热休克蛋白27改善小鼠内毒素血症心功能不全的机制

目的 探讨Hsp27的保护作用是否与激活P13K/Akt信号通路、减轻炎症反应有关.方法 (1)诱导小鼠内毒素血症心肌特异性高表达Hsp27转基因鼠(Hsp 27 Tg)和野生型对照鼠(WT)均腹腔注射内毒素(LPS,10ms/ks);(2)心功能测定 LPS注射后6 h,以心脏超声评价,n:6;(3)P13K/Akt信号通路活性测定 LPS注射后1 h收集心脏,以Western blot法测定Akt及Gsk-3(的磷酸化水平,n=4;(4)NF-B炎症通路活性检测 LPS注射后1 h以Western blot测定IκBα水平,n=4;同时在大鼠心肌细胞中进行相同实验,n:3;(5)心肌细胞凋亡LPS注射后24 h,以TUNEL法检测,,n=4.结果 (1)Hsp27显著改善LPS所致的心功能不全.与基础值相比,LPS处理使WT,Hsp27 Tg均出现心功能不全,但Hsp 27 Tg的心功能不全程度显著轻于WT(P<0.01或P<0.05).(2)Hsp27显著抑制LPS所致的IκBα降解.与WT比较,高表达Hsp27显著减轻了LPS诱导的小鼠心肌IκBα降解[(41.43±24.10)%vs.(72.92±9.20)%,P<0.05],培养的大鼠心肌细胞实验结果与之类似.(3)Hsp27可增强激活P13K/Akt信号通路.LPS处理后,WT和Hsp27Tg鼠心肌组织中磷酸化Akt水平分别为(3.11±0.83)和(5.13±0.73),磷酸化GSK-30水平分别为(3.19±1.04)和(5.71±1.20).与WT比较,Hsp27Tg心肌组织中的磷酸化Akt和GSK-3β水平均显著提高(P<0.05).类似结果在体外培养的细胞实验中也被证实.(4)Hsp27显著抑制LPS所致的心肌细胞凋亡.LPS处理后24 h,WT和Hsp27Tg心肌细胞凋亡率分别为(6.46±1.74)%和(2.88±0.91)%,与WT比较,心肌细胞凋亡在Hsp27Tg中被显著减轻(P<0.01).结论 高表达Hsp27对小鼠内毒素血症心功能不全有显著改善作用,其保护作用与激活P13K/Akt信号通路、抑制NFκB;依赖性炎症反应有关.     

内毒素预处理对内毒素血症大鼠肝损伤的保护作用

目的 探讨内毒素预处理对内毒素血症大鼠肝损伤的保护作用.方法 采用直接注射内毒素的方法建市大鼠急性内毒素血症模型.雄性Wistar人鼠72只,随机分成三组:生理盐水对照组(N组,n=24只)、内素脂多糖(IPS)(L组,n=24只)和LPS预处理组(P组,n=24只),每组义按时间分为2 h组、4 h组、6 h组、12 h组4个亚组,每个亚组6只.P组:首次经腹腔注射LPS 0.25mg/kg;24 h后再经腹腔注射IPSO.5 mg/kg,其余两组给予等容量生理盐水.第二次腹腔注射72 h后,L组和P组经尾静脉一次注射LPS 10 mg/kg,N组给予等量生理盐水,L组和P组在注射LPS后2,4,6,12 h,N组在注射最后一次NS后2,4,6,12 h,各取6只取肝组织制成匀浆检测Toll样受体4(Toll likereceptor-4,TLR-4)、核因子-кB(nuclear factor kappa B,NF-кB)、肿瘤坏死因子-α(tulnor necrosis factor-a,TNF-α)和丙二醛(malondiahtehyde,MDA),抽血检测谷丙转氨酶(ALT),谷草转氨酶(ASF),取肝脏(肝左上叶)用10%的中性甲醛固定.采用SPSS 13.0统汁软件进行单因素方差分析.结果 内毒素血症时符时间点肝脏组织TLR-4,NF-кB和TNF-α浓度较对照组显著升高,而内毒素预处理后则有明显下降,其中内毒素血症时4 h组肝脏组织TLR-4,NF-кB和TNF-α分别为(38.76±0.67),(170.82±31.40),(293.16±49.49)和(6.263±0.351),显著高于对照组(P＜0.05),而内毒素预处理后上述指标降至(22.32±1.35),(135.55±26.44)和(234.23±44.96),差异有统计学意义(P＜0.05).结论 内毒素预处理可减轻内毒素血症时的肝损伤,其机理可能与肝组织TLR-4,NF-кB和TNF-α的表达减少有关.     

