Bacterial Lipopolysaccharide Research Articles

Exploring the Significance of Gut Microbiota in Diabetes Pathogenesis and Management-A Narrative Review.

Type 2 diabetes is a disease with significant health consequences for the individual. Currently, new mechanisms and therapeutic approaches that may affect this disease are being sought. One of them is the association of type 2 diabetes with microbiota. Through the enteric nervous system and the gut-microbiota axis, the microbiota affects the functioning of the body. It has been proven to have a real impact on influencing glucose and lipid metabolism and insulin sensitivity. With dysbiosis, there is increased bacterial translocation through the disrupted intestinal barrier and increased inflammation in the body. In diabetes, the microbiota's composition is altered with, for example, a more abundant class of Betaproteobacteria. The consequences of these disorders are linked to mechanisms involving short-chain fatty acids, branched-chain amino acids, and bacterial lipopolysaccharide, among others. Interventions focusing on the gut microbiota are gaining traction as a promising approach to diabetes management. Studies are currently being conducted on the effects of the supply of probiotics and prebiotics, as well as fecal microbiota transplantation, on the course of diabetes. Further research will allow us to fully develop our knowledge on the subject and possibly best treat and prevent type 2 diabetes.

In vitro analysis of the expression of inflammasome, antiviral, and immune genes in an Oreochromis niloticus liver cell line following stimulation with bacterial ligands and infection with tilapia lake virus.

The inflammasome is a multimeric protein complex that plays a vital role in the defence against pathogens and is therefore considered an essential component of the innate immune system. In this study, the expression patterns of inflammasome genes (NLRC3, ASC, and CAS-1), antiviral genes (IFNγ and MX), and immune genes (IL-1β and IL-18) were analysed in Oreochromis niloticus liver (ONIL) cells following stimulation with the bacterial ligands peptidoglycan (PGN) and lipopolysaccharide (LPS) and infection with TiLV. The cells were stimulated with PGN and LPS at concentrations of 10, 25, and 50 µg/ml. For viral infection, 106 TCID50 of TiLV per ml was used. After LPS stimulation, all seven genes were found to be expressed at specific time points at each of the three doses tested. However, at even higher doses of LPS, NLRC3 levels decreased. Following TiLV infection, all of the genes showed significant upregulation, especially at early time points. However, the gene expression pattern was found to be unique in PGN-treated cells. For instance, NLRC3 and ASC did not show any response to PGN stimulation, and the expression of IFNγ was downregulated at 25 and 50 µg of PGN per ml. CAS-1 and IL-18 expression was downregulated at 25 µg of PGN per ml. At a higher dose (50 µg/ml), IL-1β showed downregulation. Overall, our results indicate that these genes are involved in the immune response to viral and bacterial infection and that the degree of response is ligand- and dose-dependent.

Influence of nuclear factor κB and adenosine monophosphate-activated protein kinase on the vascular bed of the liver under the conditions of modeling chronic alcoholic hepatitis

Chronic alcohol use activates the transcription factor κB (NF-κB) in liver sinusoidal endothelial cells (LSECs), macrophages and other cells of the liver parenchyma, which controls the regulation of the expression of pro-inflammatory cytokines that activate signaling pathways of immune regulation of liver inflammation and vascular tone. AMP-activated protein kinase (AMPK) is an important immunometabolic regulatory factor in macrophages and, as a metabolic sensor, AMPK in vascular cells may be involved in the regulation of blood flow. The aim of the work was to find out the effect of modulators of the transcription factor κB and AMP-activated protein kinase on the morphometric parameters of the vascular bed of the liver of rats under the conditions of modeling chronic alcoholic hepatitis. Simulation of chronic alcoholic hepatitis leads to a violation of the exchange of metabolites between the central and interlobular veins, which creates prerequisites for the development of hypoxic damage to hepatocytes, as evidenced by a decrease in the diameters of lobular arterioles and venules. Administration of ammonium pyrrolidinedithiocarbamate and bacterial lipopolysaccharide, which affect NF-κB signaling under the conditions of modeling chronic alcoholic hepatitis, helps to restore the exchange of metabolites between the central and interlobular veins, which is evidenced by an increase in the lumens of lobular arterioles and venules. The introduction of phenformin and doxorubicin, which affect AMPK under the conditions of modeling chronic alcoholic hepatitis, prevents the development of hypoxic damage to hepatocytes, as evidenced by an increase in the diameters of lobular arteries and veins, and leads to intensification of interlobular blood circulation.

