔn)將患者分為AMI組、UAP組和對(duì)照組。入選標(biāo)準(zhǔn):年齡小于85歲擬行CAG術(shù)者,UAP診斷標(biāo)準(zhǔn)參照美國(guó)心臟病學(xué)會(huì)/美國(guó)心臟協(xié)會(huì)(AHA/AHA)2007年診斷指南[5],AMI診斷標(biāo)準(zhǔn)參照2012年AHA/AHA/ESC/WHF聯(lián)合發(fā)布的第3版《全球心肌梗死新定義》[6],有胸痛癥狀而CAG完全正常為對(duì)照組。排除標(biāo)準(zhǔn):合并先天性心臟病、擴(kuò)張型心肌病、瓣膜性心臟病、Killip分級(jí)Ⅲ-Ⅳ級(jí)、肝腦腎重要器官功能障礙、血液系統(tǒng)疾病、結(jié)締組織疾病、腫瘤的患者。

臨床常規(guī)檢測(cè) 研究方案經(jīng)復(fù)旦大學(xué)附屬華山醫(yī)院倫理委員會(huì)批準(zhǔn)(倫理審查號(hào):KY2015-244),所有入組患者簽署知情同意書(shū),分別于CAG當(dāng)天術(shù)前(-0 d)和術(shù)后1天(+1 d)晨起6:00空腹抽取靜脈全血(需急診PCI患者術(shù)前臨時(shí)抽血),后序血樣本檢測(cè)在2 h內(nèi)進(jìn)行。其中,-0 d全血2 mL用于流式細(xì)胞術(shù)檢測(cè)外周血單核細(xì)胞亞群,-0 d和+1 d全血4 mL經(jīng)792×g離心5 min后,上層血漿用于ELISA檢測(cè)MCP-1和FKN。常規(guī)測(cè)肌鈣蛋白T (troponin T,cTnT)、肌酸激酶MB型同工酶(creatine kinase isoenzyme-MB,CK-MB)、肌紅蛋白(myoglobin,MYO)、尿酸(uric acid,UA)。 常規(guī)檢測(cè)外周血中性粒細(xì)胞(neutrophil,N)比例、淋巴細(xì)胞(lymphocyte,L)比例、單核細(xì)胞(monocyte,M)比例、紅細(xì)胞分布寬度(red cell distribution width,RDW)、血小板平均體積(mean platelet volume,MPV)、膽固醇(total cholesterol)、三酰甘油(triglyceride,TG)、高密度脂蛋白(HDL-C)、低密度脂蛋白(LDL-C)、超敏C反應(yīng)蛋白(high-sensitivity C-reactive protein,Hs-CRP)水平。記錄所有患者CAG檢查結(jié)果,包括GENSINI評(píng)分和冠狀動(dòng)脈各支病變情況:左冠狀動(dòng)脈(left anterior descending,LAD)、左主干動(dòng)脈(left main coronary artery,LM)、右冠狀動(dòng)脈(right coronary artery,RCA)、左室后支(posterior branch of left ventricle,PBLV)、左旋支(left circumflex,LCX)、鈍緣支(obtuse marginal,OM)、對(duì)角支(diagonal branches,DI)。

主要試劑與設(shè)備 異硫氰酸熒光素(fluorescein isothiocyanate,FITC)標(biāo)記鼠抗人CD14、別藻青蛋白(allophycocyanin,APC)標(biāo)記鼠抗人CD16、PBS 緩沖液、淋巴細(xì)胞分離液(美國(guó)BD公司);人CX3CL1/FKN ELISA試劑盒,(R&D Cat:DCX310)和人CCL2/MCP-1 ELISA試劑盒(R&D Cat:DCP00)(美國(guó)R&D公司)。TRIzol試劑(INVITROGEN公司),逆轉(zhuǎn)錄試劑(日本Takara公司),PCR試劑(國(guó)藥集團(tuán)),引物合成由上海右升生物科技有限公司完成。

流式細(xì)胞術(shù)檢測(cè)外周血單核細(xì)胞亞群

提外周血單個(gè)核細(xì)胞 取2 h內(nèi)新鮮抽取的晨起空腹外周靜脈血2 mL,EDTA抗凝,792×g離心5 min,將下層血細(xì)胞(約1 mL)用1 mL 1×PBS稀釋1倍,輕輕吹打,將2 mL血細(xì)胞稀釋液加入含2 mL淋巴細(xì)胞分離液的15 mL離心管中,靜置2 min。792×g設(shè)置緩升緩降離心20 min。離心后血細(xì)胞清晰分為4層,上層為血漿層,中層為分離液層,用吸管將兩者之間薄層云霧狀的外周血單個(gè)核細(xì)胞(peripheral blood mononuclear cell,PBMC) PBMC層輕輕吸出,并收集到1.5 mL EP管中。

