細(xì)菌脂蛋白耐受對(duì)膿毒癥老齡及成年小鼠心臟功能保護(hù)作用的對(duì)比

王 磊，周 靜，周蘇明

?

細(xì)菌脂蛋白耐受對(duì)膿毒癥老齡及成年小鼠心臟功能保護(hù)作用的對(duì)比

王 磊，周 靜，周蘇明*

（南京醫(yī)科大學(xué)第一附屬醫(yī)院老年重癥監(jiān)護(hù)室，南京 210029）

探討細(xì)菌脂蛋白（BLP）耐受對(duì)老齡小鼠心肌細(xì)胞的保護(hù)作用，并比較其對(duì)老齡小鼠與成年小鼠心功能影響的差異。選取86只健康雄性C57BL/6老齡小鼠（SPF級(jí)，24月齡）和55只成年雄性C57BL/6小鼠（6～8周齡），用小劑量BLP預(yù)處理老齡小鼠及成年小鼠誘導(dǎo)BLP耐受，分別比較正常對(duì)照組、假手術(shù)（sham）組、膿毒癥（剖腹手術(shù)行盲腸結(jié)扎穿孔致膿毒癥，CLP）組、BLP耐受＋CLP組，其中成年小鼠在盲腸結(jié)扎穿孔0，2，6，12h四個(gè)時(shí)間點(diǎn)做小鼠二維超聲心動(dòng)圖，老齡小鼠在0，3，6，12h四個(gè)時(shí)間點(diǎn)做小鼠二維超聲心動(dòng)圖，選擇左心室短軸縮短率（FS）、射血分?jǐn)?shù)（EF）、左心室舒張末期內(nèi)徑（LVIDd）等做為觀測(cè)指標(biāo)。老齡小鼠CLP組與BLP耐受＋CLP組的FS、EF、LVIDd自手術(shù)后均呈下降趨勢(shì)，但是BLP耐受＋CLP組的降幅較CLP組小。成年小鼠BLP耐受＋CLP組0～6h時(shí)段FS、EF、LVIDd的變化趨勢(shì)與CLP組相同，但各時(shí)間點(diǎn)數(shù)值均低于CLP組；而6h后BLP耐受＋CLP組的3項(xiàng)指標(biāo)轉(zhuǎn)而上升，至12h時(shí)高于CLP組，其中FS、EF差異均有統(tǒng)計(jì)學(xué)意義（＜0.05）。BLP耐受＋CLP組老齡小鼠的FS呈現(xiàn)出進(jìn)行性下降趨勢(shì)，而在成年小鼠中6h后為進(jìn)行性增高趨勢(shì)，12h明顯高于假手術(shù)組和CLP組（＜0.05）。成年小鼠CLP組在術(shù)后EF上升，6h后出現(xiàn)下降的趨勢(shì)，12h低于術(shù)前水平；而老齡小鼠CLP組術(shù)后EF進(jìn)行性下降。成年小鼠BLP耐受＋CLP組術(shù)后出現(xiàn)的EF值呈逐漸上升趨勢(shì)，相反，老齡組則進(jìn)行性下降。老齡小鼠BLP耐受+CLP組的LVIDd術(shù)后進(jìn)行性下降，而成年小鼠BLP耐受＋CLP組術(shù)后2h時(shí)低于術(shù)前水平（＜0.05），6h后逐漸回升。膿毒癥對(duì)FS、EF、LVIDd有抑制作用，老齡小鼠心臟收縮功能和舒張功能所受抑制更加嚴(yán)重；BLP耐受對(duì)心功能有一定的保護(hù)作用，此作用在成年小鼠較明顯，在老齡小鼠未見明顯保護(hù)作用。

休克，膿毒性；細(xì)菌脂蛋白耐受；心功能；保護(hù)

