The endothelium: physiological functions and role in microcirculatory failure during severe sepsis

The endothelium is a highly dynamic cell layer that is involved in a multitude of physiological functions, including the control of vascular tone, the movement of cells and nutrients, the maintenance of blood fluidity and the growth of new vessels. During severe sepsis, the endothelium becomes proadhesive, procoagulant, antifibrinolytic and is characterized by alterations of vasomotor regulation. Most of these functions have been discovered using in vitro and animal models, but in vivo exploration of endothelium in patients remains difficult. New tools to analyze endothelial dysfunction at bedside have to be developed.     

Clinical review: Role of triggering receptor expressed on myeloid cells-1 during sepsis

Triggering receptor expressed on myeloid cells (TREM)-1 is a recently identified molecule that is involved in monocytic activation and in the inflammatory response. It belongs to a family related to the natural killer cell receptors and is expressed on neutrophils, mature monocytes and macrophages. The inflammatory response mediated by Toll-like receptor-2 and -4 stimulation is amplified by the engagement of TREM-1. The expression of membrane-bound TREM-1 is greatly increased on monocytes during sepsis. Moreover, infection induces the release of a soluble form of this receptor, which can be measured in biological fluid and may be useful as a diagnostic tool. Modulation of the TREM-1 signalling pathway by the use of small synthetic peptides confers interesting survival advantages during experimental septic shock in mice, even when this teatment is administered late after the onset of sepsis.     

Effects of dopexamine on rat cardiorenal functions during lipopolysaccharide-induced experimental sepsis

We aimed to test the protective effect of dopexamine on renal function and systemic haemodynamics in rats with induced sepsis. Female Sprague-Dawley rats were randomized into three equal groups: group 1 (control, received 3% creatinine throughout the experiment); group 2 (sepsis, received 3% creatinine and Escherichia coli lipopolysaccharide [LPS] endotoxin [8 mg/kg per h]); and group 3 (sepsis plus dopexamine, received 3% creatinine, E. coli LPS and dopexamine [1 microgram/kg per min]). Time-adjusted heart rate, systolic, diastolic and mean arterial pressures, urine volume and glomerular filtration rate (from creatinine clearance) were recorded. After bacterial infusion heart rate increased and mean arterial pressure decreased; the fall in mean arterial pressure was less pronounced with dopexamine (group 3) than without (group 2). Dopexamine also induced significant and moderate increases in urine volume and heart rate, respectively. We concluded that dopexamine has some positive inotropic-chronotropic effects and induces favourable responses in renal function.     

肝脏缺血预处理过程中蛋白激酶C活性的变化

目的:观察在肝脏缺血预处理过程中蛋白激酶C的活性改变及细胞内信号转导机制。方法:实验于2004-03/2005-03在南方医科大学珠江医院中心实验室完成。实验分组:36只大鼠随机分成6组,每组6只。①假手术组。②缺血再灌注组。③缺血预处理组。④缺血再灌注 豆蔻酸佛波酰乙脂组。⑤缺血预处理 白屈菜季铵碱组。⑥缺血预处理 PD98059组。实验干预:①假手术组仅分离肝十二指肠韧带,不阻断肝门,不进行其他干预处理。②缺血再灌注组在第一肝门用小血管夹阻断尾状叶及左肝叶血流40min松开血管夹肝脏再灌注3h,再灌注开始后关腹。③缺血预处理组先行3个循环的缺血预处理,阻断第一肝门10min,开放再灌注10min为1个循环,随后操作同缺血再灌注组。④缺血再灌注 豆蔻酸佛波酰乙脂组:按4μg/kg共0.5mL蛋白激酶C激动剂豆蔻酸佛波酰乙脂溶液,经静脉通道缓慢推注,推注10min,推注后等待10min,余处理同缺血再灌注组。⑤缺血预处理 白屈菜季铵碱组:按5mg/kg共0.5mL蛋白激酶C抑制剂白屈菜季铵碱溶液,同样用10min经静脉通道缓慢推注,推注后等待10min,其余处理同缺血预处理组。⑥缺血预处理 PD98059组:按5mg/kg共0.5mL丝裂原激活的蛋白激酶抑制剂PD98058溶液,同上速度缓慢静脉推注,推注后等待10min,余处理同缺血预处理组。实验评估:①在ECHNICONRA-1000全自动生化分析仪上采用速率法检测血清谷草转氨酶和血清谷丙转氨酶活性。②按蛋白激酶C活性测定试剂盒操作步骤检测肝组织蛋白激酶C活力。③Westernblot免疫印迹法检测肝组织磷酸P44/42MAPKs活性。结果:36只大鼠均进入结果分析,中途无脱落和死亡。①血清谷草转氨酶和血清谷丙转氨酶活性:缺血预处理组低于缺血再灌注组(P<0.01)。缺血预处理 白屈菜季铵碱组、缺血预处理 PD98059组显著高于缺血预处理组(P<0.01)。缺血再灌注 豆蔻酸佛波酰乙脂组显著低于缺血再灌注组(P<0.01)。②蛋白激酶C活力:缺血预处理组高于缺血再灌注组[(1877.2±81.0),(713.1±37.0)pkat/g,P<0.01];缺血再灌注 豆蔻酸佛波酰乙脂组亦显著高于缺血再灌注组[(2758.4±454.3),(713.1±37.0)pkat/g,P<0.01];缺血预处理 白屈菜季铵碱组明显低于缺血预处理组[(567.9±46.2),(1877.2±81.0)pkat/g,P<0.01]。③磷酸P44/42MAPKs活性:与假手术组比较,其在缺血再灌注组的表达量轻度升高;缺血预处理组较缺血再灌注组升高显著;缺血预处理 白屈菜季铵碱组较缺血预处理组明显降低,缺血预处理 PD98059组表达量下降更为显著;缺血再灌注 豆蔻酸佛波酰乙脂组表达量较缺血再灌注组明显升高。结论:缺血预处理保护效应中,蛋白激酶C对P44/42MAPKs信号通路的激活是细胞保护效应的一个重要环节。     

