Hemorrhagic transformation in patients with large-artery atherosclerotic stroke is associated with the gut microbiota and lipopolysaccharide

Hemorrhagic transformation is a major complication of large-artery atheroscle rotic stroke(a major ischemic stro ke subtype)that wo rsens outcomes and increases mortality.Disruption of the gut microbiota is an important feature of stroke,and some specific bacteria and bacterial metabolites may contribute to hemorrhagic transformation pathogenesis.We aimed to investigate the relationship between the gut microbiota and hemorrhagic transformation in largearte ry atheroscle rotic stro ke.An observational retrospective study was conducted.From May 2020 to September 2021,blood and fecal samples were obtained upon admission from 32 patients with first-ever acute ischemic stroke and not undergoing intravenous thrombolysis or endovascular thrombectomy,as well as 16 healthy controls.Patients with stro ke who developed hemorrhagic transfo rmation(n=15)were compared to those who did not develop hemorrhagic transformation(n=17)and with healthy controls.The gut microbiota was assessed through 16S ribosomal ribonucleic acid sequencing.We also examined key components of the lipopolysaccharide pathway:lipopolysaccharide,lipopolysaccharide-binding protein,and soluble CD14.We observed that bacterial diversity was decreased in both the hemorrhagic transformation and non-hemorrhagic transfo rmation group compared with the healthy controls.The patients with ischemic stro ke who developed hemorrhagic transfo rmation exhibited altered gut micro biota composition,in particular an increase in the relative abundance and dive rsity of members belonging to the Enterobacteriaceae family.Plasma lipopolysaccharide and lipopolysaccharide-binding protein levels were higher in the hemorrhagic transformation group compared with the non-hemorrhagic transfo rmation group.lipopolysaccharide,lipopolysaccharide-binding protein,and soluble CD14 concentrations were associated with increased abundance of Enterobacte riaceae.Next,the role of the gut microbiota in hemorrhagic transformation was evaluated using an experimental stroke rat model.In this model,transplantation of the gut microbiota from hemorrhagic transformation rats into the recipient rats triggered higher plasma levels of lipopolysaccharide,lipopolysaccharide-binding protein,and soluble CD14.Ta ken togethe r,our findings demonstrate a noticeable change in the gut microbiota and lipopolysaccharide-related inflammatory response in stroke patients with hemorrhagic transformation.This suggests that maintaining a balanced gut microbiota may be an important factor in preventing hemorrhagic transfo rmation after stro ke.

Endotoxin-induced alterations of adipose tissue function:a pathway to bovine metabolic stress

During the periparturient period, dairy cows exhibit negative energy balance due to limited appetite and increased energy requirements for lactogenesis. The delicate equilibrium between energy availability and expenditure puts cows in a state of metabolic stress characterized by excessive lipolysis in white adipose tissues(AT), increased production of reactive oxygen species, and immune cell dysfunction. Metabolic stress, especially in AT, increases the risk for metabolic and inflammatory diseases. Around parturition, cows are also susceptible to endotoxemia. Bacterial-derived toxins cause endotoxemia by promoting inflammatory processes and immune cell infiltration in different organs and systems while impacting metabolic function by altering lipolysis, mitochondrial activity, and insulin sensitivity. In dairy cows, endotoxins enter the bloodstream after overcoming the defense mechanisms of the epithelial barriers, particularly during common periparturient conditions such as mastitis, metritis, and pneumonia, or after abrupt changes in the gut microbiome. In the bovine AT, endotoxins induce a pro-inflammatory response and stimulate lipolysis in AT, leading to the release of free fatty acids into the bloodstream. When excessive and protracted, endotoxin-induced lipolysis can impair adipocyte's insulin signaling pathways and lipid synthesis. Endotoxin exposure can also induce oxidative stress in AT through the production of reactive oxygen species by inflammatory cells and other cellular components. This review provides insights into endotoxins' impact on AT function, highlighting the gaps in our knowledge of the mechanisms underlying AT dysfunction, its connection with periparturient cows' disease risk, and the need to develop effective interventions to prevent and treat endotoxemia-related inflammatory conditions in dairy cattle.

