Inflammatory Tissue Injury Research Articles

Antioxidant Activity and Suppression of Intracellular Radical Generation of Streptomyces Strains and Genome Analysis of Strain ET3-23

Actinomycetes, predominantly found in soil, represent a remarkable source of natural products. The natural antioxidants obtained from them have been employed for human infectious diseases. In particular, Streptomyces strains serve as significant sources of natural antioxidants with demonstrated health benefits. The in vitro antioxidant activity and the inhibition of intracellular radical generation by crude extracts from various Streptomyces strains were evaluated and the genome sequence of selected strains was analyzed. Strains CT2-10, NE1-12, and ET3-23 showed 16S rRNA gene sequence similarity to Streptomyces capoamus JCM 4734T (98.94%), Streptomyces nigra 452T (99.78%), and Streptomyces morookaense LMG 20074T (99.49%), respectively. The genome analysis of strain ET3-23 had 106 contigs, with a total length of 8121874 bp and an average G+C content of 71.46%. Ethyl acetate extracts of strains CT2-10, ET3-23, and NE1-12 exhibited, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity with IC50 555.9 µg/mL, 66.5 µg/mL, and 463.3 µg/mL and showed NO scavenging activity, except for CT2-10. Strain NE1-12 significantly inhibited ROS production induced by hydrogen peroxide on macrophage cells, while strains NE1-12 and CT2-10 inhibited NO production induced by lipopolysaccharides with IC50 82.4 µg/mL and 2.3 µg/mL, respectively. These findings suggest their ability to modulate NO production which is crucial in inflammatory responses and tissue injury. The antioxidant activities of Streptomyces strains indicate their potential as valuable sources of bioactive natural products and effective antioxidant agents, warranting further investigation for therapeutic applications.

RBM25 is required to restrain inflammation via ACLY RNA splicing-dependent metabolism rewiring.

Spliceosome dysfunction and aberrant RNA splicing underline unresolved inflammation and immunopathogenesis. Here, we revealed the misregulation of mRNA splicing via the spliceosome in the pathogenesis of rheumatoid arthritis (RA). Among them, decreased expression of RNA binding motif protein 25 (RBM25) was identified as a major pathogenic factor in RA patients and experimental arthritis mice through increased proinflammatory mediator production and increased hyperinflammation in macrophages. Multiomics analyses of macrophages from RBM25-deficient mice revealed that the transcriptional enhancement of proinflammatory genes (including Il1b, Il6, and Cxcl10) was coupled with histone 3 lysine 9 acetylation (H3K9ac) and H3K27ac modifications as well as hypoxia inducible factor-1α (HIF-1α) activity. Furthermore, RBM25 directly bound to and mediated the 14th exon skipping of ATP citrate lyase (Acly) pre-mRNA, resulting in two distinct Acly isoforms, Acly Long (Acly L) and Acly Short (Acly S). In proinflammatory macrophages, Acly L was subjected to protein lactylation on lysine 918/995, whereas Acly S did not, which influenced its affinity for metabolic substrates and subsequent metabolic activity. RBM25 deficiency overwhelmingly increased the expression of the Acly S isoform, enhancing glycolysis and acetyl-CoA production for epigenetic remodeling, macrophage overactivation and tissue inflammatory injury. Finally, macrophage-specific deletion of RBM25 led to inflammaging, including spontaneous arthritis in various joints of mice and inflammation in multiple organs, which could be relieved by pharmacological inhibition of Acly. Overall, targeting the RBM25-Acly splicing axis represents a potential strategy for modulating macrophage responses in autoimmune arthritis and aging-associated inflammation.

Damage sensing through TLR9 Regulates Inflammatory and Antiviral Responses During Influenza Infection.

