Increased Lipopolysaccharide Levels Research Articles

Gut-derived bacterial LPS attenuates incubation of methamphetamine craving via modulating microglia

BackgroundThe microbiota-gut-brain axis plays a critical role in the pathophysiology of neuropsychiatric disorders, and the compositions of gut microbiota are altered by addictive drugs. However, the role of gut microbiota in the incubation of methamphetamine (METH) craving remains poorly understood. Methods16S rRNA gene sequencing was performed to assess the richness and diversity of gut microbiota in METH self-administration model. Hematoxylin and eosin staining was performed to evaluate the integrity of intestinal barrier. Immunofluorescence and three-dimensional reconstruction were performed to assess the morphologic changes of microglia. Serum levels of lipopolysaccharide (LPS) were determined using the rat enzyme-linked immunosorbent assay kits. Quantitative real-time PCR was performed to assess transcript levels of dopamine receptor, glutamate ionotropic AMPA receptor 3 and brain-derived neurotrophic factor. ResultsMETH self-administration induced gut microbiota dysbiosis, intestinal barrier damage and microglia activation in the nucleus accumbens core (NAcc), which was partially recovered after prolonged withdrawal. Microbiota depletion via antibiotic treatment increased LPS levels and induced a marked change in the microglial morphology in the NAcc, as indicated by the decreases in the lengths and numbers of microglial branches. Depleting the gut microbiota also prevented the incubation of METH craving and increased the population of Klebsiella oxytoca. Furthermore, Klebsiella oxytoca treatment or exogenous administration of the gram-negative bacterial cell wall component LPS increased serum and central LPS levels, induced microglial morphological changes and reduced the dopamine receptor transcription in the NAcc. Both treatments and NAcc microinjections of gut-derived bacterial LPS significantly decreased METH craving after prolonged withdrawal. ConclusionsThese data suggest that LPS from gut gram-negative bacteria may enter circulating blood, activate microglia in the brain and consequently decrease METH craving after withdrawal, which may have important implications for novel strategies to prevent METH addiction and relapse.

Long-term LDR exposure may induce cognitive impairments: A possible association through targeting gut microbiota-gut-brain axis

Environmental and occupational low-dose radiation (LDR) exposure may be harmful for health but the previous reports regarding effect of LDR on cognition are contradictory. Here we investigated the effect of long-term LDR exposure on cognition. In this study, male Balb/c mice’ cognitive functions were tested at 15 weeks after being exposed to 0.5 Gy LDR in 10 fractions at each dose of 0.05 Gy. The results demonstrated that long-term LDR exposure increases escape latency and the time spent in finding exits in mice compared with non LDR exposure. Meanwhile, the inflammation-related proteins including NFκB and p38 also increased. Lipopolysaccharide (LPS) increased and short-chain fatty acid (SCFA) levels decreased following long term LDR exposure. Treatment with microbiota-derived LPS and SCFAs reversed these effects in mice. Furthermore, the gut barrier integrity was damaged in a time-dependent manner with the decreased expression of intestinal epithelial-related biomarkers such as ZO-1 and occludin. Mechanistically, long after exposure to LDR, increased LPS levels may cause cognitive impairment through the regulation of Akt/mTOR signaling in the mouse hippocampus. These findings provide new insight into the clinical applications of LDR and suggest that the gut microbiota–plasma LPS and SCFAs–brain axis may underlie long-term LDR-induced cognition effects.

Gut microbiota changes and biological mechanism in hepatocellular carcinoma after transarterial chemoembolization treatment.

