Protein Levels Of Inflammatory Cytokines Research Articles

Ursolic acid alleviates heat stress-induced lung injury by regulating endoplasmic reticulum stress signaling in mice

Acute lung injury has been reported to be associated with heat stress in various animals. Ursolic acid is a natural pentacyclic triterpenoid compound with multiple bioactivities. However, it remains unknown whether ursolic acid supplementation alleviates heat stress-induced lung injury. In the present study, male Institute of Cancer Research mice were left untreated under a normal temperature condition (23±1°C), receiving orally administrated with vehicle (phosphate buffered saline) or ursolic acid (40 mg/kg BW−1·d−1 for 2 d), and then were subjected to high temperature (41±1°C) for 2 h. Histological alterations, activities of antioxidative enzymes, apoptosis, generation of reactive oxygen species, abundance of inflammatory cytokines, and endoplasmic reticulum stress-related proteins were analyzed. Compared with the controls, heat stress treatment led to enhanced apoptosis, increased H2O2 production, and upregulated protein levels of inflammatory cytokines in the serum, including tumor necrosis factor alpha, interleukin-6, and interleukin-1 beta. Activities of malondialdehyde, lactate dehydrogenase, and myeloperoxidase were increased, while the activities for superoxide dismutase and catalase were reduced in lung tissues of mice. All these alterations were significantly prevented by ursolic acid administration. Further study showed that heat stress led to activation of protein kinase-like ER kinase eukaryotic initiation factor 2 alpha -the transcription factor CCAAT-enhancer-binding protein homologous protein (CHOP) signaling, which was attenuated by ursolic acid supplementation. These findings indicated that ursolic acid pretreatment protected lung tissues against heat stress-induced injury by regulating inflammatory cytokines and unfolded protein response in mice. Ursolic acid supplementation might be a therapeutic strategy to alleviate high temperature-induced lung injury in humans and animals.

The Protective Effects of VVN001 on LPS-Induced Inflammatory Responses in Human RPE Cells and in a Mouse Model of EIU.

To investigate protective effects of VVN001 on lipopolysaccharide (LPS)-induced inflammatory response in human retinal pigment epithelial (RPE) cells and in a mouse model of endotoxin-induced uveitis (EIU), and to explore the underlying mechanisms. Human primary RPE (hRPE) and ARPE-19 cells were pretreated with or without VVN001 for 1h followed by 10μg/mL LPS stimulation for 24h. mRNA, and protein levels of inflammatory cytokines were analyzed with real-time PCR, western blotting, and ELISA. EIU was induced by intravitreal injection of 125ng LPS in female BALB/c mice. VVN001 eye drops (1%) were locally administrated every 4h for 24h after LPS injection. Clinical scores were assessed with a slit lamp. mRNA and protein levels of inflammatory cytokines were investigated simultaneously. Compared with the LPS group, VVN001 pretreatment significantly reduced mRNA expressions of intercellular adhesion molecule-1 (ICAM-1), IL-6, IL-8, TNF-α, IL-1β, IL-18, caspase-1 in hRPE, and ARPE-19 cells. Protein overproduction of ICAM-1, TNF-α, IL-1β, NLRP3, caspase-1 P20, and p-IκBα/IκBα stimulated by LPS was suppressed by VVN001 pretreatment. In vivo, VVN001 significantly reduced the average clinical score from 5.0 to 1.3 in EIU mice. Furthermore, overproduction of ICAM-1, IL-1β, NLRP3, caspase-1 P20, and p-IκBα/IκBα at mRNA and protein levels were remarkably suppressed by VVN001. VVN001 alleviated the inflammatory response induced by LPS both in vitro and in vivo. The effect of anti-inflammation is associated with inhibiting the overproduction of ICAM-1 and blocking the activation of NLRP3 inflammasome and the NF-κB signaling pathway.

Hyaluronic Acid/Parecoxib-Loaded PLGA Microspheres for Therapy of Temporomandibular Disorders.