洗滌并染色 將PBMC用PBS緩沖液1 mL洗滌,792×g離心5 min;重復(fù)洗滌、離心1次,以 100 μL PBS 緩沖液重懸細(xì)胞。加入 20 μL CD14-異硫氰酸熒光素(fluorescein isothiocyanate,FITC)、5 μL CD16-別藻青蛋白(allophycocyanin,APC)單克隆抗體 ,振蕩混勻,避光室溫下孵育15 min;300×g離心5 min,棄上清;PBS 緩沖液1 mL洗滌;后重懸細(xì)胞。

檢測(cè)方法 前向角散射光(forward scatter,FSC)和側(cè)向角散射光(side scatter,SSC)雙參數(shù)點(diǎn)圖檢測(cè)外周血各群細(xì)胞;以 FSC為橫坐標(biāo),SSC為縱坐標(biāo),建立FSC和SSC 散點(diǎn)圖,正向設(shè)門法圈出單核細(xì)胞群(圖 1 A);根據(jù)單核細(xì)胞上CD14 和 CD16 表達(dá) 水平分為CD14+CD16-Mon (Mon1)、CD14+CD16+Mon (Mon2)及 CD14-CD16+Mon (Mon3) 3個(gè)亞群(圖1 B);用 Cell Quest和BD Accuri C6軟件獲取并分析數(shù)據(jù)。

ELISA檢測(cè)血漿FKN和MCP-1表達(dá)水平 將新鮮抽取的晨起空腹外周靜脈血4 mL,792×g離心5 min,取上層血漿置于2個(gè)1.5 mL EP管中,-80 ℃冷凍。其中一管按照人CX3CL1/FKN ELISA試劑盒(R&D Cat:DCX310)操作檢測(cè)FKN:依次加100 μL Assay Diluent RD1-88到每孔;分別加100 μL標(biāo)品、樣本和空白對(duì)照到適當(dāng)?shù)目變?nèi),貼膜,4 ℃孵育3 h;洗脫:PBS洗3次,第4次洗完后,充分拍干出去孔內(nèi)液體;加200 μL預(yù)冷指示劑人的FKN 結(jié)合到每個(gè)孔,封膜,4 ℃孵育3 h;重復(fù)洗脫步驟;每孔加200 μL 底物顯色劑,室溫30 min避光孵育;每孔加50 μL 終止液,充分反應(yīng),顏色從藍(lán)逐漸變黃,在450 nm下測(cè)定吸光度值。同樣,另一EP管血漿用于檢測(cè)MCP-1,按人CCL2/MCP-1 ELISA試劑盒(R&D Cat:DCP00)操作在450 nm下測(cè)定吸光度值(D)。

A:FSC/SSC(forward scatter/sideward scatter) dot-plot,show every peripheral blood cell population.M:Square micron.B:Monocytes were gated in a FSC/SSC dot-plot.CD14+CD16-Mon,CD14-CD16+Mon and CD14+CD16+Mon were defined according to the expression levels of CD14 and CD16 in monocytes.

圖1 流式細(xì)胞術(shù)檢測(cè)單核細(xì)胞亞群

Fig 1 Flow cytometry analysis monocyte subgroups

結(jié) 果

一般資料 本臨床試驗(yàn)共入選70例患者,其中AMI組30例,年齡(65.04±2.48)歲;UAP組25例,年齡(66.41±2.30)歲;有胸痛癥狀而CAG完全正常者(對(duì)照組) 15例,年齡(65.64±1.43)歲。性別、高血壓、糖尿病、血脂、血常規(guī)、冠狀動(dòng)脈病變支數(shù)等資料見(jiàn)表1。

臨床與實(shí)驗(yàn)室檢查AMI組RDW為12.86±0.380,UAP組RDW為12.12±0.410,明顯高于正常對(duì)照組18.22±3.455,差異有統(tǒng)計(jì)學(xué)意義(P<0.05,表1)。AMI組術(shù)后心肌標(biāo)志物(包括cTnT、MYO、CK-MB)均高于其他兩組(P<0.05,表1)。冠脈病變血管支數(shù)(簡(jiǎn)寫為NumberdCV)AMI組(2.077±0.175)和UAP組(1.632±0.205)均高于正常對(duì)照組(P<0.05,表1),而AMI、UAP兩組之間差異無(wú)統(tǒng)計(jì)學(xué)意義。

表1 不同組別患者一般臨床資料與實(shí)驗(yàn)室數(shù)據(jù)的組間比較

(1)P<0.05,AMI groupvs.UAP group;(2)P<0.05,AMI groupvs.control group;(3)P<0.05,UAP groupvs.control group.