血管再生在小鼠脓毒症心肌损伤中的作用

目的 对脓毒症小鼠发生心肌损伤时心肌血管再生和细胞凋亡情况及相关机制的研究.方法 将40只8周左右的雄性C57BL/6小鼠随机(随机数字法)分为两组,实验组(n=20)和对照组(n=20),实验组用脂多糖(LPS)腹腔注射(10 mg/kg),对照组给以生理盐水(10mg/kg),6h后超声观察两组小鼠心功能(n=40),后取两组小鼠心脏、肺脏、肾脏组织石蜡包埋切片做HE染色(n=6)观察病理学变化鉴定模型,免疫组化(n=3)PECAM -1、α-SAM染色观察心肌血管再生,心肌细胞凋亡染色(TUNLE) (n=3)观察细胞凋亡情况,提取心脏组织RNA(n=6)通过RT-PCR技术对HIF-1α等血管再生因子进行检测,所得数据处理均采用独立样本t检验.结果 实验组与对照组相比小鼠心脏左室舒张期前壁厚度(t=-4.60,P＜0.05)、左室收缩期前壁厚度增厚(t=-3.24,P＜0.05),左心室舒张期内径减小(t=3.57,P＜0.01),每搏输出量下降(t=5.51,P＜0.01),免疫组化染α-SAM抗体显示实验组心脏新生血管数增加(t=-11.00,P＜0.01),凋亡染色(TUNEL)存在心肌细胞凋亡[实验组比对照组:(191.31±5.41) vs.(52.24±4.32)],RT-PCR显示HIF-1α表达显著增高(t=-8.12,P＜0.05).结论 脓毒症小鼠存在明显心肌血管再生、细胞凋亡及心功能障碍,导致血管再生的通路有低氧诱导因子( HIF -1α)的参与.     

应激性心肌损伤发病机制的实验研究

目的 探讨应激性心肌损伤的发病机制.方法 30只雄性Wistar大鼠按随机数字表法分成正常对照组、制动后冰泳应激组(每日制动6h,第13日开始制动后冰泳5 min)、内毒素组[腹腔注射脂多糖(LPS)10 mg/kg]3组,每组10只.取心肌组织,光镜下观察心肌组织病理变化;电镜下观察心肌超微结构;用酶联免疫吸附试验(ELISA)检测血清心肌肌钙蛋白Ⅰ(cTnI)含量;用原位末端缺刻标记法(TUNEL)检测心肌细胞凋亡并计算凋亡指数;用免疫组化法检测心肌组织天冬氨酸特异性半胱氨酸蛋白酶(caspase-8和caspase-3)的表达;并分析caspase与细胞凋亡指数的相关性.结果 与正常对照组相比,制动冰泳应激组和内毒素应激组在光镜和电镜下均有不同程度的心肌细胞损伤表现,血清cTnI含量(μg/L)均明显增加(0.63±0.12、0.74±0.08比0.53±0.03,P＜0.05和P＜0.01);心肌细胞凋亡指数有不同程度增加[(7.91±1.71 )％、(12.94±2.00)％比0],心肌组织中caspase-8和caspase-3表达增加(caspase-8灰度值:126.65±3.13、114.82±8.67比156.99±9.66; caspase-3灰度值:130.20±2.96、108.58±5.72比160.51±5.25,均P＜0.01),且内毒素应激组各指标均明显高于制动冰泳应激组(P＜0.05或P＜0.01).相关分析显示,制动冰泳应激组中caspase-8、caspase-3与细胞凋亡指数呈显著正相关(r=0.914,P1=0.002;r2=0.929,P2=0.001);内毒素应激组中caspase-8、caspase-3与细胞凋亡指数呈显著正相关(r1=0.956,P1 =0.000;r2=0.916,P2=0.001).结论 应激可能通过增加caspase-8和caspase-3蛋白的表达诱导大鼠心肌细胞凋亡,从而导致心肌损伤.     

