高迁移率族蛋白B1在失血性休克致急性肺损伤中的作用

目的 探讨高迁移率族蛋白 B1(high mobility group box1,HNGB1)在失血性休克(hemarrhag-ic shock,HS)所致急性肺损伤(acute lung injury,ALI)小鼠体内的变化和意义.方法健康雄性BALB/c小鼠42只,随机分为对照组、HS致ALI组和抗HMGB1抗体干预组.心脏穿刺抽血复制小鼠HS致ALI模型,酶联免疫吸附(ELISA)法检测肺组织IL-1β,TNF-α,Westemblot检测肺组织HMGB1,比色法检测肺组织髓过氧化物酶(MPO)、伊文蓝(EBD),观察肺脏病理变化.统计学采用方差分析及LSD-t检验.结果 HS后2 h即可见肺血管通透性增高,肺脏出现弥漫件炎细胞侵润.肺绀织IL-1β和TNF-α在HS诱导的ALI早期(4 h)升高[IL-1β(333.83±31.18)vs.(284.83±30.49),P=0.014;TNF-α(805.00±114.67)vs.(584.17±181.17),P=0.023];肺组织HMCB1在ALI 16～48 h显著升高[HMGB1/β-actin(0.99±0.16)vs.(0.50±0.18),P=0.003].ALI组小鼠肺MPO及EBD水平显著增高,分别于ALI 4 h和24 h达峰值[MPO(38.33±3.88)vs.(8.00±0.89),P<0.01;EBD(20.05±2.79)vs.(4.63±0.43),P<0.01];抗HMGB1抗体干预组肺组织MPO及EBD峰值显著下降[MPO(25.67±1.63)vs.(38.33±3.88),P<0.01;EBD(5.68±0.53)vs.(20.05±2.79),P<0.01],抗体干预组小鼠肺组织病理切片见肺部弥漫性炎细胞浸润减轻,抗体干预24 h组尤为明显.结论 HNGB1是HS所致ALI的晚期炎症介质,HNGBl拮抗治疗能减轻小鼠HS所致ALI病理牛理改变.     

抑制核因子-κB对创伤休克大鼠肝脏热休克蛋白70的影响

目的 探讨抑制核因子-κB(NF-κB)后热休克蛋白70(HSP70)在创伤失血性休克肝组织中的变化及其对肝脏结构和功能的影响.方法 雄性健康Wistar大鼠66只,采用双侧股骨骨折伴失血性休克创伤模型,随机分成正常对照组6只,创伤休克组30只,NF-κB抑制伴创伤休克组30只,NF-κB抑制采用致伤前1 h腹腔注射二硫代氨基甲酸吡咯烷(PDTC)200mg/kg.动态观察伤后0.5、2、4、6、8h大鼠肝组织NF-κB、HSP70、肝脏病理、肝功能、TNF-α、IL-6等变化.NF-κB采用EMSA法测定结合活性,HSP70采用免疫印迹法测定其蛋白含量,并进行计算机图像分析.数据采用SPSS 12.0软件分析,两组间比较采用成组资料的t检验.结果 NF-κB的活性伤后迅速升高,伤后6 h达到高峰;HSP70伤后2 h较正常对照相比[(10.8±1.1)、vs.(4.7±0.5),P＜0.01],伤后6 h达到高峰,和正常组相比[(23.0±1.7)vs.(4.7±0.5),P＜0.01].TNFα、IL-6伤后逐渐升高,并于伤后6 h达到高峰,和正常组相比[TNF-α(173.7±12.1)vs.(30.8±1.8)pg/ml,P＜0.01;IL-6(175.5±12.5)vs.(10.4±0.7)pg/ml,P＜0.01];伤后8 h光镜下可见肝窦内淤血明显,有大量炎性细胞浸润;血清ALT、TB伤后4 h开始增高,8 h达到峰值,和创伤组相比[ALT(640.6±80.2)vs.(536.8±60.0)nmol-1·L-1,P＜0.01;TB(4.7±1.1)vs.(1.6±0.2)mol/L,P＜0.01].抑制NF-κB再致伤后,HSP70在肝组织中表达仍然较高,但在伤后各个时相点的表达均较未抑制NF-κB创伤性休克伤组明显回落;伤后6 h和创伤组相比[(16.9±4.4)vs.(23.0±1.7),P＜0.05].TNF-α、IL-6伤后各个时相点均迅速回落,伤后6 h,和创伤组相比[TNF-α(135.2±10.2)vs.(173.7±12.1)pg/ml,P＜0.05;IL-6(113.0 4±10.8)vs.(175.5±12.5)pg/ml,P＜0.05];肝脏大体淤血、肿胀明显减轻;伤后8 h光镜下可见肝细胞变性明显好转,肝窦内淤血减轻,仅见少许淋巴细胞及中性粒细胞浸润;伤后4 h,血清ALT、TB即明显下降,和未抑制组相比[ALT(540.8±66.2)vs.(640.6±80.2)nmol-1·L-1,P＜0.05;TB(2.3±0.3)vs.(4.7±1.1)mol/L,P＜0.05].结论 NF-κB、HSP70参与了严重创伤失血性休克后肝损伤与抗损伤的发生,抑制NF-κB的活性有助于减轻创伤失血性休克后肝脏的急性损害,NF-κB、HSP70可作为反映创伤休克后肝脏损害程度的重要应激指标.     