Extracellular vesicles derived from mesenchymal stem cells ameliorate cognitive impairment caused by neuroinflammation in young but not aged mice

Aim: The aim of this work was to study the effects of extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) on inflammation-impaired cognitive functions and the brain of mice. Methods: Young mice (~3-month-old) and aged mice (~18-month-old) were injected with bacterial lipopolysaccharide (LPS) and obtained intravenously donor 106 human umbilical cord MSCs, EVs isolated from a similar amount of MSCs or conditioned medium (CM) of MSCs. Subsequently, the mice were examined in behavioral tests and the mouse brains were analyzed for the levels of pro-inflammatory cytokines, α7 nicotinic acetylcholine receptors (α7 nAChRs) and amyloid beta 1-42 (Aβ1-42). Results: EVs prevented LPS-induced memory impairment in mice, whereas CM provided a weaker and temporal effect. Both EVs and MSCs injected once after regular injections of LPS stably improved memory of young mice. In contrast, both cells and EVs provided only transient effect in aged mice injected with LPS. The brains of aged LPS-treated mice contained elevated amounts of IL-1β and IL-6; both MSCs and EVs decreased them significantly. The brains of non-treated aged mice contained decreased levels of α7 nAChRs and increased levels of Aβ1-42 and α7-bound Aβ1-42 compared to the brains of young mice. LPS treatment decreased α7 nAChRs in both young and aged mice, while both MSCs and EVs restored them up to the control level. In young mice, LPS treatment increased the level of Aβ1-42 and α7-bound Aβ1-42, whereas MSCs and EVs decreased it. In contrast, neither LPS nor MSCs/EVs influenced the elevated level of Aβ1-42 but increased α7-bound Aβ1-42 in the brains of aged mice. Conclusions: Regenerative potential of MSCs and MSC-derived EVs is sufficient to support cognitive functions of LPS-treated young mice but is quite poor for aged animals, possibly, due to decreased levels of α7 nAChRs and accumulated Aβ1-42 in their brains.

NP-12 peptide functionalized nanoparticles counteract the effect of bacterial lipopolysaccharide on cultured osteoblasts

ObjectiveTo evaluate whether nanoparticles (NPs) functionalized with Tideglusib (TDg, NP-12), and deposited on titanium surfaces, would counteract the effect of bacterial lipopolysaccharide (LPS) on osteoblasts. MethodsExperimental groups were: (a) Titanium discs (TiD), (b) TiD covered with undoped NPs (Un-NPs) and (c) TiD covered with TDg-doped NPs (TDg-NPs). Human primary osteoblasts were cultured onto these discs, in the presence or absence of bacterial LPS. Cell proliferation was assessed by MTT-assay and differentiation by measuring the alkaline phosphatase activity. Mineral nodule formation was assessed by the alizarin red test. Real-time quantitative polymerase chain reaction was used to study the expression of Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-β1, VEGF, TGF-βR1, TGF-βR2, and TGF-βR3 genes. Osteoblasts morphology was studied by Scanning Electron Microscopy. One-way ANOVA or Kruskal-Wallis and Bonferroni multiple comparisons tests were carried out (p < 0.05). ResultsTDg-NPs enhanced osteoblasts proliferation. Similarly, this group increased ALP production and mineral nodules formation. TDg-NPs on titanium discs resulted in overexpression of the proliferative genes, OSC and OSX, regardless of LPS activity. In the absence of LPS, TDg-NPs up-regulated Runx2, COL-I, ALP, BMP2 and BMP7 genes. OPG/RANKL gene ratios were increased about 2500 and 4,000-fold by TDg-NPs, when LPS was added or not, respectively. In contact with the TDg-NPs osteoblasts demonstrated an elongated spindle-shaped morphology with extracellular matrix production. SignificanceTDg-NPs on titanium discs counteracted the detrimental effect of LPS by preventing the decrease on osteoblasts proliferation and mineralization, and produced an overexpression of proliferative and bone-promoting genes on human primary osteoblasts.

A Simple and Rapid Microscale Method for Isolating Bacterial Lipopolysaccharides.