内毒素耐受大鼠肺部iNOS和NO表达的动态变化和意义

目的 研究内毒素血症时内毒素耐受大鼠肺部诱导型一氧化氮合酶(iNOS)、一氧化氮(NO)的动态变化.方法 72只SD大鼠随机分为对照组(NC组)和内毒素耐受组(ET组).腹腔注射小剂量内毒素(LPS),第1天0.1 mg/只;第2～5天0.5 mg/只,建立内毒素耐受模型.NC组腹腔注射同等剂量生理盐水.最后一次注射LPS 72 h后,两组均腹腔注射大剂量LPS(10 mg/kg).各组按注射LPS前(0 h),注射后2、4、6、12、24 h分为六小组(n=6).通过比色法(即Griess法)测量肺组织iNOS、NO、丙二醛(MDA)及髓过氧化物酶(MPO)含量.同时在第6、12 h取大鼠左下肺组织行组织病理形态学观察.用SPSS13.0统计软件进行统计分析.结果 NC组大鼠在大剂量LPS刺激后肺组织iNOS和NO在4 h开始升高,6～12 h达到高峰(P＜0.05);MDA和MPO含量在2 h即开始升高,4～12 h达到高峰(P＜0.05).ET组大鼠在大剂量LPS刺激前肺组织iNOS、NO、MDA和MPO即有微量升高(P＞0.05),大剂量LPS刺激后肺组织iNOS、NO、MDA和MPO亦有升高,但与NC组相比明显降低(P＜0.05).病理学检查显示,ET组肺组织损伤明显较NC组减轻.结论 内毒素耐受时,大剂量内毒素血症引起的大鼠肺部损伤减轻与肺部iNOS、NO低表达有关.     

核因子-κB促使水肿型胰腺炎向坏死型胰腺炎转化

目的 探讨核因子-κB(NF-κB)在肠源性内毒素血症促使急性水肿型胰腺炎(AEP)向急性坏死型胰腺炎(ANP)转化中的作用。方法 C57BL／6小鼠随机分5组：正常组(n=5)、脂多糖(LPS)组(n=20)、AEP组(n=20)、AEP LPS组(n=20)、PDTC干预组(AEP LPS PDTC)组(n=20)。腹腔内间隔1h共7次注射雨蛙素(50ug／kg)诱导小鼠AEP。第6h胃管内灌入LPS溶液(5mg／kg)。PDTC干预组于雨蛙素注射前腹腔内注射PDTC，剂量为100mg／kg。监测12h、24h、48h和5d血清淀粉酶、乳酸脱氢酶(LDH)活性；免疫组化观察胰组织Mac-1(CD11b／CD18)、E选择素、P选择素和ICAM-1表达；Southern印迹检测胰组织TNF-a、IL-1βmRNA变化。结果 LPS单独并不引起胰组织明显病理变化和血清淀粉酶、LDH活性改变，但使小鼠AEP胰腺组织损伤加重，血清淀粉酶和LDH活性显著增加；胰腺组织Mac-1表达增强；PDTC处理组血清淀粉酶和LDH活性显著降低，胰腺组织病理改善，细胞因子TNF-a、IL1βmRNA表达下调。结论 肠源性内毒素血症可促使AEP转化为ANP。PDTC通过选择性抑制NF-KB阻断内毒素信号通路，抑制内毒素介导的高细胞因子反应及其继发的中性粒细胞活化，最终使胰腺炎程度减轻。提示NF-kB在肠源性内毒素血症促使AEP转化为ANP发展的过程中起着重要的介导作用。     