Quercetin Alleviates Lipopolysaccharide-Induced Cardiac Inflammation via Inhibiting Autophagy and Programmed Cell Death

Objective The aim of this study is to explore the potential modulatory role of quercetin against Endotoxin or lipopolysaccharide(LPS)induced septic cardiac dysfunction.Methods Specific pathogen-free chicken embryos(n=120)were allocated untreated control,phosphate buffer solution(PBS)vehicle,PBS with ethanol vehicle,LPS(500 ng/egg),LPS with quercetin treatment(10,20,or 40 nmol/egg,respectively),Quercetin groups(10,20,or 40 nmol/egg).Fifteenday-old embryonated eggs were inoculated with abovementioned solutions via the allantoic cavity.At embryonic day 19,the hearts of the embryos were collected for histopathological examination,RNA extraction,real-time polymerase chain reaction,immunohistochemical investigations,and Western blotting.Results They demonstrated that the heart presented inflammatory responses after LPS induction.The LPS-induced higher mRNA expressions of inflammation-related factors(TLR4,TNFα,MYD88,NF-κB1,IFNγ,IL-1β,IL-8,IL-6,IL-10,p38,MMP3,and MMP9)were blocked by quercetin with three dosages.Quercetin significantly decreased immunopositivity to TLR4 and MMP9 in the treatment group when compared with the LPS group.Quercetin significantly decreased protein expressions of TLR4,IFNγ,MMP3,and MMP9 when compared with the LPS group.Quercetin treatment prevented LPS-induced increase in the mRNA expression of Claudin 1 and ZO-1,and significantly decreased protein expression of claudin 1 when compared with the LPS group.Quercetin significantly downregulated autophagyrelated gene expressions(PPARα,SGLT1,APOA4,AMPKα1,AMPKα2,ATG5,ATG7,Beclin-1,and LC3B)and programmed cell death(Fas,Bcl-2,CASP1,CASP12,CASP3,and RIPK1)after LPS induction.Quercetin significantly decreased immunopositivity to APOA4,AMPKα2,and LC3-II/LC3-I in the treatment group when compared with the LPS group.Quercetin significantly decreased protein expressions of AMPKα1,LC3-I,and LC3-II.Quercetin significantly decreased the protein expression to CASP1 and CASP3 by immunohistochemical investigation or Western blotting in treatment group when compared with LPS group.Conclusion Quercetin alleviates cardiac inflammation induced by LPS through modulating autophagy,programmed cell death,and myocardiocytes permeability.

Natural Products Improve Organ Microcirculation Dysfunction Following Ischemia/Reperfusion-and Lipopolysaccharide-Induced Disturbances:Mechanistic and Therapeutic Views

Microcirculatory disturbances are complex processes caused by many factors,including abnormal vasomotor responses,decreased blood flow velocity,vascular endothelial cell injury,altered leukocyte and endothelial cell interactions,plasma albumin leakage,microvascular hemorrhage,and thrombosis.These disturbances involve multiple mechanisms and interactions among mechanisms that can include energy metabolism,the mitochondrial respiratory chain,oxidative stress,inflammatory factors,adhesion molecules,the cytoskeleton,vascular endothelial cells,caveolae,cell junctions,the vascular basement membrane,neutrophils,monocytes,and platelets.In clinical practice,aside from drugs that target abnormal vasomotor responses and platelet adhesion,there continues to be a lack of multi-target drugs that can regulate the complex mechanistic links and interactions underlying microcirculatory disturbances.Natural products have demonstrated obvious positive therapeutic effects in treating ischemia/reperfusion(I/R)-and lipopolysaccharide(LPS)-induced microcirculatory disturbances.In recent years,numerous research papers on the improvement of microcirculatory function by natural products have been published in international journals.In 2008 and 2017,the first listed author of this review was invited to publish reviews in the journal of Pharmacology&Therapeutics on the improvement of microcirculatory disturbances and organ injury induced by I/R using Salvia miltiorrhiza ingredients and other natural components of compounded Chinese medicine,respectively.This review systematically summarizes the effects,targets of action,and mechanisms of natural products regarding improving I/R-and LPSinduced microcirculatory disturbances and tissue injury.Based on this summary,scientific proposals are suggested for the discovery of new drugs to improve microcirculatory disturbances in disease.