Bacterial endotoxins (lipopolysaccharides (LPSs)) are important mediators of inflammatory processes induced by Gram-negative microorganisms. LPSs are the key inducers of septic shock due to a Gram-negative bacterial infection; thus, the structure and functions of LPSs are of specific interest. Often, highly purified bacterial endotoxins must be isolated from small amounts of biological material. Each of the currently available methods for LPS extraction has certain limitations. Herein, we describe a rapid and simple microscale method for extracting LPSs. The method consists of the following steps: ultrasonic destruction of the bacterial material, LPS extraction via heating, LPS purification with organic solvents, and treatment with proteinase K. LPSs that were extracted by using this method contained less than 2-3% protein and 1% total nucleic acid. We also demonstrated the structural integrity of the O-antigen and lipid A via the sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) methods, respectively. We demonstrated the ability of the extracted LPSs to induce typical secretion of cytokines and chemokines by primary macrophages. Overall, this method may be used to isolate purified LPSs with preserved structures of both the O-antigen and lipid A and unchanged functional activity from small amounts of bacterial biomass.

Effect of dietary macronutrients and immune challenge on gut microbiota, physiology and feeding behaviour in zebra finches.

Macronutrients play a vital role in host immunity and can influence host-pathogen dynamics, potentially through dietary effects on gut microbiota. To increase our understanding of how dietary macronutrients affect physiology and gut microbiota and investigate whether feeding behaviour is influenced by an immune threat, we conducted two experiments. First, we determined whether zebra finches (Taeniopygia guttata) exhibit shifts in physiology and gut microbiota when fed diets differing in macronutrient ratios. We found the type and amount of diet consumed affected gut microbiota alpha diversity, where microbial richness and Shannon diversity increased with caloric intake in birds fed a high-fat diet and decreased with caloric intake in birds fed a high protein diet. Diet macronutrient content did not affect physiological metrics, but lower caloric intake was associated with higher complement activity. In our second experiment, we simulated an infection in birds using the bacterial endotoxin lipopolysaccharide (LPS) and quantified feeding behaviour in immune challenged and control individuals, as well as birds housed near either a control pair (no immune threat), or birds housed near a pair given an immune challenge with LPS (social cue of heightened infection risk). We also examined whether social cues of infection alter physiological responses relevant to responding to an immune threat, an effect that could be mediated through shifts in feeding behaviour. LPS induced a reduction in caloric intake driven by a decrease in protein, but not fat consumption. No evidence was found for socially induced shifts in feeding behaviour, physiology or gut microbiota. Our findings carry implications for host health, as sickness-induced anorexia and diet-induced shifts in the microbiome could shape host-pathogen interactions.

Poking holes in the blood-brain barrier

Bacterial lipopolysaccharide (LPS) is implicated in disrupting the blood-brain barrier (BBB). In a recent issue of Nature, Wei etal. now show that LPS activates the inflammatory caspases (4, 5, and 11) and gasdermin D (GSDMD) in brain endothelial cells, which triggers their pyroptotic cell death and disrupts the BBB.

Risk factors for severe COVID-19 disease increase SARS-CoV-2 infectivity of endothelial cells and pericytes.

Coronavirus disease 2019 (COVID-19) was initially considered a primarily respiratory disease but is now known to affect other organs including the heart and brain. A major route by which COVID-19 impacts different organs is via the vascular system. We studied the impact of apolipoprotein E (APOE) genotype and inflammation on vascular infectivity by pseudo-typed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viruses in mouse and human cultured endothelial cells and pericytes. Possessing the APOE4 allele or having existing systemic inflammation is known to enhance the severity of COVID-19. Using targeted replacement human APOE3 and APOE4 mice and inflammation induced by bacterial lipopolysaccharide (LPS), we investigated infection by SARS-CoV-2. Here, we show that infectivity was higher in murine cerebrovascular pericytes compared to endothelial cells and higher in cultures expressing APOE4. Furthermore, increasing the inflammatory state of the cells by prior incubation with LPS increased infectivity into human and mouse pericytes and human endothelial cells. Our findings provide insights into the mechanisms underlying severe COVID-19 infection, highlighting how risk factors such as APOE4 genotype and prior inflammation may exacerbate disease severity by augmenting the virus's ability to infect vascular cells.

Bacterial sepsis triggers stronger transcriptomic immune responses in larger primates.