單核細(xì)胞亞群水平 AMI組Mon1百分比最高(0.615 5± 0.066 8),與UAP組(0.357 5±0.057 4)和對(duì)照組(0.390 5±0.078 9)比較差異有統(tǒng)計(jì)學(xué)意義(P<0.05,t=2.908,t=2.189,圖2 A )。但UAP組和對(duì)照組兩組間差異無(wú)統(tǒng)計(jì)學(xué)意義。Mon3百分比水平同樣在AMI組中最高,但Mon3在3組間差異尚無(wú)統(tǒng)計(jì)學(xué)意義。AMI組患者M(jìn)on3/Mon1比值(0.677 5±0.205 5)低于UAP組(1.178± 0.247 0)和對(duì)照組(1.599±0.530 9),差異有統(tǒng)計(jì)學(xué)意義(P<0.05,t=2.972,圖 2 B)。結(jié)果提示Mon1比例早期即會(huì)快速上升,Mon3比例也有所上升,但升高幅度低于Mon1。提示斑塊不穩(wěn)定時(shí)Mon1已經(jīng)升高,而Mon3升高尚不明顯。

趨化因子水平 AMI組患者血漿MCP-1水平(130.0±10.14)明顯高于對(duì)照組(99.70±7.552)且差異有統(tǒng)計(jì)學(xué)意義(P<0.05,t=2.401,圖2 C);UAP組MCP-1水平(119.6±7.960)高于對(duì)照組(P=0.085,t=1.751,圖2 C),AMI組UAP組間MCP-1水平差異無(wú)統(tǒng)計(jì)學(xué)意義。AMI組患者血漿FKN水平(0.329 6±0.033 0)高于正常對(duì)照組(0.024 73±0.025 7)(P=0.051 2,t=1.995,圖2 D),UAP組血漿FKN水平(0.368 6±0.0276)明顯高于對(duì)照組(P<0.05,t=3.196,圖2 D)。AMI組FKN水平與UAP組水平相比差異無(wú)統(tǒng)計(jì)學(xué)意義。提示ACS時(shí),Mon1與Mon3所對(duì)應(yīng)的血漿趨化因子水平均會(huì)升高,從而發(fā)揮招募相應(yīng)單核細(xì)胞的功能。

相關(guān)性分析 FKN與Mon3比例、單核細(xì)胞、淋巴細(xì)胞呈正相關(guān)(P<0.05,表2),FKN的受體CX3CR1不僅在Mon3上表達(dá),同時(shí)在淋巴細(xì)胞等PBMC上少量表達(dá),FKN與單核細(xì)胞百分比和淋巴細(xì)胞百分比呈正相關(guān)。FKN同時(shí)與CK-MB和LDL-C呈正相關(guān)(P<0.05,表2)。FKN與RDW和TG呈負(fù)相關(guān),FKN與TG和RDW呈負(fù)相關(guān)(P<0.05,表2)。而Mon1、 Mon3、 MCP-1、GENSINI評(píng)分與各指標(biāo)間未發(fā)現(xiàn)明顯相關(guān)性。

圖2 單核細(xì)胞亞群及其趨化因子在AMI、UAP和Control組間的水平比較

FKN:Fractalkine;Mon3:Monocyte3;M:Monocyte;L:Lymphocyte;CK-MB:Creatine kinase isoenzyme MB;TG:Triglyceride;LDL-C:Low-density lipoprotein-cholesterol;HDL-C:High-density lipoprotein-cholesterol;RDW:Red cell distribution width.