Foeniculum vulgare Mill.inhibits lipopolysaccharide-induced microglia activation and ameliorates neuroinflammation-mediated behavioral deficits in mice

Objective:To investigate the effect of Foeniculum vulgare extract against lipopolysaccharide(LPS)-induced microglial activation in vitro as well as cognitive behavioral deficits in mice.Methods:LPS-activated BV-2 cell viability was measured using MTT assay and reactive oxygen species(ROS)was studied using DCF-DA assay.The antioxidative enzymes and pro-inflammatory mediators were analyzed using respective ELISA kits and Western blotting.For in vivo testing,LPS(1 mg/kg,i.p.)was given daily for five days in male Swiss albino mice to produce chronic neuroinflammation.Cognitive and behavioral tests were performed using open-field,passive avoidance,and rotarod experiments in LPS-induced mice.Results:Foeniculum vulgare extract(25,50 and 100μg/mL)significantly attenuated the LPS-activated increase in nitric oxide(NO),ROS,cyclooxygenase-2,inducible NO synthase,IL-6,and TNF-alpha(P<0.05).Moreover,LPS-induced oxidative stress and reduced antioxidative enzyme levels were significantly improved by Foeniculum vulgare extract(P<0.05).The extract also regulated the NF-κB/MAPK signaling in BV-2 cells.In an in vivo study,Foeniculum vulgare extract(50,100,and 200 mg/kg)markedly mitigated the LPS-induced cognitive and locomotor impairments in mice.The fingerprinting analysis showed distinctive peaks with rutin,kaempferol-3-O-glucoside,and anethole as identifiable compounds.Conclusions:Foeniculum vulgare extract can ameliorate LPS-stimulated neuroinflammatory responses in BV-2 microglial cells and improve cognitive and locomotor performance in LPS-administered mice.

Antidepressant effects of Peiyuan Jieyu formula in a mouse model of chronic stress in conjunction with lipopolysaccharide-induced depression

Objective:To explore the mechanism of the Peiyuan Jieyu formula in treating depression by assessing its impact on a lipopolysaccharide-induced(LPS-induced)depression mouse model.Methods:We created a mouse model of depression by exposing mice that had previously received chronic stress to intraperitoneal LPS injections.The mice were divided into the following groups:control,model,fluoxetine,Tiansi Yin,Sini powder,and low-,medium-,and high-dose Peiyuan Jieyu formula groups.Forced swim and tail suspension tests were used to assess the efficacy of the depression(despair)model,and weight gain rates were also measured.Furthermore,serum levels of various depression and inflammation-associated molecules,including tumor necrosis factor-a(TNF-a),interferon-γ(IFN-γ),tryptophan,5-hydroxytryptamine,kynurenine(KYN),and kynurenic acid(KA)were assessed.Furthermore,the expression levels of ionic calcium-binding adaptor molecule-1(IBA-1)and indoleamine 2,3-dioxygenase(IDO)mRNA in hippocampal microglia were measured.Results:The model group displayed greater despair-associated immobility,which was shortened in response to various doses of Peiyuan Jieyu formula.Furthermore,formula administration significantly reduced serum TNF-a levels and hippocampal IDO mRNA expression.The high formula dose also reduced IFN-γand IBA-1 levels,the latter was also decreased in response to the medium formula dose.However,the low formula dose reduced serum KYN level and KYN/tryptophan(TRP)and KYN/KA ratios.Conclusion:The Peiyuan Jieyu formula holds immense potential in treating depression in a mouse model,potentially inhibiting inflammation and improving TRP-KYN metabolic disorders.