膿毒癥是感染引起的全身炎癥反應(yīng)綜合征，具有高患病率和死亡率，心臟是易受損傷的靶器官，近50%膿毒癥患者出現(xiàn)不同程度心肌抑制。膿毒癥的主要病因是細(xì)菌感染過程中其胞壁成分，如細(xì)菌脂蛋白（bacterial lipoprotein，BLP）、內(nèi)毒素脂多糖（lipopolysaccharide，LPS）等激活宿主炎癥細(xì)胞而導(dǎo)致炎癥細(xì)胞呈高度活化狀態(tài)，炎癥細(xì)胞因子被過度合成釋放，引起失控的全身性炎癥反應(yīng)。BLP不但可活化宿主炎癥免疫細(xì)胞，誘導(dǎo)炎癥細(xì)胞因子釋放，導(dǎo)致休克甚至死亡，而且可以誘導(dǎo)自我耐受[1]。目前有文獻(xiàn)報(bào)道BLP耐受對(duì)膿毒癥小鼠的心功能有保護(hù)作用[2]。本文將通過體外實(shí)驗(yàn)探討老齡小鼠與成年小鼠對(duì)BLP耐受的心功能差異。

1 材料與方法

1.1 實(shí)驗(yàn)動(dòng)物和分組

86只健康雄性C57BL/6老齡小鼠，SPF級(jí)，24月齡，體質(zhì)量26～41g；55只成年雄性C57BL/6小鼠，6～8周齡，體質(zhì)量20～25g（均上海斯萊克實(shí)驗(yàn)動(dòng)物有限公司）。飼養(yǎng)于清潔級(jí)小鼠飼養(yǎng)室，環(huán)境溫度18～25℃，相對(duì)濕度45%～55%。適應(yīng)性喂養(yǎng)1周后，均隨機(jī)分為正常對(duì)照（control）組，假手術(shù)（sham）組，剖腹手術(shù)行盲腸結(jié)扎穿孔致膿毒癥（cecal ligation and puncture，CLP）組，BLP耐受＋CLP組（BLP＋CLP組）。

1.2 動(dòng)物模型

術(shù)前禁食12h，自由飲水，使用10%水合氯醛對(duì)實(shí)驗(yàn)小鼠進(jìn)行腹腔麻醉，胸、腹部皮膚去毛，常規(guī)消毒腹部皮膚，取下腹正中行長約1.5cm切口，找到盲腸，游離腸系膜。假手術(shù)組盲腸不結(jié)扎不穿孔，將盲腸還納腹腔，0號(hào)線逐層縫合肌肉和皮膚。CLP組輕輕牽出盲腸，尋找盲腸與回腸和結(jié)腸交界處，用3-0號(hào)絲線環(huán)行結(jié)扎盲腸根部，再用18G針頭穿刺盲腸2個(gè)孔，從孔中擠出糞便少許，然后將盲腸送回腹腔，逐層關(guān)腹。BLP＋CLP組是按10mg/kg劑量，予小鼠腹腔內(nèi)注射BLP，24h后制造盲腸結(jié)扎穿孔的膿毒癥模型。術(shù)后均立即給小鼠皮下注射林格液（Ringer’s solution）25ml/kg抗休克，并且讓其隨意進(jìn)食進(jìn)水。以小鼠造模后出現(xiàn)精神萎靡、腹脹，進(jìn)食、進(jìn)水及活動(dòng)減少，開腹后有血性及膿性滲出、盲腸粘連腫脹、腸脹氣等表現(xiàn)作為造模成功標(biāo)志。

1.3 主要試劑與儀器

BLP（Pam3CSK4·3HCl）（美國Alexis公司）。配置方法：加雙蒸水，超聲振蕩混勻，分別制成1g/L和0.1g/L兩種稀釋液，-40℃保存。小鼠超聲心動(dòng)圖儀（美國GE公司）Vivid 7 Dimension。

1.4 觀察指標(biāo)