This study aimed to fabricate Hyaluronic Acid (HA)/parecoxib-loaded PLGA microspheres for the treatment of Temporomandibular Disorders (TMD) and investigate the in vitro and in vivo effect of the microsphere system to solve the issues of poor drug delivery and short duration on drug concentration in conventional TMD therapy. The microspheres were prepared by the double emulsion (w/o/w) method. Various formulations were compared in terms of particle size, drug loading rate and encapsulation rate. Scanning Electron Microscopy (SEM), Differential Scanning Calorimetry (DSC) and FT-IR spectroscopy were performed to evaluate physicochemical properties. The drug release behavior of microspheres and toxicity assay on synovial cells were investigated. The in vitro anti-inflammatory effect on inflammatory markers, such as IL-1β, TNF-α and COX-2, was assessed by real-time PCR. Then, the in vivo therapeutic effect of microspheres was investigated using mechanically-induced rat synovitis model. Protein levels of inflammatory cytokines (IL-1β, TNF-α and COX-2) from TMJ periarticular tissues were quantified by Enzyme-Linked Immunosorbent Assay (ELISA). The results showed that microspheres were morphologically regular, smooth and non-cohesive. The average particle size of the microspheres was (25.32 ± 1.01) μm. The drug loading rate of parecoxib was 17.12%-20.95% with encapsulation efficiency reaching 51.9%-54.7%. In vitro drug release tests showed a successful sustained release over 28 days with a burst of 19.98% of the total drug substance. Treatment with HA/parecoxib-loaded PLGA microspheres declined the mRNA expression of IL-1β, TNF-α and COX-2 induced by LPS in articular synovial cells. Moreover, in vivo results demonstrated that the intra-articular microspheres significantly reduced protein levels of inflammatory cytokines (IL-1β, TNF-α and COX-2) for more than two weeks and stopped the mechanically-induced synovitis in its tracks in rat models. The study presented new and potential insights into treatments of TMD using PLGA microspheres loaded with HA and parecoxib as a successful drug delivery system.

Nec-1 attenuates inflammation and cytotoxicity induced by high glucose on THP-1 derived macrophages through RIP1

ObjectivesThis research aimed to study whether necrostain-1 (Nec-1) could alleviate inflammatory injury induced by high glucose upon THP-1 derived macrophages through RIP1. DesignFirstly, THP-1 derived macrophages were incubated with 5.5 mM glucose (normal glucose, NG), 25 mM glucose (high glucose, HG), and mannitol as the high osmotic pressure group (5.5 mM glucose+19.5 mM mannitol) for 24, 48, and 72 h respectively. TNF-α, IL-1β, IL-6, and IL-8 levels were measured by ELISA. Secondly, macrophages were exposed to NG, HG, or HG plus 5 μM necrostatin-1 (Nec-1) for 72 h. mRNA expression of inflammatory cytokine was measured by RT-PCR, and protein levels of inflammatory cytokines and LDH leakage were determined by ELISA. RIP1 expression was determined by RT-PCR and WB. Thirdly, macrophages were transfected with si-RIP1 or negative control (si-NC). Wild type and RIP1-silenced macrophages were incubated with NG or HG, and TNF-α, IL-1β, IL-6, IL-8, and LDH levels were measured again by ELISA. Results1) TNF-α, IL-1β, IL-6, and IL-8 levels were elevated in the HG group, as compared with that the NG group. Inflammation remained unchanged in the mannitol group. 2) Inflammatory response and LDH levels in the HG plus Nec-1 group were remarkably lower than in the HG group. 3) Inflammatory injury in the si-NC group was more severe than in the si-RIP1 group. ConclusionsCurrent results indicated that Nec-1 could alleviate HG-caused inflammatory injury on THP-1 derived macrophages by regulating RIP1. These findings could help cast light on the relationships between diabetes and periodontitis.

Alpha-pinene exerts neuroprotective effects via anti-inflammatory and anti-apoptotic mechanisms in a rat model of focal cerebral ischemia-reperfusion

BackgroundIschemic stroke is a severe neurological disorder that affected millions of people worldwide. Neuro-inflammation and apoptosis play an essential role in the pathogenesis of neuronal death during ischemic stroke. Alpha-pinene is a bicyclic terpenoid with anti-inflammatory and anti-apoptotic activities. Accordingly, the main purpose of this study was to assess the protective effect of α-pinene in ischemic stroke. Materials and MethodsTo induce ischemic stroke in male Wistar rats, the middle cerebral artery was occluded for 60 min followed by 24 h reperfusion. Alpha-pinene was injected intraperitoneally at the beginning of reperfusion. A day after reperfusion, the neurological deficits, volume of infarct area, and blood-brain barrier (BBB) permeability were evaluated. The mRNA expression of inflammatory cytokines as well as pro- and anti-apoptotic genes was assessed by using reverse transcription-polymerase chain reaction. The protein levels of inflammatory cytokines were also measured by ELISA method. ResultsThe results showed that α-pinene (50 and 100 mg/kg) significantly improved sensorimotor function and decreased the volume of infarct area in the brain. The high permeability of BBB was also alleviated by α-pinene (50 and 100 mg/kg) in ischemic areas. Besides, α-pinene (100 mg/kg) attenuated neuro-inflammation through decreasing both the gene and protein expression of TNF-α and IL-1β in the hippocampus, cortex, and striatum. Besides, α-pinene (100 mg/kg) suppressed apoptosis via downregulation of the pro-apoptotic Bax mRNA expression with a concomitant upregulation of anti-apoptotic Bcl-2 gene expression. ConclusionsOverall, it was concluded that α-pinene exerts neuroprotective effect during ischemic stroke through attenuating neuroinflammation and inhibition of apoptosis.

