Prostaglandin D2 is involved in the regulation of inflammatory response in Staphylococcus aureus-infected mice macrophages.

Staphylococcus aureus (S. aureus) is one of the most infamous and widespread bacterial pathogens, causing a hard-to-estimate number of uncomplicated skin infections and probably hundreds of thousands to millions of more severe, invasive infections globally per year. S. aureus may also be acquired from animals, especially in the livestock industry. The interaction mechanism of host and S. aureus has significance for finding ways to against S. aureus infection and control inflammatory response of host, while the molecular biological activities after S. aureus infection, particular in inflammatory and immune cells are not fully clear. The present study aimed to explore whether pattern recognition receptors (PRRs) mediate prostaglandin D2 (PGD2) synthesis and PGD2 participates in the regulation of inflammatory response in macrophages during S. aureus infection or synthetic bacterial lipopeptide (Pam2CSK4) stimulation. PGD2 secretion level was enhanced by mice peritoneal macrophages infected with the S. aureus. The results indicated that PGD2 secretion was impaired in S. aureus infected-macrophages from toll-like receptors 2 (TLR2)-deficient and NLR pyrin domain-containing 3 (NLRP3)-deficient mice. PGD2 synthetase (hematopoietic PGD synthase, HPGDS) inhibitors could reduce the activation of macrophage mitogen-activated protein kinase (MAPK)/nuclear factor-κ-gene binding (NF-κB) signaling pathways. HPGDS inhibition impaired cytokines (TNF-α, IL-1ß, IL-10 and RANTES) secretion and macrophage phagocytosis during S. aureus infection. In addition, inhibition of endogenous PGD2 synthesis was unable to affect the TLR2 and NLRP3 expression in S. aureus-infected macrophages. Taken together, macrophage PGD2 secretion after S. aureus infection depended on receptors TLR2 and NLRP3, and the induced PGD2 participated in the regulation of inflammatory response in S. aureus-infected macrophages. Interestingly, it was found that exogenous PGD2 down-regulated the cytokines secretion and had no effect on phagocytosis in the S. aureus-infected macrophages.

Prostaglandin D2 regulates Escherichia coli-induced inflammatory responses through TLR2, TLR4, and NLRP3 in macrophages.

Prostaglandin D2 (PGD2) synthesis is closely associated with the innate immune response mediated by pattern recognition receptors (PPRs). We determined PGD2 synthesis whether mediated by Toll-like receptor 2 (TLR2), TLR4 and Nod-like receptor pyrin domain-containing protein 3 (NLRP3) in Escherichia coli (E. coli)-, lipopolysaccharide (LPS)- and Braun lipoprotein (BLP)-stimulated macrophages. Our data demonstrate that TLR2, TLR4, and NLRP3 could regulate the synthesis of PGD2 through cyclo-oxygenase-2 (COX-2) and hematopoietic PGD synthase (H-PGDS) in E. coli-, LPS- or BLP-stimulated macrophages, suggesting that TLR2, TLR4, and NLRP3 are critical in regulating PGD2 secretion by controlling PGD2 synthetase expression in E. coli-, LPS- or BLP-stimulated macrophages. The H-PGDS (a PGD2 specific synthase) inhibitor pre-treatment could down-regulate the secretion of TNF-α, RANTES and IL-10 in LPS- and E. coli-stimulated macrophage. Meanwhile, H-PGDS inhibitor could down-regulate the secretion of TNF-α, while up-regulated RANTES and IL-10 secretion in BLP-stimulated macrophages, suggesting that PGD2 could regulate the secretion of cytokines and chemokines in E. coli-, LPS- or BLP-stimulated macrophages. Furthermore, exogenous PGD2 regulates the secretion of cytokines and chemokines through activation of MAPK and NF-κB signaling pathways after E. coli-, LPS- or BLP stimulation in macrophages. Taken together, PGD2 is found able to regulate E. coli-induced inflammatory responses through TLR2, TLR4, and NLRP3 in macrophages.

Besides TLR2 and TLR4, NLRP3 is also involved in regulating Escherichia coli infection-induced inflammatory responses in mice.