Host response aimed at eliminating the infecting pathogen, as well as the pathogen itself, can cause tissue injury. Tissue injury leads to the release of a myriad of cellular components including mitochondrial DNA, which the host senses through pattern recognition receptors. How the sensing of tissue injury by the host shapes the anti-pathogen response remains poorly understood. In this study, we utilized mice that are deficient in toll-like receptor-9 (TLR9), which binds to unmethylated CpG DNA sequences such as those present in bacterial and mitochondrial DNA. To avoid direct pathogen sensing by TLR9, we utilized the influenza virus, which lacks ligands for TLR9, to determine how damage sensing by TLR9 contributes to anti-influenza immunity. Our data show that TLR9-mediated sensing of tissue damage promotes an inflammatory response during early infection, driven by the epithelial and myeloid cells. Along with the diminished inflammatory response, the absence of TLR9 led to impaired viral clearance manifested as a higher and prolonged influenza components in myeloid cells including monocytes and macrophages rendering them highly inflammatory. The persistent inflammation driven by infected myeloid cells led to persistent lung injury and impaired recovery in influenza-infected TLR9-/- mice. Further, we show elevated TLR9 activation in the plasma samples of patients with influenza and its association with the disease severity in hospitalized patients, demonstrating its clinical relevance. Overall, we demonstrate an essential role of damage sensing through TLR9 in promoting anti-influenza immunity and inflammatory response.

Endogenous stem cell mobilization and localized immunosuppression synergistically ameliorate DSS-induced Colitis in mice

BackgroundStem cell therapy is a promising alternative for inflammatory diseases and tissue injury treatment. Exogenous delivery of mesenchymal stem cells is associated with instant blood-mediated inflammatory reactions, mechanical stress during administration, and replicative senescence or change in phenotype during long-term culture in vitro. In this study, we aimed to mobilize endogenous hematopoietic stem cells (HSCs) using AMD-3100 and provide local immune suppression using FK506, an immunosuppressive drug, for the treatment of inflammatory bowel diseases.MethodsReactive oxygen species (ROS)-responsive FK506-loaded thioketal microspheres were prepared by emulsification solvent-evaporation method. Thioketal vehicle based FK506 microspheres and AMD3100 were co-administered into male C57BL6/J mice with dextran sulfate sodium (DSS) induced colitis. The effect of FK506-loaded thioketal microspheres in colitis mice were evaluated using disease severity index, myeloperoxidase activity, histology, flow cytometry, and gene expression by qRT-PCR.ResultsThe delivery of AMD-3100 enhanced mobilization of HSCs from the bone marrow into the inflamed colon of mice. Furthermore, targeted oral delivery of FK506 in an inflamed colon inhibited the immune activation in the colon. In the DSS-induced colitis mouse model, the combination of AMD-3100 and FK506-loaded thioketal microspheres ameliorated the disease, decreased immune cell infiltration and activation, and improved body weight, colon length, and epithelial healing process.ConclusionThis study shows that the significant increase in the percentage of mobilized hematopoietic stem cells in the combination therapy of AMD and oral FK506 microspheres may contribute to a synergistic therapeutic effect. Thus, low-dose local delivery of FK506 combined with AMD3100 could be a promising alternative treatment for inflammatory bowel diseases.

Effects and mechanism of Qingke Pingchuan granules against influenza virus infection.

Qingke Pingchuan granules (QPGs), which contain Houttuynia cordata Thunb, Fritillaria cirrhosa, fired licorice, and fired bitter almonds, among other components, can clear heat and ventilate the lungs, relieving cough and asthma. Clinically, QPGs are mainly used to treat cough, asthma, fever and other discomforts caused by acute or chronic bronchitis. In this study, the antiviral activity of QPGs against respiratory syncytial virus (RSV), influenza A virus A/FM/1/47 (H1N1), oseltamivir-resistant H1N1, A/Beijing/32/92 (H3N2), Sendai virus, and human adenovirus type 3 in Hep-2 or MDCK cells was evaluated using the CCK-8 method, and the cytotoxicity of QPGs to these two cell lines was tested. The effect of QPGs on mice infected with influenza A virus A/FM/1/47 (H1N1) was evaluated by measuring body weight, survival time, and survival rate, as well as virus titers and lesions in the lungs and levels of inflammatory factors in serum. In addition, the expression of TLR-7-My88-NF-κB signaling pathway-related proteins in lung tissues was analyzed by Western blotting and qRT-PCR. The results showed that QPGs had a potent inhibitory effect on the six viruses tested in vitro. Interestingly, QPGs also displayed particularly pronounced antiviral activity against H1N1-OC, similar to that of oseltamivir, a well-known antiviral drug. QPGs effectively protected mice from infection by H1N1, as indicated by significantly increased body weights, survival times, and survival rates and reduced lung virus titers of inflammatory factors and lung tissue injury. The levels of TLR-7-MyD88-NF-κB-pathway-related proteins in the lung tissue of infected mice were found to be decreased after QPG treatment, thereby alleviating lung injury caused by excessive release of inflammatory factors. Taken together, these findings indicate that QPGs have satisfactory activity against influenza virus infection.