選用Vivid 7 Dimension彩色超聲診斷儀，線陣探頭頻率為13MHz。小鼠于前述麻醉劑半量腹腔注射，麻醉后取仰臥位，剃須刀剃去胸部毛，再用脫毛劑脫去剩余皮毛。將超聲心動(dòng)圖探頭置于小鼠左胸前，探頭示標(biāo)向左側(cè)并與胸骨中線保持70°～80°，于左室乳頭肌水平短軸切面測(cè)量。分別在0h、造模后2，6，12h 4個(gè)時(shí)間點(diǎn)對(duì)成年小鼠做二維超聲心動(dòng)圖，在0h、造模后3，6，12h 4個(gè)時(shí)間點(diǎn)對(duì)老齡小鼠做二維超聲心動(dòng)圖，選擇左心室短軸縮短率（fractional shortening，F(xiàn)S）、射血分?jǐn)?shù)（ejection fraction，EF）、左心室舒張末期內(nèi)徑（left ventricular internal dimension at end-diastole，LVIDd）等做為觀測(cè)指標(biāo)，每組原始數(shù)據(jù)取連續(xù)3個(gè)心動(dòng)周期的平均值，保留圖像用于脫機(jī)分析。

1.5 統(tǒng)計(jì)學(xué)處理

2 結(jié) 果

2.1 老齡小鼠與成年小鼠的左心室FS

老齡小鼠對(duì)照組與BLP＋CLP組在0h相比，差異無統(tǒng)計(jì)學(xué)意義（＞0.05）；6h時(shí)BLP＋CLP組與CLP相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05），CLP組與假手術(shù)組相比，差異亦有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h時(shí)BLP＋CLP組及CLP組與假手術(shù)組相比，差異均有統(tǒng)計(jì)學(xué)意義（＜0.05；表1）。

成年小鼠0h BLP＋CLP組與相同時(shí)間點(diǎn)正常對(duì)照組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；6h及12h BLP＋CLP組與相同時(shí)間點(diǎn)假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h BLP＋CLP組與相同時(shí)間點(diǎn)CLP組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05），12h CLP組與假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05；表2）。

2.2 老齡小鼠與成年小鼠的EF變化

老齡小鼠0h各組數(shù)據(jù)相比，差異無統(tǒng)計(jì)學(xué)意義（＞0.05）；3h各組數(shù)據(jù)相比，差異無統(tǒng)計(jì)學(xué)意義（＞0.05）；6h時(shí)BLP耐受＋CLP組與CLP組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05），CLP組與假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h時(shí)BLP耐受＋CLP組與假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05），CLP組與假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05；表3）。

成年小鼠0h時(shí)BLP+CLP組與相同時(shí)間點(diǎn)正常對(duì)照組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；6h及12h時(shí)BLP＋CLP組與相同時(shí)間點(diǎn)假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h時(shí)BLP＋CLP組與相同時(shí)間點(diǎn)CLP組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h時(shí)CLP組與相同時(shí)間點(diǎn)假手術(shù)組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05；表4）。

2.3 老齡小鼠與成年小鼠的LVIDd變化

老齡小鼠6h時(shí)CLP組與正常對(duì)照組相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；12h時(shí)BLP＋CLP組及CLP組與正常對(duì)照組相比，差異均有統(tǒng)計(jì)學(xué)意義（＜0.05），BLP＋CLP組及CLP組與假手術(shù)組相比，差異均有統(tǒng)計(jì)學(xué)意義（＜0.05；表5）。

成年小鼠0h時(shí)BLP＋CLP組與相同時(shí)間點(diǎn)正常對(duì)照組相比，差異無統(tǒng)計(jì)學(xué)意義（＞0.05）；2，6及12h時(shí)BLP＋CLP組與假手術(shù)組對(duì)比，差異均有統(tǒng)計(jì)學(xué)意義（＜0.05）；6，12h時(shí)CLP組與假手術(shù)組對(duì)比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05）；另外2，6h時(shí)BLP＋CLP組與CLP組相比，差異亦有統(tǒng)計(jì)學(xué)意義（＜0.05；表6）。

表1 不同處理后老齡小鼠左心室FS的變化

FS: fractional shortening; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with sham group,*＜0.05; compared with CLP group,#＜0.05

表2 不同處理后成年小鼠左心室FS的變化

FS: fractional shortening; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with control group,*＜0.05; compared with sham group,#＜0.05; compared with CLP group,△＜0.05

表3 不同處理后老齡小鼠EF的變化

EF: ejection fraction; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with sham group,*＜0.05; compared with CLP group,#＜0.05

表4 不同處理后成年小鼠EF的變化

EF: ejection fraction; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with control group,*＜0.05; compared with sham group,#＜0.05; compared with CLP group,△＜0.05