胸部开放伤后胸腔海水浸泡致犬急性肺损伤的救治研究

目的 探讨肺保护性通气联合己酮可可碱对胸部开放伤后胸腔内海水致急性肺损伤（ALI）的救治疗效。方法 用胸部开放伤后胸腔内灌注海水制备犬ALI模型。将24条犬随机分为未救治组（A组）、普通救治组（B组）、肺保护性通气组（C组）和肺保护性通气＋己酮可可碱组（D组）。分别于致伤前、致伤6h及救治2h和4h监测各组动物的动脉血气分析、血流动力学、血浆渗透浓度和血钠、血氯的变化；收集外周静脉血和支气管肺泡灌洗液（BALF）检测肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）和IL-8含量。结果 B组救治2h和4h后动脉血氧分压（PaO2）及氧合指数（PaO2／FiO2）仍低于正常水平，C组和D组恢复正常；B、C和D组血流动力学指标以及高渗、高钠、高氯血症在救治2h和4h后较A组显著改善。C组外周血TNF—α及D组外周血TNF—α、IL～8和BALF中IL-6、IL-8水平在救治后显著低于A组和B组；D组外周血TNF-α及BALF中IL-8水平亦显著低于C组。结论 应用肺保护性机械通气治疗胸腔海水致ALI可以起到良好的呼吸支持作用，己酮可可碱能有效的抑制肺内、外炎症反应。     

肿瘤坏死因子-α、白介素-6、丙二醛、超氧化物歧化酶在大鼠腹腔海水浸泡伤中的表达

目的:探讨肿瘤坏死因子-α(tumor necrosis factor-alpha,TNF-α)、白介素6(IL-6)、丙二醛(MDA)及超氧化物歧化酶(SOD)在大鼠腹部开放合并海水浸泡中的表达.方法:采用腹部开放伤合并海水浸泡大鼠致伤模型.在光镜下观察肠黏膜损伤程度;利用酶联免疫吸附测定(ELISA)试剂盒检测血浆TNF-α、IL-6含量的变化;分光光度法检测小肠组织MDA、SOD的含量.结果:腹部开放伤合并海水浸泡后,大鼠小肠病理学检查出现小肠充血、淋巴细胞浸润及肠上皮细胞间紧密连接破坏等改变;血中TNF-α、IL-6水平升高(P＜0.05或P＜0.01):肠组织MDA水平升高,SOD活性降低(P＜0.05或P＜0.01).结论:腹部开放合并海水浸泡可致大鼠肠黏膜屏障功能受损,TNF-α、IL-6、氧自由基参与了腹部开放海水浸泡伤后继发性炎症反应过程.     

急性肺损伤早期肺常规树突状细胞的动态变化

目的 观察急性肺损伤(ALI)早期肺常规树突状细胞(cDCs)数量及成熟程度的动态改变及其对ALI早期炎症反应和肺损伤的影响.方法 C57BL/6小鼠18只随机(随机数字法)分为健康对照组、6 h-ALI组和24 h-ALI组,气管内注射LPS复制ALI模型后6h及24h分别处死小鼠,留取肺组织,采用流式细胞术检测肺cDCs的数量及成熟程度的动态变化;ELISA检测肺组织IL-6、TNF-α水平评价肺部炎症反应;PCR检测肺组织转录因子T-bet及GATA-3mRNA的表达评价Th1/Th2亚群漂移;ELISA检测肺组织IFN-γ、IL-4水平反映Th1/Th2型细胞因子的平衡;计算肺湿质量/体质量比评价肺水肿程度;肺组织HE染色行组织病理学检查及肺损伤评分反映肺损伤程度.组间比较采用单因素方差分析.结果 ALI组小鼠肺IL-6和TNF-α水平、肺湿质量体质量比及肺损伤评分均较健康对照组显著增高(P＜0.01);6 h-ALI组肺cDCs占肺单个核细胞比例(％)较健康对照组升高[(2.25±0.25)vs(0.93±0.40),P＜0.01],其肺cDCs表达MHC Ⅱ的比例(％)也高于健康对照组[(55.51±11.72) vs.(6.67±0.87),P＜0.01];而24 h-ALI组肺cDCs占肺单个核细胞比例(％)较6h-ALI组显著降低[(1.01±0.28)vs.(2.25±0.25),P＜0.01],其肺cDCs表达MHC Ⅱ的比例(％)也低于6h-ALI组[(10.94±2.55) vs.(55.51±11.72),P＜0.01];此外24 h-ALI组肺Th1亚群特异性转录因子T-bet相对表达水平较健康对照组升高[(2.94±0.30)vs(1.00±0.12),P＜0.01],其Th1型细胞因子IFN-γ的水平(Pg·mg-1)也高于健康对照组[(48.17±2.43)vs(16.46±1.68),P＜0.01].结论 ALI早期即存在肺cDCs数量和成熟程度的增加,并可能通过增强Th1型免疫反应及促炎性细胞因子的分泌启动及加剧ALI早期肺部炎症反应.     

海水浸泡对犬弹烧复合伤血浆电解质的影响

目的　研究海水浸泡对弹烧复合伤动物血浆电解质的影响。方法　成功建立海水浸泡弹烧复合伤模型后 ,依模型将 15只成年杂种犬随机分为单纯弹烧复合伤组 (简称非浸泡组 ) 7只和海水浸泡弹烧复合伤组 (简称浸泡组 ) 8只 ,动态监测伤前及伤后 4、7、10、2 0、2 8h共 6个时相点外周动脉血钠、钾、氯、钙及血浆总渗透压的变化。结果　弹烧复合伤后 ,动物血浆电解质及渗透压呈现缓慢上升 ;而海水浸泡后 ,动物血浆电解质及渗透压明显较单纯弹烧复合伤组升高 ,并且贯穿在整个实验过程。结论　海水浸泡能明显加重机体血浆电解质及渗透压的紊乱 ,出现严重的高钠、高氯、高渗血症 ,从而明显加重动物伤情。     

腹部开放伤后海水浸泡大鼠肠屏障功能障碍与细胞因子过度表达的关系

目的:探讨腹部开放伤后海水浸泡大鼠肠屏障功能变化,讨论细胞因子的过度表达及核因子-KB(nuclear factor-kB,NF-kB)的调控作用.方法:观察腹部开放伤后海水浸泡大鼠小肠黏膜组织病理损伤程度;采用酶联免疫吸附测定(ELISA)法检测血肿瘤坏死因子-α(TNF-α)、白介素-6(IL-6)含量;免疫组化技术检测NF-kB在外周血淋巴细胞(PBL)和小肠组织的表达情况.结果:腹部开放伤后海水浸泡大鼠小肠病理组织学检查显示肠黏膜屏障受损;血TNF-α、IL-6水平显著升高:PBL和小肠组织NF-kB表达的阳性率显著升高,与对照组比较差异有统计学意义(P＜0.05或P＜0.01).结论:腹部开放伤合并海水浸泡可致大鼠肠屏障功能障碍,NF-kB活性明显升高,NF-kB活化可上调TNF-α、IL-6的表达,参与腹部开放伤合并海水浸泡后继发性炎症反应过程.     