Empirical data relating body mass to immune defence against infections remain limited. Although the metabolic theory of ecology predicts that larger organisms would have weaker immune responses, recent studies have suggested that the opposite may be true. These discoveries have led to the safety factor hypothesis, which proposes that larger organisms have evolved stronger immune defences because they carry greater risks of exposure to pathogens and parasites. In this study, we simulated sepsis by exposing blood from nine primate species to a bacterial lipopolysaccharide (LPS), measured the relative expression of immune and other genes using RNAseq, and fitted phylogenetic models to determine how gene expression was related to body mass. In contrast to non-immune-annotated genes, we discovered hypermetric scaling in the LPS-induced expression of innate immune genes, such that large primates had a disproportionately greater increase in gene expression of immune genes compared to small primates. Hypermetric immune gene expression appears to support the safety factor hypothesis, though this pattern may represent a balanced evolutionary mechanism to compensate for lower per-transcript immunological effectiveness. This study contributes to the growing body of immune allometry research, highlighting its importance in understanding the complex interplay between body size and immunity over evolutionary timescales.

The In Vitro Antioxidant and Anti-Inflammatory Activities of Selected Australian Seagrasses.

Recent studies have shown that seagrasses could possess potential applications in the treatment of inflammatory disorders. Five seagrass species (Zostera muelleri, Halodule uninervis, Cymodocea rotundata, Syringodium isoetifolium, and Thalassia hemprichii) from the Great Barrier Reef (QLD, Australia) were thus collected, and their preliminary antioxidant and anti-inflammatory activities were evaluated. From the acetone extracts of five seagrass species subjected to 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging antioxidant assay, the extract of Z. muelleri had the highest activity (half minimal concentration of inhibition (IC50) = 138 µg/mL), with the aerial parts (IC50 = 119 µg/mL) possessing significantly higher antioxidant activity than the roots (IC50 ≥ 500 µg/mL). A human peripheral blood mononuclear cells (PBMCs) assay with bacterial lipopolysaccharide (LPS) activation and LEGENDplex cytokine analysis showed that the aerial extract of Z. muelleri significantly reduced the levels of inflammatory cytokines tumour necrosis factor alpha (TNF-α), interleukin (IL)-1β, and IL-6 by 29%, 74%, and 90%, respectively, relative to the LPS treatment group. The aerial extract was thus fractionated with methanol (MeOH) and hexane fraction, and purification of the MeOH fraction by HPLC led to the isolation of 4-hydroxybenzoic acid (1), luteolin (2), and apigenin (3) as its major constituents. These compounds have been previously shown to reduce levels of TNF-α, IL-1β, and IL-6 and represent some of the major bioactive components of Z. muelleri aerial parts. This investigation represents the first study of the antioxidant and anti-inflammatory properties of Z. muelleri and the first isolation of small molecules from this species. These results highlight the potential for using seagrasses in treating inflammation and the need for further investigation.

Glaucoma and the Human Microbiome.

To explore a view of the human microbiome as an interconnected, functional, dynamic system that may be linked to the pathogenesis and progression of glaucoma. A literature review was undertaken that included publications from 1966 to 2023. Bacterial lipopolysaccharides (LPS) activate toll-like receptors (TLR) and mediate the human immune response. The LPS-TLR4 pathway is a potential avenue for the ocular, gut, and oral microbiomes to interface and/or influence ocular disease. Studies of gut dysbiosis have shown that alterations in the healthy microbiota can predispose the host to immune-mediated inflammatory and neurodegenerative conditions, while oral and ocular surface dysbiosis has been correlated with glaucoma. While developmental exposure to commensal microflora has shown to be necessary for the autoimmune and neurodegenerative responses to elevated intraocular pressure to take place, commensal bacterial products like short-chain fatty acids have regulatory effects protective against glaucoma. Alterations to human microbiotas have been associated with changes in intestinal permeability, gene regulation, immune cell differentiation, and neural functioning, which may predispose the host to glaucoma. Select microbes have been highlighted for their potential contributions to glaucoma disease progression or protection, raising the potential for microbiota-based treatment modalities. Current topical glaucoma treatments may disrupt the ocular surface microbiota, potentially having ramifications on host health. Further study of the relationships between human microbiome and glaucoma is needed.

Effects of Akt Activator SC79 on Human M0 Macrophage Phagocytosis and Cytokine Production.

