Liposaccharide Research Articles

N-formylmethionine-leucyl-phenylalanine protects against irradiation-induced damage to hematopoiesis and intestines

BackgroundIonizing radiation (IR), including radiotherapy, can exert lasting harm on living organisms. While liposaccharide (LPS) offers resistance to radiation damage, it also induces toxic responses. Thankfully, an LPS analogue called N-formylmethionine-leucyl-phenylalanine (fMLP) holds the potential to mitigate this toxicity, offering hope for radiation protection.MethodsSurvival of C57BL/6 mice exposed to IR after administration with fMLP/LPS/WR-2721 or saline was recorded. Cell viability and apoptosis assay of bone marrow (BMC), spleen and small intestinal epithelial (HIECs) cells were tested by Cell Counting Kit-8 (CCK-8) and flow cytometry assay. Tissue damage was evaluated by Hematoxilin and Eosin (H&E), Ki-67, and TUNEL staining. RNA sequencing was performed to reveal potential mechanisms of fMLP-mediated radiation protection. Flow cytometry and western blot were performed to verify the radiation protection mechanism of fMLP on the cell cycle.ResultsThe survival rates of C57BL/6 mice exposed to ionizing radiation after administering fMLP increased. fMLP demonstrated low toxicity in vitro and in vivo, maintaining cell viability and mitigating radiation-induced apoptosis. Moreover, it protected against tissue damage in the hematopoietic and intestinal system. RNA sequencing shed light on fMLP’s potential mechanism, suggesting its role in modulating innate immunity and cell cycling. This was evidenced by its ability to reverse radiation-induced G2/M phase arrests in HIECs.ConclusionfMLP serves as a promising radioprotective agent, preserving cells and radiosensitive tissues from IR. Through its influence on the cell cycle, particularly reversing radiation-induced arrest in G2/M phases, fMLP offers protection against IR’s detrimental effects.

Design, synthesis and bioactivity evaluation of 4-hydroxycoumarin derivatives as potential anti-inflammatory agents against acute lung injury and colitis

Acute lung injury (ALI) and inflammatory bowel disease (IBD) are common inflammatory illnesses that seriously affect people's health. Herein, a series of 4-hydroxylcoumarin (4-HC) derivatives were designed and synthesized. The inhibitory effects of these compounds on LPS-induced interleukin-6 (IL-6) release from J774A.1 cells were then screened via ELISA assay, compound B8 showed 3 times more active than the lead compound 4-HC. The most active compound B8 had the IC50 values of 4.57 μM and 6.51 μM for IL-6 release on mouse cells J774A.1 and human cells THP-1, respectively. Furthermore, we also found that B8 could act on the MAPK pathway. Based on the target prediction results of computer virtual docking, kinase inhibitory assay was carried out, and it revealed that targeting IRAK1 was a key mechanism for B8 to exert anti-inflammatory activity. Moreover, B8 exerted a good therapeutic effect on the dextran sulfate sodium (DSS)-induced colitis model and liposaccharide (LPS)-induced ALI mouse models. The acute toxicity experiments indicated that high-dose B8 caused no adverse reactions in mice, confirming its safety in vivo. Additionally, the preliminary pharmacokinetic (PK) parameters of B8 in SD rats were also examined, revealing a bioavailability (F) of 28.72 %. In conclusion, B8 is a potential candidate of drug for the treatment of ALI and colitis.

Unravelling the Link between the Gut Microbiome and Autoimmune Kidney Diseases: A Potential New Therapeutic Approach.

The gut microbiota and short chain fatty acids (SCFA) have been associated with immune regulation and autoimmune diseases. Autoimmune kidney diseases arise from a loss of tolerance to antigens, often with unclear triggers. In this review, we explore the role of the gut microbiome and how disease, diet, and therapy can alter the gut microbiota consortium. Perturbations in the gut microbiota may systemically induce the translocation of microbiota-derived inflammatory molecules such as liposaccharide (LPS) and other toxins by penetrating the gut epithelial barrier. Once in the blood stream, these pro-inflammatory mediators activate immune cells, which release pro-inflammatory molecules, many of which are antigens in autoimmune diseases. The ratio of gut bacteria Bacteroidetes/Firmicutes is associated with worse outcomes in multiple autoimmune kidney diseases including lupus nephritis, MPO-ANCA vasculitis, and Goodpasture's syndrome. Therapies that enhance SCFA-producing bacteria in the gut have powerful therapeutic potential. Dietary fiber is fermented by gut bacteria which in turn release SCFAs that protect the gut barrier, as well as modulating immune responses towards a tolerogenic anti-inflammatory state. Herein, we describe where the current field of research is and the strategies to harness the gut microbiome as potential therapy.