大蒜素对脓毒症大鼠肠黏膜屏障的保护作用

目的 研究大蒜素对脓毒症大鼠肠黏膜屏障功能的保护作用,并初步探讨其机制.方法 24只雄性SD大鼠随机(随机数字法)分为假手术组、脓毒症模型组、大蒜素治疗组,每组8只.脓毒症模型采用盲肠结扎穿孔(cecum ligation and punctue,CLP),6h及12 h后用大蒜素(30 mg/kg,ip)干预,假手术组及模型组于同时间点给予等量生理盐水.24 h后处死大鼠检测血清D-乳酸、二胺氧化酶(diamine oxidase,DAO)活性、荧光异硫氰酸盐葡聚糖(fluorescence isothiocyanate dextran,FITC-Dextran,FD-40)水平;检测肠组织肿瘤坏死因子[α(tumor necrosis factor alpha,TNF-α)、白介素-6(interleukin-6,IL-6)、丙二醛(malondialdehyde,MDA)水平及超氧化物歧化酶(superoxidedismutase,SOD)活性,并取部分小肠组织进行组织病理学分析.统计方法采用单因素方差分析.结果 与假手术组比较,CLP组大鼠血清D[乳酸、DAO活性及FD40水平明显升高[D-乳酸(nmol/mL):(599.4±101.1) vs.(149.2±20.63),t=11.84,P＜0.01;DAO (ng/mL):(302.1±64.56)vs.(76.57±14.76),t=9.433,P＜0.01;FD-40 (ng/mL):(6664.0±1437.0)vs.(1446.0±205.0),t=9.704,P＜0.01];病理切片示CLP组大鼠肠道形态学损伤明显;肠组织中TNF-α、IL-6、MDA水平明显升高[TNF-α (pg/mL):(186.35±20.43)vs.(58.76±8.94),t=17.23,P＜0.01;IL-6 (pg/mL):(763.25±85.23)vs.(125.36±14.37),t=22.54,P＜0.01;MDA(nmol/mg prot):(29.36±3.27)vs.(7.24 ±0.85),t=16.61,P＜0.01],SOD活性降低[SOD(U/mg prot):(35.75 ±6.53)vs.(73.26±8.35),t=10.57,P＜0.01].大蒜素显著减少脓毒症诱导的血清中D哥酸、DAO活性及FD-40的升高[D-乳酸(nmol/mL):(330.1±81.77)vs.(599.4±101.1),t=7.086,P＜0.01;DAO (ng/mL):(171.8±49.70)vs.(302.1±64.56),t=5.45,P＜0.01;FD-40(ng/mL):(3349.0±1 167.0)vs.(6664.0±1437.0),t=6.165,P＜0.01];病理切片显示大蒜素减轻肠道形态学损伤;大蒜素明显抑制肠组织中TNF-α、IL-6、MDA水平[TNF-α (pg/mL):(95.37±12.68)vs.(186.35±20.43),t=12.29,P＜0.01;IL-6 (pg/mL):(354.27±46.27)vs.(763.25 ±85.23),t=14.45,P＜0.01;MDA (nmol/mgprot):(16.27±3.14)vs.(29.36±3.27),t=9.831,P＜0.01],增加SOD活性[SOD (U/mg prot):(55.35±6.23)vs.(35.75±6.53),t=5.522,P＜0.01].结论 大蒜素对CLP诱导的肠黏膜屏障功能具有保护作用,其机制可能与抑制炎症和氧化应激有关.     

核因子-κB、IκB在创伤失血性休克大鼠肝损伤中的作用

目的 从组织受体水平探讨核因子-κB(NF-κB)、IκB在创伤失血性休克后肝组织中的变化及其在肝损伤中的作用机制.方法 雄性健康Wistar大鼠36只,采用双侧股骨骨折伴失血性休克模型,随机分成正常对照组6只,双侧股骨骨折伴失血性休克组30只.动态观察伤后0.5、2、4、6、8 h大鼠肝组织核因子-κB、IκB、肝脏病理、肝功能、TNF-α、IL-6等变化.肝组织NF-κB采用EMSA法测定结合活性、IκB采用免疫印迹法测定其蛋白含量,并进行计算机图像分析.数据采用SPSS 12.0软件分析,两组间比较采用配对资料的t检验.结果 NF-κB的活性伤后迅速升高,伤后2 h与正常对照相比,差异具有统计学意义,[(8.4±0.7) vs (2.3±0.4),P＜0.01];伤后6 h达到高峰,和正常组相比,P＜0.01[(43.4±4.6) vs (2.3±0.4),P＜0.01].IκB伤后迅速降低,伤后2 h和正常对照相比,差异具有统计学意义[(17.0±2.0) vs (26.4±2.2),P＜0.01].伤后6 h继续下降至比较低的水平,和正常组相比,P＜0.01[(6.5±1.1) vs (26.4±2.2),P＜0.01].TNF-α、IL-6伤后逐渐升高,并于伤后6 h达到高峰:和正常组相比,P＜0.01[TNF-α,(173.7±12.1) vs (30.8±1.8)pg/ml,P＜0.01;IL-6,(175.5±12.5) vs (10.4±0.7)pg/ml,P＜0.01].光镜下伤后4～8 h肝窦内有少许淤血,有散在炎性细胞浸润;血清ALT、TB伤后4 h开始增高,和正常组相比,P＜0.01[ALT,(640.6±80.2) vs (536.8±60.0)nmol·L-1,P＜0.01;TB,(4.7±1.1) vs (1.6±0.2)mol/L,P＜0.01];白蛋白伤后4 h明显下降,和正常组相比,P＜0.01[(13.3±1.6) vs (20.6±3.4)g/L,P＜0.01].结论 NF-κB及其抑制蛋白IκB参与了严重创伤失血性休克后肝损伤的发生;且NF-κB增高越多、IκB减少越明显,肝损害越重.提示NF-κB及其IκB在严重创伤休克后肝组织细胞损伤与抗损伤机制方面起着重要作用.     