Protective effects of rolipram on endotoxic cardiac dysfunction via inhibition of the inflammatory response in cardiac fibroblasts

BackgroundCardiac fibroblasts, regarded as the immunomodulatory hub of the heart, have been thought to play an important role during sepsis-induced cardiomyopathy (SIC). However, the detailed molecular mechanism and targeted therapies for SIC are still lacking. Therefore, we sought to investigate the likely protective effects of rolipram, an anti-inflammatory drug, on lipopolysaccharide (LPS)-stimulated inflammatory responses in cardiac fibroblasts and on cardiac dysfunction in endotoxic mice.MethodCardiac fibroblasts were isolated and stimulated with 1 μg/ml LPS for 6 h, and 10 μmol/l rolipram was administered for 1 h before LPS stimulation. mRNA levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in fibroblasts and their protein concentrations in supernatant were measured with real-time PCR (rt-PCR) and enzyme-linked immunosorbent assay, respectively. The expression of dual specificity phosphatase 1 (DUSP1), an endogenous negative regulator that inactivates MAPK-mediated inflammatory pathways, was also measured by rt-PCR and western blotting. DUSP1-targeted small interfering RNA (siRNA) was used to examine the specific role of DUSP1. To evaluate the role of rolipram in vivo, an endotoxic mouse model was established by intraperitoneal injection of 15 mg/kg LPS, and 10 mg/kg rolipram was intraperitoneally injected 1 h before LPS injection. mRNA and protein levels of inflammatory cytokines and DUSP1 in heart, inflammatory cell infiltration and cardiac function were all examined at 6 h after LPS injection.ResultsThe results showed that LPS could increase the expression and secretion of inflammatory cytokines and decrease the transcription and expression of DUSP1 in cardiac fibroblasts. However, rolipram pretreatment significantly reversed the LPS-induced downregulation of DUSP1 and inhibited LPS-induced upregulation and secretion of TNF-α and IL-6 but not IL-1β. Moreover, DUSP1-targeted siRNA experiments indicated that the protective effect of rolipram on inflammatory response was specific dependent on DUSP1 expression. Moreover, rolipram could further reduce inflammatory cell infiltration scores as shown by pathological analysis and increase the ejection fraction (EF) detected with echocardiography in the hearts of endotoxic mice.ConclusionsRolipram could improve endotoxin-induced cardiac dysfunction by upregulating DUSP1 expression to inhibit the inflammatory response in cardiac fibroblasts, which may be a potential treatment for SIC.

The Role of the Actin‐Binding Protein Cortactin in the Onset and Development of Sepsis

IntroductionSepsis is a devastating disease, which occurs as a result of an exacerbated immune response to systemic infection. Major hallmarks of sepsis include increased vascular permeability and leukocyte recruitment to secondary organs such as lung and kidney, often leading to organ dysfunction and, in severe cases, organ failure. Cortactin is an actin‐binding protein, expressed ubiquitously with the exception of neutrophils. Endothelial cortactin regulates vascular permeability and neutrophil recruitment by controlling activation of small GTPases. Therefore, endothelial cortactin could play an important role during sepsis pathogenesis by controlling neutrophil influx into affected organs such as lungs.Aim/ObjectiveTo elucidate the effects of cortactin deficiency on the onset and development of sepsis, which is a disease characterized by massive systemic neutrophil extravasation.Experimental StrategySepsis was induced by cecal ligation and puncture (CLP) in age‐matched male cortactin‐deficient (CTTN KO) and littermate wild‐type (WT) mice; and five day survival was monitored. Blood and lung tissue samples were collected 24 h after CLP for protein and gene expression analysis. Lung histology was analyzed after hematoxylin/eosin staining of paraffin‐crossections of lung tissue. Oxidative stress was assessed in lung cross‐sections using dihydroethidium. mRNA and protein levels of inflammatory cytokines were assessed by quantitative real‐time PCR (lung tissue) and bead‐based milliplex assay (peripheral blood) respectively. Neutrophil numbers were determined in blood and peritoneum by flow cytometry.Results and ConclusionsCortactin deficiency improved survival of mice subjected to CLP. Of note, lung tissue of septic KO animals showed better preserved architecture of the alveoli, and absence of oedema, hemorrhage and mucus deposition. Gene expression and secretion of the inflammatory mediators tumor necrosis factor‐α (TNFα) and interleukin‐1β in KO mice were reduced in comparison to septic WT mice. Moreover, we observed less oxidative stress in the lungs of septic KO mice in comparison to septic WT mice. Cortactin deficiency also protected against sepsis‐induced neutropenia. Additionally, in the absence of cortactin, less neutrophils were recruited to the peritoneum without compromising bacterial clearance. In summary, cortactin deficiency attenuates both local and systemic inflammatory responses during sepsis, protects against sepsis‐induced neutropenia and protects from lung tissue damage. Thus, cortactin could be an interesting target as novel treatment strategy for sepsis.Cortactin deficiency protects against sepsis‐induced lung damageFigure 1Expression of inflammatory cytokines in the lung are reduced in the absence of cortactinFigure 2

Anti-inflammation Effects of Sinomenine on Macrophages through Suppressing Activated TLR4/NF-κB Signaling Pathway.