The host Toll-like Receptor-2 (TLR2) and Toll-like Receptor-4 (TLR4) play critical roles in defense against Escherichia coli (E. coli) infection is well-known. The NLR pyrin domain-containing 3 (NLRP3) inflammasome is also an important candidate during the host-recognized pathogen, while the roles of NLRP3 in the host inflammatory response to E. coli infection remains unclear. This study aimed to explore the roles of NLRP3 in regulating the inflammatory response in E. coli infection-induced mice. Our result indicated that compared to wild-type mice, the TLR2-deficient (TLR2-/-), TLR4-deficient (TLR4-/-), and NLRP3-deficient (NLRP3-/-) mice had significant decrease in liver damage after stimulation with Lipopolysaccharide (LPS, 1 µg/mL), Braun lipoprotein (BLP, 1 µg/mL), or infected by WT E. coli (1 × 107 CFU, MOI 5:1). Meanwhile, compared with wild-type mice, the TNF-α and IL-1ß production in serum decreased in TLR2-/-, TLR4-/-, and NLRP3-/- mice after LPS, BLP treatment, or WT E. coli infection. In macrophages from NLRP3-/- mice showed significantly reduced secretion of TNF-α and IL-1ß in response to stimulation with LPS, BLP, or WT E. coli infection compared with macrophages from wild-type mice. These results indicate that besides TLR2 and TLR4, NLRP3 also plays a critical role in host inflammatory responses to defense against E. coli infection, and might provide a therapeutic target in combating disease with bacterium infection.

Braun Lipoprotein Protects against Escherichia coli-Induced Inflammatory Responses and Lethality in Mice.

Escherichia coli (E. coli), a Gram-negative bacterium, is an important pathogen that causes several mammalian diseases. The outer membrane components of E. coli, namely, lipopolysaccharide (LPS) and bacterial lipoprotein, can induce the host innate immune response through pattern recognition receptors (PRRs). However, the detailed roles of the E. coli Braun lipoprotein (BLP) in the regulation of host inflammatory response to E. coli infection remain unclear. In this study, we sought to determine the effects of BLP on E. coli-induced host inflammatory response and lethality using mouse models. Experiments using the E. coli DH5α strain (BLP-positive), E. coli JE5505 strain (BLP-negative), and E. coli JE5505 strain combined with BLP indicated that the presence of BLP could alleviate mortality and organ (liver and lung) damage and decrease proinflammatory cytokine (tumor necrosis factor alpha [TNF-α] and interleukin-1ß [IL-1ß]) and chemokine (regulated on activation normal T-cell expressed and secreted [RANTES]) production in mouse serum and organs. Conversely, E. coli JE5505, E. coli DH5α strain, and E. coli JE5505 combined with BLP treatment induce enhanced anti-inflammatory cytokine (interleukin 10 [IL-10]) production in mouse serum and organs. In addition, BLP could regulate the secretion of proinflammatory cytokines (TNF-α and IL-1ß), chemokines (RANTES), and anti-inflammatory factors (IL-10) through mitogen-activated protein kinase (MAPK) and nuclear factor-kappaB (NF-κB) signaling pathways in macrophages. Altogether, our results demonstrate that the bacterial component BLP plays crucial and protective roles in E. coli-infected mice, which may influence the outcome of inflammation in host response to E. coli infection. IMPORTANCE In this study, we investigated the roles of bacterial outer membrane component BLP in regulating inflammatory responses and lethality in mice that were induced by a ubiquitous and serious pathogen, Escherichia coli. BLP could alleviate the mortality of mice and organ damage, as well as decrease proinflammatory cytokines and chemokine production and enhance anti-inflammatory cytokine production in mouse serum and organs. Overall, our results demonstrate that the bacterial component BLP plays crucial and protective roles in E. coli-infected mice through regulating the production of an inflammatory mediator, which may influence the outcome of inflammation in host response to E. coli infection. Our findings provide new information about the basic biology involved in immune responses to E. coli and host-bacterial interactions, which have the potential to translate into novel approaches for the diagnosis and treatment of E. coli-related medical conditions, such as bacteremia and sepsis.

Prostaglandin E receptor 2 mediates the inducible effects of prostaglandin E2 on expression of growth factors and enzymes in cattle endometrial epithelial cells and explants.

Prostaglandin E2 (PGE2 ) is able to induce the expression of several growth factors and enzymes in cattle endometria. However, the specific type of PGE2 receptors which mediates this effect is not fully clear. In this study, the role of prostaglandin E receptor 2 (PTGER2) in PGE2 -mediated induction of growth factors and enzymes expression in cattle endometrial explants and epithelial cells were investigated. PTGER2 was blocked by a PTGER2 antagonist, AH6809, before PGE2 treatment, then the mRNA and protein expression levels of several growth factors and enzymes were compared with that in PGE2 alone treatment group by real-time RT-PCR and Western blotting analysis in endometrial epithelial cells and explants. Results indicated that PGE2 significantly increased the mRNA and protein levels of these growth factors and enzymes, while the rates of increment in the expression of these growth factors and enzymes were inhibited by AH6809. In addition, a PTGER2 agonist, butaprost, significantly increased the expression levels of these growth factors and enzymes, and the effect could be blocked by AH6809. In conclusion, PTGER2 was found to be one dominant receptor mediating the inducible effects of PGE2 on the expression of these growth factors and enzymes in cattle endometrial explants and epithelial cells.

