TNF-α Protein Research Articles

Biancaea decapetala (Roth) O.Deg. extract exerts an anti-inflammatory effect by regulating the TNF/Akt/NF-κB pathway

BackgroundBiancaea decapetala (Roth) O.Deg. (Fabaceae) is used to treat colds, fever, and rheumatic pain caused by inflammation. However, the mechanism underlying its anti-inflammatory properties remains unclear. PurposeThis study aimed to evaluate the anti-inflammatory activity of Biancaea decapetala extract (BDE) in vitro and in vivo and explore the possible underlying mechanism and potential targets. MethodsThe release of nitric oxide (NO) and inflammatory cytokines in LPS-stimulated RAW264.7 cells and rats were measured using Griess reagent and enzyme-linked immunosorbent assay (ELISA). Hematoxylin and eosin (H&E) staining was employed to examine the pathology of animal tissues. Transcriptome analysis was performed to screen the pathways related to BDE-mediated inhibition of inflammation, and the expression of related proteins was measured using real-time quantitative polymerase chain reaction (RT-qPCR), western blotting, ELISA, and immunofluorescence methods. Surface Plasmon Resonance (SPR) and the Drug Affinity Reaction Target Stability (DARTS) method were used to verify whether BDE binds to TNF-α target protein, while a L929 cell model and NF-κB gene reporter systematic method were used to investigate the inhibitory effect of BDE on the activity of TNF-α protein. ResultsBDE inhibited the expression of TNF-α, IL-1β, IL-6, and NO in RAW264.7 cells and rats, and improved the pathological changes in lung tissue. RNA-seq showed that BDE may regulate the TNF/Akt/NF-κB pathway to inhibit inflammation onset. BDE significantly downregulated the mRNA expression of TNF-α, IL-6, IL-1β, and that of relevant proteins, including TNF-α, p-p65, p-Akt, p-IκBα. Furthermore, BDE inhibited the nuclear translocation of NF-κB (p65) and the activation of the Akt pathway by SC79. The L929 cell model, luciferase reporter gene analysis, DARTS, and SPR experiments showed that BDE may bind to TNF-α and inhibit the TNF-α-NF-κB pathway. ConclusionBDE may target TNF-α to inhibit the TNF/Akt/NF-κB pathway, thereby attenuating inflammation. These findings reveal the anti-inflammatory effects and mechanisms of BDE and provide a theoretical basis for the further development and utilization of BDE.

TLR4, MyD88, and TNF-α Expression in the Lungs of Akkaraman and Romanov Lambs in Response to LPS and LTA.

Toll-like receptors are involved in the recognition of bacterial toxins, which cause infection in the respiratory system. This study aimed to evaluate microanatomical and histological alterations in the lungs of 24 healthy Akkaraman and Romanov lambs after the administration of lipoteichoic acid (LTA), lipopolysaccharide (LPS), and LTA + LPS and investigate the gene, protein, and immune expression levels of TLR4, MyD88, and TNF-α molecules, known to have immune functions. Microanatomical examinations showed thickened peribronchial and alveolar walls in the lungs of groups LTA, LPS, and LTA + LPS of both breeds due to immune cell infiltration. TLR4, MyD88, and TNF-α immunoexpressions were positive to varying degrees in the cytoplasm and nucleus of the bronchial and bronchiolar luminal epithelial cells, alveolar epithelial cells, and alveolar macrophages. TLR4 and TNF-α protein expressions were statistically different in the LPS-treated Romanov lambs, compared to the other groups. Among the Akkaraman lambs, TLR4 gene expression was significantly higher in group LPS, and among the Romanov lambs, TLR4, MyD88, and TNF-α gene expressions were significantly higher in group LTA + LPS. Therefore, TLR4, MyD88, and TNF-α molecules, involved in the immune response, were found to be expressed at different levels against LTA and LPS in the lungs of two different sheep breeds.

Dyrk1a Phosphorylation of α-Synuclein Mediating Apoptosis of Dopaminergic Neurons in Parkinson's Disease.