Temporal dynamics of the multi-omic response to endurance exercise training

Regular exercise promotes whole-body health and prevents disease, but the underlying molecular mechanisms are incompletely understood1–3. Here, the Molecular Transducers of Physical Activity Consortium4 profiled the temporal transcriptome, proteome, metabolome, lipidome, phosphoproteome, acetylproteome, ubiquitylproteome, epigenome and immunome in whole blood, plasma and 18 solid tissues in male and female Rattus norvegicus over eight weeks of endurance exercise training. The resulting data compendium encompasses 9,466 assays across 19 tissues, 25 molecular platforms and 4 training time points. Thousands of shared and tissue-specific molecular alterations were identified, with sex differences found in multiple tissues. Temporal multi-omic and multi-tissue analyses revealed expansive biological insights into the adaptive responses to endurance training, including widespread regulation of immune, metabolic, stress response and mitochondrial pathways. Many changes were relevant to human health, including non-alcoholic fatty liver disease, inflammatory bowel disease, cardiovascular health and tissue injury and recovery. The data and analyses presented in this study will serve as valuable resources for understanding and exploring the multi-tissue molecular effects of endurance training and are provided in a public repository (https://motrpac-data.org/).

A Novel IRAK4 Inhibitor DW18134 Ameliorates Peritonitis and Inflammatory Bowel Disease.

IRAK4 is a critical mediator in NF-κB-regulated inflammatory signaling and has emerged as a promising therapeutic target for the treatment of autoimmune diseases; however, none of its inhibitors have received FDA approval. In this study, we identified a novel small-molecule IRAK4 kinase inhibitor, DW18134, with an IC50 value of 11.2 nM. DW18134 dose-dependently inhibited the phosphorylation of IRAK4 and IKK in primary peritoneal macrophages and RAW264.7 cells, inhibiting the secretion of TNF-α and IL-6 in both cell lines. The in vivo study demonstrated the efficacy of DW18134, significantly attenuating behavioral scores in an LPS-induced peritonitis model. Mechanistically, DW18134 reduced serum TNF-α and IL-6 levels and attenuated inflammatory tissue injury. By directly blocking IRAK4 activation, DW18134 diminished liver macrophage infiltration and the expression of related inflammatory cytokines in peritonitis mice. Additionally, in the DSS-induced colitis model, DW18134 significantly reduced the disease activity index (DAI) and normalized food and water intake and body weight. Furthermore, DW18134 restored intestinal damage and reduced inflammatory cytokine expression in mice by blocking the IRAK4 signaling pathway. Notably, DW18134 protected DSS-threatened intestinal barrier function by upregulating tight junction gene expression. In conclusion, our findings reported a novel IRAK4 inhibitor, DW18134, as a promising candidate for treating inflammatory diseases, including peritonitis and IBD.

Formulation and Evaluation of Diclofenac Transdermal Gel

Diclofenac sodium gel is a widely used nonsteroidal anti-inflammatory drug (NSAID) formulation designed for topical application, primarily targeting local pain and inflammation. This wok aims to provide a comprehensive analysis of the anti-inflammatory activity of diclofenac sodium gel, focusing on its mechanism of action, efficacy, safety profile, and clinical applications. Several clinical studies have investigated the efficacy of diclofenac sodium gel in various inflammatory conditions, including osteoarthritis, rheumatoid arthritis, and soft tissue injuries. Results consistently demonstrate significant reductions in pain, swelling, and functional impairment, validating the gel's anti-inflammatory properties. The safety profile of diclofenac sodium gel is generally favorable, with local skin reactions being the most commonly reported adverse events. Systemic absorption is low, reducing the risk of gastrointestinal complications and other systemic side effects associated with oral NSAIDs. However, caution should be exercised in individuals with a history of hypersensitivity reactions or skin disorders.

