脂多糖致感染性脑水肿大鼠星形胶质细胞的活化和凋亡

目的通过观察脂多糖（Lipopolysaccharide,LPS）致大鼠感染性脑水肿后星形胶质细胞的凋亡、活化及iNOS表达情况,探讨在LPS致脑水肿中星形胶质细胞的作用。方法84只雄性SD大鼠,随机分为3组：空白对照纽（C组,n=12）;生理盐水对照纽（S组,n=12）;感染性水肿组（L组,n=60）。感染性脑水肿组又按颈内动脉注射脂多糖后6h、12h、24h、48h、72h分为5个亚组（n=12）。向颈内动脉注射脂多糖LPS150μg（0．15ml）建立大鼠感染性脑水肿模型。采用HE染色、流式细胞检测和免疫纽化染色分别观察脑组织病理改变、星形胶质细胞凋亡、胶质纤维酸性蛋白（GFAP）和iNOS表达情况。结果与C组和S组比较,L组大鼠星形胶质细胞胞体变大、突起增多;细胞内GFAP和iNOS表达增强,12h达高峰,除72h组,其余亚组均有统计学差异（P〈0．05）;各亚组星形胶质细胞凋亡增多（P〈0．05）,24h凋亡最显著。C组和S纽比较无统计学意义（P〉0．05）。结论感染性脑水肿后星形胶质细胞凋亡增加,异常活化,分泌iNOS,参与感染性脑水肿的形成,在脑水肿的发生发展中发挥重要作用。     

Bacterial lipopolysaccharide (LPS) enhances concanavalin A reactivity of thymocytes from the low-LPS-responder mouse strain C3H/Hej.

The capacity of LPS to enhance Con A reactivity of thymocytes was studied comparatively in the low-LPS-responder C3H/Hej mice and the high-LPS-responder CBA mice. The extent of synergism LPS + Con A was found similar in both strains.     

Reactivity in vitro of the leukocytes of the earthworm Eisenia foetida Sav. to several mitogenic substances]

Earthworm leukocytes were stimulated by various mitogens including LPS, PHA and Con A. Con A could stimulate leukocytes when they were cultivated in totality. After separation into sub-populations, small-non-adherent-leukocytes and large-adherent ones showed specific reactions with mitogens. Both PHA and LPS reacted with small leukocytes but LPS had no action on large adherent leukocytes.     

Lipopolysaccharide-binding molecules: transporters, blockers and sensors

Lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria, can be beneficial to the host by activating the innate immune system, or harmful, by inducing inflammation, disseminated intravascular coagulation, multiple organ failure, shock and often death. On the bacteria, and in host biological fluids and cells, LPS is never free but constantly attached to cognate-binding proteins. Understanding how LPS is transported and further recognized by sensors able to deliver a signal, or by inactivating molecules able to neutralize its biological effects, is an important goal. This review describes the large panel of peptides and proteins reported to associate with LPS, and provides information on their origin, their structure and the location of amino acid residues involved in their interaction with LPS. A better understanding of the mode of recognition of LPS by cognate proteins prompted many laboratories to design on a rational basis synthetic molecules which can be used to detect low amounts of endotoxin, or to act as efficient blockers of in vitro and in vivo responses to LPS.Received 15 January 2004; received after revision 20 February 2004; accepted 25 February 2004     

Bacterial lipopolysaccharide (LPS) enhances concanavalin A reactivity of thymocytes from the low-LPS-responder mouse strain C3H/Hej

Summary The capacity of LPS to enhance Con A reactivity of thymocytes was studied comparatively in the low-LPS-responder C3H/Hej mice and the high-LPS-responder CBA mice. The extent of synergism LPS+Con A was found similar in both strains.This work was supported by a grant from the INSERM (CRL: 76-5-101-1).     

The Sushi peptides: structural characterization and mode of action against Gram-negative bacteria

The compositional difference in microbial and human cell membranes allows antimicrobial peptides to preferentially bind microbes. Peptides which specifically target lipopolysaccharide (LPS) and palmitoyl-oleoyl-phosphatidylglycerol (POPG) are efficient antibiotics. From the core LPS-binding region of Factor C, two 34-mer Sushi peptides, S1 and S3, were derived. S1 functions as a monomer, while S3 is active as a dimer. Both S1 and S3 display detergent-like properties in disrupting LPS aggregates, with specificity for POPG resulting from electrostatic and hydrophobic forces between the peptides and the bacterial lipids. During interaction with POPG, the S1 transitioned from a random coil to an α-helix, while S3 resumed a mixture of α-helix and β-sheet structures. The unsaturated nature of POPG confers fluidity and enhances insertion of the peptides into the lipid bilayer, causing maximal disruption of the bacterial membrane. These parameters should be considered in designing and developing new generations of peptide antibiotics with LPS-neutralizing capability. Received 2 October 2007; received after revision 2 November 2007; accepted 4 December 2007 J. L. Ding, B. Ho: Co-senior authors.     