Background and aimsIntestinal flora is closely associated with the occurrence and development of hepatocellular carcinoma (HCC). However, gut microbial changes and biological mechanisms in HCC after transarterial chemoembolization (TACE) treatment are rarely reported.MethodsWe evaluated changes in intestinal flora after TACE in rabbit HCC models and assessed the impact of these changes on the disease. Twenty-four rabbit VX2 HCC models were established and intestinal flora structures, intestinal barrier function, changes in blood lipopolysaccharide (LPS) levels, Toll-like receptor 4 (TLR4), Cyclooxygenase-2 (COX-2), and p-signal transducer and activator of transcription 3(p-STAT3) protein expression levels were studied after TACE treatment.ResultsCompared with healthy rabbits, the intestinal flora in HCC models exhibited structural changes; intestinal barrier function was decreased, and increased LPS levels entered the circulation. A short-term follow-up after TACE showed the procedure partially reversed the intestinal microflora disorder caused by the tumor: intestinal barrier and liver functions were improved, intestinal LPS levels in the blood were reduced, and liver metabolism toward LPS was enhanced. Correlation analyses of the first 75 significantly changed bacteria with clinical factors showed that harmful bacteria had decreased and beneficial bacteria increased. Blood LPS levels and downstream signaling molecule TLR4, COX-2, and p-STAT3 protein expression levels were reduced, which correlated with tumor drug resistance and invasion capabilities.ConclusionsWe first characterized gut microbiota changes and biological mechanisms in HCC after TACE treatment. Our data provide a theoretical research basis for TACE combined with an intestinal flora intervention and systemic chemotherapy.

Lipopolysaccharide from the commensal microbiota of the breast enhances cancer growth: role of S100A7 and TLR4.

The role of commensal bacterial microbiota in the pathogenesis of human malignancies has been a research field of incomparable progress in recent years. Although breast tissue is commonly assumed to be sterile, recent studies suggest that human breast tissue may contain a bacterial microbiota. In this study, we used an immune‐competent orthotopic breast cancer mouse model to explore the existence of a unique and independent bacterial microbiota in breast tumors. We observed some similarities in breast cancer microbiota with skin; however, breast tumor microbiota was mainly enriched with Gram‐negative bacteria, serving as a primary source of lipopolysaccharide (LPS). In addition, dextran sulfate sodium (DSS) treatment in late‐stage tumor lesions increased LPS levels in the breast tissue environment. We also discovered an increased expression of S100A7 and low level of TLR4 in late‐stage tumors with or without DSS as compared to early‐stage tumor lesions. The treatment of breast cancer cells with LPS increased the expression of S100A7 in breast cancer cells in vitro. Furthermore, S100A7 overexpression downregulated TLR4 and upregulated RAGE expression in breast cancer cells. Analysis of human breast cancer samples also highlighted the inverse correlation between S100A7 and TLR4 expression. Overall, these findings suggest that the commensal microbiota of breast tissue may enhance breast tumor burden through a novel LPS/S100A7/TLR4/RAGE signaling axis.

Transplantation of fecal microbiota from patients with inflammatory bowel disease and depression alters immune response and behavior in recipient mice

Gut dysbiosis is closely associated with the occurrence of inflammatory bowel disease (IBD) and psychiatric disorder. Here, to understand the difference of gut microbiota composition and physiological effect between IBD patients with (IBD/D+) or without depression (IBD/D−), we analyzed the fecal microbiota composition of patients with IBD with (/D+) or without depression (/D−) and healthy volunteers (HVs) and examined the effects of these fecal microbiota transplantations (FMTs) on the occurrence of systemic inflammation and anxiety/depression in mice. FMTs from patients with IBD/D+ or IBD/D− caused IBD-like colitis in the transplanted mice: they increased the myeloperoxidase activity, IL-1β and IL-6 expression, and NF-κB+/CD11c+ cell population in the colon. Transplantation of the IBD/D+ patient feces (IBD/D+-F) caused IBD-like colitis more strongly than that of IBD/D−-F. FMTs from patients with IBD/D+ also caused anxiety-/depression-like behaviors, increased the NF-κB+/Iba1+ and lipopolysaccharide (LPS)+/Iba1+ cell populations, and decreased the BDNF+/NeuN+ cell population in the hippocampus. They increased LPS levels in the blood. FMTs from patients with IBD/D− caused anxiety-like, but not depression-like, behaviors. α-/β-diversities and composition of gut microbiota in IBD-F were different from those of HV feces (HV-F). The Enterobacteriaceae and Enterococcaceae populations and LPS levels were higher in the IBD-F than in the HV-F. The Enterococcaceae population was higher in IBD/D+-F vs. IBD/D−-F. However, the transplantation of HV-F into mice previously transplanted with IBD/D+-F significantly reduced depression-like behaviors, NF-κB+/Iba1+ and LPS+/Iba1+ cell populations in the hippocampus, LPS levels in the feces and blood, and IL-1β expression in the colon. These findings suggest that the outbreak of depression/anxiety may be dependent on the systemic inflammation with a leaky gut through the gut dysbiosis-attributable overproduction of bacterial LPS and suppression of tight junction protein expression in patients with IBD.