腺苷对体外循环下心脏手术患者血浆细胞因子的影响

[目的]探讨腺苷对体外循环心脏手术患者围术期血浆细胞因子的影响.[方法]回顾性分析60例体外循环心脏手术患者的临床资料:A组(33例)为腺苷(cAMP处理组,B组(27例)为等量生理盐水处理组;分别于术前(T0)、主动脉开放10 min(T1)、12 h(T2)、24 h(T3)采集患者动脉血标本,用ELISA法检测围术期血浆TNF-α、IL-8、IL-10、cTnT水平;用自动生化仪测量围术期血浆CK-MB水平.术毕采集右房组织标本,镜下观察肺组织炎症反应.[结果]两组患者术中、后各时点TNF-α、IL-8、IL-10水平均较术前升高(P＜0.05),在T0、T1两组间无明显差异(P＞0.05);在T3、T4两各时点TNF-α、IL-8水平A组较B组明显降低(P＜0.05),而IL-10水平A组较B组明显升高(P＜0.05);术后T2、T3两个时点CK-MB、cTnT水平较术前(T0)明显升高(P＜0.05),但A组较B组明显降低(P＜0.05).镜下观察两组病人均有部分心肌结构破坏,炎症反应,但A组病人较B组轻.[结论]腺苷对缺血再灌注的心肌组织具有一定的保护作用.     

硫化氢对急性肺损伤大鼠白细胞介素的调节

目的 研究气体信号分子硫化氢(H2S)在油酸诱导的大鼠急性肺损伤(ALI)中的作用及其在ALI中对炎症反应的调节.方法 采用鼠尾静脉缓慢注射油酸(OA,0.01 ml/kg)复制大鼠急性肺损伤模型,将SD大鼠49只随机分为三组:对照组、OA组及OA+硫氢化钠(NaHS)组.OA组和OA+NaHS组分为2 h、4 h、6 h三个观察点.对照组鼠尾静脉注射生理盐水(0.01 ml/kg),观察6 h.测定动脉血氧分压(PaO2)、肺湿干重比(W/D)、肺泡灌洗液中性粒细胞百分比(PMN%),光镜下半定量肺损伤评分(IQA),用敏感硫电极法检测血浆、肺组织匀浆H2S含量,用双抗体夹心ELISA法测定血浆、肺组织匀浆IL-6、IL-8和IL-10的含量.多组样本的均数的比较采用单因素差分析.结果 油酸鼠尾静脉注射可引起肺组织明显的形态学改变;与对照组比,OA组可见PaO2下降,W/D、PMN%、IQA增加(P＜0.01),在2 h、4 h、6 h时,血浆中H2S[对照:(36.58±6.80)μmol/L,2 h:(21.30±2.75)μmol/L,4 h:(20.63±1.26)μmol/L,6 h:(20.00±1.60)μmol/L,P＜0.01]以及肺匀浆中H2S[对照:(27.61±2.20)μmol/L,2 h:(20.67±1.37)μmol/L,4 h:(20.79±1.10)μmol/L,6 h:(18.92±0.75)μmol/L,P=0.000]均明显降低,血浆中IL-6[对照:(73.95±14.68)pg/ml,2 h:(186.70±23.85)pg/ml,4 h:(238.50±26.46)pg/ml,6 h:(215.95±25.86)pg/ml,P＜0.01]、肺匀浆中IL-6[对照:(60.58±12.91)pg/ml,2 h:(160.32±24.57)pg/ml,4 h:(195.27±46.28)pg/ml,6 h:(185.66±17.42)pg/ml,P＜0.01]均显著增加,血浆IL-8[对照:(80.69±20.42)pg/ml,2 h:(184.11±19.51)pg/ml,4 h:(286.20±53.34)pg/ml,6 h:(241.30±45.85)pg/ml,P＜0.01]、肺匀浆中IL-8[对照:(69.14±15.96)pg/ml,2 h:(174.10±20.36)pg/ml,4 h:(249.02±31.17)pg/ml,6 h:(237.74±34.18)pg/ml,P＜0.01]均显著增加,血浆IL-10[对照:(39.78±8.97)pg/ml,2 h:(111.18±11.46)pg/ml,4 h:(115.60±13.91)pg/ml,6 h: (102.41±9.93)pg/ml,P＜0.01]、肺匀浆中IL-10[对照:(71.86±14.19)pg/ml,2 h:(126.96±18.72)pg/ml,4 h:(151.88±27.61)pg/ml,6 h:(137.28±14.22)pg/ml,P＜0.01]含量均显著增加.而预先给予NaHS能减轻油酸引起的肺损伤,显著提高血浆和肺匀浆中H2S含量[4 h,血浆:(26.67±3.44)μmol/L vs(20.63±1.26)μmol/L,P=0.042;肺匀浆:(23.20±1.48)μmol/L vs(20.79±1.10)μmol/L,P=0.011;6 h,血浆:(26.98±4.93)μmol/L vs(20.00±1.60)μmol/L,P=0.022;肺匀浆:(21.43±1.79)μmol/L vs(18.92±0.75)μmol/L,P=0.016),同时血浆IL-6(185.37±21.98 pg/ml vs 238.50±26.46 pg/ml,4 h,P＜0.01;(124.22±21.84)pg/ml vs(215.95±25.86)pg/ml,6 h,P＜0.01 ]、IL-8[(199.40±34.56)pg/ml vs(286.20±53.34)pg/ml,4h,P＜0.01;(146.58±20.23)pg/ml vs(241.30±45.85)pg/ml,6 h,P＜0.01]含量明显降低,而IL-10含量增高[(154.48±18.08)pg/ml vs(115.60±13.91)pg/ml,4 h,P=0.000;(138.06±20.01)pg/ml vs(102.41±9.93)pg/ml,6 h,P=0.1300].结论 油酸诱导的大鼠ALI内源性H2S生成减少.外源性H2S通过抑制炎症因子IL-6和IL-8的表达,增加抗炎因子IL-10的表达,进而改变炎症/抗炎因子之间的比例,对油酸诱导的大鼠急性肺损伤起保护作用.     