To investigate the role of aberrant Dyrk1a expression in phosphorylation modification at the α-synuclein serine 129 (Ser129) site to analyze its molecular mechanism in mediating apoptosis of PD. The protein level of P-α-synuclein (Ser129), α-synuclein, Bcl-2, Bax, active caspase 3, GSK3β, PI3K, AKT, and cyclinD1 were detected. The mRNA transcript levels of Dyrk1a and DAT and protein levels of IL-1β, IL-6, COX-2, and TNF-α were detected. P-α-synuclein (Ser129), α-synuclein, Bax, active caspase 3, GSK3β, and cyclinD1 expressions were decreased in Dyrk1a-AAV-ShRNA (P < 0.05), and Bcl-2, AKT, and PI3K expressions were increased (P < 0.05). Increased TH protein expression was shown in Dyrk1a-AAV-ShRNA (P < 0.05). Dyrk1a mRNA was decreased in the Dyrk1a-AAV-ShRNA group (P < 0.05), and DAT mRNA was increased (P < 0.05). IL-1β, IL-6, COX-2, and TNF-α protein levels were decreased in Dyrk1al-AAV-Sh-RNA (P < 0.05). Transcriptome sequencing showed that Fam220a, which was expected to activate STAT family protein binding activity and participate in the negative regulation of transcription through RNA polymerase II and protein dephosphorylation showed differentially upregulated expression. The untargeted metabolome showed that the major compounds in the Dyrk1a-AAV-ShRNA group were hormones and transmission mediators and the most metabolism-related pathways. Fam220a showed differentially upregulated expression, and differentially expressed genes were enriched for the neuroactive ligand-receptor interaction, vascular smooth muscle contraction, and melanogenesis-related pathways. Abnormal Dyrk1a expression can affect α-synuclein phosphorylation modifications, and dyrk1a knockdown activates the PI3K/AKT pathway and reduces dopaminergic neuron apoptosis. It provides a theoretical basis for the group to further investigate the molecular mechanism.

HWJMSCs inhibit inflammation and apoptosis in an ARDS cell model

Acute respiratory distress syndrome (ARDS) is a type of lung failure caused by fluids and hypoxemia. Mesenchymal stem cells (MSCs) have been shown to decrease levels of pro-inflammatory mediators and inflammatory cells. These cells have anti-inflammatory, anti-apoptotic, and anti-microbial activity, and protect against lung injury. ObjectiveThis research evaluated the potential of human Wharton's jelly MSCs (hWJMSCs) to inhibit inflammation and apoptosis in lipopolysaccharide (LPS)-induced rat lung cells (L2). MethodshWJMSC treatment in LPS-induced rat lung cells was performed with 1:1, 1:5, 1:10, or 1:25 ratios of hWJMSCs to L2 cells. The gene expression of angiotensin-converting enzyme-2 (ACE-2), receptor for advanced glycation end products (RAGE), nuclear factor kappa B (NFκB), and C-X-C motif chemokine ligand-9 (CXCL-9) was quantified with RT-PCR, and the levels of C-reactive protein (CRP), interleukin-12 (IL-12), and tumor necrosis factor-alpha (TNF-α) were measured with ELISA. ResultshWJMSCs increased ACE-2 gene expression, and decreased CXCL-9, NFκB, and RAGE gene expression. The treatment also suppressed CRP, TNF-α, and IL-12 levels, and increased the percentage of live cells, but decreased the percentages of necrotic cells and apoptotic cells in inflammatory rat lung cells, which served as an ARDS cell model. ConclusionCo-culture of hWJMSCs and L2 cells mitigated inflammation through increasing ACE-2 gene expression, and decreasing CXCL-9, NFκB, and RAGE gene expression; decreasing TNF-α and CRP protein levels; and decreasing necrosis, and early and late apoptosis. A co-culture ratio of 1:1 was most effective.

Associations between blood inflammatory markers and bone mineral density and strength in the femoral neck: findings from the MIDUS II study

Chronic and systematic inflammation have been related to increased risks of osteopenia and related fracture. However, studies concerning the association between low-grade inflammation and the bone mineral density (BMD) and strength of the femoral neck are still few and inconsistent. This study aimed to examine the relationships between blood inflammatory biomarkers and BMD and femoral neck strength in an adult-based cohort. We retrospectively analyzed a total of 767 participants included in the Midlife in the United States (MIDUS) study. The blood levels of inflammatory markers, including interleukin-6 (IL6), soluble IL-6 receptor, IL-8, IL-10, TNF-α and C-reactive protein (CRP), in these participants were measured, and their associations with the BMD and strength of the femoral neck were determined. We analyzed these 767 subjects with data concerning the BMD, bending strength index (BSI), compressive strength index (CSI), and impact strength index (ISI) in the femoral neck and inflammatory biomarkers. Importantly, our results suggest that strongly negative associations exist between the blood soluble IL6 receptor levels and the BMD (per SD change, Sβ = −0.15; P < 0.001), CSI (per SD change, Sβ = −0.07; P = 0.039), BSI (per SD change, Sβ = −0.07; P = 0.026), and ISI (per SD change, Sβ = −0.12; P < 0.001) in the femoral neck after adjusting for age, gender, smoked cigarettes regularly, number of years drinking, BMI and regular exercise. However, the inflammatory biomarkers, including blood IL-6 (per SD change, Sβ = 0.00; P = 0.893), IL-8 (per SD change, Sβ = −0.00; P = 0.950), IL-10 (per SD change, Sβ = −0.01; P = 0.854), TNF-α (per SD change, Sβ = 0.04; P = 0.260) and CRP (per SD change, Sβ = 0.05; P = 0.137), were not strongly associated with the BMD in the femoral neck under the same conditions. Similarly, there was no significant difference in the relationships between the inflammatory biomarkers (IL-6, IL-8, IL-10, TNF-α and CRP) and the CSI, BSI, and ISI in the femoral neck. Interestingly, in concomitant inflammation-related chronic diseases, only arthritis affected the soluble IL-6 receptor and the CIS (interaction P = 0.030) and SIS (interaction P = 0.050) in the femoral neck. In this cross-sectional analysis, we only observed that high blood levels of soluble IL-6 receptor were strongly associated with reduced BMD and bone strength in the femoral neck. The independent associations between the other inflammatory indicators, including IL-6, IL-8, IL-10, TNF-α and CRP, and the BMD and femoral neck strength in an adult-based cohort were not significant.