Predictive effect of lipopolysaccharide-stimulated inflammatory cytokines on symptoms of generalized anxiety disorder

BACKGROUND Generalized anxiety disorder(GAD)is a relatively common mental disorder.Recently,inflammation,an important factor for the development of depression,has attracted increasing attention.Several studies have shown that inflammatory cytokines can affect the pathophysiological processes of several nervous system diseases.We hypothesized that there is a correlation between the levels of lipopolysaccharide(LPS)-stimulated inflammatory cytokines and the clinical symptoms of GAD.AIM To investigate the predictive effect of LPS-stimulated inflammatory cytokines on symptoms of GAD.METHODS This was a cross-sectional study in which 89 patients with GAD diagnosed at The First Hospital of Hebei Medical University from January 2022 to December 2022 and 70 individuals without anxiety and depression(controls)during the same period were included.Fasting venous blood was collected from all the subjects in heparin tubes,and another 3 ml of blood was supplemented with LPS(10 ng/ml).The plasma levels of 12 cytokines[Interleukin(IL)-1β,IL-2,IL-4,IL-5,IL-6,IL-8,IL-10,tumor necrosis factor(TNF)-α,interferon(IFN)-γ,IL-17A,IL-12p70,and IFN-α]were detected.RESULTS Post-LPS stimulation,the levels of IL-1β,IL-6,IL-8,IL-10,and TNF-αin both the control and GAD groups were significantly elevated above those in the nonstimulated groups,with IL-6 and IL-8 showing marked increases.Increases in IL-8 and TNF-αwere statistically significant in the GAD group(P<0.05).IL-1β,IL-6,IL-8,IL-10,and TNF-αwere found to be significantly correlated with Hamilton Anxiety Rating Scale(HAMA)scores(P<0.05).A negative correlation was observed between IL-10 levels and HAMA scores.Further analysis revealed that TNF-αwas associated with mental anxiety,whereas IL-1β,IL-8,and IL-10 were associated with physical anxiety symptoms,with IL-10 showing a negative correlation with physical anxiety.IL-6 was associated with both mental and physical aspects of anxiety.CONCLUSION The physical symptoms of GAD are related to inflammatory factors.IL-1β,IL-8,IL-10,and TNF-a can be used as predictors of physical or mental anxiety in patients with GAD.

Structural Properties of the Lipid A Component of the Lipopolysaccharide(Endotoxin)Isolated from Bacteriodes fragilis NCTC 9343

Various chemical and gas-liquid chromatographic analyses indicate that the lipid A,backbone of the lipopolysaccharide(LPS)isolated from B.fragilis NCTC 9343 is chemically con-stitutεd by a β1,6-interlinked D-glucosamine disaccharide.It is phosphorylated at its 1-positionby a glvcosidic-linkage while the ester-linked phosphate at 4’—position present generally in otherlipid A is depleted.The lipid A is lower fatty acylated in the amount of 5.2 fatty acids/lipid Amolecule(of which 0.73 forms 3-acyloxyacyl groups).The LPS containing such an incompletelipid A component has been shown endowina with a weaker endotoxicitv.

Yeast hydrolysate attenuates lipopolysaccharide-induced inflammatory responses and intestinal barrier damage in weaned piglets