Protective effects of aerobic exercise on acute lung injury induced by LPS in mice

Introduction

The regular practice of physical exercise has been associated with beneficial effects on various pulmonary conditions. We investigated the mechanisms involved in the protective effect of exercise in a model of lipopolysaccharide (LPS)-induced acute lung injury (ALI).

Methods

Mice were divided into four groups: Control (CTR), Exercise (Exe), LPS, and Exercise + LPS (Exe + LPS). Exercised mice were trained using low intensity daily exercise for five weeks. LPS and Exe + LPS mice received 200 µg of LPS intratracheally 48 hours after the last physical test. We measured exhaled nitric oxide (eNO); respiratory mechanics; neutrophil density in lung tissue; protein leakage; bronchoalveolar lavage fluid (BALF) cell counts; cytokine levels in BALF, plasma and lung tissue; antioxidant activity in lung tissue; and tissue expression of glucocorticoid receptors (Gre).

Results

LPS instillation resulted in increased eNO, neutrophils in BALF and tissue, pulmonary resistance and elastance, protein leakage, TNF-alpha in lung tissue, plasma levels of IL-6 and IL-10, and IL-1beta, IL-6 and KC levels in BALF compared to CTR (P ≤0.02). Aerobic exercise resulted in decreases in eNO levels, neutrophil density and TNF-alpha expression in lung tissue, pulmonary resistance and elastance, and increased the levels of IL-6, IL-10, superoxide dismutase (SOD-2) and Gre in lung tissue and IL-1beta in BALF compared to the LPS group (P ≤0.04).

Conclusions

Aerobic exercise plays important roles in protecting the lungs from the inflammatory effects of LPS-induced ALI. The effects of exercise are mainly mediated by the expression of anti-inflammatory cytokines and antioxidants, suggesting that exercise can modulate the inflammatory-anti-inflammatory and the oxidative-antioxidative balance in the early phase of ALI.     

高浓度氧对未成年大鼠肺部炎症反应的影响

目的 探讨高浓度氧对未成年大鼠肺部炎症反应的影响.方法 将40只出生21 d的SD大鼠按随机数字表法分为空气对照组及高氧暴露12、24、48、72 h组,每组8只,分别将大鼠置于空气和常压高氧箱(氧含量达92%～94%)中.于相应时间点采用放血法处死大鼠后取肺组织,并行支气管肺泡灌洗.采用硫代巴比妥酸法和比色法分别测定肺组织丙二醛(MDA)含量及髓过氧化物酶(MPO)活性;采用酶联免疫吸附法(ELISA)检测支气管肺泡灌洗液(BALF)中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)和IL-10含量;观察肺组织病理改变,并进行肺损伤评分.结果 与空气对照组比较,高氧暴露12 h肺组织MDA含量(mmol/g)即显著升高(2.24±0.43比1.57±0.31),MPO活性(U/g)于高氧暴露24 h显著升高(1.24±0.25比0.69±0.22),并均随高氧暴露时间延长逐渐增加(P<0.05或P<0.01).BALF中TNF-α、IL-6和IL-10含量于高氧暴露24 h时较空气对照组显著增加[TNF-α(ng/L):135.2±44.0比94.5±22.3,IL-6(ng/L):73.1±14.2比55.7±17.3,IL-10(ng/L):67.9±21.7比48.2±7.6,P<0.05或P<0.01];但高氧暴露48 h时较24 h时显著降低(48 h时BALF中TNF-α、IL-6、IL-10分别为105.4±17.0,54.3±17.4,50.9±6.9,均P<0.05).高氧暴露12 h时肺损伤评分(分)即较空气对照组显著升高(4.5±1.4比1.3±0.5),并随高氧暴露时间延长进一步升高(P<0.05或P<0.01).结论 高浓度氧可引起未成年大鼠肺部炎症损伤;炎症细胞因子的出现高峰均在高氧暴露24 h.     