Akt is an important kinase in metabolism. Akt also phosphorylates and activates endothelial and neuronal nitric oxide (NO) synthases (eNOS and nNOS, respectively) expressed in M0 (unpolarized) macrophages. We showed that e/nNOS NO production downstream of bitter taste receptors enhances macrophage phagocytosis. In airway epithelial cells, we also showed that the activation of Akt by a small molecule (SC79) enhances NO production and increases levels of nuclear Nrf2, which reduces IL-8 transcription during concomitant stimulation with Toll-like receptor (TLR) 5 agonist flagellin. We hypothesized that SC79's production of NO in macrophages might likewise enhance phagocytosis and reduce the transcription of some pro-inflammatory cytokines. Using live cell imaging of fluorescent biosensors and indicator dyes, we found that SC79 induces Akt activation, NO production, and downstream cGMP production in primary human M0 macrophages. This was accompanied by a reduction in IL-6, IL-8, and IL-12 production during concomitant stimulation with bacterial lipopolysaccharide, an agonist of pattern recognition receptors including TLR4. Pharmacological inhibitors suggested that this effect was dependent on Akt and Nrf2. Together, these data suggest that several macrophage immune pathways are regulated by SC79 via Akt. A small-molecule Akt activator may be useful in some infection settings, warranting future in vivo studies.

#1995 Runt-related Transcription Factor 1 (RUNX1) is a mediator of acute kidney injury

Abstract Background and Aims A better understanding of the pathogenesis of acute kidney injury (AKI) may lead to new therapeutic approaches. Kidney transcriptomics analyses of murine folic acid-induced AKI (FA-AKI) identified Runx1 as the most upregulated RUNX family gene. RUNX1 is a key regulator of hematopoiesis, associated to leukemias and other non-immunological cancers and modulates myoblast proliferation during muscle regeneration and cardiac remodelling after myocardial infarction. In kidney diseases, RUNX1 is overexpressed in polycystic kidney disease and kidney cancer and favours fibrosis. Method We examined the expression of RUNX1 in folic acid-AKI (FA-AKI), in bacterial lipopolysaccharide (LPS)-induced cytokine storm-AKI (CS-AKI) and in human AKI. For this, female 12- to 14-week-old C57BL/6J wild type (WT) mice, received a single intraperitoneal (i.p) injection of acid folic (250 mg/kg), LPS (5 mg/kg) or vehicle. In cultured murine tubular MCT cells, we explored the expression and role of RUNX1 in response to the cytokine TWEAK or LPS. The chemical inhibitor of RUNX1 Ro5-3335 and a specific small interfering RNA (siRNA) against RUNX1 were used in cultured cells to explore the role of RUNX1 in TWEAK and LPS-induced inflammation. Ro5-3335 was also used in murine AKI (FA-AKI and CS-AKI) to test its potential as a therapeutic target. In all experimental models, plasma samples were collected to assess renal function. Kidneys were collected for RNA (RT-PCR), protein (Western blot, ELISA, immunohistochemistry) and histopathologic studies. Results Kidney transcriptomics identified Runx1 as the most upregulated Runx gene in FA-AKI at 24 h and the most overactive transcription factor in upstream regulator analysis. RUNX1 overexpression in FA-AKI was validated at mRNA and protein levels and localized mainly to tubular cell nuclei. CS-AKI also upregulated kidney RUNX1 in nuclei. In a transcriptomics array of murine tubular cells stimulated with TWEAK for 6 h, Runx1 was upregulated compared with non-stimulated cells, while Runx2 and Runx3 were not. RT-PCR confirmed an early upregulation of Runx1 (3 h) followed by a progressive return to baseline by 24 h. TWEAK also increased total and nuclear RUNX1 protein levels from 6 h onwards. Since kidney RUNX1 is also increased in CS-AKI, we tested whether LPS regulated RUNX1 in cultured tubular cells. LPS transiently increased Runx1 mRNA expression and it also increased whole cell and nuclear RUNX1 protein levels. Mechanistically, RUNX1 bound to the Il-6 gene promoter in response to TWEAK and LPS, and RUNX1 targeting with the chemical inhibitor Ro5-3335 or a specific siRNA prevented the TWEAK- and LPS-induced upregulation of IL-6. NFκB1 p50 pathway is involved in the RUNX1/IL-6 axis since the specific p50 inhibitor SN50 decreased the nuclear accumulation of RUNX1 and IL-6 expression in TWEAK- or LPS-stimulated MCT cells. In vivo, preventive Ro5-3335 improved kidney function and reduced inflammation in FA-AKI and CS-AKI. Moreover, in FA-AKI, Ro5-3335 also prevented the expression of profibrotic genes. However, Ro5-3335 administration after the insult only improved kidney function in CS-AKI. Kidney transcriptomics identified inflammatory genes and transcription factors such as Yap1 and p53 as key targets of Ro5-3335 in CS-AKI. Conclusion RUNX1 contributes to AKI by driving the expression of genes involved in inflammation and represents a novel therapeutic target in AKI.