Increased LPS levels coexist with systemic inflammation and result in monocyte activation in severe COVID-19 patients

Mucosal barrier alterations may play a role in the pathogenesis of several diseases, including COVID-19. In this study we evaluate the association between bacterial translocation markers and systemic inflammation at the earliest time-point after hospitalization and at the last 72 h of hospitalization in survivors and non-survivors COVID-19 patients. Sixty-six SARS-CoV-2 RT-PCR positive patients and nine non-COVID-19 pneumonia controls were admitted in this study. Blood samples were collected at hospital admission (T1) (Controls and COVID-19 patients) and 0–72 h before hospital discharge (T2, alive or dead) to analyze systemic cytokines and chemokines, lipopolysaccharide (LPS) concentrations and soluble CD14 (sCD14) levels. THP-1 human monocytic cell line was incubated with plasma from survivors and non-survivors COVID-19 patients and their phenotype, activation status, TLR4, and chemokine receptors were analyzed by flow cytometry. COVID-19 patients presented higher IL-6, IFN-γ, TNF-α, TGF-β1, CCL2/MCP-1, CCL4/MIP-1β, and CCL5/RANTES levels than controls. Moreover, LPS and sCD14 were higher at hospital admission in SARS-CoV-2-infected patients. Non-survivors COVID-19 patients had increased LPS levels concomitant with higher IL-6, TNF-α, CCL2/MCP-1, and CCL5/RANTES levels at T2. Increased expression of CD16 and CCR5 were identified in THP-1 cells incubated with the plasma of survivor patients obtained at T2. The incubation of THP-1 with T2 plasma of non-survivors COVID-19 leads to higher TLR4, CCR2, CCR5, CCR7, and CD69 expression. In conclusion, the coexistence of increased microbial translocation and hyperinflammation in patients with severe COVID-19 may lead to higher monocyte activation, which may be associated with worsening outcomes, such as death.

The concomitant use of cannabis and cocaine coexists with increased LPS levels and systemic inflammation in male drug users

Illicit drug use can cause a variety of effects including alterations in the immune system. The aim of this study was to investigate the effects of illicit drugs on circulating lipopolysaccharide (LPS), systemic inflammation and oxidative stress markers in drug users. We evaluated the levels of soluble CD14 (sCD14), LPS, inflammatory (TNF-α and IL-6) and regulatory (IL-10) cytokines, as well as C-reactive protein (CRP), lipid peroxidation (TBARS) and total thiols in the peripheral blood of 81 men included in groups of cannabis (n = 21), cocaine (n = 12), cannabis-plus-cocaine users (n = 27), and non-drug users (n = 21). The use of cannabis plus cocaine leads to higher systemic levels of LPS, CRP, IL-6 and higher IL-6/IL-10 ratio, characterizing a proinflammatory profile. In contrast, a regulatory profile as viewed by lower systemic TNF-α and IL-6 levels and lower TNF-α/IL-10 ratio were observed in cannabis users compared to the control group. Moreover, cocaine users presented a lower content of non-enzymatic antioxidant thiol compared to control group, cannabis group and cannabis plus cocaine group. In conclusion, our results indicate that the use of cannabis contributes to an anti-inflammatory/or regulatory profile while the concomitant cannabis plus cocaine consumption coexists with increased circulating amounts of LPS and proinflammatory status.