Background Intestinal inflammation is the main risk factor causing intestinal barrier dysfunction and lipopolysaccharide(LPS)can trigger inflammatory responses in various eukaryotic species.Yeast hydrolysate(YH)possesses multibiological effects and is received remarkable attention as a functional ingredient for improving growth performance and promoting health in animals.However,there is still inconclusive on the protective effects of dietary YH supplementation on intestinal barrier of piglets.This study was conducted to investigate the attenuate effects of YH supplementation on inflammatory responses and intestinal barrier injury in piglets challenged with LPS.Methods Twenty-four piglets(with an average body weight of 7.42±0.34 kg)weaned at 21 days of age were randomly assigned to one of two dietary treatments(12 replications with one pig per pen):a basal diet or a basal diet containing YH(5 g/kg).On the 22nd d,6 piglets in each treatment were intraperitoneally injected with LPS at 150μg/kg BW,and the others were injected with the same amount of sterile normal saline.Four hours later,blood samples of each piglet were collected and then piglets were euthanized.Results Dietary YH supplementation increased average daily feed intake and average daily gain(P<0.01),decreased the ratio of feed intake to gain of piglets(P sponse,evidenced by the increase o=0.048).Lipopolysaccharide(LPS)injection induced systemic inflammatory ref serum concentrations of haptoglobin(HP),adrenocorticotropic hormone(ACTH),cortisol,and interleukin-1β(IL-1β).Furthermore,LPS challenge resulted in inflammatory intestinal damage,by up-regulation of the protein or mRNA abundances of tumor necrosis factor-α(TNF-α),IL-1β,toll-like receptors 4(TLR4)and phosphor-nuclear factor-κB-p65(p-NFκB-p65)(P<0.01),and down-regulation of the jejunal villus height,the protein and mRNA abundances of zonula occludens-1(ZO-1)and occludin(OCC;P<0.05)in jejunal mucosa.Dietary YH supplementation decreased the impaired effects of ACTH,cortisol,HP,IL-1βand diamine oxidase in serum(P<0.05).Moreover,YH supplementation also up-regulated the jejunal villus height,protein and mRNA abundances of ZO-1 and OCC(P<0.05),down-regulated the mRNA expressions of TNF-αand IL-1βand the protein abundances of TNF-α,IL-1β,TLR4 and p-NFκB-p65 in jejunal mucosa in LPS-challenged pigs(P<0.01).Conclusion Yeast hydrolysate could attenuate inflammatory response and intestinal barrier injury in weaned piglets challenged with LPS,which was associated with the inhibition of TLR4/NF-κB signaling pathway activation.

Physico-chemical characterization of the S-and R-lipid A derived from the natural endotoxin(lipopolysaccharide) preparations

The natural Salmonella abortus equi endotoxin(lipopolysaccharide) preparation was fractionated with organic solvents into smooth (S-) and rough(R-) fractions to prepare their corresponding lipid A moieties. The lipid A obtained from S-fraction(S-lip A) wa

Physico-chemical Characterization of the Lipopolysaccharide (Endotoxin) Isolated from Bacteriodes fragilis NCTC 9343

The lipopolysaccharide (LPS, endotoxin) isolated from Bacteriodes fragilis NCTC 9343 ex-hibted an endogenous heterogencity consisting of R (rough) and SR (semi-rough ) LPS fractionsupon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The LPS was chemically consti-tuted by D-glucosamine, D-glucose. L-rhamnose, D-galactose. fatty acids and phosphate in amolar ratio of 2: 1: 1: 4: 5. 4.: 2.7 The fatty acids were predominated by 3-hydroxypentadecanoic(3-OH-15: 0), 3-hydroxyhcxadecanoic (3-OH-16: 0), 3-hydroxyheptadecanoic (3-OH-17: 0), 3-hydroxy-15-methyl hexadecanoic (3-OH-15-Me-16: 0) and 13-methyl-letradecanoic acid (13-Mc-14:0). Approximately 1 mol of 2-Keto-3-deoxy-D-mannooctonatc (KDO) was detected after treat-ment of LPS with 48% of hydrofluoric acid (HF), indicating it was present as a phosphorylatedmolecule.

Effect of dual targeting procyanidins nanoparticles on metabolomics of lipopolysaccharide-stimulated inflammatory macrophages

Inflammation plays an important role in the occurrence and development of many inflammatory diseases.The purpose of this study was to evaluate the anti-inflammatory effect and metabolic behavior of the dual targeting procyanidins(PC)nanoparticles on lipopolysaccharide(LPS)-stimulated inflammatory macrophages by metabolomics method.The double-targeting PC nanoparticles could specifi cally target both the CD44 receptor and mitochondria,while the single targeting PC-loaded nanoparticles that could target the CD44 receptor on the surface of macrophages.The double-targeting PC nanoparticles had better inhibitory effect than single-targeting PC nanoparticles on the leakage of lactate dehydrogenase and reactive oxygen species overexpression induced by LPS.Amino acid metabolism,energy metabolism and purine metabolism were disordered in LPS-treated group,and metabolic pathway analysis indicated that the double-targeting PC nanoparticles reversed some of LPS impacts.The changes of these potential biomarkers and their corresponding pathways are helpful to further understand the mechanism of PC nanoparticles in alleviating inflammation,and promote their application in nutrition intervention.