討 論

本研究檢測(cè)了ACS患者CAG術(shù)前單核細(xì)胞亞群含量及比例,及術(shù)前和術(shù)后趨化因子水平,旨在探討MCP-1-Mon1和FKN-Mon3兩條邏輯線在ACS組中相對(duì)CAG正常組的變化及其相關(guān)性。首次證明Mon1在AMI組中所占比例明顯高于UAP組和對(duì)照組,并證明AMI患者M(jìn)on3/Mon1比例降低;還發(fā)現(xiàn)MCP-1在ACS組中增加,提示Mon1通過(guò)MCP-1途徑被募集到AS炎癥部位和心肌梗死部位,促進(jìn)斑塊不穩(wěn)定和清除壞死物質(zhì);同時(shí)發(fā)現(xiàn)FKN在ACS組中均增加,提示FKN表達(dá)增加可通過(guò)募集Mon3,發(fā)揮促斑塊破裂并誘發(fā)ACS的作用。FKN與Mon3呈正相關(guān)(P<0.05,r=0.650 2),驗(yàn)證了其招募關(guān)系。ACS早期以趨化因子增高為主,可能與單核細(xì)胞亞群分化尚且滯后有關(guān)。

動(dòng)物實(shí)驗(yàn)已證實(shí)MCP-1-Mon1和FKN-Mon3在ACS中發(fā)揮作用的靶點(diǎn)主要有2個(gè)微環(huán)境(動(dòng)脈粥樣斑塊中和梗死的心肌組織中)和4個(gè)階段(AS斑塊慢性缺氧階段、AS斑塊炎癥階段、心肌梗死后的炎癥階段、缺血心肌的增殖修復(fù)階段),且其在ACS發(fā)生發(fā)展中的作用隨不同微環(huán)境和不同發(fā)展階段而變化[3]。在AS斑塊慢性炎癥階段和心肌梗死后的炎癥階段,血管內(nèi)皮細(xì)胞和心肌細(xì)胞以表達(dá)MCP-1為主,募集Mon1占優(yōu)勢(shì),發(fā)揮促炎作用。而在AS斑塊慢性缺血缺氧階段和缺血心肌的增殖修復(fù)階段,血管內(nèi)皮細(xì)胞或心肌細(xì)胞由表達(dá)MCP-1轉(zhuǎn)變?yōu)楸磉_(dá)FKN,后者介導(dǎo)Mon3募集到損傷部位,與淋巴細(xì)胞、肥大細(xì)胞等代替早期的Mon1,并通過(guò)分泌IL-10、纖維生長(zhǎng)因子等發(fā)揮抗炎作用,并促進(jìn)血管新生、成纖維細(xì)胞增殖及膠原纖維沉積,心臟組織修復(fù)[7]。但是,斑塊內(nèi)的血管新生可促進(jìn)無(wú)癥狀纖維粥樣斑塊變?yōu)橐灼屏训囊讚p斑塊[3],易增加斑塊破裂的風(fēng)險(xiǎn),MCP-1-Mon1通過(guò)此途徑發(fā)揮促進(jìn)斑塊不穩(wěn)定的不良效果。既往國(guó)外關(guān)于單核細(xì)胞亞群或趨化因子的基礎(chǔ)和臨床研究結(jié)果還未達(dá)成一致結(jié)論[8-12]。有研究證實(shí)敲除兔的MCP-1基因可降低易損斑塊破裂風(fēng)險(xiǎn)[13]。Ikejima等[14]證明FKN表達(dá)上調(diào)可促進(jìn)UAP患者斑塊破裂,這些發(fā)現(xiàn)均與本研究結(jié)果相符。

本臨床試驗(yàn)首次同步研究了ACS早期階段MCP-1-Mon1和FKN-Mon3的表達(dá)特點(diǎn)及相互關(guān)系,對(duì)根據(jù)單核細(xì)胞亞群及其趨化因子的表達(dá)特點(diǎn)早期診斷ACS具有很強(qiáng)的前瞻性意義。MCP-1和FKN招募不同單核細(xì)胞亞群到不同微環(huán)境并發(fā)揮相應(yīng)功能,本實(shí)驗(yàn)檢測(cè)結(jié)果提示人周血單核細(xì)胞亞群及其相應(yīng)趨化因子二者的表達(dá)時(shí)程相一致,并且在ACS組中均有增加趨勢(shì),因而推測(cè)MCP-1-Mon1和FKN-Mon3在ACS中可能發(fā)揮很大的作用。

另外,在AS斑塊慢性缺氧環(huán)境和梗死的心肌組織中,Mon3是通過(guò)FKN-CX3CR1-RhoA信號(hào)[2]通路促進(jìn)微環(huán)境內(nèi)血管新生的[15]。而在心肌梗死后,與Mon1早期即開(kāi)始增加不同,Mon3在心梗發(fā)生2周后才顯著增加[9],并且單核細(xì)胞分化與極化的平衡對(duì)心梗后心臟修復(fù)的影響應(yīng)進(jìn)行進(jìn)一步的基礎(chǔ)和臨床研究[16]。