Sinomenine (SN) has been used in the clinical treatment of systemic lupus erythematosus and rheumatoid arthritis for many years. Studies showed that SN held protective effects such as anti-inflammation, scavenging free radicals and suppressing immune response in many autoimmune diseases. The purpose of the present study is to explore the mechanism of anti-inflammation of SN on lipopolysaccharide (LPS)-induced macrophages activation and investigate whether the TLR4/NF-κB signaling pathway participated in. Macrophages isolated from mouse peritoneal cavity were stimulated by 1 µg/mL LPS for 24 h. And then the cells were treated with various concentrations of SN, TLR4 inhibitor respectively for additional 48 h. Drug toxicity was detected by MTT assay and Transwell experiment was used to assess chemotaxis. Furthermore, TLR4 and MyD88 mRNA levels were detected by real-time PCR. Western blotting was used to examine TLR4, MyD88 and phosphorylated IκB protein expression in macrophages. Immunofluorescence assay was applied to observe p65 NF-κB protein expression in macrophage nucleus. We extracted macrophages with high purity and activity from the abdominal cavity of mice. SN remarkably inhibited the chemotaxis and secretion function of LPS-stimulated macrophages. It also down-regulated both the protein levels of inflammatory cytokines (TNF-α, IL-1β and IL-6) and the RNA and protein levels of the key factors (TLR4, MyD88, P-IκB) in TLR4 pathway. The expression of p65 NF-κB protein in nuclei was down-regulated, which was correlated with a similar decrease in P-IκB protein level. In conclusion, SN can inhibit the LPS induced immune responses in macrophages by blocking the activated TLR4/NF-κB signaling pathway. These results may provide a therapeutic approach to regulate inflammatory responses.

Leukocyte Apheresis Using a Fiber Filter Suppresses Colonic Injury Through Calcitonin Gene-Related Peptide Induction.

The aim of this study was to address whether the therapeutic effect of leukocytapheresis (LCAP) depends on calcitonin gene- related peptide (CGRP) induction. An HLA-B27 transgenic rat model was treated with an LCAP column. The effects of LCAP on clinical, endoscopic, and histologic disease activity, the colony-forming ability of colony-forming unit (CFU)-granulocyte macrophages (GMs), colonic blood flow, and tissue expression of tumor necrosis factor (TNF)-α and CGRP were examined. Changes in the effects of LCAP after pretreatment with the CGRP antagonist CGRP8-37 were also observed. A dextran sulfate sodium-induced colitis rat model included treatment with CGRP, and the effect was assessed based on clinical, endoscopic, and histologic disease activity, colonic blood flow, the colony-forming ability of CFU-GMs, and tissue expression of inflammatory cytokines and CGRP receptor families. LCAP improved disease activity, enhanced colonic blood flow, and induced the bone marrow colony-forming ability of CFU-GMs with an increase in CGRP mRNA levels. These effects were abolished by pretreatment with CGRP8-37. The administration of CGRP suppressed colitis, promoting colonic blood flow, inducing bone marrow-derived cells, downregulating inflammatory cytokines, and upregulating receptor activity-modifying protein-1. The mRNA and protein levels of inflammatory cytokines in lipopolysaccharide-stimulated mononuclear cells were also decreased after CGRP treatment. The therapeutic effects of LCAP depend on CGRP induction. CGRP can effectively suppress colitis through the downregulation of inflammatory events and upregulation of protective events.

Long non-coding RNA zinc finger antisense 1 expression associates with increased disease risk, elevated disease severity and higher inflammatory cytokines levels in patients with lumbar disc degeneration.