Juvenile peripheral LPS exposure overrides female resilience to prenatal VPA effects on adult sociability in mice

Autism spectrum disorder (ASD) exhibits a gender bias, with boys more frequently affected than girls. Similarly, in mouse models induced by prenatal exposure to valproic acid (VPA), males typically display reduced sociability, while females are less affected. Although both males and females exhibit VPA effects on neuroinflammatory parameters, these effects are sex-specific. Notably, females exposed to VPA show increased microglia and astrocyte density during the juvenile period. We hypothesized that these distinct neuroinflammatory patterns contribute to the resilience of females to VPA. To investigate this hypothesis, we treated juvenile animals with intraperitoneal bacterial lipopolysaccharides (LPS), a treatment known to elicit brain neuroinflammation. We thus evaluated the impact of juvenile LPS-induced inflammation on adult sociability and neuroinflammation in female mice prenatally exposed to VPA. Our results demonstrate that VPA-LPS females exhibit social deficits in adulthood, overriding the resilience observed in VPA-saline littermates. Repetitive behavior and anxiety levels were not affected by either treatment. We also evaluated whether the effect on sociability was accompanied by heightened neuroinflammation in the cerebellum and hippocampus. Surprisingly, we observed reduced astrocyte and microglia density in the cerebellum of VPA-LPS animals. These findings shed light on the complex interactions between prenatal insults, juvenile inflammatory stimuli, and sex-specific vulnerability in ASD-related social deficits, providing insights into potential therapeutic interventions for ASD.

Zerumbone exhibits anti-inflammatory effects by suppressing eicosanoid signaling: Evidence from LPS-induced peripheral blood leukocytes

Zerumbone, a sesquiterpene isolated from Zingiber zerumbet, has many bioactivities, exhibiting anti-inflammatory properties. However, the effect of zerumbone on the eicosanoid signaling pathway has yet to be examined. Here, we deciphered the anti-eicosanoid properties of zerumbone isolated from ginger. The molecular interaction between zerumbone and eicosanoid metabolizing enzymes (COX-2, 5-LOX, FLAP, and LTA4-hydrolase) and receptors (EP-4, BLT-1, and ICAM-1) along with NOS-2 were assessed using Auto-Dock 4.2 and visualized by chimera and Liggplot+ software. Further, the leukocytes were treated with zerumbone (1–20 μM) and activated using bacterial lipopolysaccharide (LPS-10 nM). The oxidative stress (OS) markers, antioxidant enzymes, and the eicosanoid pathway mediators such as COX-2, 5-LOX, BLT-1, and EP-4 were assessed. The molecular interaction of zerumbone with eicosanoids showed a higher binding affinity with mPGES-1, followed by NOS-2, FLAP, COX-2, LTA-4-hydrolase, and BLT-1. The concentration of 5 μM zerumbone effectively prevented the generation of reactive oxygen species (ROS) and nitric oxide (NO). Likewise, zerumbone significantly (p<0.05) inhibited COX-2, 5-LOX, NOS-2, EP-4, BLT-1, and ICAM-1 expression in LPS-induced peripheral blood leukocytes from rats. Further, the zerumbone treatment on the human PBMCs activated with LPS showed significant inhibition in the expression of ICAM1, COX-2, 5-LOX, and the generation of inflammatory cytokines compared to the control. Overall, the data presented infers that zerumbone positively modulates critical enzymes and receptors of eicosanoids in leukocytes activated with lipopolysaccharides. Thus, zerumbone can be a potential anti-eicosanoid drug in managing inflammation.

Top-Down Characterization of Bacterial Lipopolysaccharides and Lipooligosaccharides Using Activated-Electron Photodetachment Mass Spectrometry.