Ochratoxin A promotes chronic enteritis and early colorectal cancer progression by targeting Rinck signaling.

Mycotoxins, such as aflatoxin and ochratoxin A (OTA), are found at measurable levels in many staple foods; the health implications of long-term exposure of such toxins are poorly understood. Increasing evidence has confirmed the important role of OTA in upregulation of oxidative stress- and inflammatory response-induced tissue injury. However, it remains unknown whether ochratoxin A can promote chronic colitis and its associated colon cancer (CRC) development, and potential molecular mechanism. Additionally, RING finger-interacting protein with C kinase (RINCK) is a ubiquitin ligase and mediates immune response. Unfortunately, the potential molecular function of RINCK on regulation of colitis is still largely unknown. This study aims to provide mechanistic evidence that the role of RINCK in colitis and early colorectal cancer progression in response to OTA treatment via targeting nuclear factor erythroid 2-related factor 2 (NRF2). The Cancer Genome Atlas (TCGA) database, GEO database, human subjects with CC phenotype and CC cell lines were used in this work. Pathological links between OTA, RINCK and treatment of CC are revealed through comprehensive means such as biological information analysis, clinical experiments, RNA-seq, and verification experiments. In this study, under oxidative stress in setting of colitis, we first identified RINCK as a key regulatory factor and a novel endogenous suppressor of nuclear factor erythroid 2-related factor 2 (NRF2), and we also confirm that RINCK is a NRF2 partner protein that catalyses its ubiquitination and degradation in intestinal epithelial cells (IECs). Notably, in vivo study, pathological phenotypes triggered by OTA pretreatment, accompanied by post-treatment of dextran sulfate sodium (DSS)-induced colitis was significantly mitigated by IEC-specific deficiency of Rinck, IEC-Rinck(KO) and adenovirus-associated virus (AAV)-triggered suppression of Rinck in rodent model, and lentivirus (LV)-mediated downregulation of Rinck (LV-shRinck) in rabbit model, as determined by decreased endogenous reactive oxygen species (ROS) production, pro-inflammatory cytokines contents, improved body weights, reduced survival rates, restored colon length, assuasive DAI and histological scores. Inversely, transgenic mice by IEC-specific Rinck overexpression, IEC-Rinck(OE) accelerated colitis in acute or chronic colitis rodent models and in vitro experiments. Moreover, we found that OTA pretreatment-promoted azoxymethane (AOM)/DSS-induced colitis-associated early colorectal cancer (CRC) was also dramatically reduced by IEC-Rinck(KO), indicated by the decreased tumor number and corresponding KI-67 levels. Clinical samples analysis revealed that RINCK levels were greatly increased in tumor tissues of patients with CRC phenotypes. In parallel, RINCK deletion remarkably retarded the proliferation of colon cancer and tumor growth in vitro and in vivo, respectively. Mechanistically, in response to onset of colitis, RINCK directly interacts with NRF2 and promotes ubiquitin-proteasome degradation via increasing K48-linkage ubiquitin chain, thus leads in suppression of NRF2 nuclear translocation and its downstream cascade inactivation, which retards antioxidant defense. The findings suggested that oral sub-chronic exposure of OTA significantly facilitates DSS-induced colitis and colitis-associated CRC development. These results further elucidated the potential role of RINCK in colitis progression by mediating NRF2 degradation, and could be considered as a therapeutic target for the treatment of such disease.