Molecular docking analysis of natural compounds as TNF-α inhibitors for Crohn's disease management.

Crohn's disease (CD) is a type of inflammatory bowel disease that is immune-mediated and affects the gastrointestinal tract. The chronic and severe nature of this condition leads to diminished health-related life quality, and frequent hospitalization. While medications such as sulfasalazine, corticosteroids, and immuno-suppressants are used to manage the condition, there are no definite treatments for pain and inflammation associated with CD. TNF-α is a prominent target, and medicines such as infliximab and adalimumab have pharmacological efficacy; however, they also have significant toxicity. Here, the natural compound library (2706 compounds) was screened against TNF-α to find natural TNF-α inhibitors to combat CD. The compounds namely ZINC5223934, ZINC6482465, ZINC4098633, ZINC1702729, and ZINC4649679 had higher binding affinity and interaction with the TNF-α protein than the positive control. Furthermore, these compounds had promising drug-like properties, indicating their potential for future exploration and optimization as TNF-α inhibitors for the treatment of CD.

Etanercept Ameliorated cerebral Damage During Global Cerebral Ischemia-Reperfusion Injury in Male Rats

Background: Ischemic cerebrovascular conditions lead to brain tissue oxygen depletion, triggering activation and infiltration of inflammatory cells and immunological reactions. Reperfusion induces the release of reactive oxygen species (ROS), triggered by therapeutic recanalization or natural thrombus disintegration. Method: Rats were divided into four groups after a week of acclimatization: Sham group: No occlusion of common carotid arteries after general anesthesia. Ischemic-reperfused control group: General anesthesia, followed by 30 minutes of common carotid artery occlusion and one hour of reperfusion without drugs. Vehicle group: Similar to control group but with intraperitoneal (i.p.) administration of DMSO 30 minutes prior to ischemia. Etanercept group: Identical procedure as control, with additional intraperitoneal (i.p.) administration of etanercept (50 mg/kg) 30 minutes before ischemia. Results: Etanercept groups showed significant reduction in NF-kB and TLRs (2 and 4) compared to control (P&lt;0.05). Histologically, brain tissue lesions were reduced in etanercept groups compared to control and vehicle groups (P&lt;0.05). Conclusion: Etanercept exhibits neuroprotective effect against cerebral I/R injury by down-regulating TNF-α protein and inflammation TLRs.

Effect of Adipose Derived Stromal Vascular Fraction on The Early Inflammatory Phase of Burn Wound Healing in Male Rat Model

Objectives: Severe burn treatment remains challenging. Research has investigated the efficacy of stem cells and stromal vascular fraction (SVF) in many types of injuries. SVF has advantages for its heterogenous population of cells and avoidance of culturing and ethical issues. SVF can be isolated either enzymatically or mechanically. Enzymatically isolated SVF reduced inflammation and enhanced neovascularization and re-epithelization in the treatment of burn injury. This study highlighted the efficacy of applying mechanically isolated SVF on the early inflammatory phase of deep partial-thickness burn by investigating toll-like receptor 4 (TLR4) signaling pathway.&#x0D; Methods: 30 male Wistar rats with deep partial-thickness burns were assigned into: the control group, silver sulfadiazine (SSD) cream group, and mechanically isolated SVF group. Morphological, histopathological evaluation of inflammation, and immunohistochemical analysis for (TLR4, TNF-α, IL‐1β, and IL-6) were evaluated for the first eight days post-treatment.&#x0D; Results: Morphologically, the SVF group significantly reduced edema and increased wound bed dryness on day one compared to the control group (P=0.0001) and to the SSD group (P=0.001). Histopathologically, the SVF group significantly reduced inflammation four days post-treatment compared with the control group (P=0.045). The SVF group significantly reduced TLR4 protein expression on days four and eight post-treatment compared with the control group (P= 0.001, P= 0.042, respectively). The TNF-α protein expression in the SVF group was significantly lower on days four and eight post-treatment compared with the control group (P= 0.046, P= 0.046, respectively) and with the SSD group (P= 0.008, P= 0.001, respectively). The IL‐1β expression was significantly reduced in the SVF group compared to the control group on day four post-treatment (P= 0.017). There were no significant differences in IL-6 expressions between all groups on both days.&#x0D; Conclusion: The mechanical isolation of SVF has an early anti-inflammatory impact on deep partial-thickness burn injury. This effect could be through inhibiting TLR4, TNF-α, and IL‐1β pathways. This could partially explain the mechanism behind SVF efficacy in the healing process of burn injury.