Lack of pyrogenic tolerance transmission between brain and periphery in the rabbit

Summary Successive injections of lipopolysaccharide (LPS) either intravenously (i.v.) or intracerebroventricularly (i.c.v.) induced pyrogenic tolerance to LPS in rabbits. Tolerance was shown by a decrease of the magnitude of the fever response to repeated doses of LPS, irrespective of the route of pyrogen administration. A significantly greater and more dramatic decrease of the fever index, however, was observed in rabbits made tolerant to pyrogen given i.v. than when the pyrogen was given i.c.v. Transmission of the pyrogenic toleraance between brain and peripheral tissues, however, has not been ascertained.     

Lack of pyrogenic tolerance transmission between brain and periphery in the rabbit

Successive injections of lipopolysaccharide (LPS) either intravenously (i.v.) or intracerebroventricularly (i.c.v.) induced pyrogenic tolerance to LPS in rabbits. Tolerance was shown by a decrease of the magnitude of the fever response to repeated doses of LPS, irrespective of the route of pyrogen administration. A significantly greater and more dramatic decrease of the fever index, however, was observed in rabbits made tolerant to pyrogen given i.v. than when the pyrogen was given i.c.v. Transmission of the pyrogenic tolerance between brain and peripheral tissues, however, has not been ascertained.     

An MD2-derived peptide promotes LPS aggregation,facilitates its internalization in THP-1 cells,and inhibits LPS-induced pro-inflammatory responses

MD2, a 160-residue accessory glycoprotein, is responsible for the recognition and binding of Gram-negative bacterial membrane component, lipopolysaccharide (LPS). Internalization of pathogen inside the mononuclear phagocytes has also been attributed to MD2 which leads to the clearance of pathogens from the host. However, not much is known about the segments in MD2 that are responsible for LPS interaction or internalization of pathogen inside the defense cells. A 16-residue stretch (MD54) from MD2 protein has been identified that possesses a short heptad repeat sequence and four cationic residues enabling it to participate in both hydrophobic and electrostatic interactions with LPS. An MD54 analog of the same size was also designed in which a leucine residue at a heptadic position was replaced with an alanine residue. MD54 but not its analog, MMD54 induced aggregation of LPS and aided in its internalization within THP-1 monocytes. Furthermore, MD54 inhibited LPS-induced nuclear translocation of NF-κB in PMA-treated THP-1 and TLR4/MD2/CD14-transfected HEK-293T cells and the production of pro-inflammatory cytokines. In addition, in in vivo experiments, MD54 showed marked protection and survival of mice against LPS-induced inflammation and death. Overall, we have identified a short peptide with heptad repeat sequence from MD2 that can cause aggregation of LPS and abet in its internalization within THP-1 cells, resulting in attenuation of LPS-induced pro-inflammatory responses in vitro and in vivo.     

Change in sensitivity to lipopolysaccharide during the differentiation of human monocytes to macrophages in vitro

Mononuclear phagocytes in distinct differentiation stages and cultured under different conditions were tested for their sensitivity towards lipopolysaccharide (LPS), using procoagulant activity (PCA) expression and tumor necrosis factor (TNF) production as indices. The response of mature monocyte-derived macrophages differed from that of freshly isolated monocytes 1) by higher levels of constititive PCA, 2) by responding to approximately 1,000-fold lower concentrations of LPS with PCA and TNF production, and 3) by a faster rise in PCA and TNF production. Due to the high constitutive level of PCA expression, the PCA stimulation index for LPS was low in macrophages when compared with that in monocytes. Thus, during differentiation to macrophages, human monocytes acquire increased sensitivity to LPS (2 orders of magnitude more sensitive than a sensitive turbidimetricLimulus amoebocyte lysate assay). This exquisite sensitivity to LPS is expressed regardless of whether LPS is offered in the presence or absence of lipopolysaccharide binding protein-containing serum. This points to as yet uncharacterized pathways of high affinity interaction between LPS and macrophages.     