The liver in sepsis: shedding light on the cellular basis of hepatocyte dysfunction

Liver dysfunction is believed to contribute to the metabolic derangements of critical illness. The cellular basis of liver dysfunction is poorly understood and its consequences largely unknown. Recent work by Gonnert and colleagues sheds additional light. Using two imaging techniques to track the clearance of biotransformed dyes by the liver in a rat model of intra-abdominal infection, the authors show that the predominant defect in sepsis lies in the excretion of biotransformed molecules from the hepatocyte into the bile canaliculi. Their work both points to a new aspect of hepatic dysfunction through focus on a role in the metabolic derangements of sepsis and suggests a possible strategy to diagnose and monitor this process in critically ill patients.A new report by Gonnert and co-workers from the Center for Sepsis Control and Care in Jena Germany, recently appearing in Critical Care, provides important new insight into the nature and mechanisms of hepatocellular dysfunction in sepsis [1]. Since Bywaters first described the phenomenon of jaundice following severe trauma in 1946 [2], liver dysfunction has been considered a prominent feature of the Multiple Organ Dysfunction Syndrome and has been defined predominantly by hyperbilirubinemia and clinical jaundice [3-6]. The liver, however, is an enormously complex organ whose function is difficult to characterize succinctly or comprehensively. The liver plays a critical role in protein synthesis, in intermediary metabolism, and in the excretion of toxins and wastes, including the breakdown products of senescent red cells; this latter role is reflected in the widespread use of bilirubin levels as a measure of hepatic dysfunction. However, the liver is also involved in hematopoiesis during embryologic development, in innate immunity, and in digestion through the production of bile.Yet we understand little about how liver function is altered during critical illness, and how these alterations are best measured. Hyperbilirubinemia, the classic marker of hepatic dysfunction, provides little insight into what is happening within the liver. The prevalence of hyperbilirubinemia appears to be on the decline, perhaps because of declining use of total parenteral nutrition and of transfusion, although it is an independent risk factor for increased mortality [7,8]. Moreover, the presence and prognostic import of hyperbilirubinemia appear to derive more from the fact that this marker delineates a population of patients with irreversible end-stage liver disease rather than a transient and potentially reversible process that might be considered a component of the Multiple Organ Dysfunction Syndrome. Greater insight into other aspects of hepatic dysfunction that might be both deranged in critical illness and measured in critically ill patients is clearly needed.The Jena group is making substantial strides in addressing this need, and is bringing attention to striking abnormalities in hepatic biotransformation and excretion that arise in severely ill patients [9]. They have shown previously that hepatocellular transport at the canalicular pole of the hepatocyte is impaired in sepsis, with the result that a spectrum of hepatotoxic materials accumulates within the septic liver. They now extend these important observations [1].Gonnert and colleagues use a combination of elegant imaging techniques to detect clearance of two dyes that are metabolized by the liver - indocyanine green and DY635 - in a lethal rat model of intra-abdominal infection resulting from intraperitoneal administration of feces. They show that dye is cleared from the circulation and taken up readily by liver cells, but that its appearance in bile is delayed, indicating that defective cellular function primarily involves excretion of transformed molecules from the hepatocyte into the biliary canaliculi, rather than its delivery to, or uptake by, liver cells. Impaired excretion of dye is further evidenced as sustained persistence of