Codonopsis pilosula polysaccharides attenuate Escherichia coli-induced acute lung injury in mice.

Acute lung injury (ALI) is an inflammatory lung disease that is caused by bacterial infection. Lipopolysaccharide (LPS), a prototype pathogen-associated molecular pattern (PAMP) from Gram-negative bacteria such as Escherichia coli (E. coli), is an essential risk factor for ALI. LPS and E. coli induced the activation of mitogen-activated protein kinase (MAPK) and nuclear factor kappaB (NF-κB) signaling pathways, which led to the increasing immune molecule transcription, including pro-inflammatory cytokine and chemokine secretion. Codonopsis pilosula polysaccharides (CPPS) exhibit various biological activities and pharmacological effects. However, the effect of CPPS on ALI caused by LPS stimulation or E. coli infection remains unclear. Our results showed that CPPS (6.25, 12.5, 25, or 50 µg mL-1) could attenuate the secretion of TNF-α and IL-1ß and impair the phosphorylation of ERK, p38 and p65 in E. coli-infected macrophages without causing toxic reactions. In addition to regulating the secretion of pro-inflammatory cytokines and the activation of MAPK and NF-κB signaling pathways, CPPS could enhance bacterial phagocytosis and intracellular killing in macrophages, and inhibit the bacterial growth of E. coli. In vivo experiments showed that CPPS attenuated LPS- and E. coli-induced lung damage in mice, which was characterized by decreased pro-inflammatory cytokine (TNF-α, IL-1ß and IL-6) and chemokine (RANTES) production and production of the biomarkers of tissue damage (HABP2 and HMGB1) in the lungs. Altogether, this study demonstrated that CPPS have a protective effect on the lungs in LPS- and E. coli-induced ALI mouse models, suggesting that CPPS could be a potential drug for the treatment of ALI.

TLR2, TLR4, and NLRP3 mediated the balance between host immune-driven resistance and tolerance in Staphylococcus aureus-infected mice.

Staphylococcus aureus (S. aureus) is a gram-positive pathogen that can cause infectious diseases in mammals. S. aureus-induced host innate immune responses have a relationship with Toll-like receptor 2 (TLR2), TLR4, and Nod-like receptor pyrin domain-containing protein 3 (NLRP3). However, the detailed roles of TLR2, TLR4, and NLRP3 in regulating the host inflammatory response to S. aureus infection remain unclear. Our data indicated that the S. aureus-induced mortality was aggravated by deficiency of TLR2, TLR4, and NLRP3 in mice. In the subsequent experiment, we found that during S. aureus infection, the roles of TLR2, TLR4, and NLRP3 seemed to be different at multiple timepoints. The deficiency of TLR2, TLR4, or NLRP3 attenuated the expression of High-mobility group box protein 1 (HMGB1) and Hyaluronic acid-binding protein 2 (HABP2), which is accompanied by decreased proinflammatory cytokine (TNF-α), chemokine (RANTES), and anti-inflammatory cytokine (IL-10) production in lungs and serum at 3 h and 6 h post-infection. However, with S. aureus infection prolonged (24 h post-infection), the trend was diametrically opposite. The results showed that deficiency of TLR2, TLR4, or NLRP3 aggravated HABP2 and HMGB1 expression, which is accompanied by enhanced proinflammatory cytokine (TNF-α), chemokine (RANTES), and anti-inflammatory cytokine (IL-10) production in lungs and serum. These results were consistent with the data observed in S. aureus-infected bone marrow-derived macrophages (BMDMs). All these results suggested that during S. aureus infection, TLR2, TLR4, and NLRP3 has time-dependent effect in regulating the balance between immune-driven resistance and tolerance.

Occurrence and exposure risk assessment of phthalate esters in edible plant oils with a high-frequency import rate in west China.