Lipopolysaccharides (LPS) and lipooligosaccharides (LOS) are located in the outer membrane of Gram-negative bacteria and are comprised of three distinctive parts: lipid A, core oligosaccharide (OS), and O-antigen. The structure of each region influences bacterial stability, toxicity, and pathogenesis. Here, we highlight the use of targeted activated-electron photodetachment (a-EPD) tandem mass spectrometry to characterize LPS and LOS from two crucial players in the human gut microbiota, Escherichia coli Nissle and Bacteroides fragilis. a-EPD is a hybrid activation method that uses ultraviolet photoirradiation to generate charge-reduced radical ions followed by collisional activation to produce informative fragmentation patterns. We benchmark the a-EPD method for top-down characterization of triacyl LOS from E. coli R2, then focus on characterization of LPS from E. coli Nissle and B. fragilis. Notably, a-EPD affords extensive fragmentation throughout the backbone of the core OS and O-antigen regions of LPS from E. coli Nissle. This hybrid approach facilitated the elucidation of structural details for LPS from B. fragilis, revealing a putative hexuronic acid (HexA) conjugated to lipid A.

Hide-and-sick: How bacteria manipulate a neural circuit that makes you sick

Hide-and-sick: How bacteria manipulate a neural circuit that makes you sick

Lysophosphatidylcholine Acetyltransferase 2 (LPCAT2) Influences the Gene Expression of the Lipopolysaccharide Receptor Complex in Infected RAW264.7 Macrophages, Depending on the E. coli Lipopolysaccharide Serotype.

Escherichia coli (E. coli) is a frequent gram-negative bacterium that causes nosocomial infections, affecting more than 100 million patients annually worldwide. Bacterial lipopolysaccharide (LPS) from E. coli binds to toll-like receptor 4 (TLR4) and its co-receptor's cluster of differentiation protein 14 (CD14) and myeloid differentiation factor 2 (MD2), collectively known as the LPS receptor complex. LPCAT2 participates in lipid-raft assembly by phospholipid remodelling. Previous research has proven that LPCAT2 co-localises in lipid rafts with TLR4 and regulates macrophage inflammatory response. However, no published evidence exists of the influence of LPCAT2 on the gene expression of the LPS receptor complex induced by smooth or rough bacterial serotypes. We used RAW264.7-a commonly used experimental murine macrophage model-to study the effects of LPCAT2 on the LPS receptor complex by transiently silencing the LPCAT2 gene, infecting the macrophages with either smooth or rough LPS, and quantifying gene expression. LPCAT2 only significantly affected the gene expression of the LPS receptor complex in macrophages infected with smooth LPS. This study provides novel evidence that the influence of LPCAT2 on macrophage inflammatory response to bacterial infection depends on the LPS serotype, and it supports previous evidence that LPCAT2 regulates inflammatory response by modulating protein translocation to lipid rafts.

Abstract 3048: The Role Of 17:0(2r-oh) Ceramide In Lps Activated Macrophages

Macrophages play a crucial role in the initiation and progression of atherosclerosis, contributing to the buildup of fatty deposits which results in the plaque formation in the walls of arteries. Macrophages modulate the mediators of inflammation and immune response in atherosclerosis. Bacterial lipopolysaccharide (LPS) exerts significant effects on these macrophages including secretion of pro-inflammatory cytokines. The current study is designed to examine the role of sphingosine intermediate N-(2'-(R)-hydroxyheptadecanoyl)-D-erythro-sphingosine (17:0(2R-OH) in the macrophage’s inflammation potential. Couple of studies have indicated a protective role for sphingosine however 17:0(2R-OH) didn’t receive adequate attention. Study design: Mouse J774 macrophages were cultured in Dulbecco’s Modified Eagle’s medium with 4.5g/L glucose and sodium pyruvate without L-glutamine containing 10% fetal bovine serum and 1% penicillin/streptomycin in a humidified atmosphere with 5% CO 2 at 37 °C. The cells were treated in 6-well plates at a density of 5.5х10 6 cells/well. Sphingosine was dissolved in DMSO prior to the treatment. The cells were treated with the 17:0(2R-OH) with 3 different concentrations (5, 25, 50 μg) in duplicates with or without LPS except for the controls. The cells and media were collected at 3 time points (12,24,36) hours. Cells were harvested in Trizol reagent, RNA was then extracted from cells and genes expression were determined in RTPCR using SYBR Green. Results and Conclusion: Our initial microscopic observations suggest that 17:0(2R-OH) potentially modulates inflammation in macrophages, this appears to be dose-dependent. 17:0(2R-OH) has been identified in marine and freshwater fish, marine oils, seaweed, sea urchins, sea cucumbers and may have dietary benefits.