Recipient Platelet-Derived Extracellular Vesicles (PEV) Suppress Antibody-Mediated Transfusion-Related Acute Lung Injury (TRALI)

Transfusion-related acute lung injury (TRALI) is a leading cause of transfusion-related fatalities with a 5 to 14% mortality rate. Symptoms range from mild dyspnea, cough, fever to full-blown respiratory failure and severe hypoxemia within 6 hours from the transfusion. To date, the pathogenesis is not fully understood, but it appears that TRALI requires two hits in order to be initiated; the first-hit is the patient's clinical condition often reflected by a state of inflammation while the second hit is the transfusion itself (often containing anti-donor antibodies). Several reports have described platelet involvement in antibody-mediated TRALI, however, there are significant controversies that have arisen as to whether intact platelets actually play a role or not. On the other hand, platelet extracellular vesicles (PEV) have been found to accumulate in stored platelet units and are positively correlated with significant adverse transfusion reactions. Furthermore, in a murine antibody-independent model of TRALI, it was recently demonstrated that PEV increase in stored murine platelet concentrates and are associated with TRALI-induction via excess ceramide production. We have used an in vivo murine model of TRALI where liposaccharide (LPS)-primed BALB/c mice are injected with a murine monoclonal anti-MHC class I antibody (34-1-2s). This antibody-mediated TRALI model exhibits severe acute lung injury that closely mimics the human condition. We attempted to understand what role recipient PEV may play in antibody-dependent TRALI. Male BALB/c mice were administered LPS ip (0.1 mg/kg) 18 hrs prior to initiating TRALI. At time zero, the mice are injected iv with 34-1-2s (1 mg/kg) and subsequently, TRALI manifestations were monitored every 30 min, and at 90 min post-injection, blood and lungs were harvested for the indicated measurements. Results show that compared with isotype-control treated mice, animals administered 34-1-2s had significantly elevated lung Wet/Dry (W/D) ratios (measure of lung edema, Figure 1a) and pulmonary neutrophil accumulation. The TRALI mice had significant thrombocytopenia and elevated levels of PEV in their plasma as detected by High-sensitivity Flow Cytometry (Hs-FCM). To determine the role of in vivo generated PEV in antibody-mediated TRALI, mice were first administered amitriptyline (AMI), a potent inhibitor of PEV formation. AMI treatment significantly reduced plasma PEV levels, but after administration of 34-1-2s, TRALI responses (lung edema) were significantly enhanced in the AMI-treated mice (Figure 1). To determine if this effect was due to PEV-derived serotonin, mice were next treated with Prozac to deplete platelet/PEV stores of serotonin. Prozac-treated mice had normal levels of PEV after 34-12s administration and TRALI induction was not affected. Our results suggest that in antibody-mediated TRALI, recipient generated PEV play a suppressive role that is independent of serotonin. Further studies are underway to determine the mechanism(s) of PEV-mediated TRALI suppression. Thus, modulating PEV formation in recipients may be an effective therapy for reducing TRALI reactions.

Association of microbiota‐derived products with astrogliosis and neurodegeneration in patients with Alzheimer’s Disease

AbstractBackgroundMicrobiota‐derived short chain fatty acids (SCFA) and liposaccharide (LPS) could modulate astrocyte and neuronal activity. Glial fibrillary acidic protein (GFAP) and neurofilament light (NfL) are markers of astroglial activation and neuronal damage, can be measured in blood samples and have been associated with Alzheimer’s disease (AD) pathology. The aim of the present study was to assess the association of SCFAs and LPS with GFAP and NfL in 13 amyloid‐β‐negative cognitively unimpaired individuals (CU), 38 patients with cognitive impairment and no brain amyloidosis (CI‐NAD) and 34 patients with cognitive impairment and amyloid‐β positivity (CI‐AD).MethodBlood levels of 4 SCFAs (acetate, propionate, valerate and butyrate) were measured by GC‐FID gaschromatography, LPS by ELISA, and GFAP and NfL by single‐molecule array technology.ResultPlasma valerate, LPS and GFAP concentrations were increased in CI‐AD compared with CI‐NAD and CU (p<0.034), acetate was increased in CI‐AD as well as decreased in CI‐NAD as compared with CU (p<0.018) and NfL was increased in both CI groups (p<0.023). In the pooled group of CI‐AD and CU, there were significant associations between plasma LPS and valerate with higher GFAP and NfL (p<0.032). Conversely, in the pooled group of CI‐NAD and CU, there were associations between plasma acetate with lower level of GFAP and NfL (p<0.026).ConclusionValerate and LPS were associated with astrogliosis and neuronal damage only in patients with AD. These results suggest that microbiota‐derived molecules might be mediators along the gut‐brain axis and contribute to AD pathology by modulating microglial and neuronal activity.