dye within the liver, and its reduced appearance within the lumen of the gastrointestinal tract.This body of work and the ongoing evolution of the authors'' investigations hold exciting and important possibilities. First, the work provides valuable insight into the nature of hepatocellular dysfunction in critical illness, shifting the focus from microcirculatory impairments or abnormalities in uptake of hepatically transformed compounds to their excretion into the biliary tree and gastrointestinal tract. The significance of accumulation of a spectrum of compounds - toxins, drugs, bilirubin, and other metabolic substances - within the hepatocyte is uncertain, and is fertile ground for future study. Does their accumulation subsequently result in impaired uptake from the liver microvasculature? If so, then toxins or drugs may accumulate in the circulation, their normal metabolism impeded by failure of their excretion from the hepatocyte. If this is the case, then those alterations may be modifiable by altering drug doses or by extracorporeal removal, and eventually by pharmacologic interventions that correct the excretory derangement. Does impaired hepatocellular excretion affect the normal synthetic function of the hepatocyte, and therefore impair its ability to synthesize proteins such as acute phase reactants or coagulation factors? Does impaired excretion alter the ability of the liver to take up glucose and store it as glycogen, and therefore may hepatocellular dysfunction emerge as a component of the syndrome of insulin resistance that is so commonly seen in sepsis? Do alterations in hepatocyte function result in altered Kupffer cell function, and therefore contribute to the immunologic alterations of sepsis?Moreover, as techniques of near-infrared fluorescence imaging improve, the technologies described by Gonnert and colleagues may well find a clinical role as a noninvasive test of hepatic dysfunction in the critically ill patient, and so provide a measure of liver function that is more sensitive, sophisticated, and relevant than the serum bilirubin level.Despite its fundamental and varied role in normal homeostasis, to the intensivist the liver remains a frustratingly opaque black box whose inner workings are obscure. Studies like those of Gonnert and colleagues are beginning to lift the lid, if only a little, and to shine a light on a largely unexplored area in the support of the critically ill patient.     

Time-course of sTREM (soluble triggering receptor expressed on myeloid cells)-1, procalcitonin, and C-reactive protein plasma concentrations during sepsis

OBJECTIVE: To describe the course of plasma sTREM (soluble triggering receptor expressed on myeloid cells)-1, procalcitonin (PCT), and C-reactive protein (CRP) concentrations during sepsis and their clinical informative value in predicting outcome. DESIGN: Prospective, noninterventional study. SETTING: Medical adult intensive care unit at a university hospital in France. PATIENTS: Sixty-three critically ill patients with sepsis, severe sepsis, or septic shock. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Soluble TREM-1 concentrations were significantly lower at admission in nonsurvivors (n = 21) than in survivors (n = 42) (94 [30-258] vs. 154 [52-435] pg/mL, p = .02), whereas PCT levels were higher among nonsurvivors (19.2 [0.3-179] vs. 2.4 (0-254) pg/mL, p = .001). CRP levels did not differ between the two groups of patients. Plasma PCT and CRP decreased during the 14-day period of study in both survivors and nonsurvivors. Conversely, sTREM-1 plasma concentrations remained stable or even increased in nonsurviving patients and decreased in survivors. An elevated baseline sTREM-1 level was found to be an independent protective factor with an odds of dying of 0.1 (95% confidence interval, 0.1-0.8). CONCLUSION: A progressive decline of plasma sTREM-1 concentration indicates a favorable clinical evolution during the recovery phase of sepsis. In addition, baseline sTREM-1 level may prove useful in predicting outcome of septic patients.     