This study aimed to investigate the correlation of long noncoding RNA zinc finger antisense 1 (lncRNA ZFAS1) expression with disease risk, disease severity and inflammatory cytokines levels in lumbar disc degeneration (LDD) patients.83 LDD patients underwent surgery and 28 traumatized, non-LDD patients underwent lumbar disc surgery (controls) were consecutively enrolled in this case-control study. Lumbar disc tissue was obtained during surgery and herniated nucleus pulposus (HNP) was isolated to detect lncRNA ZFAS1 expression and inflammatory cytokines mRNA levels by RT-qPCR, and determine protein levels of inflammatory cytokines by western blot.HNP lncRNA ZFAS1 expression in LDD patients was up-regulated compared with controls (P < .001), and receiver operating characteristic (ROC) curve showed lncRNA ZFAS1 expression disclosed a good predictive value for LDD risk with area under curve (AUC) 0.753 (95% CI 0.646–0.859). And after adjustment by age, gender and body mass index (BMI), lncRNA ZFAS1 (P = .017) remained to be an independent predictive factor for higher LDD risk. In addition, lncRNA ZFAS1 expression was positively associated with Modified Pfirrmann Grade (P = .015). As to inflammatory cytokines, lncRNA ZFAS1 expression was observed to be positively correlated with TNF-α (P = .002), IL-1β (P = .007) and IL-6 (P = .015) mRNAs expressions while reversely associated with IL-10 mRNA level (P = .014); and lncRNA ZFAS1 expression was also positively correlated with protein levels of TNF-α (P = .038) and IL-6 (P = .027) while reversely associated with IL-10 protein expression (P = .039).lncRNA ZFAS1 expression associates with increased risk, elevated disease severity and higher inflammatory cytokines levels in LDD patients.

Long non-coding RNA MEG3 regulates CSE-induced apoptosis and inflammation via regulating miR-218 in 16HBE cells

Chronic obstructive pulmonary disease (COPD) is a prevalent disease worldwide, mainly caused by cigarette smoking. Maternally expressed gene 3 (MEG3) functions as the lncRNA and is upregulated in COPD patients and human bronchial epithelial cells after fine particulate matter (PM2.5) treatment. However, the molecular mechanism of MEG3 in COPD remains unknown. The expression of MEG3 and miR-218 in COPD tissues and cigarette smoke extract (CSE)-treated 16HBE cells was detected by RT-qPCR. The effects of MEG3 and miR-218 on proliferation and apoptosis in (CSE)-treated 16HBE cells were analyzed by CCK-8 and flow cytometry assay, respectively. The protein levels of inflammatory cytokines (IL-1β IL-6 and TNF-α) were detected in 16HBE cells by ELISA. MEG3 and miR-218 binding interaction was predicted by LncBase Predicted v.2 and further confirmed by dual luciferase reporter assay and RNA Immunoprecipitation (RIP) assay. MEG3 was upregulated in COPD tissues and inversely related to FEV1%. MEG3 was upregulated in (CSE)-treated 16HBE cells, and knockdown of MEG3 mitigated CSE-repressed proliferation and CSE-triggered apoptosis or inflammation. MiR-218 was demonstrated as a target miRNA of MEG3. MiR-218 was downregulated in COPD tissues and (CSE)-treated or MEG3 overexpressed 16HBE cells. MiR-218 overexpression attenuated CSE-blocked proliferation and CSE-induced apoptosis or inflammation. Deficiency of MEG3 counteracted CSE-blocked proliferation CSE-induced apoptotic rate and inflammatory cytokine (IL-1β IL-6 and TNF-α) levels, while introduction of anti-miR-218 reversed these effects. MEG3 regulated CSE-inhibited proliferation and CSE-induced apoptosis or inflammation by targeting miR-218, providing a possible therapeutic target for treatment of CSE-induced COPD.

IL-35 alleviates inflammation progression in a rat model of diabetic neuropathic pain via inhibition of JNK signaling

BackgroundEmerging evidence has demonstrated that inflammation is involved in the occurrence and development of diabetic neuropathic pain (DNP). The anti-inflammatory property of interleukin (IL)-35 makes it a promising candidate to block the pain perception. The present study was undertaken to investigate whether IL-35 could attenuate DNP in streptozotocin (STZ)-induced rat model and its potential mechanism.MethodsThe rat model of DNP was established by a single STZ injection followed by measurements of fasting blood glucose and insulin. Fourteen days after STZ injection, DNP rats were intrathecally injected with IL-35, c-Jun N-terminal kinase (JNK) inhibitor or activator or dimethylsulfoxide (DMSO) as vehicle control, respectively. The mechanical allodynia was assayed to evaluate the therapeutic effect of IL-35. In mechanism study, the serum and protein levels of inflammatory cytokines using ELISA and western blotting and the activation of JNK signaling were further evaluated by quantitative reverse transcription PCR (qRT-PCR). Histopathologic changes were evaluated by Nissl staining. Apoptosis was examined using TUNEL staining.ResultsDNP rats exhibited increased fasting blood glucose and insulin levels and reduced insulin sensitivity index (ISI). Intrathecal injection of IL-35 reduced accumulation of pro-inflammatory cytokines in the spinal cord of DNP rats. Furthermore, IL-35 displayed anti-inflammatory and anti-apoptotic effects via inhibition of JNK pathway.ConclusionIL-35 treatment mitigated DNP via downregulating JNK signaling pathway.

Losartan, an Angiotensin II Type 1 Receptor Antagonist, Alleviates Mechanical Hyperalgesia in a Rat Model of Chemotherapy-Induced Neuropathic Pain by Inhibiting Inflammatory Cytokines in the Dorsal Root Ganglia.