PRMT5 inhibition ameliorates inflammation and promotes the osteogenic differentiation of LPS‑induced periodontal stem cells via STAT3/NF‑κB signaling.

It has been reported that protein arginine methyltransferase 5 (PRMT5) serves a significant role in osteogenic differentiation and inflammatory response. Nevertheless, its role in periodontitis as well as its underlying mechanism remain to be elucidated. The aim of the present study was to explore the role of PRMT5 in periodontitis and whether PRMT5 could reduce liposaccharide (LPS)-induced inflammation of human periodontal ligament stem cells (hPDLSCs) and promote osteogenic differentiation through STAT3/NF-κB signaling. In the current study, the expression levels of PRMT5 were determined in LPS-induced hPDLSCs by reverse transcription-quantitative PCR and western blot analysis. ELISA and western blot analysis were employed to assess the secretion and expression levels of inflammatory factors, respectively. The osteogenic differentiation and mineralization potential of hPDLSCs were evaluated using alkaline phosphatase (ALP) activity assay, Alizarin red staining and western blot analysis. Additionally, western blot analysis was applied to determine the expression levels of the STAT3/NF-κB signaling pathway-related proteins. The results showed that the expression levels of PRMT5 were significantly enhanced in LPS-induced hPDLSCs. Additionally, PRMT5 knockdown reduced the contents of IL-1β, IL-6, TNF-α, inducible nitric oxide synthase and cyclooxygenase-2. PRMT5 depletion also enhanced ALP activity, improved the mineralization ability and upregulated bone morphogenetic protein 2, osteocalcin and runt-related transcription factor 2 in LPS-induced hPDLSCs. Furthermore, PRMT5 knockdown inhibited inflammation and promoted the osteogenic differentiation of hPDLSCs via blocking the activation of the STAT3/NF-κB signaling pathway. In conclusion, PRMT5 inhibition suppressed LPS-induced inflammation and accelerated osteogenic differentiation in hPDLSCs via regulating STAT3/NF-κB signaling, thus providing a potential targeted therapy for the improvement of periodontitis.

STAT3/NF‑κB decoy oligodeoxynucleotides inhibit atherosclerosis through regulation of the STAT/NF‑κB signaling pathway in a mouse model of atherosclerosis.

Atherosclerosis is a progressive chronic inflammatory condition that is the cause of most cardiovascular and cerebrovascular diseases. The transcription factor nuclear factor‑κB (NF‑κB) regulates a number of genes involved in the inflammatory responses of cells that are critical to atherogenesis, and signal transducer and activator of transcription (STAT)3 is a key transcription factor in immunity and inflammation. Decoy oligodeoxynucleotides (ODNs) bind to sequence‑specific transcription factors and limit gene expression by interfering with transcription invitro and invivo. The present study aimed to investigate the beneficial functions of STAT3/NF‑κB decoy ODNs in liposaccharide (LPS)‑induced atherosclerosis in mice. Atherosclerotic injuries of mice were induced via intraperitoneal injection of LPS and the mice were fed an atherogenic diet. Ring‑type STAT3/NF‑κB decoy ODNs were designed and administered via an injection into the tail vein of the mice. To investigate the effect of STAT3/NF‑κB decoy ODNs, electrophoretic mobility shift assay, western blot analysis, histological analysis with hematoxylin and eosin staining, Verhoeff‑Van Gieson and Masson's trichrome staining were performed. The results revealed that STAT3/NF‑κB decoy ODNs were able to suppress the development of atherosclerosis by attenuating morphological changes and inflammation in atherosclerotic mice aortae, and by reducing pro‑inflammatory cytokine secretion through inhibition of the STAT3/NF‑κB pathway. In conclusion, the present study provided novel insights into the antiatherogenic molecular mechanism of STAT3/NF‑κB decoy ODNs, which may serve as an additional therapeutic intervention to combat atherosclerosis.