Lipopolysaccharide-induced Trigeminal Ganglion Nerve Fiber Damage is Associated with Autophagy Inhibition

Objective This study aimed to determine whether lipopolysaccharide(LPS)induces the loss of corneal nerve fibers in cultured trigeminal ganglion(TG)cells,and the underlying mechanism of LPS-induced TG neurite damage.Methods TG neurons were isolated from C57BL/6 mice,and the cell viability and purity were maintained for up to 7 days.Then,they were treated with LPS(1µg/mL)or the autophagy regulator(autophibib and rapamycin)alone or in combination for 48 h,and the length of neurites in TG cells was examined by the immunofluorescence staining of the neuron-specific proteinβ3-tubulin.Afterwards,the molecular mechanisms by which LPS induces TG neuron damage were explored.Results The immunofluorescence staining revealed that the average length of neurites in TG cells significantly decreased after LPS treatment.Importantly,LPS induced the impairment of autophagic flux in TG cells,which was evidenced by the increase in the accumulation of LC3 and p62 proteins.The pharmacological inhibition of autophagy by autophinib dramatically reduced the length of TG neurites.However,the rapamycin-induced activation of autophagy significantly lessened the effect of LPS on the degeneration of TG neurites.Conclusion LPS-induced autophagy inhibition contributes to the loss of TG neurites.

川藏香茶菜庚素的体外抗炎作用与分子机制研究

为探究川藏香茶菜庚素(Pseurata H)对巨噬细胞炎症模型的抗炎作用及分子机制,利用LPS刺激RAW264.7细胞建立体外炎症模型,并分别以浓度为10、20和40μmol/L的Pseurata H干预该细胞模型。利用CCK-8法检测Pseurata H对细胞增殖活性的影响;Griess法和ELISA试剂盒分别检测细胞上清液中NO、IL-6和TNF-α的含量;Western Blot检测COX-2、iNOS蛋白、p-NF-κB和p-ERK蛋白的表达变化;以RT-qPCR法检测药物干预前后细胞中iNOS、TNF-α、IL-6和IL-1β的mRNA基因表达变化。结果表明:浓度为10、20和40μmol/L的Pseurata H对细胞没有毒性,不同浓度的Pseurata H对LPS诱导RAW264.7细胞炎症模型中NO、IL-6和TNF-α分泌具有很好的下调作用,对iNOS、COX-2、NF-κB、p-NF-κB、ERK和p-ERK蛋白的表达具有良好的抑制作用,且具有浓度效应。Pseurata H具有明显的抗炎作用,其作用机制可能是通过抑制NF-κB/MAPK信号通路而发挥抗炎作用。

Value of combining the serum D-lactate, diamine oxidase, and endotoxin levels to predict gut-derived infections in cancer patients

Objective:This is a retrospective observational cohort study.The objective of this retrospective observational cohort study was to evaluate the value of the combined serum D-lactic acid,diamine oxidase(DAO),and endotoxin levels to predict intestinal barrier impairment and gut-derived infection(GDI)in cancer patients.Methods:Cancer patients receiving chemotherapy or palliative care treatment at our hospital were enrolled in the study.The serum concentrations of DAO,D-lactic acid,and endotoxin were determined using the intestinal barrier function biochemical index analysis system.The patients'infection information came from the hospital's Medicom Prescription Automatic Screening System and themedical records.Three hundred fifty-three cancer patients were included in the study(53.8%female,73.7%cancer stage IV,27.8%had bowel obstruction).Results:The total incidence of GDI was 33.4%(118/353).The median length of hospital stay was 16 days for patients with GDI,compared with 7 days for patients without GDI(P<0.001).The media hospitalization costs were¥27,362.35 for patients with GDI compared with¥11,614.08 for patients without GDI(P<0.001).The serum concentrations of DAO,D-lactic acid,and endotoxin were significantly higher in patients with GDI.As malignant bowel obstruction(MBO)worsened,the concentrations of DAO,D-lactic acid,and endotoxin increased.Multivariate logistic regression models revealed that the DAO,endotoxin,IL-6,and C-reactive protein levels were significantly associated with an increased risk of GDI.In addition,we also found a fivefold increased risk of infection in patients withMBO compared with those without bowel obstruction(OR=6.210,P<0.001).All of the areas under the receiver operating characteristic curve(AUCs)for DAO,D-lactate,and endotoxin to predict GDI were<0.7(AUC=0.648,P<0.001;AUC=0.624,P<0.01;AUC=0.620,P<0.01,respectively).However,when the parameters were combined(DAO+D-lactate+endotoxin),the predictive power increased significantly(AUC=0.797,P<0.001).Moreover,combining these intestinal barrier indicators and the presence of MBO had better power to predict GDI than either alone(AUC=0.837,P<0.001).Conclusions:Combining the serum DAO,D-lactic acid,and endotoxin levels was a better predictor of GDI than any of the indicators alone,and combining these with the diagnosis of MBO could further improve the efficacy for predicting GDI.