Chemotherapy-induced peripheral neuropathy (CIPN) adversely impacts quality of life and a challenge to treat with existing drugs used for neuropathic pain. Losartan, an angiotensin II type 1 receptor (AT1R) antagonist widely used to treat hypertension, has been reported to have analgesic effects in several pain models. In this study, we assessed losartan's analgesic effect on paclitaxel-induced neuropathic pain (PINP) in rats and its mechanism of action in dorsal root ganglion (DRG). Rats received intraperitoneal injections of 2mg/kg paclitaxel on days 0, 2, 4, and 6 and received single or multiple intraperitoneal injections of losartan potassium dissolved in phosphate-buffered saline at various times. The mechanical thresholds, protein levels of inflammatory cytokines, and cellular location of AT1R and interleukin 1β (IL-1β) in the DRG were assessed with behavioral testing, Western blotting, and immunohistochemistry, respectively. Data were analyzed by two-way repeated-measures analysis of variance for the behavioral test or the Mann-Whitney U test for the Western blot analysis and immunohistochemistry. Single and multiple injections of losartan ameliorated PINP, and losartan delayed the development of PINP. Paclitaxel significantly increased, and losartan subsequently decreased, the expression levels of inflammatory cytokines, including IL-1β and tumor necrosis factor α (TNF-α), in the lumbar DRG. AT1R and IL-1β were expressed in both neurons and satellite cells and losartan decreased the intensity of IL-1β in the DRG. Losartan ameliorates PINP by decreasing inflammatory cytokines including IL-1β and TNF-α in the DRG. Our findings provide a new or add-on therapy for CIPN patients.

Deletion of Pregnancy Zone Protein and Murinoglobulin-1 Restricts the Pathogenesis of West Nile Virus Infection in Mice.

West Nile virus (WNV) is an enveloped positive-stranded RNA virus that causes meningitis, encephalitis, and acute flaccid paralysis in humans. There are no therapeutic agents available for use against WNV infection. Alpha-2 macroglobulin (A2M) is a major plasma proteinase inhibitor that also has important role in immune modulation. In mice, pregnancy zone protein (PZP) and murinoglobulin-1 (MUG-1) are two close homologous of human A2M. In this study, we investigated the role of PZP and MUG-1 proteins in the pathogenesis of WNV infection in mice. Adult C57BL/6J wild-type and PZP/MUG-1 double knockout (DKO) mice were inoculated subcutaneously with WNV and mortality, virus burden, and immune responses were analyzed. Infection of wild-type (WT) mice with WNV resulted in significantly high morbidity and mortality. In comparison, no mortality was observed in DKO mice, suggesting that PZP and MUG-1 play a deleterious role in WNV infection. Increased survival in WNV-infected DKO mice was associated with significantly low viral burden in serum, spleen, kidney, and brain compared to WT mice. In addition, significantly reduced levels of type 1 interferon and WNV-specific antibodies were observed in the DKO mice compared to WT mice. We further demonstrated that protein levels of inflammatory cytokines and chemokines in the serum, spleen, and brain were significantly reduced in DKO mice compared to WT mice. Collectively our data demonstrate that lack of PZP and MUG-1 restricts the pathogenesis of WNV infection in mice.

Sodium Fluoride (NaF) Induces Inflammatory Responses Via Activating MAPKs/NF-κB Signaling Pathway and Reducing Anti-inflammatory Cytokine Expression in the Mouse Liver.

At present, no reports are focused on fluoride-induced hepatic inflammatory responses in human beings and animals. This study aimed to investigate the mRNA and protein levels of inflammatory cytokines and signaling molecules for evaluating the effect of different doses (0, 12, 24, and 48mg/kg) of sodium fluoride (NaF) on inflammatory reaction in the mouse liver by using methods of experimental pathology, quantitative real-time polymerase chain reaction (qRT-PCR), and western blot analysis. We found that NaF in excess of 12mg/kg caused the hepatic inflammatory responses, and the results showed that NaF activated the mitogen-activated protein kinases (MAPKs) signaling pathway by markedly increasing (p < 0.01 or p<0.05) mRNA and protein levels of apoptosis signal-regulating kinase 1 (ASK1), mitogen-activated protein kinase kinases 1/2 (MEK1/2), extracellular signal-regulated protein kinases 1/2 (Erk1/2), mitogen-activated protein kinase kinases 4/7 (MEK4/7), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38) and mitogen-activated protein kinase kinases 3/6 (MEK3/6), and the nuclear factor-kappa B (NF-κB) signaling pathway by increasing (p < 0.01 or p<0.05) the production of NF-κB and inhibitor of nuclear factor kappa-B kinase subunit beta (IKK-β) and reducing (p < 0.01 or p<0.05) the production of the inhibitory kappa B (IκB). Thus, NaF that caused the hepatic inflammatory responses was characterized by increasing (p < 0.01 or p<0.05) the production of pro-inflammatory mediators such as interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-8 (IL-8), monocyte chemotactic protein 1 (MCP-1), and cyclooxygenase-2 (COX-2) via the activation of MAPKs and NF-κB pathways, and by significantly inhibiting (p < 0.01 or p<0.05) the production of anti-inflammatory mediators including interleukin-4 (IL-4) and transforming growth factor beta (TGF-β).