Chicoric Acid Presented NLRP3-Mediated Pyroptosis through Mitochondrial Damage by PDPK1 Ubiquitination in an Acute Lung Injury Model.

Chicoric acid (CA), a functional food ingredient, is a caffeic acid derivative that is mainly found in lettuce, pulsatilla, and other natural plants. However, the anti-inflammatory effects of CA in acute lung injury (ALI) remain poorly understood. This study was conducted to investigate potential drug usage of CA for ALI and the underlying molecular mechanisms of inflammation. C57BL/6 mice were given injections of liposaccharide (LPS) to establish the in vivo model. Meanwhile, BMDM cells were stimulated with LPS+ATP to build the in vitro model. CA significantly alleviated inflammation and oxidative stress in both the in vivo and in vitro models of ALI through the inhibition of NLR family pyrin domain-containing 3 (NLRP3)-mediated pyroptosis. In addition, CA attenuated mitochondrial damage to suppress NLRP3-mediated pyroptosis in the in vivo and in vitro models of ALI by suppressing the production of reactive oxygen species (ROS) via inhibiting the Akt/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. CA inhibited the interaction between Akt at T308 and phosphoinositide-dependent kinase-1 (PDPK1) at S549, thus promoting the phosphorylation of the Akt protein. Furthermore, CA directly targeted the PDPK1 protein and accelerated PDPK1 ubiquitination, indicating that 91-GLY, 111-LYS, 126-TYR, 162-ALA, 205-ASP, and 223-ASP might be responsible for the interaction between PDPK1 and CA. In conclusion, CA from Lettuce alleviated NLRP3-mediated pyroptosis in the ALI model through ROS-induced mitochondrial damage by activating Akt/Nrf2 pathway via PDPK1 ubiquitination. The present study suggests that CA might be a potential therapeutic drug to treat or preventALI in pneumonia or COVID-19.

In vitro immunomodulatory effects of novel strontium and zinc-containing GPCs.

Glass polyalkenoate cements (GPCs) are bio-adhesives which consist of ionomeric glass particles embedded in a poly-salt matrix. These materials have been used in dentistry and orthodontics extensively but are presently being optimized as bone putties for orthopedic applications. This study utilized a patented ionomeric glass (mole fraction: SiO2:0.48, ZnO:0.36, CaO:0.12, SrO:0.04) to formulate two GPCs: GPC A (<45μm particle size glass) and GPC B (45μm-63μm). These formulations were previously assessed for their effect on osteoblast viability and osteogenic function. However, the immunomodulatory effects of GPC A and B have not previously been investigated. Non-toxic concentrations of (a) GPC dissolution products and (b) fragmented GPC particles were tested for their ability to affect the secretion of cytokines (TNF-α, IL-1β, IL-6 and IL-10) by rat peripheral blood mononuclear cells (PBMCs), in the presence or absence of the stimulant liposaccharide (LPS). Additionally, the ionic concentrations of Sr, Zn, Ca, and Si were measured in GPC ionic extracts, and the size, shape and concentration of fragmented GPC particles in deionized water were characterized using an optical microscope-based particle analyzer. The results showed that GPC A ionic products reduced the concentration of TNF-α secreted by stimulated cells compared with cells stimulated in the absence of GPC products. Interestingly, the particles released from GPC A significantly increased the secretion of both TNF-α and IL-6 from unstimulated cells, compared to control cells. Neither GPC B ionic products nor released particles were found to be biologically active with respect to PBMC cytokine secretion.

Ion channel Piezo1 activation promotes aerobic glycolysis in macrophages.