表5 不同處理后老齡小鼠LVIDd

LVIDd: left ventricular internal dimension at end-diastole; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with control group,*＜0.05; compared with sham group,#＜0.05

表6 不同處理后成年小鼠LVIDd

LVIDd: left ventricular internal dimension at end-diastole; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with sham group,*＜0.05; compared with CLP group,#＜0.05

2.4 老齡小鼠和成年小鼠在EF、FS及LVIDd的比較

BLP＋CLP組老齡小鼠的FS呈現(xiàn)出進(jìn)行性下降趨勢(shì)，其中BLP＋CLP組下降幅度較之CLP組??；而在成年小鼠中BLP＋CLP組6h后為進(jìn)行性增高趨勢(shì)，12h明顯高于CLP組，差異有統(tǒng)計(jì)學(xué)意義（＜0.05；圖1）。

成年小鼠CLP組在術(shù)后EF上升，6h后出現(xiàn)下降的趨勢(shì)，12h低于術(shù)前水平；而老齡小鼠CLP組，術(shù)后EF為進(jìn)行性下降趨勢(shì)。另外，成年小鼠BLP＋CLP組術(shù)后出現(xiàn)的EF值呈逐漸上升趨勢(shì)，相反，老齡組則進(jìn)行性下降，無代償性增加（圖2）。

老齡小鼠BLP＋CLP組的LVIDd在術(shù)后出現(xiàn)進(jìn)行性下降，術(shù)后3h時(shí)較0h相比，差異有統(tǒng)計(jì)學(xué)意義（＜0.05），而成年小鼠BLP＋CLP組術(shù)后2h時(shí)低于術(shù)前水平（＜0.05），6h后逐漸回升（圖3）。

2.5 小鼠腹腔解剖所見

假手術(shù)組小鼠腹腔無明顯炎癥改變。CLP組小鼠腹腔內(nèi)可見濃性或血性滲出液，結(jié)扎端盲腸腫脹，與周圍組織粘連，形成盲腸周圍膿腫，腸管擴(kuò)張。BLP＋CLP組小鼠腹腔亦可見膿性或血性滲出液，腸管明顯擴(kuò)張、水腫及充血，結(jié)扎端盲腸腫脹明顯，與周圍組織粘連，肝充血腫大。

3 討 論

膿毒癥是病原菌引起的全身性炎癥反應(yīng)[3]，合并感染性休克時(shí)病死率＞25%，是目前重癥監(jiān)護(hù)病房（intensive care unit，ICU）患者死亡的重要原因[4]。而疾病的進(jìn)展是由于機(jī)體自身免疫系統(tǒng)過度激活，導(dǎo)致炎癥通路的激活及級(jí)聯(lián)放大的瀑布效應(yīng)，從而導(dǎo)致全身炎癥反應(yīng)綜合征及多器官功能不全綜合征[5]，其中大約有40%的患者會(huì)發(fā)生心肌功能障礙[6]。1951年Waisbren最早報(bào)道了膿毒癥患者出現(xiàn)心功能損害，如心臟擴(kuò)大、LVEF下降、左室收縮峰壓/左室舒張末期壓下降、對(duì)容量負(fù)荷收縮反應(yīng)差等表現(xiàn)。

BLP是革蘭陽性和革蘭陰性細(xì)菌外膜中最豐富的蛋白，其特征是在蛋白的N端含有獨(dú)特的脂?；?氨基酸結(jié)構(gòu)。BLP可由生長或裂解的細(xì)菌釋放，通過抑制細(xì)胞因子等炎癥介質(zhì)的釋放來減輕炎癥反應(yīng)，使其在人體免疫中發(fā)揮重要作用。機(jī)體對(duì)細(xì)菌的胞壁成分有耐受現(xiàn)象，即機(jī)體接觸過的某些細(xì)菌胞壁成分后，當(dāng)再次接觸大劑量的相同成分或細(xì)菌時(shí)，會(huì)出現(xiàn)暫時(shí)性敏感性下降的現(xiàn)象，使致炎因子產(chǎn)生減少，表現(xiàn)為免疫耐受。BLP耐受的重要特征是耐受細(xì)胞中的核因子?κB（nuclear factor-κB，NF-κB）活化受到抑制，從而使腫瘤壞死因子?α（tumor necrosis factor-α，TNF-α）、白細(xì)胞介素（interleukin，IL）-1、IL-6等NF-κB依賴基因的表達(dá)明顯減少[7,8]。有實(shí)驗(yàn)表明BLP耐受可使小鼠抵抗活細(xì)菌或盲腸結(jié)扎穿孔造成的膿毒癥而免于死亡[2,9]。此外，有臨床資料表明，膿毒癥患者脂蛋白濃度明顯降低與死亡率增加密切相關(guān)，以及與入住ICU患者炎癥加重相關(guān)[10]。