EXTRACORPOREAL CARDIOPULMONARY RESUSCITATION WITH THERAPEUTIC HYPOTHERMIA MITIGATES KIDNEY INJURY AFTER CARDIAC ARREST IN RATS.

Many patients with cardiac arrest (CA) experience severe kidney injury after the return of spontaneous circulation. This study aimed to compare the renal protective effect of conventional cardiopulmonary resuscitation (CCPR), extracorporeal cardiopulmonary resuscitation (ECPR), and ECPR with therapeutic hypothermia (ECPR+T) in a CA rat model. Twenty-four adult male Sprague-Dawley rats were randomly and equally allocated into the sham, CCPR, ECPR, and ECPR+T groups. The sham group underwent basic surgical procedures without asphyxia-induced CA. The other three groups were treated with asphyxiation to establish the CA model. Subsequently, they were rescued using three different therapeutic methods. The end points were 1 h after return of spontaneous circulation or death. Renal injury was evaluated by histopathology. Oxidative stress, endoplasmic reticulum stress, necroptosis, inflammatory, and apoptosis-related genes, and proteins were detected using western blotting, ELISA, and assay kit. Compared with CCPR, ECPR and ECPR+T alleviated oxidative stress by upregulating nuclear factor erythroid 2-related factor 2, superoxide dismutase, glutathione and downregulating heme oxygenase-1, and malondialdehyde. Expression of endoplasmic reticulum stress-related proteins, glucose-regulated protein 78, and CCAAT/enhancer-binding protein homologous protein was lower in ECPR and ECPR+T groups than that in the CCPR group, along with levels of TNF-α, IL-6, and IL-β, and necroptosis proteins (receptor-interacting serine/threonine kinases 1 and 3). Furthermore, the ECPR and ECPR+T groups had significantly increased B-cell lymphoma 2 and decreased B-cell lymphoma 2-associated X levels compared with the CCPR group. Extracorporeal cardiopulmonary resuscitation and ECPR+T alleviate kidney damage after CA in rats compared with CCPR. Furthermore, ECPR+T had a better renal protective effect.

The Protective Effect of Nutraceuticals on Hepatic Ischemia-Reperfusion Injury in Wistar Rats.

Nutraceuticals are bioactive compounds present in foods, utilized to ameliorate health, prevent diseases, and support the proper functioning of the human body. They have gained attention due to their ability to hit multiple targets and act as antioxidants, anti-inflammatory agents, and modulators of immune response and cell death. Therefore, nutraceuticals are being studied to prevent and treat liver ischemia-reperfusion injury (IRI). This study evaluated the effect of a nutraceutical solution formed by resveratrol, quercetin, omega-3 fatty acid, selenium, ginger, avocado, leucine, and niacin on liver IRI. IRI was performed with 60 min of ischemia and 4 h of reperfusion in male Wistar rats. Afterward, the animals were euthanized to study hepatocellular injury, cytokines, oxidative stress, gene expression of apoptosis-related genes, TNF-α and caspase-3 proteins, and histology. Our results show that the nutraceutical solution was able to decrease apoptosis and histologic injury. The suggested mechanisms of action are a reduction in gene expression and the caspase-3 protein and a reduction in the TNF-α protein in liver tissue. The nutraceutical solution was unable to decrease transaminases and cytokines. These findings suggest that the nutraceuticals used favored the protection of hepatocytes, and their combination represents a promising therapeutic proposal against liver IRI.

Amentoflavone mitigates doxorubicin-induced cardiotoxicity by suppressing cardiomyocyte pyroptosis and inflammation through inhibition of the STING/NLRP3 signalling pathway