Targeting the programmed cell death 1: programmed cell death ligand 1 pathway reverses T cell exhaustion in patients with sepsis

Introduction

A major pathophysiologic mechanism in sepsis is impaired host immunity which results in failure to eradicate invading pathogens and increased susceptibility to secondary infections. Although many immunosuppressive mechanisms exist, increased expression of the inhibitory receptor programmed cell death 1 (PD-1) and its ligand (PD-L1) are thought to play key roles. The newly recognized phenomenon of T cell exhaustion is mediated in part by PD-1 effects on T cells. This study tested the ability of anti-PD-1 and anti-PD-L1 antibodies to prevent apoptosis and improve lymphocyte function in septic patients.

Methods

Blood was obtained from 43 septic and 15 non-septic critically-ill patients. Effects of anti-PD-1, anti-PD-L1, or isotype-control antibody on lymphocyte apoptosis and interferon gamma (IFN-γ) and interleukin-2 (IL-2) production were quantitated by flow cytometry.

Results

Lymphocytes from septic patients produced decreased IFN-γ and IL-2 and had increased CD8 T cell expression of PD-1 and decreased PD-L1 expression compared to non-septic patients (P<0.05). Monocytes from septic patients had increased PD-L1 and decreased HLA-DR expression compared to non-septic patients (P<0.01). CD8 T cell expression of PD-1 increased over time in ICU as PD-L1, IFN-γ, and IL2 decreased. In addition, donors with the highest CD8 PD-1 expression together with the lowest CD8 PD-L1 expression also had lower levels of HLA-DR expression in monocytes, and an increased rate of secondary infections, suggestive of a more immune exhausted phenotype. Treatment of cells from septic patients with anti-PD-1 or anti-PD-L1 antibody decreased apoptosis and increased IFN-γ and IL-2 production in septic patients; (P<0.01). The percentage of CD4 T cells that were PD-1 positive correlated with the degree of cellular apoptosis (P<0.01).

Conclusions

In vitro blockade of the PD-1:PD-L1 pathway decreases apoptosis and improves immune cell function in septic patients. The current results together with multiple positive studies of anti-PD-1 and anti-PD-L1 in animal models of bacterial and fungal infections and the relative safety profile of anti-PD-1/anti-PD-L1 in human oncology trials to date strongly support the initiation of clinical trials testing these antibodies in sepsis, a disorder with a high mortality.     

对高迁移率族蛋白B1作用的新认识

高迁移率族蛋白B1(high mobility grouo box 1 protein，HMGB1)是存在于真核生物细胞内的一类非组蛋白染色体结合蛋白，曾经作为一种转录N子和促生长因子而被广泛研究。新近研究发现。HMGB1可分泌到胞浆乃至胞外，并可与白细胞介素-1(IL-1)、IL-6、肿瘤坏死因子-α(TNF-α)等重要炎症因子相互诱生。循环HMGB1在脓毒症后期升高，有可能作为一种“晚期”炎症介质参与脓毒症的发生与发展过程。     

Endothelial protein C receptor polymorphisms and risk of severe sepsis in critically ill patients

Purpose

Endothelial protein C receptor (EPCR) is expressed mainly in endothelial cells and is involved in regulation of the cytoprotective and anticoagulant pathways of protein C. We assessed whether haplotypes in the EPCR gene modify the risk of severe sepsis and/or septic shock (SS/SS) development in critically ill patients.

Methods

Three polymorphisms in the EPCR gene were genotyped in 389 Caucasian critically ill patients, hospitalized in the intensive care units of two major hospitals in Athens, Greece. Multivariate logistic regression analysis controlling for age, acute physiology and chronic health evaluation (APACHE) II and sequential organ failure assessment (SOFA) scores, sex, and diagnosis was performed to determine the effect of haplotypes H1 and H3 in the EPCR gene on the development of SS/SS.

Results

H2 carriers versus all other genotypes combined had a nonsignificant excess of SS/SS (p = 0.087). SS/SS occurred in 38.8 % of critically ill patients carrying minor alleles belonging to both H1 and H3 haplotypes, in 58.0 % of H1 carriers, 64.3 % of H3 carriers, and 65.2 % of patients carrying all common alleles (H2). Compared with H2 carriers, the odds ratios (OR) for developing SS/SS were 0.34 [95 % confidence interval (CI) 0.16–0.76, p = 0.008] for simultaneous H1 and H3 carriers, 0.65 (95 % CI 0.37–1.13, p = 0.123) for H1 carriers, and 0.82 (95 % CI 0.39–1.70, p = 0.590) for H3 carriers.