内毒素血症幼鼠小肠黏膜组织学及血浆、肠组织二胺氧化酶、血浆D-乳酸的变化

目的 观察内毒素血症幼鼠小肠黏膜组织学及血浆、肠组织二胺氧化酶(DAO)、血浆D-乳酸的变化.方法 18日龄wistar大鼠48只,随机分为内毒素血症组(n=40),正常对照组(n=8),内毒素血症组根据注射脂多糖(LPS)后取标本时间分为1.5 h、6 h、24 h、72 h、7 d共5个亚组,各组于各时间点分别取血浆及小肠匀浆,测定血浆DAO活性及D-乳酸、小肠匀浆DAO值.结果 (1)内毒素血症组呈小肠黏膜损伤的组织学改变;(2)内毒素血症1.5 h、6 h、24 h、72 h亚组血浆DAO较正常对照组增高(P<0.05);而7 d亚组较正常对照组偏低;(3)内毒素血症各亚组小肠组织DAO低于正常对照组DAO;(4)内毒素血症组血浆D-乳酸较正常组增高(P<0.05);(5)血浆DAO与小肠组织DAO含量呈负相关(r=-0.392,P=0.006).结论 腹腔注射LPS可以引起小肠上皮细胞受损,紧密连接破坏,肠通透性增加,提示临床上可以通过测定血浆DAO及D-乳酸来及时了解肠道屏障功能.     

内毒素血症小鼠骨骼Toll样受体4基因的变化

目的 通过检测内毒素血症模型动物骨骼相关基因表达变化,进一步明确内毒素对机体的损害机制,为下一步研究骨骼变化对脏器产生何种影响奠定基础.方法 腹腔注射内毒素脂多糖(LPS)制作内毒素血症小鼠模型.将48只小鼠随机分为正常组和LPS处理4、6、8、12、24、48和72 h组,每组6只.采尾血计数全血白细胞;取眼眶血检测肝、肾功能;用逆转录-聚合酶链反应(RT-PCR)检测骨骼中TLR4 mRNA表达;苏木素-伊红(HE)染色,镜下观察骨骼病理学变化.结果 与正常组比较,LPS 4 h组白细胞计数显著下降(P＜0.01),之后逐渐升高,于72 h时显著高于正常组(P＜0.05);LPS 4 h和6 h组丙氨酸转氨酶(ALT)水平显著升高(P均＜0.05),于8 h降至正常水平(P＞0.05);LPS 6 h组血尿素氮(BUN)水平逐渐升高(P＜0.05),于8 h达峰值(P＜0.05),然后下降,12 h趋于正常(P＞0.05);LPS 6 h组TLR4 mRNA表达增加(P＜0.01),24 h达峰值(P＜0.01),然后逐渐下降,72 h仍处在较高水平(P＜0.05).LPS作用于骨骼后,HE染色显示骨骼无明显改变.结论 内毒素血症中骨骼TLR4 mRNA表达量显著增加.     

降钙素基因相关肽对内毒素急性肺损伤的保护作用

目的:探讨降钙素基因相关肽(CGRP),对内毒素急性肺损伤(ALI)的保护作用.方法:应用内毒素(LPS,6 mg/kg,iv)复制大鼠ALI模型.选用SD大鼠24只,随机分成盐水对照组(NS,n=8)、ALI模型组(LPS,n=8)、CGRP干预组(n=8).CGRP干预组按2μg/kg经腹腔注入CGRP,30 min后再经颈静脉注入LPS;LPS组以生理盐水代替CGRP;NS组中,CGRP和LPS均以等量生理盐水代替.各组大鼠均于注射LPS或生理盐水6 h后测定动脉血氧分压(PaO2)、血清肿瘤坏死因子(TNF-α)及白介素-8(IL-8)含量,随后处死动物,观察肺病理改变,测定肺湿/干重比(W/D)及肺组织匀浆髓过氧化物酶(MPO)浓度.结果:注射LPS 6 h后,W/D、肺组织匀浆MPO、血清TNF-α、IL-8含量较对照组均有显著升高(P＜0.01),PaO2显著降低;而预先给予CGRP(2μg/kg)干预可显著缓解上述变化,PaO2改善,MPO含量较ALI模型组降低(P＜0.05),肺组织病理改变明显减轻.结论:小剂量CGRP能改善大鼠ALI时气体交换功能,抑制炎症介质的释放,对内毒素诱导的ALI有保护作用.     