Alteration of Intestinal Microbiota in 3-Deoxyglucosone-Induced Prediabetic Rats.

Our previous research suggests that 3-deoxyglucosone (3DG), formed in the caramelization course and Maillard reactions in food, is an independent factor for the development of prediabetes. Since the relationship between type 2 diabetes (T2D) and intestinal microbiota is moving from correlation to causality, we investigated the alterations in the composition and function of the intestinal microbiota in 3DG-induced prediabetic rats. Rats were given 50 mg/kg 3DG by intragastric administration for two weeks. Microbial profiling in faeces samples was determined through the 16S rRNA gene sequence. The glucagon-like peptide 2 (GLP-2) and lipopolysaccharide (LPS) levels in plasma and intestinal tissues were measured by ELISA and Limulus test, respectively. 3DG treatment did not significantly change the richness and evenness but affected the composition of intestinal microbiota. At the phylum level, 3DG treatment increased the abundance of nondominant bacteria Proteobacteria but did not cause the change of the dominant bacteria. Meanwhile, the abundance of the Prevotellaceae family and Parasutterela genus and the Alcaligencaeae family and Burkholderiales order and its attachment to the Betaproteobacteria class were overrepresented in the 3DG group. The bacteria of Candidatus Soleaferrea genus, Gelria genus, and Thermoanaerobacteraceae family and its attachment to Thermoanaerobacterales order were apparently more abundant in the control group. In addition, 45 KEGG pathways were altered after two-week intragastric administration of 3DG. Among these KEGG pathways, 13 KEGG pathways were involved in host metabolic function related to amino acid metabolism, carbohydrate metabolism, metabolism of cofactors and vitamins, and metabolism of terpenoids and polyketides. Moreover, the increased LPS levels and the decreased GLP-2 concentration in plasma and intestinal tissues were observed in 3DG-treated rats, together with the impaired fasting glucose and oral glucose tolerance. The alterations in composition and function of the intestinal microbiota were observed in 3DG-treated rats, which provides a possible mechanism linking exogenous 3DG intake to the development of prediabetes.

Dysbiosis of Gram-negative gut microbiota and the associated serum lipopolysaccharide exacerbates inflammation in type 2 diabetic patients with chronic kidney disease.

Lipopolysaccharide (LPS), a potent endotoxin present in the outer membrane of Gram-negative bacteria, causes chronic immune responses associated with inflammation. In the present study, the association between LPS and the dysbiosis of Gram-negative bacteria in the gut microbiome was determined in patients with type 2 diabetes mellitus (T2DM) and chronic kidney disease (T2DM-CKD; stages 4 and 5, not on dialysis) compared with healthy individuals. Microbiome diversity was analyzed in patients with T2DM-CKD and healthy controls by sequencing the hypervariable sub-regions of the 16S ribosomal RNA gene from stool samples. Serum samples were assayed by ELISA for LPS, C-reactive protein (CRP), tumor necrosis factor-α (TNFα), interleukin-6 (IL6) and endothelin-1. A total of four gut Gram-negative phyla (Bacteroidetes, Proteobacteria, Fusobacteria and Verrucomicrobia) were identified in the gut microbiome of the T2DM-CKD and control groups. Proteobacteria, Verrucomicrobia and Fusobacteria exhibited significantly increased relative abundance in patients with T2DM-CKD when compared with controls (P<0.05). The levels of serum LPS were significantly increased in patients with T2DM-CKD compared with controls (P<0.05). Elevated serum LPS was significantly correlated with increased levels of TNFα, IL6 and CRP. The dysbiosis of Gram-negative bacteria in the gut microbiome of patients with T2DM-CKD may contribute to the elevated serum levels of LPS and the consequential effects on the inflammatory biomarkers in these patients. The association between the dysbiosis of Gram-negative bacteria in the gut microbiome of patients with T2DM-CKD, increased LPS levels and the effects on inflammatory biomarkers may provide insight into potential diagnostic and therapeutic approaches in the treatment of T2DM-CKD.