大承气汤对急性呼吸窘迫综合征大鼠肺泡巨噬细胞NF-κB活性及其炎性细胞因子表达的影响

目的 研究大承气汤(Dachengqi Decoction,DD)对ARDS大鼠肺泡巨噬细胞NF-κB活性及其炎性细胞因子表达影响,探讨大承气汤抗炎作用的分子机制.方法雄性Wistar大鼠65只,随机分为对照组(n=12)、ARDS模型组(n=21)、大承气汤治疗组(n=16)、地塞米松治疗组(n=16).对照组用0,5mL生理盐水替代大肠杆菌脂多糖(LPS)尾静脉缓慢注射;模型组LPS按1 mg/kg·0.5mL)腹腔注射,16 h后按5 mg/(kg·0.5 mL)经气管缓慢滴入,6 h后动脉血气分析验证造模成功,连续观察3 d;大承气汤治疗组造模成功后DD按生药2.31 g/(kg·d)连续灌胃3 d;地塞米松治疗组造模成功后开始腹腔内注射地塞米松2mg/,kg,连续治疗3 d.各组72 h后行动脉血气分析、病理学检验和评分观察疗效;运用酶联免疫吸附法(enzyme linked immunosorbent assay,ELLSA)检测血浆和支气管肺泡灌洗液中肿瘤坏死因子-α(TNF-α)、白细胞介素-1(IL-1)和IL-10水平,评价全身和肺部促炎因子/抗炎因子平衡状态;BCA法测定PAN核蛋白浓度,蛋白免疫印迹法(western Blotting)检测肺泡巨噬细胞(pul-monary alveolar macrophage,PAM)核因子-κB(nucleus factor-κH;NF-κB)活性,评价叻对PAM炎性细胞因子转录活性影响.所有试验数据均用采用SPSS 13.0统计分析软件进行处理,计量指标以均数±标准差(x±s)表示,组间比较采用方差分析(LDS-t检验),以P<0.05为差异具有统计学意义.结果 ①DD降低ARDS大鼠血浆TNF-α作用不显著[(510.97±76.20)pg/mL,(476.16±98.03)pg/mL,P>0.05],降低IL-1作用显著[(381.99±34.30)pg/mL,(300.69±50.99)pg/mL,P<0.05],但对IL-10水平无影响[(345.30±78.52)pg/mL,(345.916±67.72)pg/mL,P>0.05];DD显著降低ARDS大鼠BALF中TNF-α水平(130.94±33.51 pg/mL,(106.59±26.64)pg/mL,P<0.05)和IL-1水平[(82.5±25.36)pg/mL,(63.89±22.96)pg/mL,P<0.05],使IL-10水平显著升高[(77.09±26.05)pg/mL,(148.05±53.50)pg/mL,P<0.01].DD显著降低ARDS大鼠PAM核蛋白水平[(5.35±2.44)μg/μL(3.54± 2.01)μg/μL,P<0.05],对NF-κB活性(电泳条带光密度值×面积/参照物比值)具显著抑制作用[(1.45±0.71),(1.11±0.28),P<0.05].结论DD主要通过降低IL-1水平来调节ARDS大鼠全身促炎介质/抗炎介质平衡;对局部肺损伤的调节作用主要通过抑制前炎症介质(TNF-α)的生成,同时又显著降低IL-1水平和升高IL-10水平重新建立局部促炎因子/抗炎因子平衡.DD可以抑制ARDS大鼠PAM中NF-κB活性,从而抑制促炎介质(TNF-α,IL-1)的产生.DD这种多靶点双向调节作用在调节免疫平衡,保护靶器官免受过度损伤发挥积极作用.     

骨髓间充质干细胞对烟雾吸入性损伤兔早期炎症因子分泌的影响

目的 探讨骨髓间充质干细胞(MSCs)对烟雾吸入性损伤早期外周血及肺组织中肿瘤坏死因子-α(TNF-α)、白细胞介素(IL-1β、IL-6、IL-10)分泌的影响.方法 全骨髓培养法体外培养兔MSCs,用流式细胞术鉴定.将56只健康新西兰大耳白兔按随机数字表法分为正常对照组(C组,n=8)、烟雾吸入性损伤组(S组,n=24)、烟雾吸入性损伤+MSCs移植组(M组,n=24),后两组再分为伤后2、4、6 h亚组,每组8只.采用酶联免疫吸附法(ELISA)检测血浆及肺组织匀浆液中促炎因子TNF-α、IL-1β、IL-6及抗炎因子IL-10的含量.结果 与C组比较,S组各时间点血浆促炎、抗炎因子均显著升高;各时间点肺组织促炎因子显著升高,抗炎因子无明显变化.与S组比较,M组各时间点血浆促炎因子显著下降,抗炎因子显著升高[6 h时TNF-α(μg/L):1.7±1.7比4.1±1.6,IL-1β(ng/L):9.9±1.7比21.2±2.6,IL-6(μg/L):1.0±0.3比1.3±0.2,IL-10(ng/L):15.2±4.4比7.9±3.5,均P＜0.05];各时间点肺组织促炎因子显著降低,而抗炎因子仅在4 h、6 h显著升高[6 h时TNF-α(ng/L):503.0±156.4比587.7±171.2,IL-1β(ng/L):0.4±0.2比0.6±0.2,IL-6(ng/L):155.2±13.7比350.2±20.3,IL-10(ng/L):23.3±5.4比11.0±5.6,均P＜0.05].结论 MSCs移植能降低烟雾吸入性损伤早期促炎因子水平,升高抗炎因子水平,改善全身炎症反应,对烟雾吸入性损伤肺组织具有一定的保护作用.