Conclusions

Our results indicate that simultaneous carriers of minor alleles belonging to both the H1 and H3 haplotypes may be at reduced risk of developing SS/SS in this cohort of critically ill patients.     

Protective functions of intracellular heat-shock protein (HSP) 70-expression in patients with severe sepsis

Intracellular expression of heat-shock-protein 70 (HSP70) arose early in evolutionary development as a tool to protect cellular homeostasis. HSP70 detects proteins that are incorrectly folded or denatured. They form a complex with such proteins which can lead to correct folding, compartmentalization in organelles, or to proteolytic degradation. HSP70 also appears to protect proteins from degeneration. Intracellular HSP70-expression is induced by a wide variety of stimuli including heat, fever, hypoxia, oxygen radicals, endotoxins, cytokines, and heavy metal ions. Pre-emptive induction of HSP70-expression reduces organ dysfunction and mortality in animal models of sepsis.     

Cholecystokinin-1 receptor protein up-regulation during pancreatic regeneration after acute haemorrhagic pancreatitis in rats

BACKGROUND: Cholecystokinin (CCK) plays an important role in regeneration after acute pancreatitis in rats. The present study was aimed to elucidate the role of CCK-1 receptor (CCK-1R) in acute pancreatitis. We investigated the serial changes in CCK-1R mRNA and protein levels and their immunolocalization after acute haemorrhagic pancreatitis induced in male Wistar rats by retrograde intraductal infusion of 4% sodium taurocholate (100 micro L 100 g(-1) body weight). METHODS: Histological changes were evaluated by haematoxylin and eosin staining. Pancreatic CCK-1R mRNA was determined by Northern blot analysis. Pancreatic CCK-1R protein was evaluated by immunoblot analysis and immunohistochemistry with a polyclonal antibody against rat CCK-1R protein. RESULTS: Histological findings revealed that newly formed acinar cells were detected at the periphery of tubular complexes on day 14, and normal architecture of lobules was observed focally on day 21. Pancreatic CCK-1R mRNA peaked on day 3 and thereafter gradually decreased. Cholecystokinin-1R protein rapidly increased after induction of pancreatitits, reaching a maximal level on day 3. On day 3, intense immunoreactivity for CCK-1R protein was observed in both the cytoplasm of vacuolized acinar cells and the tubular complexes. In the regenerative process after acute haemorrhagic pancreatitis in rats, the expression of pancreatic CCK-1R mRNA and protein increased, and intense immunoreactivity for CCK-1R protein was observed in tubular complexes in the cytoplasm of regenerated acinar cells. CONCLUSION: These results suggest that CCK-1R contributes to pancreatic regeneration after acute haemorrhagic pancreatitis and that tubular complexes are involved in the process of acinar cell regeneration following pancreatic injury.     

激活素受体相互作用蛋白ARIPzip真核表达载体的构建及表达

目的构建激活素受体相互作用蛋白 （aetivin receptor interacting protein zip, ARIPzip） 真核表达载体,以便进一步探讨其生物学作用。方法从小鼠成纤维细胞3T3中提取HNA,并利用ARIPzip特异性引物通过RT-PCR方法扩增了ARIPzip cDNA,并克隆入pcDNA-Flag载体中,构建重组载体pcDNA-F-ARIPzip。将重组载体转染入HEK293细胞中,并通过免疫印迹方法检测ARIPzip的表达。结果酶切及测序结果显示,重组质粒构建成功;免疫印迹结果显示,瞬时转染重组质粒的HEK293细胞中可有效表达ARIPzip。结论本研究成功构建了激活素受体相互作用蛋白ARIPzip真核表达载体,并能够在真核细胞中进行表达,为进一步研究ARIPzip的生物学功能奠定了基础。     

Activation of mitogen-activated protein kinases during granulocyte apoptosis in patients with severe sepsis