Toll样受体4在脓毒症大鼠胰岛中的表达

目的 研究Toll样受体4(TLR4)在脓毒症大鼠胰岛中的表达变化及其对血糖的影响.方法 SPF级SD雄性大鼠腹腔内注射脂多糖(LPS)5 mg/kg,间隔3h注射第2次,制备脓毒症模型.大鼠被随机(随机数字法)分为正常对照组(n=5)、LPS组(n=5)、抗TLR4抗体组(n=5)和抗TLR4抗体+LPS组(n=5),分别采用RT-PCR法、Western-blot法和免疫组化法检测大鼠胰岛TLR4的表达.大鼠给药处理6h后行葡萄糖静脉试验(IVGTT)测定静脉全血葡萄糖和胰岛素水平,计算胰岛素、血糖曲线下面积(AUC).结果 正常组大鼠胰岛细胞有TIR4表达,LPS处理大鼠6h后胰岛组织中TLR4蛋白和mRNA的表达升高,与对照组比较差异具有统计学意义(P＜0.01),注射抗TLR4抗体预处理后用LPS刺激,LPS诱导的TLR4蛋白及mRNA水平的升高均被抑制(P＜0.01).LPS处理后与正常对照组比较,大鼠IVGTT 10、30、60、120 min血糖明显升高,30、60、120 min胰岛素分泌降低(P＜0.01).抗TLR4抗体干预后能降低30 ～ 120min血糖的升高,改善LPS对胰岛素分泌的抑制(P＜0.01).结论 脓毒症大鼠胰岛细胞TLR4表达升高,LPS-TLR4系统可能是应激性高血糖的发生机制.     

颈脊髓切断对内毒素休克大鼠全身炎症反应及预后的影响

目的 研究颈脊髓切断对内毒素休克大鼠全身炎症反应及预后的影响.方法 92只SD大鼠被随机分为正常对照组(n=8)、内毒素休克模型组(n=42)及内毒素休克+颈脊髓切断组(n=42),后两组各取10只大鼠于制模后观察48 h存活率,其余大鼠再按制模后3、6、12和48 h分为4个亚组,每亚组8只动物.切断大鼠C7脊髓制备颈脊髓切断动物模型;腹腔注射内毒素脂多糖(LPS)10 mg/kg造成动物内毒素休克.处死动物开胸取血,采用高效液相色谱法(HPLC)测定血浆中去甲肾上腺素(NE)含量,用双抗体夹心酶联免疫吸附法(ELISA)检测血浆中白细胞介素-10(IL-10)及IL-6的含量.结果 模型组血浆NE含量逐渐升高,而切断组逐渐下降.但均较对照组显著升高(P均＜0.05);自6 h起切断组显著低于模型组(P均＜0.05).模型组血浆IL-10含量于3 h和6 h时显著低于对照组(P均＜0.05),之后逐渐升高(P均＞0.05);切断组于各时间点显著高于模型组,且于12 h和48 h显著高于对照组(P均＜0.05).模型组各时间点血浆IL-6含量均高于对照组(P均＜0.05);切断组3、6和12 h显著低于模型组(P均＜0.05),且与对照组差异无统计学意义,而在48 h时较对照组显著升高(P＜0.05).切断组大鼠48 h存活率高于模型组(70%比20%).结论 颈脊髓切断可增加内毒素休克大鼠血浆中IL-10的含量,抑制IL-6生成,并能提高48 h存活率,对内毒素休克大鼠的全身炎症反应及预后具有改善的作用.     