Abstract：

Objective To explore the effect of bone marrow mesenchymal stem cells (MSCs) engraftment on secretion of tumor necrosis factor-α (TNF-α), interleukins (IL-1β, IL-6, IL-10) in peripheral blood and lung homogenates in the early stages of smoke inhalation injury. Methods MSCs were proliferated by the method of whole marrow culture and identified by flow cytometry. Fifty-six healthy New Zealand rabbits were randomly divided into control group (C group, n=8), smoke inhalation injury group (S group, n=24)and smoke inhalation injury+MSCs engraftment group (M group, n=24). The latter two groups were subdivided into 2, 4, 6 hours after injury subgroups, with 8 rabbits in each group. The levels of TNF-α,IL-1β, IL-6 and IL-10 in peripheral blood and lung homogenates were measured by enzyme-linked immunosorbent assay (ELISA). Results Compared with C group, concent of pro-inflammatory and anti-inflammatory cytokines in peripheral blood at each time point in S group were increased significantly.The concent of pro-inflammatory cytokines in lung homogenate at each time point in S group was significantly higher than thoae in C group, and that of anti-inflammatory cytokines showed no significant changes.Compared with the S group, concent of pro-inflammatory cytokines in peripheral blood in M group was decreased significantly, and that of anti-inflammatory cytokines was increased significantly [6 hours TNF-α(μg/L):1.7±1.7 vs. 4.1±1.6, IL-1β (ng/L): 9.9±1.7 vs. 21.2±2.6, IL-6 (μg/L): 1.0±0.3 vs.1.3 ± 0. 2, IL-10 (ng/L): 15. 2 ± 4. 4 vs. 7. 9 ± 3.5, all P＜0.05]. Concent of pro-inflammatory cytokines at each time point in M group was decreased significantly when compared with S group in lung homogenate,while only anti-inflammatory cytokine at 4 hours and 6 hours was increased significantly [6 hours TNF-α (ng/L): 503. 0±156. 4 vs. 587.7±171.2, IL-1β (ng/L): 0.4±0.2 vs. 0.6±0.2, IL-6 (ng/L): 155.2±13.7 vs. 350.2±20.3, IL-10 (ng/L): 23.3±5.4 vs. 11.0±5.6, all P＜0.05]. Conclusion MSCs engraftment could decrease pro-inflammatory cytokines and increase anti-inflammatory cytokines in the early stages of smoke inhalation injury, thus amelioratea inflammatory reaponse, which confers protective effect on smoke inhalation injury.     

血红素加氧酶-1表达对大鼠呼吸机相关性肺损伤的作用及机制研究

目的 观察呼吸机相关性肺损伤(VILI)模型大鼠肺组织中血红素加氧酶-1(HO-1)表达的变化,探讨HO-1诱导剂血晶素拮抗VILI的作用机制.方法 56只雄性SD大鼠按照随机数字表法分成对照组(C组),VILI模型组(M组),诱导剂血晶素1、2、3、4组(H1、 H2、H3、H4组,制模前24 h分别腹腔注射血晶素40、80、120、160 μmol/kg)和抑制剂Z组[制模前24 h腹腔注射锌原卟啉(ZnPP)10 μmol/kg].除C组外各组机械通气4 h后处死大鼠,收集支气管肺泡灌洗液(BALF),测定总蛋白、肿瘤坏死因子-α(TNF-α)和白细胞介素-10(IL-10)含量;取肺组织,测定肺湿/干重(W/D)比值、乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)和丙二醛(MDA)水平及HO-1蛋白表达;光镜下行肺组织病理观察.结果 与C组相比,M组大鼠肺病理损伤严重,BALF中总蛋白、TNF-α、IL-10,肺组织W/D比值、MDA、LDH及HO-1蛋白表达均明显增加, VILI模型复制成功.与M组比较,随着血晶素剂量的增加,H1、H2、H3组总蛋白(g/L)显著下降(0.74±0.06、0.73±0.07、0.70±0.07比0.84±0.08,均P<0.01);W/D比值下降(4.93±0.27、4.91±0.24、4.87±0.23比5.53±0.48,均P<0.01);SOD活性(U/mg)显著升高(85±9、82±15、93±11比55±12,均P<0.01);MDA含量(nmol/mg)显著降低(15±3、15±3、13±2比18±4,P<0.05或P<0.01);IL-10含量(pg/L)逐渐升高(0.42±0.06、0.46±0.06、0.47±0.05比0.36±0.07),TNF-α含量(pg/L)逐渐降低(0.18±0.07、0.14±0.03、0.10±0.07比0.23±0.06),但只有H2、H3组差异有统计学意义(均P<0.01);LDH活性(U/g)降低(11 353±1 317、11 516±1 613、9 631±1 520比12 361±1 841),但仅H3组差异有统计学意义(P<0.01);HO-1蛋白表达[吸光度(A)值]逐渐增强(0.164±0.010、0.190±0.149、0.205±0.018比0.122±0.016,均P<0.01);肺病理损伤逐渐减轻.而随着剂量进一步增加,H4组肺组织损伤较H1、H2、H3组加重.给予HO-1抑制剂ZnPP后HO-1的保护作用消失.结论 血晶素诱导HO-1适度表达可以减轻VILI,其适度表达的最佳剂量为120 μmol/kg,其机制可能通过抗炎和抗氧化应激发挥对肺组织的保护作用.     

Elevated expression of inducible nitric oxide synthase and inflammatory cytokines in the alveolar macrophages after esophagectomy

OBJECTIVE: To evaluate the role of inducible nitric oxide (NO) synthase (iNOS) and inflammatory cytokines in alveolar macrophages (AMs) after esophagectomy in the pathogenesis of acute lung injury. DESIGN: Prospective, exploratory, open-labeled clinical study. SETTING: Intensive care unit and operating room in a university hospital. PATIENTS: Thirteen patients receiving esophagectomy with carcinoma of the esophagus (postesophagectomy group), ten patients just before the surgery (preoperation group), and seven patents receiving surgery less invasive than esophagectomy (other surgery group) were selected. INTERVENTIONS: Bronchoalveolar lavage fluid (BALF) and blood samples were obtained from study groups. MEASUREMENTS AND MAIN RESULTS: The AMs in the BALF collected from each group were stained immunohistochemically with antibodies against iNOS, interleukin (IL)-6, and IL-8. The intensities of these expressions were determined by semiquantitative cytofluorometric system. NOx (NO2- + NO3-), IL-6, and IL-8 levels in the BALF and plasma were measured concurrently. The expressional intensities of iNOS, IL-6, and IL-8 in AMs obtained from the postesophagectomy group were maximal 24 hrs after the skin incision and significantly more evident than those from other groups. The IL-6, IL-8, and NOx levels in BALF and IL-6 and IL-8 levels in plasma in the postesophagectomy patients were also elevated. The intensities of iNOS and inflammatory cytokines expressions in AMs were closely related to postoperative respiratory failure. CONCLUSIONS: The activation of topical alveolar macrophages may be involved in the pathogenesis of pulmonary complications in the postoperative period after esophagectomy.     