圖1 老齡小鼠和成年小鼠FS的比較

Figure 1 The comparison of FS between the aged and adult mice FS: fractional shortening; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with CLP/Adult group,*＜0.05

圖2 老齡小鼠和成年小鼠EF的比較

Figure 2 The comparison of EF between the aged and adult mice EF: ejection fraction; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein

圖3 老齡小鼠和成年小鼠LVIDd的比較

Figure 3 The comparison of LVIDd between the aged and adult mice LVIDd: left ventricular internal dimension at end-diastole; CLP: cecal ligation and puncture; BLP: bacterial lipoprotein. Compared with 0h in BLP+CLP/Aged group,*＜0.05; compared with 0h in BLP+CLP/Adult group,#＜0.05

本研究采用老齡及成年小鼠CLP模型，誘發(fā)廣泛的全身性炎性反應(yīng)，表現(xiàn)出與臨床相似的早期高動(dòng)力循環(huán)、高代謝和晚期低動(dòng)力循環(huán)狀態(tài)。心肌功能不全是膿毒癥常見的并發(fā)癥，對(duì)膿毒癥的預(yù)后有著重要影響。本研究就老齡及成年小鼠膿毒癥時(shí)心功能的變化進(jìn)行了研究，結(jié)果顯示，老齡小鼠CLP組與BLP＋CLP組的FS、EF、LVIDd自手術(shù)后均呈下降趨勢(shì)，但是BLP＋CLP組的降幅較CLP組小。成年小鼠BLP＋CLP組0～6h時(shí)段FS、EF、LVIDd的變化趨勢(shì)與CLP組相同，各時(shí)間點(diǎn)的數(shù)值均低于CLP組。統(tǒng)計(jì)學(xué)分析顯示，0h時(shí)BLP＋CLP組的FS、EF的值均明顯低于正常對(duì)照組（＜0.05），說明小劑量BLP注射入體內(nèi)后24h，引起的輕度炎癥反應(yīng)對(duì)心臟功能有抑制作用；而6h后BLP＋CLP組的3項(xiàng)指標(biāo)轉(zhuǎn)而上升，至12h時(shí)高于CLP組，其中FS、EF均有統(tǒng)計(jì)學(xué)差異（＜0.05），說明BLP耐受對(duì)心功能有保護(hù)作用。