BackgroundDoxorubicin (DOX) is a potent anticancer chemotherapeutic agent whose clinical application is substantially constrained by its cardiotoxicity. The pathophysiology of DOX-induced cardiotoxicity manifests as cardiomyocyte pyroptosis and inflammation. Amentoflavone (AMF) is a naturally occurring biflavone possessing anti-pyroptotic and anti-inflammatory properties. However, the mechanism through which AMF alleviates DOX-induced cardiotoxicity remains undetermined. PurposeThis study aimed at investigating the role of AMF in alleviating DOX-induced cardiotoxicity. Study design and methodsTo assess the in vivo effect of AMF, DOX was intraperitoneally administered into a mouse model to induce cardiotoxicity. To elucidate the underlying mechanisms, the activities of STING/NLRP3 were quantified using the NLRP3 agonist nigericin and the STING agonist amidobenzimidazole (ABZI). Primary cardiomyocytes isolated from neonatal Sprague-Dawley rats were treated with saline (vehicle) or DOX with or without AMF and/or ABZI. The echocardiogram, haemodynamics, cardiac injury markers, heart/body weight ratio, and pathological alterations were monitored; the STING/NLRP3 pathway-associated proteins were detected by western blot and cardiomyocyte pyroptosis was analysed by immunofluorescence staining of cleaved N-terminal GSDMD and scanning electron microscopy. Furthermore, we evaluated the potential of AMF in compromising the anticancer effects of DOX in human breast cancer cell lines. ResultsAMF substantially alleviated cardiac dysfunction and reduced heart/body weight ratio and myocardial damage in mice models of DOX-induced cardiotoxicity. AMF effectively suppressed DOX-mediated upregulation of IL-1β, IL-18, TNF-α, and pyroptosis-related proteins, including NLRP3, cleaved caspase-1, and cleaved N-terminal GSDMD. The levels of apoptosis-related proteins, namely Bax, cleaved caspase-3, and BCL-2 were not affected. In addition, AMF inhibited STING phosphorylation in DOX-affected hearts. Intriguingly, the administration of nigericin or ABZI dampened the cardioprotective effects of AMF. The in vitro anti-pyroptotic effect of AMF was demonstrated in attenuating the DOX-induced reduction in cardiomyocyte cell viability, upregulation of cleaved N-terminal GSDMD, and pyroptotic morphology alteration at the microstructural level. AMF exhibited a synergistic effect with DOX to reduce the viability of human breast cancer cells. ConclusionAMF alleviates DOX-induced cardiotoxicity by suppressing cardiomyocyte pyroptosis and inflammation via inhibition of the STING/NLRP3 signalling pathway, thereby validating its efficacy as a cardioprotective agent.

Newcastle disease virus LaSota strain induces apoptosis and activates the TNFα/NF-κB pathway in canine mammary carcinoma cells.

Spontaneous canine mammary carcinomas (CMCs) have been widely considered a good research model for human breast cancers, which brings much attention to CMCs. In recent years, the oncolytic effect of Newcastle disease virus (NDV) on cancer cells has been widely studied, but its effect on CMCs is still unclear. This study aims to investigate the oncolytic effect of NDV LaSota strain on canine mammary carcinoma cell line (CMT-U27) in vivo and in vitro. The in vitro cytotoxicity and immunocytochemistry experiments showed that NDV selectively replicated in CMT-U27 cells, and inhibited cell proliferation and migration but not in MDCK cells. KEGG analysis of transcriptome sequencing indicated the importance of the TNFα and NF-κB signalling pathways in the anti-tumour effect of NDV. Subsequently, the significantly increased expression of TNFα, p65, phospho-p65, caspase-8, caspase-3 and cleaved-PARP proteins in the NDV group suggested that NDV induced CMT-U27 cells apoptosis by activating the caspase-8/caspase-3 pathway and the TNFα/NF-κB signalling pathway. Nude mice tumour-bearing experiments showed that NDV could significantly decrease the growth rate of CMC in vivo. In conclusion, our study demonstrates the effective oncolytic effects of NDV on CMT-U27 cells in vivo and in vitro, and suggests NDV as a promising candidate for oncolytic therapy.

Comparison of the effects of remimazolam and dexmedetomidine on early postoperative cognitive function in elderly patients with gastric cancer.

To compare the effects of remimazolam and dexmedetomidine on early postoperative cognitive dysfunction (POCD) in aged gastric cancer patients. From June to December 2022, 104 elderly patients (aged 65-80 years) received laparoscopic radical resection of gastric cancer at the First Affiliated Hospital of Nanchang University. Using the random number table approach, the patients were separated into three groups: remimazolam (Group R), dexmedetomidine (Group D), and saline (Group C). The primary outcome was the incidence of POCD, and secondary outcomes included TNF-α and S-100β protein concentrations, hemodynamics, VAS scores, anesthesia recovery indicators, and the occurrence of adverse events within 48 h postoperatively. At 3 and 7 days after surgery, there were no statistically significant differences in the incidence of POCD, the MMSE and MoCA scores between groups R and D (p > 0.05). However, compared to the saline group, both groups had higher MMSE and MoCA scores and decreased incidences of POCD. These differences were statistically significant (p < 0.05). Between group R and group D, there were no statistically significant changes (p > 0.05) in the levels of TNF-α and S-100β protein at the three time points (at the end of the surgery, 1 day later, and 3 days later). Even though neither group's concentration of the two factors was as high as that of the saline group, the differences were statistically significant (p < 0.05). At all three time points-following induction (T2), 30 min into the operation (T3), and at the conclusion of the surgery (T4)-the heart rate and blood pressure in group R were greater than those in groups D and C. Statistics showed that the differences were significant (p < 0.05). The incidence of intraoperative hypotension was highest in group D and lowest in group R (p < 0.05). The dose of propofol and remifentanil, group C > group R > group D. Extubation and PACU residence times did not differ statistically significantly (p > 0.05) between the three groups. There was no significant difference in VAS scores between groups R and D after 24 h postoperatively (p > 0.05), although both had lower scores than group C, and the difference was statistically significant (p < 0.05). The VAS scores between the three groups at 72 h (T6) and 7 days (T7) were not statistically significant (p > 0.05). Adverse reactions such as respiratory depression, hypotension, bradycardia, agitation, drowsiness, and nausea and vomiting had the lowest incidence in group R and the highest incidence in group C (p < 0.05). Remimazolam is similarly beneficial as dexmedetomidine in lowering the incidence of early POCD in aged patients after radical gastric cancer resection, probably due to reduced inflammatory response.