Chronic plasminogen activator inhibitor-1 (PAI-1) overexpression dampens CD25+ lymphocyte recruitment after lipopolysaccharide endotoxemia in mouse lung

BACKGROUND: Plasma plasminogen activator inhibitor-1 (PAI-1) level rises during sepsis and confers a worse prognosis. PAI-1 participation to sepsis has been poorly documented and was mainly associated with fibrin deposits. Beside fibrin deposits, increased tissue PAI-1 expression may contribute to the poor outcome of endotoxemia through other mechanisms. OBJECTIVE AND METHODS: During lipopolysaccharide (LPS) challenge, the role of PAI-1 in the early phase of inflammation was examined in the lungs of transgenic mice that either overexpress or lack the PAI-1 gene (PAI-1Tg or PAI-1(-/-)). RESULTS: Analysis of leukocytes revealed that neutrophil and macrophage infiltrations did not differ for PAI-1Tg and wild-type (WT) mice. Remarkably, CD25+ lymphocyte infiltration was totally blunted in PAI-1Tg lungs and inversely correlated with fibrin depositions. In parallel, mRNA levels of the regulatory T cell (Treg) markers FoxP3, CTLA-4, and GITR were significantly lower in PAI-1Tg than in WT lungs after LPS challenge. These data are supported by opposite results in PAI-1(-/-) lungs. The systemic compartments (spleen and peripheral blood) showed no decrease in CD25+, CD4+ CD25+ lymphocytes, and Treg markers in PAI-1Tg mice after LPS injection compared with WT mice. In addition, plasma and lung concentrations of interleukin-6 (IL-6) and macrophage inflammatory protein-1alpha (MIP-1alpha) were significantly higher in PAI-1Tg mice than WT mice. CONCLUSION: Our results suggest that chronic tissue PAI-1 overexpression influences the early phase of the inflammatory response during endotoxemia through the control of T lymphocyte traffic.     

Cardiac response to nitric oxide synthase inhibition using aminoguanidine in a rat model of endotoxemia

This study evaluates the effect of aminoguanidine, a preferential inhibitor of inducible nitric oxide synthase (iNOS), on the prevention of cardiac depression in acute endotoxemia. Cardiac performance was evaluated after 4 h of exposure to endotoxin. Rats (n = 5) were selected randomly to receive, by intraperitoneal injection, one of four treatments: saline, LPS (lipopolysaccharide, E. coli, 4 mg/kg, AG (aminoguanidine 100 mg/kg), and LPS + AG at various times. AG and saline treatments were administered 30 min before LPS and at 1 and 3 h after LPS injection. Hearts were perfused using the Langendorff isolated perfusion system and a balloon-tipped catheter was placed into the left ventricle to measure left ventricular developed pressure (LVDP). Myocyte contractile function was assessed with electrical field stimulation and video microscopy. Tissue was immunostained for the expression of iNOS and for nitrotyrosine, a byproduct of protein nitration by peroxynitrite. Perfused hearts from LPS-treated rats exhibited a 57% decrease (P < 0.05) in LVDP compared to saline-treated animals. No improvement in ventricular function was observed with the administration of AG. Similarly, cardiac myocytes prepared from LPS-treated animals demonstrated a significant (P < 0.05) reduction in percent and velocity of shortening and this effect was unaltered with the same dose of AG. AG administration significantly reduced serum nitrite/nitrate levels (P < 0.05) in endotoxemic rats to control levels. Localized expression of iNOS in the myocardium was lessened with AG treatment and was not associated with peroxynitrite formation in this model of endotoxemia. The results indicate that AG given in vivo before and after endotoxin (at a concentration sufficient to decrease NO production) did not reduce cardiac depression. We conclude that selective inhibition of iNOS and the reduction of NO production do not prevent cardiac dysfunction at an early stage in an acute model of endotoxemia.     