The amphiregulin/EGFR axis has limited contribution in controlling autoimmune diabetes

Conventional immunosuppressive functions of CD4+Foxp3+ regulatory T cells (Tregs) in type 1 diabetes (T1D) pathogenesis have been well described, but whether Tregs have additional non-immunological functions supporting tissue homeostasis in pancreatic islets is unknown. Within the last decade novel tissue repair functions have been ascribed to Tregs. One function is production of the epidermal growth factor receptor (EGFR) ligand, amphiregulin, which promotes tissue repair in response to inflammatory or mechanical tissue injury. However, whether such pathways are engaged during autoimmune diabetes and promote tissue repair is undetermined. Previously, we observed that upregulation of amphiregulin at the transcriptional level was associated with functional Treg populations in the non-obese diabetic (NOD) mouse model of T1D. From this we postulated that amphiregulin promoted islet tissue repair and slowed the progression of diabetes in NOD mice. Here, we report that islet-infiltrating Tregs have increased capacity to produce amphiregulin, and that both Tregs and beta cells express EGFR. Moreover, we show that amphiregulin can directly modulate mediators of endoplasmic reticulum stress in beta cells. Despite this, NOD amphiregulin deficient mice showed no acceleration of spontaneous autoimmune diabetes. Taken together, the data suggest that the ability for amphiregulin to affect the progression of autoimmune diabetes is limited.

The effect of bradykinin 1 receptor antagonist BI 1026706 on pulmonary inflammation after segmental lipopolysaccharide challenge in healthy smokers

BackgroundBradykinin 1 receptor (B1R) signalling pathways may be involved in the inflammatory pathophysiology of chronic obstructive pulmonary disease (COPD). B1R signalling is induced by inflammatory stimuli or tissue injury and leads to activation and increased migration of pro-inflammatory cells. Lipopolysaccharide (LPS) lung challenge in man is an experimental method of exploring inflammation in the lung whereby interference in these pathways can help to assess pharmacologic interventions in COPD. BI 1026706, a potent B1R antagonist, was hypothesized to reduce the inflammatory activity after segmental lipopolysaccharide (LPS) challenge in humans due to decreased pulmonary cell influx. MethodsIn a monocentric, randomized, double-blind, placebo-controlled, parallel-group, phase I trial, 57 healthy, smoking subjects were treated for 28 days with either oral BI 1026706 100 mg bid or placebo. At day 21, turbo-inversion recovery magnitude magnetic resonance imaging (TIRM MRI) was performed. On the last day of treatment, pre-challenge bronchoalveolar lavage fluid (BAL) and biopsies were sampled, followed by segmental LPS challenge (40 endotoxin units/kg body weight) and saline control instillation in different lung lobes. Twenty-four hours later, TIRM MRI was performed, then BAL and biopsies were collected from the challenged segments. In BAL samples, cells were differentiated for neutrophil numbers as the primary endpoint. Other endpoints included assessment of safety, biomarkers in BAL (e.g. interleukin-8 [IL-8], albumin and total protein), B1R expression in lung biopsies and TIRM score by MRI as a measure for the extent of pulmonary oedema. ResultsAfter LPS, but not after saline, high numbers of inflammatory cells, predominantly neutrophils were observed in the airways. IL-8, albumin and total protein were also increased in BAL samples after LPS challenge as compared with saline control. There were no significant differences in cells or other biomarkers from BAL in volunteers treated with BI 1026706 compared with those treated with placebo. Unexpectedly, neutrophil numbers in BAL were 30% higher and MRI-derived extent of oedema was significantly higher with BI 1026706 treatment compared with placebo, 24 h after LPS challenge. Adverse events were mainly mild to moderate and not different between treatment groups. ConclusionsTreatment with BI 1026706 for four weeks was safe and well-tolerated in healthy smoking subjects. BI 1026706 100 mg bid did not provide evidence for anti-inflammatory effects in the human bronchial LPS challenge model. Trial registrationThe study was registered on January 14, 2016 at ClinicalTrials.gov (NCT02657408).

Honokiol, an inducer of sirtuin-3, protects against non-steroidal anti-inflammatory drug-induced gastric mucosal mitochondrial pathology, apoptosis and inflammatory tissue injury.