Morin hydrate suppresses lipoteichoic acid-induced oxidative stress-mediated inflammatory events in macrophages via augmenting Nrf2/HO-1 and antioxidant defense molecules

Objectives Oxidative stress induces chronic inflammatory diseases in aerobic organisms, and antioxidants from plants represent an efficient strategy to prevent this condition. Morin hydrate (MH), a bioactive flavonoid, has a wide range of pharmacological properties, including anti-inflammatory and anti-oxidant. This study evaluated the protective effects of MH on lipoteichoic acid (LTA)-induced inflammation in RAW 264.7 macrophages by testing the main oxidative and inflammatory biomarkers and also investigating the molecular pathways involved. Methods The antioxidant and anti-inflammatory effects of MH were evaluated in a cell-free system and RAW264.7 cells. Quantitative real-time PCR (RT-qPCR) and assay kits were used to measure the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) mRNA, as well as the activity of antioxidant enzymes. The effects of MH on LTA-induced inducible nitric oxide synthase (iNOS), IL-1β, and TNF-α mRNA and protein expression were also evaluated by RT-qPCR and Western blotting. Results MH reduced DPPH and ABTS radicals in a cell-free system and LTA-induced ROS and NO production in RAW264.7 cells. MH upregulated Nrf2 and HO-1 mRNA expression and reversed LTA-mediated reduction of antioxidant enzymes, at a high concentration of 20 µM pretreated cells. MH also effectively attenuated LTA-induced iNOS, IL-1β, and TNF-α mRNA and protein expression, and these effects were reversed by ML385. Conclusions The study found that the Nrf2/HO-1 played role in the inhibition of LTA-induced oxidative stress in macrophages by MH. This study may consider to be a promising induced macrophage-targeted strategy via regulating anti-oxidative defense to control inflammatory-related disease.

HELICOBACTER AND HEPATOBILIARY DISEASES: UPDATE 2023.

•Clinical studies have shown that hepatobiliary diseases of inflammatory and neoplastic origin are associated with Helicobacter infection. •Translocation and the ascending pathway are putative mechanisms for Helicobacter spp to enter the hepatobiliary system. •H. pylori infection has a systemic effect through the activity of pro-inflammatory cytokines, TNF-α, leukotrienes, interferon-β, interferon-γ, and acute phase proteins. •Histopathological confirmation is needed to present that H. pylori eradication prevents or improves hepatobiliary disease progression. Helicobacter Pylori (H. pylori) is one of the main infectious causes of gastroduodenal diseases, however, its role in developing different extragastric diseases has been proven. The possible involvement of H. pylori in the pathogenesis of cardiovascular, metabolic, neurodegenerative, skin, and hepatobiliary diseases is suggested. The bacterium has been found in tissue samples from the liver, biliary tract, and gallstones of animals and humans. However, the role of H. pylori infection in the pathogenesis of liver and biliary diseases has not been finally established. The histopathological confirmation of the positive effect of H. pylori eradication is needed. In addition, there are discussions on the clinical significance of other Helicobacter species. The review presents the data available for and against the involvement of H. pylori in hepatobi-liary disease development and progression.

Acute cold stress induces intestinal injury via CIRP-TLR4-IRE1 signaling pathway in pre-starter broilers.

Cold stress is a common environmental stress in broiler chicks. Cold-inducible RNA-binding protein (CIRP) is a conserved cold shock protein that can regulate inflammatory response through Toll-like receptor 4 (TLR4). The mechanism that how CIRP involves in the regulation of cold stress in broilers remains unclear. In this study, 360 7-day-old healthy male SZ901 chicks were selected and randomly allocated to four groups, and then subjected to acute cold exposure at the ambient temperature of 12 ± 1°Cfor 0h, 4h, 8h, and 12h, respectively. After cold exposure, abdominall skin temperature, gene expression of CIRP-TLR4-IRE1 signaling pathway in ileum mucosa, and small intestinal structure were measured. The results showed that cold exposure decreased abdominall skin temperature, upregulated the gene expression of endoplasmic reticulum stress (ERS) markers IRE1, inflammatory factors IL-1β, IL-6, IL-10, TNF-α, and tight junction proteins ZO-1 and Occludin in ileum of chicks compared with the control group with no (0h) cold exposure. Compared with the control group, a long time cold exposure upregulated the gene expression of CIRP, TLR4, GRP78, NF-κB in ileum mucosa, and decreased the villus height and V/C of small intestine. The above results suggest that acute cold stress induces endoplasmic reticulum stress via upregulating the gene expression of CIRP-TLR4-IRE1 signaling pathway, and results in the structural damage of chick intestine.