CD200Fc Attenuates Retinal Glial Responses and RGCs Apoptosis After Optic Nerve Crush by Modulating CD200/CD200R1 Interaction.

To explore the hypothesis that CD200Fc, a CD200R1 agonist with anti-inflammatory properties, will inhibit retinal glial cells hyperactivation and retinal ganglion cells (RGCs) apoptosis after optic nerve injury. CD200Fc was immediately administered after optic nerve crush (ONC) once by intravitreal injection. Rats were euthanized at 5days after ONC. The density of RGCs was counted by immunostaining of retina flat mounts for Brn3a. TUNEL assay, immunoblotting analysis of ionized calcium-binding adapter molecule 1(iba1) (microglia marker) and glial fibrillary acidic protein (GFAP) (astrocytes and Müller cells marker), RT-PCR analysis of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor-α (TNF-α), interleukin (IL)-8 and IL-10, ELISA measure protein levels of inflammatory cytokines and western blot analysis of CD200 and CD200R1 were evaluated. CD200Fc treatment suppressed ONC-induced RGCs loss through inhibition of RGCs apoptosis. Additionally, expression of glial cells activation markers GFAP and iba1 and production of pro-inflammatory cytokines (COX-2, iNOS, MCP-1, TNF-α, IL-8) were decreased in CD200Fc treated animals after ONC. Meanwhile, anti-inflammatory cytokine IL-10 was increased by CD200Fc treatment in ONC-induced rat retina. Finally, we found that CD200Fc significantly inhibited ONC-induced increased in expression of CD200 and raised the already high basal CD200R1 expression in the rat retina after ONC. Our results demonstrated that the anti-inflammatory effects of CD200Fc in ONC rats model through inhibited the activation of retinal glial cells via the interaction between CD200 and CD200R1, and the neuroprotective effects of CD200Fc on RGCs thought inhibited its apoptosis.

Resolvin D1 attenuates imiquimod-induced mice psoriasiform dermatitis through MAPKs and NF-κB pathways

BackgroundResolvin D1 (RvD1), a pro-resolution lipid mediator derived from docosahexaenoic acid (DHA), has been described to promote several kinds of inflammatory resolution. However, the effects and anti-inflammatory mechanisms of RvD1 on psoriasis have not been previously reported. ObjectiveThe present study aimed to determine the protective effects and the underlying mechanisms of RvD1 on imiquimod (IMQ)-induced psoriasiform dermatitis. MethodsMice were topically treated with IMQ to develop psoriasiform dermatitis on their shaved back, pretreated intraperitoneally (i.p.) with or without RvD1 or tert-butoxycarbonyl Met-Leu-Phe peptide (Boc), a lipoxin A4 (ALX) receptor antagonist. The severity was monitored and graded using a modified human scoring system, the Psoriasis Area and Severity Index (PASI), histopathology, and the signature cytokines of psoriasis (IL-23, IL-17, IL-22 and TNF-α). The mRNA and protein levels of inflammatory cytokines were quantified by quantitative real-time PCR (QRT-PCR) and ELISA. The expressions of signaling proteins MAPKs and NF-κB p65 were analyzed using western blotting. Electrophoretic mobility shift assay (EMSA) was used to check NF-κB p65 DNA binding activity. ResultsOur study showed that RvD1 alleviated IMQ-induced psoriasiform dermatitis and improved skin pathological changes. RvD1 markedly inhibited IMQ-induced activation of ERK1/2, p38, JNK (c-Jun N-terminal protein kinase, a subfamily of MAPKs), and NF-κB. Furthermore, pretreatment with Boc, would not exacerbate skin inflammation of IMQ-induced mice, but significantly reversed the beneficial effects of RvD1 on IMQ-induced psoriasiform inflammation. ConclusionRvD1 can obviously improve skin inflammation in IMQ-induced mice psoriasiform dermatitis. The protective mechanisms might be related to its selective reaction with lipoxin A4 receptor/Formyl-peptide receptor 2 (ALX/FPR2), by downregulating relevant cytokines of the IL-23/IL-17 axis expression, the inhibition of MAPKs and NF-κB signaling transduction pathways. Thus, these results show that RvD1 could be a possible candidate for psoriasis therapy.