Reduction of neutrophil apoptosis represents a major cause for granulocytosis and increases the destructive potential of theses cells during systemic inflammatory response syndrome (SIRS) and sepsis. In this light, the role of protein kinases for the regulation of altered neutrophil apoptosis under infectious conditions was investigated. Neutrophils, obtained from patients with severe sepsis (n = 18), were incubated ex vivowith either LPS (1 microg/mL) or interferon-gamma (IFN-gamma; 10 ng/mL) for 16 h. Apoptosis was determined by propidium iodine (PI) staining of DNA fragments and was compared with the rate of spontaneous apoptosis. Tyrosine kinases were inhibited by herbimycin (1 microM), the mitogen-activated protein (MAP) kinase ERK was inhibited with PD98059 (50 microM), and p38 MAP kinase was inhibited with SB203580 (5 microM). Herbimycin reconstituted LPS-reduced apoptosis in neutrophils from controls (39.9 +/- 3.8%) and patients (20.8 +/- 2.8%) to levels seen in spontaneous apoptosis (70.9 +/- 2.8% and 40.7 +/- 3.7%, respectively). Inhibition of the ERK kinase yielded similar results, whereas SB203580 had no effect on LPS-reduced apoptosis. However, inhibition of p38 partially reconstituted IFN-gamma-reduced apoptosis (51.3 +/- 7.7% and 25.6 +/- 5.8%) and increased spontaneous apoptosis (82.4 +/- 3.3% and 42.0 +/- 5.8%) in controls and patients, respectively. Western blot analysis revealed phosphorylation of both MAP kinases by LPS, but not by IFN-gamma. Inhibition of MAP kinases did not augment neutrophil apoptosis in patients to the level seen in controls, indicating that other mechanisms must be involved in the regulation of neutrophil apoptosis. Although the ERK kinase regulates LPS-induced reduction of apoptosis, the p38 MAP kinase might be involved in IFN-gamma signaling and the feedback regulation of neutrophil apoptosis.     

可溶性髓样细胞触发受体-1对脓毒血症早期诊断价值的研究

目的探讨血清可溶性髓样细胞触发受体-1(sTREM-1)对脓毒血症的早期诊断价值。方法选择2010年7月至2013年6月该院急诊重症病房重症监护病房收治的81例患者,其中61例全身炎症反应综合征(SIRS)患者,根据脓毒血症诊断标准分为脓毒血症组39例与SIRS组22例,非SIRS患者20例作为对照组,检测3组血清sTREM-1和降钙素原(PCT)水平。结果脓毒血症组与SIRS组血清sTREM-1及PCT水平均明显高于对照组(P0.01);脓毒血症组血清sTREM-1及PCT水平均明显高于SIRS组(P0.01);sTREM-1和PCT在SIRS患者中早期诊断脓毒血症的受试者工作特征曲线下面积分别为0.932和0.670,sTREM-1的灵敏度和特异度分别为92.3%和86.4%,PCT的灵敏度和特异度分别为61.5%和81.8%。结论血清sTREM-1是脓毒血症早期诊断的较好指标,同时具有较高的灵敏度和特异度,优于PCT。     

Differential expression of toll-like receptor genes: sepsis compared with sterile inflammation 1 day before sepsis diagnosis

Toll-like receptors (TLRs) are critical components of innate immunity. This study was designed to evaluate differential expression of genes for TLR and associated signal transduction molecules in critically ill patients developing sepsis compared with those with sterile inflammation. Uninfected critically ill patients with systemic inflammatory response syndrome were prospectively followed daily for development of sepsis. They were divided into two groups and compared in a case-control manner: (a) preseptic patients (n = 45) who subsequently developed sepsis, and (b) uninfected systemic inflammatory response syndrome patients (n = 45) who remained uninfected. Whole blood RNA was collected (PAXGene tube) at study entry and 1, 2, and 3 days before clinical sepsis diagnosis (or time-matched uninfected control) and analyzed via Affymetrix Hg_U133 Plus 2.0 microarrays. Genes were considered differentially expressed if they met univariate significance controlled for multiple comparisons at P < 0.005. Differentially expressed probes were uploaded into the Database for Annotation, Visualization and Integrated Discovery. The TLR pathway (Kyoto Encyclopedia of Genes and Genomes-KEGG) significance was determined via Expression Analysis Systematic Explorer (EASE) scoring. A total of 2,974 Affymetrix probes representing 2,190 unique genes were differentially expressed 1 day before sepsis diagnosis. Thirty-six probes representing 25 genes were annotated to the TLR pathway (KEGG) via the Database for Annotation, Visualization and Integrated Discovery with an EASE score at P < 0.0004. Notable TLR genes demonstrating increased expression include TLR-4 (median, 1.43-fold change), TLR-5 (2.08-fold change), and MAPK14 (1.90-fold change). An additional 11 unique genes were manually annotated into the TLR pathway based on known relevance such as TLR-8 (1.54-fold change). The total 36 genes contained 28 showing increased expression and 8 showing decreased expression. Differential gene expression was noted for TLR receptors (eight genes), TLR intracellular signal transduction cascade molecules (27 genes), and TLR-related effector molecules (one gene). The TLR and downstream signaling genes are differentially expressed in critically ill patients developing sepsis compared with those with sterile inflammation. These expression differences occur before phenotypic-based diagnosis of clinical sepsis.     