Altered microenvironmental stiffness is a hallmark of inflammation. It is sensed by the mechanically activated cation channel Piezo1 in macrophages to induce subsequent immune responses. However, the mechanism by which the mechanosensitive signals shape the metabolic status of macrophages and tune immune responses remains unclear. We revealed that Piezo1-deficient macrophages exhibit reduced aerobic glycolysis in resting or liposaccharide (LPS)-stimulated macrophages with impaired LPS-induced secretion of inflammatory cytokines in vitro. Additionally, pretreatment with the Piezo1 agonist, Yoda1, or cyclical hydrostatic pressure (CHP) upregulated glycolytic activity and enhanced LPS-induced secretion of inflammatory cytokines. Piezo1-deficient mice were less susceptible to dextran sulfate sodium (DSS)-induced colitis, whereas Yoda1 treatment aggravated colitis. Mechanistically, we found that Piezo1 activation promotes aerobic glycolysis through the Ca2+-induced CaMKII-HIF1α axis. Therefore, our study revealed that Piezo1-mediated mechanosensitive signals Piezo1 can enhance aerobic glycolysis and promote the LPS-induced immune response in macrophages.

Neuroprotective effects of delavatine A on LPS-induced activation of microglia in vitro and in a rat model of ischemia-reperfusion injury

Purpose: To investigate the neuroprotective effects of deloratine A on liposaccharide (LPS)-induced microglia activation in vitro and in a rat model of ischemia-reperfusion injury.&#x0D; Methods: LPS-induced microglial activation was successfully established in mouse microglia BV2 cell line. Then, the cells were randomly divided into model group, 2.5 μM delavatine A group, 5 μM delavatine A group, and 10 μM delavatine A group. The effect of delavatine A on the release of NO, TNF-α, IL-1β and IL-6 in BV2 cells was determined. In the vivo studies, 21 male Sprague Dawley (SD) rats were used to establish a rat model of ischemia-reperfusion injury, and effect of delavatine A on neural function and cerebral infarction area was determined.&#x0D; Results: The NO content was significantly higher in LPS-induced microglial activation model than in blank control, but it was significantly lower in the 3 delavatine A groups than in model group (p &lt; 0.05). The expression levels of TNF-α, IL-1β, and IL-6 were significantly higher in model group than in blank control group, but they were significantly and dose-dependently lower in delavatine A groups than in model group (p &lt; 0.05). In the in vivo rat studies, neural function score and cerebral infarction area in the model group were significantly higher than those in the sham group, while cerebral infarction area in delavatine A groups were significantly lower than that in the model group (p &lt; 0.05).&#x0D; Conclusion: Delavatine A significantly reduces the inflammation associated with LPS-induced microglial activation, mitigates loss of neural function, and reduces cerebral infarction area in rats with ischemia-reperfusion injury. These findings may lead to the development of new neuroprotective drugs.&#x0D; Keywords: Delavatine A; LPS; Microglia activation; Ischemia-reperfusion injury; Neural function; Cerebral infarction area

Prediction of Pasteurella multocida serotypes based on whole genomic sequences

The serotypes of Pasteurella multocida were predicted based on whole genomic sequences (WGS) with specific genes of the capsular and liposaccharide (LPS) outer core polysaccharide regions as targets. A total of 56 strains were whole genomic sequenced and in addition all assembled genomes from NCBI were included for comparison. BIGSdb (Bacterial Isolate Genome Sequence Database) was installed on a Linux server and targets for capsular types A, B, D, E and F were defined as gene sequences of hyaD, bcbD, dcbF, ecbJ and fcbD, respectively and targets for LPS groups 1, 2, 3, 4, 5, 6, 7 and 8 were defined as gene sequences of pcgB, nctA, gatF, latB, rmlA, nctB, ppgB and natG, respectively. The serotypes of P. multocida were predicted from WGS by designating the capsular type and LPS group as well as subtype alleles to isolates. Comparisons between WGS predictions of capsular types and classical phenotypic typing showed correspondence in 87 % of cases whereas comparisons of WGS predictions of LPS groups to phenotypic typing corresponded for 82 % of the strains. In total 93 % and 94 % of the strains available with WGS could be capsular and LPS group typed, respectively. The server is free to access from https://ivsmlst.sund.ku.dk.

Sex differences in IL-10's anti-inflammatory function: greater STAT3 activation and stronger inhibition of TNF-α production in male blood leukocytes ex vivo.