Platelet aggregation and stimulation of leucocyte procoagulant activity by rickettsial lipopolysaccharides in rabbits and in man

Summary The effects in vitro of 4 purified lipopolysaccharide (LPS) preparations from Rickettsiae on platelets and leucocytes were studied in rabbits and in man. All LPS induced aggregation in rabbit platelet-rich plasma but to differing degrees. This activity was abolished by inactivation of complement. None of the preparations induced aggregation of human platelets. Both rabbit and human leucocytes, when incubated with each of the rickettsial LPS preparations, generated a potent procoagulant activity (tissue factor). These findings add further support to the concept that rickettsial LPS behave as typical LPS from gram-negative bacteria and may be relevant to the understanding of the mechanism(s) responsible for triggering intravascular coagulation in rickettsial diseases.     

Splenic B-cell activation in lipopolysaccharide-non-responsive C3H/HeJ mice by lipopolysaccharide ofPorphyromonas gingivalis

Porphyromonas gingivalis 381 lipopolysaccharide (LPS) definitely exhibited mitogenic activity in purified B-cells, separated from spleens of LPS-responsive C3H/HeN mice and LPS-non-responsive C3H/HeJ mice by using a magnetic cell sorting system. The mitogenic activity induced byP. gingivalis LPS was incompletely inhibited by polymyxin B.P. gingivalis LPS also induced a higher production of interleukin-6 (IL-6) in splenic B-cells of C3H/HeN mice as compared withEscherichia coli LPS. Furthermore,P. gingivalis LPS, but notE. coli LPS, induced definite IL-6 production in C3H/HeJ mice.P. gingivalis LPS increased tyrosine, serine/threonine phosphorylation of proteins with various major induced bands in splenic B-cells of both C3H/HeN and C3H/HeJ mice. Additionally, radioiodinatedP. gingivalis LPS, similarly toE. coli LPS, bound to a 73-kDa protein on C3H/HeJ as well as C3H/HeN B-cells. ThusP. gingivalis LPS may activate B-cells of C3H/HeJ as well as C3H/HeN mice via the LPS-specific binding protein on the cells.     

Course of fever response to repeated administration of sublethal doses of lipopolysaccharides, polyinosinic:polycytidylic acid and muramyl dipeptide to rabbits

The purpose of the present study was to examine the development of tolerance to three structurally dissimilar pyrogens, i.e., lipopolysaccharide (LPS), muramyl dipeptide (MDP) and polyinosinic:polycytidylic acid (poly I:C) in rabbits. The possibility of pyrogenic cross-tolerance among these agents has also been studied. It was observed that repeated injection of sublethal doses of LPS and MDP was connected with the changing of biphasic fever to monophasic. The consequence of this was a drop in the fever index. In contrast to LPS and MDP, the repeated administration of poly I:C did not result in such changes. Successive injections of this pyrogen always evoked biphasic fever. We also demonstrated that pyrogenic cross-tolerance between LPS and MDP did not occur. The cross-tolerance between LPS and MDP did not occur. The cross-tolerance among pyrogens was possible if they originated from the same class, for example endotoxin from Salmonella abortus eq. and endotoxin from Escherichia coli.     

Ikaros negatively regulates inducible nitric oxide synthase expression in macrophages: Involvement of Ikaros phosphorylation by casein kinase 2

Ikaros is known as a critical regulator of lymphocyte development. We examined the regulatory role of Ikaros in LPS/IFN-gamma-induced inducible nitric oxide synthase (iNOS) expression by macrophages. Our results showed that IK6 (Ikaros dominant negative isoform) induction increases the iNOS expression. Ikaros DNA binding activity on the iNOS promoter was decreased, and a mutation of the Ikaros-binding site on the iNOS promoter resulted in an increase in LPS/IFN-gamma-induced iNOS expression. LPS/IFN-gamma increased the histone (H3) acetylation on the Ikaros DNA binding site. These results suggest that Ikaros acts as a negative regulator on iNOS expression. Treatment with a casein kinase 2 (CK2) inhibitor reversed LPS/IFN-gamma-induced decrease in Ikaros DNA binding activity. Moreover, overexpression of kinase-inactive CK2 decreased iNOS expression and a significant amount of CK2alpha1 translocated into the nucleus in LPS/IFN-gamma-treated cells. Overall, these data indicate that LPS/IFN-gamma decreases the Ikaros DNA binding activity via the CK2 pathway, resulting in an increase of iNOS expression.     