胸部爆震伤致兔急性呼吸窘迫综合征模型的建立及相关因素分析

目的 构建室内胸部爆震伤致兔急性呼吸窘迫综合征(ARDS)模型并分析其发生机制及早期死亡原因,为研究肺爆震伤早期预警体系和治疗方法提供依据.方法 按照不同炸药量和致伤距离所产生的压强,将60只新西兰大白兔按随机数字表法分为5个致伤组和1个无致伤对照组.伤后观察存活率和组织病理学,并监测病理生理学指标、肺含水量.结果 冲击波压强低于1 210.5 mm Hg(1 mm Hg=0.133 kPa,A、B组)时,肺损伤较轻,表现为点状肺挫伤,肺简明损伤评定分级法(ALS)均在2级内,动物伤后24 h内全部恢复,长期存活无并发症.冲击波压强高于2 036.1 mm Hg(D、E组)时,肺损伤过重,表现为广泛的肺挫伤、肺门撕裂伤和肺内大血肿,AIS均大于5级,动物于伤后1 h内全部死亡.冲击波压强为1 917.3 mm Hg(C组)时,肺表现为广泛而恒定的挫伤,累及4个肺叶以上,AIS 4～5级,伤后6 h内出现动脉氧分压下降;肺组织可见肺泡壁水肿,部分肺泡壁断裂,肺泡融合;肺泡内充满大量炎性细胞,偶见透明膜形成.与对照组比较,C组兔致伤6 h肺湿/干重比值即显著升高(6.46±0.24比3.98±0.19,P＜0.01),血浆及支气管肺泡灌洗液(BALF)中肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)即明显升高[血浆TNF-α(ng/L):328.89±6.26比62.12±2.98,BALF TNF-α(ng/L):164.87±4.59比29.51±1.12;血浆IL-6(ng/L):128.51±4.13比19.32±1.53,BALF IL-6(ng/L):94.97±1.14比22.72±0.19,均P＜0.05].结论 在1 917.3 mm Hg爆炸压强的密闭环境下,冲击伤可诱导兔发生ARDS;TNF-α及IL-6参与爆震伤致ARDS的形成与发展;特定环境下,肺脏破裂致气胸为早期死亡原因,而冲击波致循环系统功能紊乱也是引起早期死亡的重要原因.

Abstract：

Objective To reproduce acute respiratory distress syndrome (ARDS) model in rabbit induced by chest blast injury and to analyze the pathogenesis and causes of early death in order to provide the basis for the early diagnosis of lung blast injury and its early-warning system to facilitate an early treatment.Methods Sixty healthy New Zealand white rabbits were divided into six groups according to the different explosion distance with the random number table method. The survival rate and its resulting pathological changes were observed and patho-physiological indexes and lung fluid content were determined at sequential time points post-explosion. Results Shock wave pressure less than 1 210. 5 mm Hg (1 mm Hg=0. 133 kPa,group A, B) resulted in limited injury to the lung within grade-2 as assessed with the abbreviated injury scale (AIS). The rabbits in these groups recovered soon and survived without any complication. Shock pressure higher than 2 036. 1 mm Hg (group D, E) caused severe injuries to the lung, including deep laceration, disruption of lung hilus and large hematoma in the lung, and the injury severity of lungs was assessed above grade-5 as assessed with AIS. All rabbits died within 1 hour post-explosion. The groups described above failed to meet the demand of an ARDS model for the present study. Shock wave pressure at 1 917. 3 mm Hg (group C) produced extensive contusion from grade-4 to grade-5 as assessed with AIS. The rabbits survived in poor general condition, and arterial partial pressure of oxygen (PaO2) lowered within 6 hours. Pathological examination showed extensive and constant multi-focal bleeding involving more than four lobes. The alveolar wall was edematous, with partial rupture and alveolar fusion in lung tissues was observed in the group C. Alveoli were filled with inflammatory cells, and hyaline membrane was formed occasionally. Compared with control group, the wet to dry weight ratio (W/D) in lungs increased obviously (6.46±0. 24 vs. 3. 98±0. 19, P＜ 0. 01) in group C within 6 hours postinjury. The contents of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in plasma and bronchoalveolar lavage fluid (BALF) were also increased distinctly compared with the control group [TNF-α (ng/L) in plasma: 328. 89± 6.26 vs.62.12±2. 98, TNF-α (ng/L) in BALF: 164.87±4.59 vs. 29. 51±1.12; IL-6 (ng/L) in plasma: 128. 51±4.13 vs. 19.32±1.53: IL-6 (ng/L) in BALF: 94.97±1.14 vs. 22.72±0. 19, all P＜0. 05]. Conclusion In an airtight environment, rabbit ARDS model can be reproduced successfully by blast injury with 1 917.3 mm Hg explosion pressure; TNF-α and IL-6 are involved in the pathogenesis and development of ARDS in blast injury. Pneumothorax as a result of lung rupture is the chief reason for early death and dysfunction of circulatory system is also an important reason in producing early death.     

急性肺损伤肺组织和血浆NO、ET-1变化的研究

目的 探讨急性肺损伤（ＡＬＩ）时肺组织和血浆一氧化氮（ＮＯ）、皮内素－１（ＥＴ－１）和白细胞介素８（ＩＬ－８）水平的变化和作用。方法 采用间隔２４ｈ两次注射大肠杆菌内毒素（ＬＰＳ）的方法,复制家兔内毒素急性肺损伤模型。结果 ＡＬＩ组浆、肺组织均浆及支气管肺泡灌洗液（ＢＡＬＦ）中ＮＯ、ＥＴ－１和ＩＬ－８水平显著升高（Ｐ〈０．０１）,ＢＡＬＦ内中性料细胞（ＰＭＮ）明显增多,肺系数（ＬＰＬ）、肺水含量