Cepharanthine Dry Powder Inhaler for the Treatment of Acute Lung Injury.

Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) induces a severe cytokine storm that may cause acute lung injury/acute respiratory distress syndrome (ALI/ARDS) with high clinical morbidity and mortality in infected individuals. Cepharanthine (CEP) is a bisbenzylisoquinoline alkaloid isolated and extracted from Stephania cepharantha Hayata. It exhibits various pharmacological effects, including antioxidant, anti-inflammatory, immunomodulatory, anti-tumor, and antiviral activities. The low oral bioavailability of CEP can be attributed to its poor water solubility. In this study, we utilized the freeze-drying method to prepare dry powder inhalers (DPI) for the treatment of acute lung injury (ALI) in rats via pulmonary administration. According to the powder properties study, the aerodynamic median diameter (Da) of the DPIs was 3.2 μm, and the in vitro lung deposition rate was 30.26; thus, meeting the Chinese Pharmacopoeia standard for pulmonary inhalation administration. We established an ALI rat model by intratracheal injection of hydrochloric acid (1.2 mL/kg, pH = 1.25). At 1 h after the model's establishment, CEP dry powder inhalers (CEP DPIs) (30 mg/kg) were sprayed into the lungs of rats with ALI via the trachea. Compared with the model group, the treatment group exhibited a reduced pulmonary edema and hemorrhage, and significantly reduced content of inflammatory factors (TNF-α, IL-6 and total protein) in their lungs (p < 0.01), indicating that the main mechanism of CEP underlying the treatment of ALI is anti-inflammation. Overall, the dry powder inhaler can deliver the drug directly to the site of the disease, increasing the intrapulmonary utilization of CEP and improving its efficacy, making it a promising inhalable formulation for the treatment of ALI.

An immunoregulator nanomedicine approach for the treatment of tuberculosis.

Introduction: A nanoparticle composed of a poly (lactic-co-glycolic acid) (PLGA) core and a chitosan (CS) shell with surface-adsorbed 1,3 β-glucan (β-glucan) was synthesized. The exposure response of CS-PLGA nanoparticles (0.1mg/mL) with surface-bound β-glucan at 0, 5, 10, 15, 20, or 25ng or free β-glucan at 5, 10, 15, 20, or 25ng/mL in macrophage in vitro and in vivo was investigated. Results: In vitro studies demonstrate that gene expression for IL-1β, IL-6, and TNFα increased at 10 and 15ng surface-bound β-glucan on CS-PLGA nanoparticles (0.1mg/mL) and at 20 and 25ng/mL of free β-glucan both at 24h and 48h. Secretion of TNFα protein and ROS production increased at 5, 10, 15, and 20ng surface-bound β-glucan on CS-PLGA nanoparticles and at 20 and 25ng/mL of free β-glucan at 24h. Laminarin, a Dectin-1 antagonist, prevented the increase in cytokine gene expression induced by CS-PLGA nanoparticles with surface-bound β-glucan at 10 and 15ng, indicating a Dectin-1 receptor mechanism. Efficacy studies showed a significant reduction in intracellular accumulation of mycobacterium tuberculosis (Mtb) in monocyte-derived macrophages (MDM) incubated with on CS-PLGA (0.1mg/ml) nanoparticles with 5, 10, and 15ng surface-bound β-glucan or with 10 and 15ng/mL of free β-glucan. β-glucan-CS-PLGA nanoparticles inhibited intracellular Mtb growth more than free β-glucan alone supporting the role of β-glucan-CS-PLGA nanoparticles as stronger adjuvants than free β-glucan. In vivo studies demonstrate that oropharyngeal aspiration (OPA) of CS-PLGA nanoparticles with nanogram concentrations of surface-bound β-glucan or free β-glucan increased TNFα gene expression in alveolar macrophages and TNFα protein secretion in bronchoalveolar lavage supernatants. Discussion: Data also demonstrate no damage to the alveolar epithelium or changes in the murine sepsis score following exposure to β-glucan-CS-PLGA nanoparticles only, indicating safety and feasibility of this nanoparticle adjuvant platform to mice by OPA.