Course of fever response to repeated administration of sublethal doses of lipopolysaccharides,polyinosinic: Polycytidylic acid and muramyl dipeptide to rabbits

Summary The purpose of the present study was to examine the development of tolerance to three structurally dissimilar pyrogens, i.e., lipopolysaccharide (LPS), muramyl dipeptide (MDP) and polyinosinic: polycytidylic acid (poly I:C) in rabbits. The possibility of pyrogenic cross-tolerance among these agents has also been studied. It was observed that repeated injection of sublethal doses of LPS and MDP was connected with the changing of biphasic fever to monophasic. The consequence of this was a drop in the fever index. In contrast to LPS and MDP, the repeated administration of poly I:C did not result in such changes. Successive injections of this pyrogen always evoked biphasic fever. We also demonstrated that pyrogenic cross-tolerance between LPS and MDP did not occur. The cross-tolerance between LPS and MDP did not occur. The cross-tolerance among pyrogens was possible if they originated from the same class, for example endotoxin fromSalmonella abortus eq. and endotoxin fromEscherichia coli.     

Form and functions of the regular surface array (S-layer) of Aeromonas salmonicida

The principal virulence factor of Aeromonas salmonicida is its S-layer (A-layer) which is comprised of tetragonally arrayed approximately 50,000 Mr protein subunits tethered to the cell surface via LPS. The detailed composition of its LPS is known, as is the primary sequence, and three-dimensional disposition of the A protein subunits. The A-layer physically protects the cell against bacteriophage, proteases, as well as immune and non-immune complement. The A-layer appears to be uniquely adapted towards binding biologically important molecules such as heme, and to various basement membrane proteins. In addition, the A-layer is required for macrophage infiltration and resistance. Specific mutants containing a disorganized A-layer are avirulent and provide significant protection to salmonids when applied by immersion.     

Lipid transport pathways in mammalian cells

Summary A major deficit in our understanding of membrane biogenesis in eukaryotes is the definition of mechanisms by which the lipid constituents of cell membranes are transported from their sites of intracellular synthesis to the multiplicity of membranes that constitute a typical cell. A variety of approaches have been used to examine the transport of lipids to different organelles. In many cases the development of new methods has been necessary to study the problem. These methods include cytological examination of cells labeled with fluorescent lipid analogs, improved methods of subcellular fractionation, in situ enzymology that demonstrates lipid translocation by changes in lipid structure, and cell-free reconstitution with isolated organelles. Several general patterns of lipid transport have emerged but there does not appear to be a unifying mechanism by which lipids move among different organelles. Significant evidence now exists for vesicular and metabolic energy-dependent mechanisms as well as mechanisms that are clearly independent of cellular ATP content.     

慢性酒精性胰腺炎中CD14、TLR4、TNFα的表达

目的 研究酒精性慢性胰腺炎组织中白细胞分化抗原14(CD14)、钟样受体4(TLR4)、肿瘤坏死因子(TNFα)的表达,探讨酒精性慢性胰腺炎的发病机制.方法 24只1月龄雄性SD大鼠随机分为对照组、脂多糖组、酒精组、酒精联合脂多糖组(以下简称联合组)各6只.酒精组和联合组饲以25％酒精,饮酒12用后联合组和脂多糖组,反复腹腔注射脂多糖2 mg/kg·w,共4次.用免疫组化及RT-PCR检测CD14、TLR4、TNF在各组的表达.结果 酒精组CD14、TLR4和TNF表达较对照组和脂多糖组增加(P＜0.05),联合组CD14、TLR4和TNF表达较对照组、脂多糖组明显增加(P＜0.01),较酒精组增加(P＜0.05).结论 酒精性慢性胰腺炎组织中CD14、TLR4和TNF表达增加,脂多糖通路可能参与了慢性胰腺炎发生发展.     

Die lokale Leukozytenreaktion bei der intraperitonealen Infektion der Maus mitE. coli

Summary Intraperitoneal aggregation of leucocytes is produced in an identical manner by intraperitoneal application of different amounts of living and dead bacteria and bacterial products. The maximum accumulation occurs with medium dosages of bacteria or their products, with high dosages no leucocytic accumulation is produced. Pretreatment with selected LPS of bacteria enhances the intraperitoneal accumulation of leucocytes also with the high amounts of bacteria, whereas the reaction in not pretreated with LPS is suppressed. Parallel to the increase of the intraperitoneal leucocytic accumulation the animals are protected against the infection with high amounts of bacteria, to which they succumb if not pretreated with LPS.     

Superiority of human complement for assaying bacterial lipopolysaccharides by their anticomplementary activity.

In assaying bacterial lipopolysaccharides (LPS) for anticomplementary activity, human complement (C) allowed detection of approximately 200 times smaller amounts of LPS than guinea-pigs C. Pig C was slightly inferior to human.