巨噬细胞极化为M1型或M2型对铁代谢的影响

目的探讨巨噬细胞极化形成M1型或M2型对铁代谢的影响。方法应用脂多糖(LPS)20 ng/mL或IL-410 ng/mL诱导RAW264.7巨噬细胞极化,电子显微镜下观察处理后的巨噬细胞极化形态特征,采用免疫荧光染色法及Western blot法检测M1型标志物TNF-α和一氧化氮合酶(iNOS)及M2型标志物精氨酸酶1(Arg-1)的表达情况;LPS或IL-4处理后的巨噬细胞与红细胞及铁剂共培养,通过瑞氏染色和铁染色,评价其铁摄入功能;ELISA法检测LPS或IL-4处理后巨噬细胞内及培养液中铁蛋白含量,以评价其铁储存功能。结果LPS处理后巨噬细胞由圆形向多形性转化;同时,TNF-α和iNOS的表达水平明显增加;可见吞噬红细胞现象,胞质中吞噬的铁颗粒明显增多。IL-4处理后巨噬细胞形态也呈多形性改变,同时Arg-1表达水平增加,但未见吞噬红细胞现象,且胞质内吞噬的铁颗粒较少。此外,LPS处理后巨噬细胞胞内铁蛋白含量较IL-4处理后明显增多。结论LPS与IL-4可诱导巨噬细胞形成不同极化亚型,其铁代谢存在明显差异。LPS处理的M1型巨噬细胞对铁的摄入及储存较IL-4处理的M2亚型巨噬细胞增加,有固铁优势。

MUC2基因沉默对脂多糖干扰Caco-2/HT-29细胞紧密连接蛋白表达的影响

肠道黏膜屏障阻碍病原物质入侵的第一道防线是由黏蛋白-2(MUC2)组成的肠道黏液层,覆盖于上皮细胞表面,由杯状细胞分泌而成。脂多糖(lipopolysaccharides, LPS)可通过损伤肠黏膜从而干扰屏障功能的正常发挥。论文对Caco-2/HT-29(3∶1)共培养模拟肠道上皮细胞,通过沉默黏蛋白-2(siMUC2)分析脂多糖对肠道黏液层屏障及其上皮紧密连接结构的影响。结果显示,与对照组相比,LPS组E-钙黏素(E-cadherin)、闭合蛋白mRNA表达量均明显降低。与LPS组相比,LPS+siMUC2组中连接蛋白(claudin)、连接附着分子(JAMA)、E-cadherin以及桥粒(desmosome) mRNA水平显著降低,细胞连接蛋白mRNA水平的下调可能影响细胞通透性进而导致屏障功能紊乱。说明黏蛋白-2作为黏液层骨架蛋白成分可以保护细胞紧密连接蛋白免受LPS的干扰。LPS可诱导Caco-2/HT-29细胞模型黏蛋白-2表达增多以及E-cadherin、闭合蛋白mRNA表达下调。当黏蛋白-2基因被沉默后,LPS可导致细胞紧密连接蛋白mRNA表达量的显著下降,提示肠道上皮细胞紧密连接的屏障功能可能被破坏。