Endotoxemia And Intestinal Barrier Disorders Associated With Overweight And Obesity

Abstract According to the World Health Organization (WHO), the widespread problem of overweight and obesity is the fifth most important risk factor for deaths in the world. The most frequently mentioned are the genetic and environmental factors that lead to the absorption of excess energy from food and to accumulate it in the form of spare adipose tissue. Another important fact is that even the use of a low-energy diet does not support the effective reduction of excessive body weight. It turns out that the cause may be intestinal microbiota, the composition of which changes in people with overweight and obesity. The intestinal microbiota dysbiosis is additionally perceived by many researchers as the cause of the development of metabolic diseases, including obesity or type 2 diabetes. On the other hand, Gram-negative bacteria constituting a component of the intestinal ecosystem are the source of lipopolysaccharide (LPS), responsible for the development of systemic inflammation and endotoxemia. Based on a literature review related to the subject, it can be concluded that intestinal microbiota disorders, intestinal barrier damage and increased LPS levels in patients adversely affect the obesity and components of the metabolic syndrome and hinder the treatment of these diseases. 1. Introduction. 2. Intestinal barrier function. 3. Intestinal barrier disorders and endotoxemia. 4. Summary

Increased circulatory levels of lipopolysaccharide (LPS) and zonulin signify novel biomarkers of proinflammation in patients with type 2 diabetes

Emerging data indicate that gut-derived endotoxin (metabolic endotoxemia) may contribute to low-grade systemic inflammation in insulin-resistant states. Specific gut bacteria seem to serve as lipopolysaccharide (LPS) sources and several reports claim a role for increased intestinal permeability in the genesis of metabolic disorders. Therefore, we investigated the serum levels of LPS and zonulin (ZO-1, a marker of gut permeability) along with systemic levels of tumor necrosis factor-α (TNF-α) and Interleukin-6 (IL-6) in patients with type 2 diabetes mellitus (T2DM) compared to control subjects. Study subjects were recruited from the Chennai Urban Rural Epidemiology Study [CURES], Chennai, India. Study group (n=45 each) comprised of a) subjects with normal glucose tolerance (NGT) and (b) patients with T2DM. LPS, ZO-1, TNF-α, and IL-6 levels were measured by ELISA. Serum levels of LPS [p<0.05], LPS activity [p<0.001], ZO-1 [p<0.001], TNFα [p<0.001], and IL-6 [p<0.001] were significantly increased in patients with T2DM compared to control subjects. Pearson correlation analysis revealed that LPS activity was significantly and positively correlated with ZO-1, fasting plasma glucose, 2h post glucose, HbA1c, serum triglycerides, TNF-α, IL-6, and negatively correlated with HDL cholesterol. Regression analysis showed that increased LPS levels were significantly associated with type 2 diabetes [odds ratio (OR) 13.43, 95% CI 1.998-18.9; p=0.003]. In Asian Indians who are considered highly insulin resistant, the circulatory LPS levels, LPS activity, and ZO-1 were significantly increased in patients with type 2 diabetes and showed positive correlation with inflammatory markers and poor glycemic/lipid control.

Characterization of meningococcal serogroup B outer membrane vesicle vaccines from strain 44/76 after growth in different media

In this study, we evaluated the effect of the growth medium on the composition and immunogenicity of meningococcal outer membrane vesicle (OMV) vaccines after cultivation of the Norwegian serogroup B 44/76 vaccine strain in either Frantz’ or modified Catlin-6 media (MC.6M). Differential proteomic analysis revealed that 97% of the OMV proteins maintained the same levels in the two preparations. However, a number of differentially expressed proteins, including TdfH, OpcA, OMP NMB0088, hypothetical NMB2134, lipoprotein NMB1126/1164 and NspA, increased significantly in OMVs produced from bacteria grown in the MC.6M. Together with increased lipopolysaccharide levels, the increased expression of these proteins was associated with significantly higher serum bactericidal titres in mice immunized with the MC.6M OMV vaccine. The high resolution two-dimensional separation of the OMVs on a large-format gel across a pH range of 3–11 resolved around 2000 protein spots from which 75 proteins were identified by mass spectrometry.