Enoxaparin sodium bone cement displays local anti-inflammatory effects by regulating the expression of IL-6 and TNF-α

ObjectiveTo explore the roles of Enoxaparin Sodium-Polymethyl methacrylate bone cement on inflammatory factors Interleukin-6 and Tumour Necrosis Factor-α in a rabbit knee replacement model. As well as the mechanisms underlying its potential effects on lipopolysaccharide-induced endothelial cell injury. MethodsA knee replacement model was established using New Zealand rabbits. Forty rabbits were randomly divided into four groups: PMMA, ES-PMMA, sham-operated, and blank control groups (n = 10 in each group). Local tissues around the incision were taken at the 30th, 60th, and 90th minute after the surgical implantation of the corresponding bone cement. Immunohistochemistry in the surgical field was used to measure the expression of local inflammatory factors IL-6 and TNF-α. In the in vitro experiments, 1 cm3 of bone cement was immersed in 3 mL of the medium for 24 h. The bone cement was discarded and diluted to 25% with normal medium. Pre-experiments were screened for the best LPS-inducing concentration of 100 mg/mL, and the most compatible LPS concentration was used for subsequent experiments simulating the primary cultures of rats’ Inferior Vena Cava Endothelial Cells. The experiments were divided into four groups: blank control group, LPS induction group, PMMA + LPS group, and ES-PMMA + LPS group. The apoptosis rate was detected by flow cytometry, and the expression levels of TNF-α and IL-6 in the cells and supernatant were measured by ELISA, western blotting, and immunofluorescence. ResultsAccording to immunohistochemical results, IL-6-positive cells were concentrated in the tissue interstitial space. In the PMMA and sham-operated groups, the number of IL-6-positive cells gradually increased over time. At all time points, IL-6 expression in the ES-PMMA group was much lower than in the PMMA and sham-operated groups. At 30 min, TNF-α positive cells in the ES-PMMA group expressed less than those in the PMMA and sham-operated groups, with no discernible difference between the PMMA and ES-PMMA groups at 60 or 90 min. Using ELISA and flow cytometry, the expression levels of IL-6 and TNF-α were improved and the apoptosis rate was magnified in the LPS-induced group (***P < 0.001) in contrast with the blank control group. Additionally, the expression levels of IL-6 and TNF-α were reduced in the ES-PMMA + LPS group compared with the LPS-induced group (*P < 0.05) and the apoptosis rate was reduced (***P < 0.001), with statistically significant variations. Western blotting and immunofluorescence analysis confirmed that IL-6 and TNF-α protein expression in cells was upregulated in the LPS-induced group compared to the blank control group (***P < 0.001), and the mean fluorescence intensity was enlarged (***P < 0.001). Meanwhile, IL-6 and TNF-α expression in the ES-PMMA + LPS group were down-regulated (**P < 0.01 or *P < 0.05) compared with the LPS-induced group and PMMA + LPS crew protein expression, and the average fluorescence intensity of IL-6 and TNF-α was lowered in the ES-PMMA + LPS group compared to the LPS-induced group (***P < 0.001). ConclusionsES-PMMA bone cement reduced the expression levels of local inflammatory factors IL-6 and TNF-α in a rabbit knee model. ES-PMMA bone cement reduced the rate of LPS-induced endothelial cell apoptosis and diminished local inflammatory damage by regulating the secretion of inflammatory factors TNF-α and IL-6.

The p,p’-DDE disturbs the M1 function without affecting the M2 phenotype nor unstimulated bone marrow-derived macrophages from BALB/c mice

DDT, a persistent organic pollutant, remains affecting human health worldwide. DDT and its most persistent metabolite (p,p'-DDE) negatively affect the immune response regulation and mechanisms involved in protecting against pathogens Such metabolite decreases the capability to limit intracellular growth of Mycobacterium microti and yeast. However, the effect on unstimulated (M0) and anti-inflammatory macrophages (M2) has been evaluated scanty. Herein, we evaluated the impact of p,p’-DDE at environmentally relevant concentrations (0.125, 1.25, 2.5, and 5 µg/mL) on bone marrow-derived macrophages stimulated with IFNγ+LPS to M1 or with IL-4 +IL-13 to M2. Thus we study whether the p,p’-DDE induces M0 to a specific phenotype or modulates activation of the macrophage phenotypes and explains, at least partly, the reported effects of p,p’-DDE on the M1 function. The p,p’-DDE did not affect the cell viability of M0 or the macrophage phenotypes. In M1, the p,p’-DDE decreased NO•- production and IL-1β secretion, but increasing cellular ROS and mitochondrial O2•-, but did not alter iNOS, TNF-α, MHCII, and CD86 protein expression nor affect M2 markers arginase activity, TGF-β1, and CD206; p,p'-DDE, did not affect marker expression in M0 or M2, supporting that its effects on M1 parameters are not dependent on M0 nor M2 modulation. The decreasing of NO•- production by the p,p’-DDE without altering iNOS levels, Arginase activity, or TNF-α, but increasing cellular ROS and mitochondrial O2 suggests that p,p’-DDE interferes with the iNOS function but not with its transcription. The p,p’-DDE decreasing of IL-1β secretion, without any effect on TNF-α, suggest that an alteration of specific targets involved in IL-1β secretion may be affected and related to ROS induction. The p,p’-DDE effect on iNOS function and the IL-1β secretion process, as the NLRP3 activation, deserves further study.