膿毒癥時(shí)循環(huán)功能的改變是雙向性的，初始為高動(dòng)力循環(huán)狀態(tài)，隨著膿毒癥的痊愈或進(jìn)展，高動(dòng)力循環(huán)狀態(tài)趨向正常或轉(zhuǎn)入低動(dòng)力循環(huán)狀態(tài)甚至導(dǎo)致死亡。成年小鼠中CLP組及BLP＋CLP組術(shù)后早期EF值均高于正常，后CLP組出現(xiàn)下降而BLP＋CLP組持續(xù)升高，而在老齡小鼠則表現(xiàn)為進(jìn)行性下降。相比于成年小鼠，老齡小鼠的心肌收縮及舒張功能下降，原因可能如下。（1）老齡小鼠心功能低下，隨著年齡的增加，心功能及其儲(chǔ)備能力下降，老齡小鼠心力衰竭的心室重塑較成年更重，目前機(jī)制尚不清；但膠原重建在心室重塑過程中起了重要的作用[11]，其中Ⅰ/Ⅲ型膠原比值過度升高，使得心室僵硬度過度增加，從而使得心功能變差。（2）機(jī)體調(diào)節(jié)對(duì)外來刺激產(chǎn)生炎癥反應(yīng)的能力出現(xiàn)了隨年齡變化，隨著年齡的增加，機(jī)體對(duì)于炎癥反應(yīng)分泌TNF-α、IL-1β、IL-6等炎癥因子的能力增強(qiáng)[12]。Vollmar等[13]發(fā)現(xiàn)內(nèi)毒素血癥模型的老齡大鼠其外周血TNF-α、IL-1β、IL-6，水平較成年大鼠高2～14倍。Marik等[14]對(duì)于NORASEPT Ⅱ臨床試驗(yàn)的結(jié)果分析發(fā)現(xiàn)，高齡膿毒癥患者外周血TNF-α明顯高于其他年齡組患者。TNF-α、IL-6作為多器官功能衰竭的中心介質(zhì)，其能夠引起心肌延展性和收縮峰值速率的下降，且兩者具有協(xié)同作用[15]。有報(bào)道IL-1β和TNF-α表達(dá)一致，說明其可能存在協(xié)同作用[16]。

上述結(jié)果表明，膿毒癥對(duì)FS、EF、LVIDd有抑制作用，BLP耐受對(duì)心功能有一定的保護(hù)作用；而在成年小鼠中此保護(hù)作用較老齡小鼠明顯。老齡小鼠因其心功能儲(chǔ)備低下，及其在膿毒癥時(shí)所釋放的致炎因子增多，其BLP耐受所帶來的心功能保護(hù)作用有限。

[1] Sato S, Nomura F, Kawai T,. Synergy and cross-tolerance between Toll-like receptor (TLR) 2- and TLR4-mediated signaling pathways[J]. J Immunol, 2000, 165(12): 7096?7101.

[2] Wang JH, Doyle M, Manning BJ,. Cutting edge: bacterial lipoprotein induces endotoxin-independent tolerance to septic shock[J]. J Immunol, 2003, 170(1): 14?18.

[3] Miyake Y, Yasunaka T, Ikeda F,. SIRS score reflects clinical features of non-acetaminophen-related acute liver failure with hepatic coma[J]. Intern Med, 2012, 51(8): 823?828.

[4] Dellinger RP, Levy MM, Carlet JM,. Surviving Sepsis Campaign: International Guidelines for Management of Severe Sepsis and Septic Shock: 2008[J]. Intensive Care Med, 2008, 34(1): 17?60.

[5] Monneret G, Payen D. Immunosuppression in sepsis: a novel understanding of the disorder and a new therapeutic approach[J]. Lancet Infect Dis, 2013, 13(3): 260?268.

[6] Fernandes CJ Jr, de Assuncao MS. Myocardial dysfunction in sepsis: a large, unsolved puzzle[J]. Crit Care Res Pract, 2012, 2012: 896430.

[7] Buckley JM, Wang JH, Redmond HP. Cellular reprogramming by Gram-positive bacterial components: a review[J]. J Leukoc Biol, 2006, 80(4): 731?741.

[8] Coldewey SM, Rogazzo M, Collino M,. Inhibition of IκB kinase reduces the multiple organ dysfunction caused by sepsis in the mouse[J]. Dis Model Mech, 2013, 6(4): 1031?1042.

[9] O’Brien GC, Wang JH, Redmond HP. Bacterial lipoprotein induces resistance to Gram-negative sepsis in TLR4-deficient miceenhanced bacterial clearance[J]. J Immunol, 2005, 174(2): 1020?1026.

[10] Van Leeuwen HJ, Heezius EC, Dallinga GM,. Lipoprotein metabolism in patients with severe sepsis[J]. Crit Care Med, 2003, 31(5): 1359?1366.

[11] Van den Borne SW, Diez J, Blankesteijn WM,. Myocardial remodeling after infarction: the role of myofibroblasts[J]. Nat Rev Cardiol, 2010, 7(1): 30?37.