Phthalate esters (PAEs) are ubiquitous pollutants in the environment with toxicological and epidemiological effects for humans. As one of the daily necessities, edible plant oil is an important exposure source of PAEs, due to the inevitable contact with PAE-containing materials and the intrinsic lipid solubility of PAEs. However, limited information is currently available on the exposure risk of PAEs in commercial plant oil. This study was aimed at investigating the occurrence and risk assessment of PAEs in plant oils with a high-frequency import rate in west China. The analysis method was referenced to the Chinese national standard for the determination of PAEs in food. Results indicated that PAEs (mainly including DBP and DEHP) were ubiquitous contaminants in imported plant oils with the detectable rate being up to 56.83% in 366 samples. The detected concentrations were in the range of 0.10-3.20 mg kg-1 (median 0.28 mg kg-1) for dibutyl phthalate (DBP) and 0.25-1.95 mg kg-1 (median 0.44 mg kg-1) for bis(2-ethylhexyl)phthalate (DEHP). Based on an integrated probabilistic analysis method, the values of non-carcinogenic risk were lower than 1 in all cases, indicating that there would be an unlikely incremental non-carcinogenic risk to humans. Generally, the carcinogenic risk of DEHP was lower than the upper acceptable carcinogenic risk level (<10-4), while 50.40% of the carcinogenic risk exceeded the lower acceptable carcinogenic risk level (>10-6). Besides, diverse health risks were obviously shown and discussed for different categories of plant oils. The obtained results in this study could provide valuable information to understand the contamination status and health risk of PAEs in plant oil and improve the relative supervision and regulation. And the proposed strategy suggests a potential application for health risk assessment of other contaminants in food or even environments.

Staphylococcus aureus lipoproteins play crucial roles in inducing inflammatory responses and bacterial internalization into bovine mammary epithelial cells.

Bovine mastitis is caused by bacterial infection and characterized by inflammatory and infectious processes. Staphylococcus aureus frequently causes subclinical mastitis in dairy cows. In this study, we aimed to investigate the roles of S. aureus lipoproteins in inducing inflammatory responses and in mediating bacterial internalization into bovine mammary epithelial cells (bMECs). The results showed that TLR2 expression in bMECs infected with S. aureus isogenic mutant deficient in lipoprotein maturation was decreased compared to that in bMECs infected with wild-type S. aureus. Lipoproteins from S. aureus and the engagement of TLR2 were essential for inducing the activation of MAPK and NF-κB signaling, and stimulating the secretion of the inflammatory mediators tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and C-X-C motif chemokine ligand 8 (CXCL8). The production of prostaglandin E2 (PGE2) and the expression of PTGS2 in S. aureus-infected bMECs were dependent on the presence of bacterial lipoproteins. Furthermore, bacterial lipoproteins contributed to S. aureus internalization into bMECs. These findings suggest the S. aureus lipoproteins are key immunobiologically active compounds that trigger inflammatory responses in bMECs and play an important role in S. aureus internalization into bMECs.

A finite element analysis on comparing the stability of different posterior fixation methods for thoracic total en bloc spondylectomy.

OBJECTIVE: To compare the spinal stability with different fixation methods after thoracic TES using finite element analysis METHODS: The spinal finite element model was established from a healthy volunteer, and the validity was verified. The models of T8 thoracic total en bloc spondylectomy (TES) with and without artificial vertebral body were established combination with different fixation methods: the first was long segment fixation with fixed segments T5-7, T9-11; the second was short segment fixation with fixed segments T6-7, T9-10; the third was modified short segment with a pair of vertebral body screws on T7 and T9 added on the basis of short segment fixation. The motions of each model in standing state were simulated in software. The range of motion (ROM) and internal fixation stress changes were analyzed. RESULTS: When anterior support was effective, the three fixation methods could effectively maintain the stability of the spine. However, when anterior support failed, the ROM of the long segment fixation group and the short segment fixation group in the flexion-extension directions was significantly higher than that of when the anterior support existed, while the modified short segment fixation group had no significant changes. Meanwhile, the stress of internal fixation in the long segment fixation group and the short segment fixation group were greatly increased. However, there were no significant changes in modified short segment fixation group. CONCLUSION: After TES, the presence of the thoracic cage gives partial anterior stabilization. When the anterior support failed, the modified short segment fixation method can provide better stability.

Simultaneous determination of 106 pesticides in nuts by LC-MS/MS using freeze-out combined with dispersive solid-phase extraction purification.

As a result of the low water content and high fat matrices in nuts, it is very difficult to simultaneously determine multi-pesticides in trace levels. Here, a sample pretreatment method was developed in which, microwave-assisted solvent extraction was firstly used to extract pesticides, and then a two-step cleanup method was conducted combining freeze-out with dispersive solid-phase extraction to remove the lipidic matrix. By this way, 106 pesticides were simultaneously determined in the complicated nut sample by using an ultra-high pressure liquid chromatography coupled with a tandem mass spectrometer. Average recoveries were 75.3-119.3% with relative standard deviations < 14% at three concentration levels. The limits of detection and quantification were in the ranges of 0.3-3.0 and 1.0-10.0 µg/kg, respectively. Furthermore, the method was successfully applied to the determination of pesticides in 180 commercial nut samples.