(+)-Borneol improves the efficacy of edaravone against DSS-induced colitis by promoting M2 macrophages polarization via JAK2-STAT3 signaling pathway

Compound edaravone injection (C.EDA), a compound preparation composed of edaravone (EDA) and (+)-Borneol with the mass ratio of 4: 1, displays a better anti-inflammatory activity than EDA. However, its precise mechanism remains to be further studied. In this work, we investigated whether (+)-Borneol could improve the efficacy of EDA against DSS-induced colitis. We found that C.EDA at 7.5 and 15mg/kg could significantly relieve the disease activity index (DAI) and reduce the loss of body weight and colon length in a dose-dependent manner, while EDA or (+)-Borneol alone only had moderate effects even at the highest dose. Additionally, ELISA revealed that C.EDA could more dramatically decrease the protein levels of inflammatory cytokines and increase the levels of anti-inflammatory cytokine than EDA or (+)-Borneol alone both in colon tissues and serum. H&E staining and IHC assay also indicated that C.EDA exhibited more prominent effects on increasing the population of M2 macrophages, decreasing M1 macrophages infiltration and protecting intestinal barrier integrity. Furthermore, in vitro studied demonstrated that C.EDA, EDA or (+)-Borneol failed in inhibiting M1 macrophages activation but could specifically induce the activation of M2 macrophages in a STAT3-dependent manner. Knockdown the expression of STAT3 successfully abolished the effect of C.EDA and EDA on promoting M2 macrophages activation. Consistent with in vivo study, C.EDA exhibited a more efficient ability of inducing M2 macrophages polarization and STAT3 activation than EDA or (+)-Borneol alone in vitro. In conclusion, we confirmed that (+)-Borneol improved the efficacy of EDA against DSS-induced colitis by promoting M2 macrophages polarization via JAK2-STAT3 signaling pathway.

Neuroprotective effect of liquiritin against neuropathic pain induced by chronic constriction injury of the sciatic nerve in mice

Managing of neuropathic pain remains clinically challenging because the existing pharmacotherapies are either ineffective or non-specific. Therefore, developing novel alternatives is essential for better treatment. Liquiritin is an active component extracted from Glycyrrhizae radix and has potential neuroprotective action. This study aimed to investigate the protective efficacy of liquiritin on chronic constriction injury (CCI)-induced neuropathic pain in mice. Liquiritin (30, 60, and 120mg/kg) and pregabalin (40mg/kg) were administered intragastrically for 7 consecutive days starting on the 8th day post-surgery. Behavioral parameters and sciatic functional index were assessed on days 0, 7, 8, 10, 12, and 14. Electrophysiological and histopathological changes were analyzed on the 14th day. Immunofluorescence and Western blot were used to evaluate the expression of glial cells and the protein levels of inflammatory cytokines in the spinal cord, respectively. Results showed that liquiritin dose-dependently reduced hyperalgesia and allodynia and increased the sciatic functional index and motor nerve conduction velocities. Moreover, liquiritin restored the injured axon and myelin sheath, inhibited the activation of astrocyte and microglia, down-regulated the pro-inflammatory cytokines including tumor necrosis factor alpha (TNF-α), interleukin (IL-6, and IL-1β), and simultaneously up-regulated the anti-inflammatory cytokine IL-10. Our study revealed that liquiritin exerted a neuroprotective effect on CCI-induced neuropathic pain, which might be attributed to its direct protective effect on damaged nerves and its anti-inflammatory activity at the level of the spinal cord. Therefore, liquiritin shows promise as a compound for the development of novel analgesic agents that can be used to effectively treat intractable neuropathic pain.

Thalidomide alleviates postoperative pain and spatial memory deficit in aged rats

Pain is a major risk factor of post-operative cognitive dysfunction (POCD) in aged population. We investigated the effects of thalidomide, an anti-inflammatory and analgesic drug, on POCD in aged rats, and also explored the underlying mechanisms. Laparotomy was performed under anesthesia in aged rats (24–25 months) to establish POCD models. Thalidomide (5–50mg/kg) was intraperitoneally administered immediately after laparotomy. Within 12h after the operation, pain symptoms were assessed by rat grimace scale (RGS). Within postoperative day (POD) 3–14, spatial memory was evaluated using performance errors in a radial arm maze. Protein levels of inflammatory cytokines and N-methyl-D-aspartate (NMDA) receptors were measured on POD 14. POCD rats treated with thalidomide showed decreased RGS and performance errors, compared with saline-treated POCD rats. Single administration of thalidomide significantly reduced production of cytokines (tumor necrosis factor (TNF)-α and interleukin (IL)-1β) in serum but not in the brain, and attenuated upregulation of NMDA receptor (NR) 2A/B subunits in the hippocampus at POD 14. MK-801 abolished thalidomide-induced attenuation of spatial memory deficits. Our results support that thalidomide could disrupt the development of post-operative memory deficit in aged rats through its long-term regulation of NMDA receptors (NRs) in the hippocampus. Therefore, thalidomide might provide a new means to prevent the development of POCD.