Interleukin-10 (IL-10) inhibits proinflammatory cytokine production in blood leukocytes-an effect mediated by signal transducer and activator of transcription 3 (STAT3) activation. To examine potential sex-based differences in IL-10's anti-inflammatory function, we treated whole blood from healthy males and females (n = 16 participants of each sex; age: 28 ± 6 yr; body mass index: 23.5 ± 2.3 kg/m2) with increasing concentrations of IL-10 (1-100 ng/mL) and quantified changes in phosphorylated STAT3 (pSTAT3) in CD14+ monocytes and CD4+ lymphocytes via flow cytometry. In parallel, liposaccharide (LPS)-stimulated whole blood cultures were used to assess sex-based differences in IL-10's ability to inhibit tumor necrosis factor (TNF)-α production. IL-10 concentration dependently increased pSTAT3 median fluorescent intensity (MFI) in CD14+ and CD4+ cells (main effects of concentration, P < 0.01) with males exhibiting larger changes in pSTAT3 MFI in both cell types (main effects of sex, P < 0.01). Accordingly, IL-10-mediated inhibition of TNF-α production was more pronounced in males (main effect of sex, P < 0.01) with changes in other monocyte-derived cytokines (IL-1β, IL-1RA, and IL-15) also supporting a sexual dimorphism in IL-10 action (P < 0.05). These sex-based differences were not explained by differences in circulating plasma IL-10 concentrations, basal IL-10 receptor expression in unstimulated CD14+ and CD4+ cells, nor the basal expression of IL-10 signaling proteins (STAT3, SHIP1, and p38 MAPK) in unstimulated peripheral blood mononuclear cells. We conclude that IL-10's anti-inflammatory function differs between male and female blood leukocytes ex vivo. This sexual dimorphism should be considered in future work investigating IL-10's anti-inflammatory action in humans as it may represent a mechanism contributing to sex differences in overall immune function.

Sesquiterpene dimers from Chloranthus fortunei and their protection activity against acute lung injury

Chloranthus fortunei (family Chloranthaceae), a perennial herb, widely distributed in south China with an altitude of 170–340 m. The whole plants were used as an anti-inflammatory agent for the treatment of cough, arthritis and tumor. Five previously unreported compounds fortulactones A-E were isolated from the aerial part of Chloranthus fortunei. Their structures were elucidated using 1D/2D NMR and HRESIMS and their absolute configuration were determined using the ECD excitron chirality method. All isolates were tested for inhibitory effects on the NO production of liposaccharide (LPS)-induced RAW 264.7 macropahges. The most potent compound 1 was further evaluated its protective activity against LPS stimulated A549 cells, the ELISA kits results showed the abnormal states of MDA and SOD were corrected to a certain extent. Meanwhile, the pro-inflammatory cytokine, such as TNF-α, IL-6 and IL-1β were also attenuated. In conclusion, these results showed that 1 exhibited therapeutic potential for ameliorating ALI.

124 The Effect of Innate Immune Receptor Activation on Differential Breed Responses to LPS in Sheep

Abstract While studying breed differences in immune response of peripheral blood mononuclear cells (PBMC) to Haemonchus contortus (Hc) antigen, RNA sequencing revealed that St. Croix (STC) PBMC upregulated expression of NLRP3 compared to PBMC from Suffolk (SUF) sheep. Nod-like receptor pyrin containing domain 3 (NLRP3) is a cytoplasmic pattern recognition receptor (PRR) with a wide array of agonists, such as PAMPs, DAMPs, ATP, bacterial product, and viral products. Stimulation of the NLRP3 inflammasome results in proteolytic activation of IL-1b and IL-18, cell pyroptosis, and classically, the induction of proinflammatory responses. Common activation of this pathway can be achieved by stimulation with liposaccharide (LPS) which binds and activates a toll-like receptor-4 (TLR4) pathway, causing production of proinflammatory cytokines. Interestingly, NLRP3 has been recently implicated as an early initiator of IL-4 production in response to helminth antigen. Thus, the objective of this study was to determine differences in breed responses to LPS and the role of NLRP3. To test this hypothesis, PBMC were collected from STC and SUF sheep and cells were incubated with LPS only, LPS in combination with pharmacological inhibitor of NLRP3 (MCC950), or complete media for 3 hrs. Gene expression of PBMC exposed to LPS revealed that Inflammatory linked genes IL-1β (P &amp;lt; 0.001), TLR4 (P &amp;lt; 0.001), TNFα (P &amp;lt; 0.001), NFκB (P &amp;lt; 0.001) were upregulated in STC PBMC. Treatment with MCC950 in combination with LPS, reduced inflammatory gene expression in STC PBMC to the level of SUF PBMC (P &amp;gt; 0.05), thus ablating inflammatory responses in STC PBMC. These data reveal a deficiency in NLPR3 function in SUF sheep and have been confirmed by SNPs found in that gene that have been predicted to alter NLRP3 protein structure.