[12] Inoue S, Sato T, Suzuki-Utsunomiya K,. Sepsis-induced hypercytokinemia and lymphocyte apoptosis in aging-accelerated Klotho knockout mice[J]. Shock, 2013, 39(3): 311?316.

[13] Vollmar B, Pradarutti S, Nickels RM,. Age-associated loss of immunomodulatory protection by granulocyte-colony stimulating factor in endotoxic rats[J]. Shock, 2002, 18(4): 348?354.

[14] Marik PE, Zaloga GP, NORASEPTⅡ Study Investigators,. The effect of aging on circulating levels of proinflammatory cytokines during septic shock. NORASEPTⅡ Study Investigators[J]. J Am Geriatr Soc, 2001, 49(1): 5?9.

[15] Vincent JL. International Sepsis Forum. Hemodynamic support in septic shock[J]. Intensive Care Med, 2001, 27 (Suppl 1): S80?S92.

[16] Wang J, Qian W, Zhu Q,. Martentoxin, a large-conductance Ca(2+)-activated K(+) channel inhibitor, attenuated TNF-α-induced nitric oxide release by human umbilical vein endothelial cells[J]. J Biomed Res, 2013, 27(5): 386?393.

（編輯: 李菁竹）

Comparison of cardiac protection of bacterial lipoprotein tolerance between aged and adult sepsis mice

WANG Lei, ZHOU Jing, ZHOU Su-Ming*

(Geriatric Intensive Care Unit, the First Affiliated Hospital, Nanjing Medical University, Nanjing 210029, China)

To determine the protective effect of bacterial lipoprotein (BLP) tolerance on myocardiocytes in aged mice, and compare its effect on cardiac function between aged and adult mice.A total of 86 healthy male aged C57BL/6 mice (SPF grade, 24 months old) and another 55 healthy male adult C57BL/6 mice (SPF grade, 6 to 8 weeks old) were employed in this study. Low-dose BLP was used to pretreat the aged and adult mice to induce BLP tolerance. Then the mice were respectively divided into control, sham operation group (sham), sepsis group (CLP, inflicted by cecal ligation and puncture) and BLP tolerance＋CLP group. Echocardiography was carried out in adult mice in 0, 2, 6 and 12h after CLP to measure left ventricle shortening fraction (FS), ejection fraction (EF), left ventricular internal diameter at end-diastole (LVIDd) in adult mice, and in aged mice at time points of 0, 3, 6 and 12h for above-mentioned indices.The FS, EF and LVIDd showed a downward trend after operation in the sepsis group and BLP tolerance＋CLP group in aged mice, with the stronger decreases in the former than in the latter group. In the time from 0 to 6h, the BLP tolerance＋CLP group in adult mice had similar tendencies in the FS, EF and LVIDd with sepsis group, but lower than the later. Then the 3 indices were increased in the BLP tolerance＋CLP group in 6h after CLP, and became higher than those of sepsis group in 12h, with FS and EF having significant differences (＜0.05). The FS in the BLP tolerance＋CLP aged mice presented a progressive decline tendency, but it was in a progressive increase trend after 6 h in adult mice, and was significantly higher than those of the sham and CLP groups at 12h (＜0.05). The EF of CLP group in adult mice increased after operation, then decreased from 6h and became lower than the preoperative level at 12h. While the EF of CLP group in aged mice showed a progressive decline tendency after surgery. The EF of BLP tolerance＋CLP group in adult mice was in a progressive increase trend; on the contrary, the corresponding group in aged mice was tending downwards. The LVIDd of the BLP tolerance＋CLP group in aged mice was progressively reduced, while that of corresponding group in adult mice was below the preoperative level in 2h after operation (＜0.05), then gradually recovered in 6h.Sepsis has inhibitory effects on FS, EF and LVIDd, and the effects on cardiac systolic and diastolic functions are stronger in aged mice. BLP tolerance exerts protective effect on cardiac function. This protection is apparently effective in adult mice than in aged mice.

shock, septic; bacterial lipoprotein tolerance; cardiac function; protection

R631.4

A

10.11915/j.issn.1671-5403.2015.04.064

2015?02?09;

2015?03?30

周蘇明, E-mail: zhousmco@yahoo.com.cn