胎羊炎症反应综合征静脉导管血流动力学变化与炎性介质关系的研究

目的 探讨胎羊内注射内毒素(LPS)后的静脉导管血流动力学变化及其与胎羊炎性介质变化的关系.方法 10只中晚孕山羊分为两组(实验组和对照组各5只),实验组于妊娠120～130 d行超声引导下胎羊脐静脉或脐-门静脉窦穿刺术,注射LPS(Escherichia coli O111:B5;L1885)约10μg/kg胎羊质量,对照组注射等量生理盐水.并分别于注射LPS前(0.5 h)、注射LPS后1 h、3 h、6 h应用彩色多普勒血流显像检测静脉导管血流频谱变化.同时抽取脐静脉血,用ELISA方法检测TNF-α、IL-6含量,并评价TNF-α、IL-6与静脉导管血流参数之间的相关性.结果 实验组给药后静脉导管搏动指数(DVPI)、心室收缩期峰值速度(S)、心室舒张期峰值流速(D)、血流量(DVQ)、静脉导管指数(DVI)、心室收缩期峰值速度/心房收缩期峰值流速(S/A)、(S-A)/D随给药时间升高,并且高于对照组(均P＜0.05);心房收缩期峰值流速(A波)下降,并且低于对照组(P＜0.05);静脉导管峡部的内径(DVD)无明显变化.实验组DVPI与血清IL-6、TNF-α呈正相关(P均＜0.05);实验组A波值与血清IL-6、TNF-α呈负相关(均P＜0.05);S波、DVI、D波、DVQ、S/A、(S-A)/D与血清IL-6呈正相关(均P＜0.05),与TNF-α无明显相关(均P＞0.05);静脉导管峡部内径与血清IL-6、TNF-α均无明显相关.结论 DVPI、A值可能成为预测胎儿全身炎症反应综合征简便而有效的指标.     

维生素D对脂多糖致急性肺损伤大鼠肺组织血管紧张素转化酶2和维生素D受体表达水平的影响

目的 观察维生素D(vitamin D,Vit D)对脂多糖(lipopolysaccharide,LPS)致Wistar大鼠急性肺损伤(acute lung injury,ALI)肺组织中血管紧张素转化酶2(angiotensinconverting enzyme 2,ACE2)和维生素D受体(vitamin D receptor,VDR)表达水平的影响.方法 采用尾静脉注射LPS方法制备大鼠ALI模型;将30只健康雄性Wistar大鼠随机(随机数字法)分为6组:正常对照组(NC组)、LPS组:尾静脉注射LPS 5mg/kg、Vit D组:给予Vit D活性形式(骨化三醇)25 μg/kg连续灌胃3d和(LPS+ Vit D) 1-3组:分别于骨化三醇1μg/kg、5μg/kg、25 μg/kg灌胃3d后尾静脉注射LPS 5mg/kg,所有大鼠于注射LPS的24 h后进行后续实验.分别观察大鼠一般情况,肺组织病理及肺干/湿重比变化、肺组织中VDR、ACE2蛋白及基因水平的表达.结果 LPS组大鼠病态表现(呼吸浅快、精神萎靡、口鼻可见血性分泌物)明显,(LPS+ VitD) 1-3组病态表现和肺组织病理损伤均较LPS组明显减轻.LPS组VDR和ACE2蛋白及基因水平的表达均较NC组和Vit D组显著降低(P＜0.05),(LPS+ VitD) 1-3组各组VDR和ACE2蛋白及基因水平的表达均较LPS组有所升高(P＜0.05),但仍显著低于Vit D组(P＜0.05),其中(LPS+ Vit D) 1-3组各组VDR蛋白及基因水平的表达差异无统计学意义(P＞0.05),ACE2的表达差异有统计学意义(P＜0.05).结论 Vit D能使LPS致ALI大鼠肺组织中ACE2和VDR蛋白及基因表达水平增加,故此推测ACE2和VDR表达增加可能对ALI的发生、发展起保护作用.     