Avenanthramide C from Oats Protects Pyroptosis through Dependent ROS-Induced Mitochondrial Damage by PI3K Ubiquitination and Phosphorylation in Pediatric Pneumonia.

Oat containing rich β-glucan, polyphenols, flavonoids, saponins, alkaloids, and other substances shows good biological activities. Therefore, the present study aimed to uncover the possible mechanism and therapeutic effect of Avenanthramide C in lessening inflammatory responses in pediatric pneumonia. Pediatric pneumonia was induced by liposaccharide (LPS) for vivo model and vitro model. Macrophage was performed to determine the mechanism and effects of Avenanthramide C in pediatric pneumonia. NLRP3 activity participated in the effects of Avenanthramide C in pediatric pneumonia. Avenanthramide C induced p-PI3K and p-Akt expressions and reduced ubiquitination of PI3K expression in pediatric pneumonia. On the other hand, Avenanthramide C integrated serine at 821 sites of the PI3K protein function. Avenanthramide C reduced ROS (reactive oxygen species)-induced mitochondrial damage by PI3K/AKT function in a model of pediatric pneumonia. Avenanthramide C protects pyroptosis in a model of pediatric pneumonia by PI3K/AKT/Nrf2/ROS signaling. Taken together, our results demonstrated that Avenanthramide C protects pyroptosis through dependent ROS-induced mitochondrial damage by PI3K ubiquitination and phosphorylation in a model of pediatric pneumonia, suggesting its potential use for the treatment of pediatric pneumonia and other inflammatory diseases.

HMBOX1 attenuates LPS-induced periodontal ligament stem cell injury by inhibiting CXCL10 expression through the NF-κB signaling pathway.

Homeobox containing 1 (HMBOX1) is a member of the homeobox transcription factor family that has been reported to serve an important role in numerous biological processes. The present study aimed to determine the role of HMBOX1 in the pathogenesis of periodontitis. Human periodontal ligament stem cells (hPDLSCs) were treated with liposaccharide (LPS) and transfected with a HMBOX1 overexpression (Ov-HMBOX1) plasmid or small interfering (si)-C-X-C motif chemokine ligand 10 (CXCL10) plasmids. The effects of Ov-HMBOX1 on cell proliferation, inflammation and apoptosis were subsequently investigated using Cell Counting Kit-8, ELISA for analysis of IL-6, TNF-α and IL-1β levels, TUNEL and western blotting assays for analysis of Bcl-2, Bax, cleaved caspase-3 and caspase-3 levels, respectively. Furthermore, the potential effects of HMBOX1 on the mRNA and protein levels of CXCL10 and the NF-κB signaling pathway were investigated by using reverse transcription-quantitative PCR and western blotting. Finally, the physiological processes of lipopolysaccharide (LPS)-induced hPDLSCs overexpressing HMBOX1 were assessed following treatment with phorbol 12-myristate 13-acetate (PMA), a NF-κB agonist. The results revealed that Ov-HMBOX1 transfection promoted proliferation whilst alleviating inflammation and apoptosis in LPS-induced hPDLSCs. Ov-HMBOX1 reduced the expression of CXCL10 by suppressing the NF-κB signaling pathway. PMA treatment inhibited the proliferation of LPS-induced hPDLSCs transfected with Ov-HMBOX1, which was reversed by transfection with si-CXCL10. In conclusion, results of the present study provided evidence that HMBOX1 can attenuate LPS-induced hPDLSC injury by downregulating CXCL10 expression via the NF-κB signaling pathway, which may provide a novel insight into the development of potentially novel treatment strategies for periodontitis.

Oxidative stress in human gingival fibroblasts from periodontitis versus healthy counterparts.

