摘要
目的 探讨苦柯胺B（KB）对脓毒症小鼠小肠炎症反应的抑制作用及其分子机制。方法 按照随机数字表法将24只雄性ICR小鼠分为对照组、模型组、KB干预组，每组8只。腹腔注射脂多糖（I^PS）20mg／kg制备脓毒症动物模型，对照组注射等量生理盐水；KB干预组于制模后4h经尾静脉注射KB20雌mg进行干预。各组于注射LPS后8h取心脏血和空肠、回肠组织，检测血浆LPS含量；采用酶联免疫吸附试验（ELISA）检测血浆及小肠组织肿瘤坏死因子-α（TNF-α,）和白细胞介素-1β（IL-1β）水平；光镜下观察小肠组织病理学改变；比色法检测小肠组织髓过氧化物酶（MPO）活性；免疫组化法观察小肠组织细胞间黏附分子-1（ICAM-1）表达；反转录-聚合酶链反应（RT—PCR）检测小肠组织诱导型-氧化氮合酶（iNOS）mRNA表达；蛋白质免疫印迹试验（WesternBlot）检测小肠组织核转录因子-kB（NF—kB）蛋白表达。结果 模型小鼠表现为肠组织微血管通透性增加，间质水肿，白细胞浸润；血浆LPS、TNF-α、IL—1β水平及小肠TNF-α、IL-1β、MPO活性、ICAM-1阳性表达、iNOS mRNA、NF—kB蛋白表达均明显升高；而经KB干预后，小肠组织微血管通透性降低，水肿程度减轻，白细胞浸润显著减少，血浆LPS、TNF-α、IL-1β及小肠TNF-α、IL-1β、MPO活性、ICAM-1阳性表达、iNOSmRNA和NF—kB蛋白表达均较模型组均明显下降[血浆LPS（kEU／L）：654．09±28．13比1155．65±47．15，TNF—α（ng／L）：12．75±0．47比30．61±0．71，IL-1β（ng／L）：53．06±5．32比64．47±2．61；空肠TNF-α（ng／L）：43．27±1．20比64．82±2．09，IL-1β（ng／L）：326．38±14．47比535．22±13．48．MPO（U，g）：0．14±0．01比0．32±0．02，iNOSmRNA（2^-△△Ct）：2．39±0．13比10．80±0．22，NF—kB蛋白（灰度值）：0．687±0．062比1．404±0．046；回肠TNF-α（ng／L）：62．75±3．9
Objective To study the role of Kukoamine B （KB） in inhibiting the inflammatory response of small intestine in septic mice and its molecular mechanisms. Methods Twenty-four male ICR mice were randomly divided into control group, model group, and KB intervention group （each, n = 8 ）. Sepsis model was reproduced by intra-peritoneal injection of 20 mg/kg tipopolysaccharide （LPS）, while equivalent normal saline was given in control group, and 20 pg/kg KB was injected through caudal vein 4 hours after LPS challenge in KB intervention group. The blood/tissue samples （jejunum and ileum ） were harvested 8 hours after LPS injection. The levels of plasma LPS, tumor necrosis factor-et （TNF-ct） and interleukin-l[3 （IL-I[3） were measured. The pathological changes in small intestine tissues were observed under light microscope, while the levels of inflammatory cytokines TNF-et and IL-I[3 in the tissue homogenates （jejunum and ileum） were assessed by enzyme linked immunosorbent assay （ELISA）. The activity of myeloperoxidase （MPO） was measured by colorimetry. The expression of intercellular adhesion molecule-1 （ICAM-1 ） was determined by immunohistochemistry. The expressions of inducible nitric oxide synthase （iNOS） mRNA was assayed by reverse transcription-polymerase chain reaction （RT-PCR）. The activation of nuclear factor-r,.B （NF-xB） was determined by Western Blot. Results The mice in model group were found to have an increase in microvascular permeability, interstitial edema, and infiltration of white blood cells, and the levels of LPS, TNF-α and IL-1β in their plasma, with an increase in concentrations of TNF-α and IL-1β, activity of MPO, positive expression of ICAM-1, expression of iNOS mRNA and NF-KB protein in small intestine （jejunum and ileum）. Compared with model group, in mice with KB intervention, microvascular permeability, interstitial edema, and infiltration of white blood cells were reduced significantly, while the levels of LPS, TNF-ct and IL-113 in pl
出处
《中华危重病急救医学》
CAS
CSCD
北大核心
2015年第2期121-126,共6页
Chinese Critical Care Medicine
基金
基金项目：国家自然科学基金（30772256,81071546,81272148）
江苏省自然科学基金（BK2012703）
关键词
脂多糖
苦柯胺B
小肠
炎症反应
Lipopolysaccharide
Kukoamine B
Small intestine
Inflammatory response