乌司他丁对脓毒症患者可溶性髓样细胞触发受体-1表达的影响

目的:探讨乌司他丁对脓毒症患者可溶性髓样细胞触发受体-1 (sTREM-1)表达的影响.方法:选择60例确诊脓毒症的患者随机分为乌司他丁治疗组和常规治疗对照组,各30例.乌司他丁治疗组在常规治疗的基础上,加用乌司他丁20万单位/次静脉注射,2次/日连续10 d,分别于治疗前,治疗前第3、7和10日时取两组患者静脉血,采用ELISA法检测血清sTREM-1浓度,采用免疫比浊法检测CRP浓度.观察两组患者APACHEⅢ评分变化,并记录治疗后3、7、10和28 d时两组患者生存或死亡情况.检测30例健康志愿者的血清sTREM-1和CRP浓度作为正常对照组.结果:治疗前两组血清sTREM-1和CRP浓度显著高于健康对照组 (P＜0.05),但两组患者血清sTREM-1和CRP浓度比较差异无统计学意义(P＞0.05).治疗3、7、10 d两组血清sTREM-1和CRP浓度较治疗前逐步下降(P＜0.05).乌司他丁治疗组较常规治疗对照组血清sTREM-I和CRP浓度下降更显著(P＜0.05).APACHEⅢ评分降低更明显(P＜0.05).结论:乌司他丁可降低脓毒症患者血清sTREM-1和CRP水平,减轻脓毒症炎症反应强度,减少MODS的发生,从而缓解脓毒症临床症状,改善预后.     

Early rise in circulating endothelial protein C receptor correlates with poor outcome in severe sepsis

Purpose  

The endothelial protein C receptor (EPCR) negatively regulates the coagulopathy and inflammatory response in sepsis. Mechanisms controlling the expression of cell-bound and circulating soluble EPCR (sEPCR) are still unclear. Moreover, the clinical impact of EPCR shedding and its potential value to predict sepsis progression and outcome remain to be established.     

Role of cAMP-response element-binding protein phosphorylation in hepatic apoptosis under protein kinase C alpha suppression during sepsis

Previous studies have shown that a decrease in protein kinase C (PKC) alpha levels contributes to hepatic failure and/or apoptosis during sepsis, and suppression of PKCalpha plays a critical role in triggering caspase-dependent apoptosis, which can modulate expression of Bcl-xL. However, the underlying molecular mechanism remains uncertain. In the present study, we examined whether a decrease in the nuclear PKCalpha levels causes hepatic apoptosis via modulation of cAMP-response element-binding protein (CREB) or nuclear factor-kappaB (NFkappaB), the crucial factors regulating the expression of prosurvival Bcl-xL. For polymicrobial sepsis induction, a cecal ligation and puncture model was used; at 9 or 18 h after CLP, experiments were terminated, referring as early or late sepsis, respectively. Additionally, PKCalpha was suppressed by stable transfection of antisense PKCalpha plasmid into a Clone-9 rat hepatic epithelial cell. The results showed that the nuclear PKCalpha was significantly decreased in the liver during sepsis, which was accompanied by decreases in phospho-CREB content, DNA-binding activity of CREB, and Bcl-xL expression. Likewise, the binding activity of NFkappaB increased significantly, which was associated with a decrease in cytosolic inhibitory-kappaBalpha content. The in vitro suppression of PKCalpha also resulted in decreases in the phospho-CREB content and DNA-binding activity, which were accompanied by down-regulation of Bcl-xL and apoptosis, but no significant alteration in NFkappaB-binding activity. The in vivo and in vitro results suggest that the suppression of PKCalpha results in a decreased CREB phosphorylation and subsequent down-regulation of Bcl-xL, which may contribute to the hepatic apoptosis during sepsis.