Elevated p53 promotes oxidative stress and production of pro-inflammatory cytokines in liposaccharide (LPS)-treated healthy human gingival fibroblasts (HGFs). This study compared oxidative stress, production of inflammatory cytokines, and p53 expression in HGFs from patients with chronic periodontitis (CP) and healthy subjects in vitro upon LPS from Porphyromonas gingivalis challenge. Human gingival fibroblasts were isolated from 6 biopsies-3 from healthy donors and 3 from diseased area in CP (Grade B, Stage III). HGFs were cultured with or without 1μg/ml 24h LPS. Oxidative stress was assessed by analyzing the level of reactive oxygen species (ROS). Mitochondrial membrane potential and respiration were determined by immunofluorescence and respirometry, respectively. Tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β were determined by enzyme-linked immunosorbent assay. P53 expression was monitored by Western blot and immunofluorescence. Human gingival fibroblasts from CP exhibited increased levels of mitochondrial p53, enhanced ROS production, decreased mitochondrial membrane potential, increased mitochondrial oxygen consumption, and increased secretion of TNF-α, IL-6, and IL-1β, as compared to HGFs from healthy donors. Moreover, LPS exacerbated these changes. Human gingival fibroblasts from CP exhibited stronger basal and LPS-inducible oxidative stress and inflammatory response as compared to HGFs from healthy subjects by increased p53 in mitochondria.

Identification of inflammatory markers in eosinophilic cells of the immune system: fluorescence, Raman and CARS imaging can recognize markers but differently

Eosinophils (Eos) play an important role in the immune system’s response releasing several inflammatory factors and contributing to allergic rhinitis, asthma, or atopic dermatitis. Since Eos have a relatively short lifetime after isolation from blood, usually eosinophilic cell line (EoL-1) is used to study mechanisms of their activation and to test therapies. In particular, EoL-1 cells are examined in terms of signalling pathways of the inflammatory response manifested by the presence of lipid bodies (LBs). Here we examined the differences in response to inflammation modelled by various factors, between isolated human eosinophils and EoL-1 cells, as manifested in the number and chemical composition of LBs. The analysis was performed using fluorescence, Raman, and coherent anti-Stokes Raman scattering (CARS) microscopy, which recognised the inflammatory process in the cells, but it is manifested slightly differently depending on the method used. We showed that unstimulated EoL-1 cells, compared to isolated eosinophils, contained more LBs, displayed different nucleus morphology and did not have eosinophilic peroxidase (EPO). In EoL-1 cells stimulated with various proinflammatory agents, including butyric acid (BA), liposaccharide (LPS), or cytokines (IL-1β, TNF-α), an increased production of LBs with a various degree of lipid unsaturation was observed in spontaneous Raman spectra. Furthermore, stimulation of EoL-1 cells resulted in alterations of the LBs morphology. In conclusion, a level of lipid unsaturation and eosinophilic peroxidase as well as LBs distribution among cell population mainly accounted for the biochemistry of eosinophils upon inflammation.

Inhibition of neuroinflammation by MIF inhibitor 3-({[4-(4-methoxyphenyl)-6-methyl-2-pyrimidinyl]thio}1methyl)benzoic acid (Z-312)

Microglial overactivation-mediated neuroinflammation contributes greatly to the pathogenesis of neurodegenerative diseases, such as Parkinson’s disease. Macrophage migration inhibitory factor (MIF) is a pleiotropic proinflammatory cytokine that is involved in the pathophysiology of various inflammatory diseases by inducing various proinflammatory cytokines. Compound 3-({[4-(4-methoxyphenyl)-6-methyl-2-pyrimidinyl]thio}methyl)benzoic acid (Z-312) is a novel small -molecule inhibitor of MIF tautomeric activity. In this study, we investigated the anti-inflammatory effects of Z-312 on liposaccharide (LPS)-induced neuroinflammation in vitro and in vivo. The results showed that Z-312 significantly decreased the production of nitric oxide (NO), interleukin (IL)-1β, tumor necrosis factor (TNF)-α and IL-6 in LPS-stimulated microglial cells. Mechanistically, nuclear translocation of the p65 subunit of nuclear factor (NF)-κB, degradation and phosphorylation of IκBα, NF-κB transcriptional activity and phosphorylation of p38 mitogen-activated protein kinase (MAPK) and JNK were markedly attenuated by pretreatment with Z-312 in BV-2 microglial cells. In addition, Z-312 suppressed the neurotoxic effects of cell culture medium of LPS-activated BV-2 microglia on cocultured mouse HT22 neuroblastoma cells. An in vivo study demonstrated that Z-312 markedly ameliorated microglial activation and subsequent DA neuron loss in an LPS-induced Parkinson’s disease (PD) mouse model. These results suggest that MIF inhibitor Z-312 may be a promising neuroprotective agent for the treatment of neuroinflammation-mediated neurological diseases.