Reply to Lester et al.: Microbial translocation and HIV pathogenesis in Africa

Lester et al. (1) suggest that the data presented in our recent report (2) “are not sufficient to conclude that microbial translocation is not an important contributor to HIV disease in Africa.” The authors support this statement by first claiming that our study did not examine levels of lipopolysaccharide (LPS) in HIV-uninfected subjects. However, this is incorrect because our study examined LPS levels in HIV seroconverters from preinfection (HIV antibody and RNA negative) to either death or long-term nonprogression and found no consistent changes in LPS levels related to HIV infection or HIV disease progression (2). In contrast, Lester et al. (3) observed in a cross-sectional analysis of female commercial sex workers in Kenya slightly increased LPS levels in HIV-positive women when compared to uninfected women. However, it is difficult to directly compare these results with ours because they did not examine their subjects in a longitudinal manner, making it impossible to infer any causal relationship between LPS levels and HIV disease.

Neutrophilic airway inflammation and association with bacterial lipopolysaccharide in children with asthma and wheezing

Asthma is a leading chronic childhood illness in the US. To gain further insight into the pathophysiology of childhood asthma, we studied markers of airway inflammation and possible triggers such as bacterial lipopolysaccharide (LPS) in 18 children with chronic asthma and persistent wheezing who underwent clinically indicated bronchoscopy and bronchoalveolar lavage (BAL). We predominantly found neutrophilic airway inflammation associated with increased levels of IL-8, metalloproteinase (MMP)-9, tissue inhibitor of metalloproteinase (TIMP-1) and MMP-9/TIMP-1 ratio. A significant correlation was found between levels of LPS in BAL and airway neutrophils in BAL from a subgroup of children who had a tendency of increased levels of MMP-9 and TIMP-1, suggesting that increased LPS levels in BAL may contribute to chronic airway inflammation and early remodeling. Our data highlight the importance of defining chronic triggers of early airway inflammation in children and characterizing their inflammation, considering the use of bronchoscopy and BAL. Increased knowledge of airway inflammation in children may help prevent a more severe asthma phenotype and lead to environmental control measures and new treatment strategies to intervene against the establishment of irreversible inflammation.

Increased intestinal permeability in obese mice: new evidence in the pathogenesis of nonalcoholic steatohepatitis

A small percentage of pathologically obese subjects with fatty livers develop histological signs of necroinflammation and fibrosis, suggesting a variety of cofactors in the pathogenesis of obesity-related liver diseases including nonalcoholic steatohepatitis. Since several observations have linked bacterial endotoxins to liver damage, the aim of this study was to determine the effect of obesity on intestinal mucosal integrity and portal blood endotoxemia in two strains of obese mice: leptin-deficient (ob/ob) and hyperleptinemic (db/db) mice. Murine intestinal mucosal barrier function was assessed using a Ussing chamber, whereas ileum tight junction proteins were analyzed by immunocytochemistry and Western blot analysis. Circulating proinflammatory cytokines and portal blood endotoxin levels were measured by ELISA and the limulus test, respectively. The inflammatory and fibrogenic phenotype of murine hepatic stellate cells (HSCs) was determined by ELISA and quantitative RT-PCR. Ob/ob and db/db mice showed lower intestinal resistance, profoundly modified distribution of occludin and zonula occludens-1 in the intestinal mucosa, and higher circulating levels of inflammatory cytokines and portal endotoxemia compared with lean control mice. Moreover, HSCs isolated from ob/ob and db/db mice showed higher membrane CD14 mRNA levels and more pronounced lipopolysaccharide-induced proinflammatory and fibrogenic responses than HSCs from lean animals. In conclusion, genetically obese mice display enhanced intestinal permeability leading to increased portal endotoxemia that makes HSCs more sensitive to bacterial endotoxins. We suggest that in metabolic syndrome, patients may likewise have a greater intestinal mucosa permeability and increased lipopolysaccharide levels in portal blood that can contribute to the liver inflammatory damage.