人参皂苷Rb1减轻脂多糖诱导急性肺损伤的作用研究

目的:探讨人参皂苷Rb1(GsRb1)对脂多糖(LPS)诱导急性肺损伤(ALI)的保护作用及其作用机制。方法:C57BL/6小鼠随机分为对照组、模型组、GsRb1给药组(低、中、高剂量组),气管内给予LPS造模,给药组在造模前3 d分别腹膜腔注射给予不同剂量GsRb1预处理,造模12 h将其麻醉,先采集肺泡灌洗液后再处死取出肺组织,计算肺湿/干比,试剂盒检测灌洗液白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)水平以及肺髓过氧化物酶(MPO)活性,Western blot检测肺组织核因子-κB(NF-κB)蛋白的表达。结果:高剂量GsRb1能明显降低肺组织湿/干比(P<0.05);中、高剂量GsRb1能明显降低肺组织MPO活性(P<0.05);高剂量GsRb1明显降低小鼠肺泡灌洗液IL-1β和TNF-α水平(P<0.05);高剂量GsRb1能明显降低肺组织NF-κB p65蛋白的表达(P<0.05)。结论:GsRb1可能通过调控NF-κB通路,减轻炎症反应,抑制LPS诱导的小鼠ALI。

益生菌发酵黄芪对LPS诱导的人正常结肠上皮细胞NCM-460氧化损伤保护作用的机制研究

探究益生菌发酵黄芪(fermented Astragali Radix,F-As)与黄芪(Astragali Radix,As)对脂多糖(lipopolysaccharide,LPS)诱导的肠道细胞NCM-460氧化损伤的保护作用及其分子机制。利用MTT法确定了LPS、As和F-As的工作浓度;分别检测并比较了LPS组、As+LPS组和F-As+LPS组,细胞中氧化应激相关指标、抗氧化相关基因在转录水平和蛋白水平表达的变化。与Con组相比,LPS组的活性氧(reactive oxygen species,ROS)和丙二醛(malondialdehyde,MDA)增加(P<0.01),而细胞的存活率、谷胱甘肽(glutathione,GSH)、超氧化物歧化酶(superoxide dismutase,SOD)、总抗氧化能力(total antioxidant capacity,T-AOC)以及细胞中的抗氧化基因在转录水平和蛋白水平均显著降低(P<0.01);而As+LPS组和F-As+LPS组,细胞中的ROS和MDA水平降低(P<0.01),细胞的存活率GSH、SOD、T-AOC以及抗氧化基因在转录和蛋白水平得到恢复(P<0.01),且与As+LPS组相比,F-As+LPS组恢复得更显著(P<0.05)。研究结果提示,As和F-As均可以通过激活抗氧化基因的表达,清除细胞内过多的ROS和MDA,从而对LPS诱导的细胞氧化损伤起到保护作用,且F-As优于As。

不同检测方法对复用手术器械清洗效果的评价研究

目的:采用细菌内毒素检测方法对复用手术器械的清洗效果进行评价,探索不同检测方法检测不同种类手术器械清洗合格率的差异。找到不同种类复用手术器械的最佳检测方法,提高清洗质量,降低成本,以防止医院感染的发生。方法:将手术器械分为咬合齿类、光滑类和管腔类三组,使用目测法、放大镜法、残留蛋白试验、三磷酸腺苷(ATP)生物荧光法、细菌内毒素法进行器械清洗后的检测。结果:用肉眼裸视法与放大镜观察法检测手术器械68份。目测法合格率为98.53%,放大镜法合格率为94.12%。其中44份样品进行了残留蛋白检测和51份样品进行了ATP生物荧光法检测,合格率分别为81.82%和86.27%。细菌内毒素法检测65份样品,合格率为75.38%,其中管腔类器械合格率最低为33.33%。结论:手术器械清洗效果的5种评价方法的灵敏度依次为细菌内毒素法、残留蛋白试验、ATP生物荧光法、放大镜法和目测法。同类器械的清洗合格率除ATP法外差异无统计学意义,不同检测方法显示内毒素法结果差异均有统计学意义。