RDW反映紅細(xì)胞體積的變異程度。凡是可以影響到紅細(xì)胞成熟的病理因素均可導(dǎo)致RDW升高。AS和ACS中的炎性反應(yīng)亦會(huì)進(jìn)一步抑制紅細(xì)胞成熟。由于AS與氧化應(yīng)激、炎性反應(yīng)密切相關(guān)。因此RDW在ACS中升高具有合理性[17]。

本研究病例樣本量偏小,屬于單中心橫斷面研究。今后我們將繼續(xù)增加樣本并進(jìn)一步延長(zhǎng)隨訪時(shí)間,增加觀察時(shí)間點(diǎn),動(dòng)態(tài)評(píng)估MCP-1-Mon1和FKN-Mon3的變化情況及其對(duì)ACS患者預(yù)后的影響。在缺血心肌和動(dòng)脈粥樣硬化斑塊兩種微環(huán)境中,單核細(xì)胞及其趨化系統(tǒng)的表達(dá)時(shí)間和空間的差異需要后期更多的研究和探索。

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Characteristic of peripheral blood monocyte subsets and chemokines in early stage of acute coronary syndrome

XU Miao-miao, SHEN Wei△, SHI Hai-ming, ZHUANG Xin-yu, LIU Xiang-xu,OU Yang, SUN Sheng-jia, WU Bang-wei, ZHU Zhi-dong, CHEN Yu-fei

(DepartmentofCardiovascularMedicine,HuashanHospital,FudanUniversity,Shanghai200040,China)

Objective To investigate the expression of monocyte subsets and their chemokine,i.e.,monocyte chemoattractant protein (MCP-1) and fractalkine (FKN), in patients with acute coronary syndrome (ACS),and to analyze their correlation. Methods Patients with the syndrome of pectoralgia and to be inspected with coronary angiography (CAG) in our hospital from Sep.to Dec.,2016 were included.Patients’ venous blood was collected on the operation day before operation,the level and proportion of monocyte (Mon) subsets,which was namely CD14+CD16-Mon (Mon1),CD14+CD16+Mon (Mon2) and CD14-CD16+Mon (Mon3) according to the expression of cluster differentiation-14 (CD14) and CD16,were detected by flow cytometry (FCM).Patients’ venous blood was collected on the operation day before operation and one day after operation,the concentrations of MCP-1 and FKN in plasma were measured by ELISA.We compared the expression levels of MCP-1-Mon1 and FKN-Mon3,and analyzed their relationship between each other respectively in different groups. Results Diagnosed according to the clinical symptoms,myocardial markers,electrocardiogram and CAG results,70 individuals were analyzed,including 30 patients with acute myocardial infarction (AMI group),25 patients with unstable angina pectoris (UAP group) and 15 patients with the chest pain symptoms and normal CAG results (control group).The percentage of Mon1 in the AMI group was higher than that in the other groups (P<0.05);no difference was observed for Mon3 among the groups (P>0.05).The Mon3/Mon1 ratio in the AMI group was lower than that in the control group (P<0.05).Moreover,the levels of FKN and MCP-1 in the ACS group were greater than those in the control group.The level of red blood cell distribution width (RDW) was significantly increased in the AMI and UAP group than that in the control group (P<0.05).There was a significant correlation between FKN and Mon3 (P<0.05,R=0.650 2). Conclusions The monocyte subset of Mon1 and Mon3 increased in the early stage of ACS,with their chemokine (FKN and MCP-1) increasing at the same time.There is a significant correlation between FKN and Mon3,which indicates MCP-1-Mon1 and FKN-Mon3 may participate in the pathophysiological process of early ACS in patients.

acute myocardium infarction; acute coronary syndromes; monocyte subsets; monocyte chemoattractant protein-1; fractalkine

上海市衛(wèi)計(jì)委中醫(yī)藥科研基金(2014JZ006A);上海市科委科研計(jì)劃項(xiàng)目中醫(yī)類引導(dǎo)項(xiàng)目(15401932000);國(guó)家自然科學(xué)基金面上項(xiàng)目(81673701,81573711)

R541.4

A

10.3969/j.issn.1672-8467.2017.04.002

2017-01-19;編輯:張秀峰)

△Corresponding author E-mail:drshenwei@aliyun.com

*This work was supported by the Traditional Chinese Medicine Research Fund from Shanghai Municipal Commission of Health and Family Planning (2014JZ006A),Science and Technology Program of Traditional Chinese Medicine Guidance Project from Science and Technology Committee of Shanghai (15401932000) and the General Program of National Natural Science Foundation of China (81673701,81573711).