Effects of cervical chordotomy on systemic inflammatory response and the outcome in rats with endotoxic shock induced by lipopolysaccharide

OBJECTIVE: To investigate the effects of cervical chordotomy on systemic inflammatory response and the outcome in rats with endotoxemia induced by lipopolysaccharide (LPS). METHODS: Ninety-two Sprague Dawley (SD) rats were randomly divided into three groups: normal control group (groupI, n=8) , endotoxemia group (groupII, n=42) and endotoxemia with cervical chordotomy ( groupIII, n=42). Endotoxemia was induced by intra-peritoneal injection of LPS 10 mg/kg. In group III, "cervical chordotomy" was attained by transection of spinal cord at C7 immediately before intra-peritoneal LPS administration. Ten rats of groupII and III each were observed for 48-hour survival. The other rats were further divided into four subgroups of 8 animals each, according to the time when the animals were sacrificed . The animals were sacrificed at 3, 6, 12, and 48 hours after intra-peritoneal LPS injection. Heart blood samples were obtained for determination of plasma concentration of norepinephrine [NE, by high performance liquid chromatography (HPLC)] and plasma concentration of interleukin-10 (IL-10) and IL-6 [by enzyme linked immunosorbent assay (ELISA)]. RESULTS: Plasma NE concentration were significantly increased after intra-peritoneal LPS injection in groupIIand III as compared with group I and were significantly lower in group III than in groupII starting from 6 hours after LPS (all P<0.05). Plasma IL-10 concentration was significantly lower at 3 hours and 6 hours while plasma IL-6 concentration was significantly higher after LPS challenge in groupII than in groupIat all time points (all P<0.05). High transection of spinal cord significantly elevated plasma IL-10 level at 12 hours and 48 hours, lowered IL-6 release at 3, 6, and 12 hours (all P<0.05), and improved 48-hour survival (20% vs. 70%) in group III as compared with group II. CONCLUSION: Transection of spinal cord at C7 level can ameliorate the systemic inflammatory response induced by endotoxemia thus improving the outcome through elevation in IL-10 level, decreases in IL-6 release, and improves 48-hour survival. This might be attributable to loss of sympathetic nerve function.     

Intravenous infusion of mesenchymal stem cells is associated with improved myocardial function during endotoxemia

Mesenchymal stem cells (MSCs) possess immunomodulatory properties and may curtail the inflammatory response that characterizes sepsis and other systemic inflammatory states. We aimed to determine whether intravenous infusion of MSCs is associated with reduced inflammation and improved myocardial function in a rat model of endotoxemia. Adult Sprague-Dawley rats were administered saline (vehicle) or LPS (5 mg/kg) via tail vein injection. Treatments, either vehicle or 2 × 10(6) MSCs, were infused 1 h later via tail vein. Animals were randomly assigned to the following groups: (a) vehicle + vehicle (control; n = 6), (b) LPS + vehicle (n = 6), or (c) LPS + MSCs (n = 6). Six hours after induction of endotoxemia, left ventricular ejection fraction (EF) and fractional shortening (FS) was assessed via parasternal short-axis M-mode echocardiography. Hearts and serum were collected for determination of cytokine levels via enzyme-linked immunosorbent assay. Animals injected with LPS + vehicle exhibited depressed cardiac function as indicated by a 26% and 37% reduction in EF and FS from baseline, respectively. Treatment with MSCs was associated with improved cardiac function compared with vehicle treatment as indicated by a reduction in EF and FS of only 10% and 17%, respectively (P < 0.05). Myocardial levels of TNF-α, IL-1β, and IL-6 were elevated in LPS-treated animals versus control. Similarly, serum levels of IL-1β, IL-6, and IL-10 were increased in LPS-treated animals. Treatment with MSCs, however, was associated with significant reductions in serum levels of IL-1β and IL-6 and in myocardial levels of TNF-α, IL-1β, and IL-6. In addition, treatment with MSCs was associated with a further increase in serum IL-10. Infusion of MSCs modulates the systemic inflammatory response and is associated with improved cardiac function during endotoxemia.