Mitochondrial oxidative stress, inflammation and apoptosis primarily underlie gastric mucosal injury caused by the widely used non-steroidal anti-inflammatory drugs (NSAIDs). Alternative gastroprotective strategies are therefore needed. Sirtuin-3 pivotally maintains mitochondrial structural integrity and metabolism while preventing oxidative stress; however, its relevance to gastric injury was never explored. Here, we have investigated whether and how sirtuin-3 stimulation by the phytochemical, honokiol, could rescue NSAID-induced gastric injury. Gastric injury in rats induced by indomethacin was used to assess the effects of honokiol. Next-generation sequencing-based transcriptomics followed by functional validation identified the gastroprotective function of sirtuin-3. Flow cytometry, immunoblotting, qRT-PCR and immunohistochemistry were used measure effects on oxidative stress, mitochondrial dynamics, electron transport chain function, and markers of inflammation and apoptosis. Sirtuin-3 deacetylase activity was also estimated and gastric luminal pH was measured. Indomethacin down-regulated sirtuin-3 to induce oxidative stress, mitochondrial hyperacetylation, 8-oxoguanine DNA glycosylase 1 depletion, mitochondrial DNA damage, respiratory chain defect and mitochondrial fragmentation leading to severe mucosal injury. Indomethacin dose-dependently inhibited sirtuin-3 deacetylase activity. Honokiol prevented mitochondrial oxidative damage and inflammatory tissue injury by attenuating indomethacin-induced depletion of both sirtuin-3 and its transcriptional regulators PGC1α and ERRα. Honokiol also accelerated gastric wound healing but did not alter gastric acid secretion, unlike lansoprazole. Sirtuin-3 stimulation by honokiol prevented and reversed NSAID-induced gastric injury through maintaining mitochondrial integrity. Honokiol did not affect gastric acid secretion. Sirtuin-3 stimulation by honokiol may be utilized as a mitochondria-based, acid-independent novel gastroprotective strategy against NSAIDs.

Berberine Alleviates Acute Lung Injury in Septic Mice by Modulating Treg/Th17 Homeostasis and Downregulating NF-κB Signaling.

A common complication of sepsis is acute lung injury (ALI), which is associated with an acute onset, rapid disease changes, and high mortality. Regulatory T (Treg) and T helper 17 (Th17) cells comprise CD4+ T cell subsets, which strongly influence inflammation during ALI. In this study, we investigated the effect of berberine (BBR), an antioxidant, anti-inflammatory, and immunomodulatory drug, on the inflammatory response and immune state in mice with sepsis. A mouse model of cecal ligation and puncture (CLP) was established. The mice were intragastrically administered 50 mg/kg BBR. We used histological techniques to evaluate inflammatory tissue injury and flow cytometry for analyzing Treg/Th17 levels. We also assessed NF-κB signaling pathways by Western blotting assays and immunofluorescence staining. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the content of cytokines. Treatment with BBR considerably mitigated lung injury while improving survival, post-cecal ligation, and puncture (CLP). Treatment with BBR ameliorated pulmonary edema and hypoxemia in septic mice and inhibited the NF-κB signaling pathway. BBR also increased Treg cells and decreased Th17 proportions in the spleen and lung tissue of CLP-treated mice. Blocking Treg cells weakened the protective effect of BBR on sepsis-associated lung injury. Overall, these results suggested that BBR is a potential therapeutic agent for sepsis.

Effects of caspase recruitment domain protein 9 gene knockout on airway inflammation in C57BL/6 mice with steroid resistant asthma

Objective: To investigate the role of caspase recruitment domain protein 9 (CARD9) in airway injury and inflammation of steroid resistant asthma in C57BL/6 mice. Methods: C57BL/6 mice were divided into A group (control group), B group (model group) and C group (dexamethasone treatment group), with 6 mouse in each group using random number table. The mouse asthma model was established in B and C group by subcutaneous injection of ovalbumin (OVA)/complete Freund adjuvant (CFA) in the abdomen and OVA aerosol challenge, the pathological change and cell count in broncho alveolar lavage fluid (BALF) were detected in order to confirm the model as steroid resistant asthma, and the lung tissue inflammatory infiltration was scored. Western blot was used to detect the changes of CARD9 protein between the group A and B; then wild-type and CARD9 knockout mice were divided into D group (wild-type control group), E group (wild-type model group), F group (CARD9 knockout control group) and G group (CARD9 knockout model group), the following indicators were observed and compared after establishing steroid resistant asthma model separately: HE staining was used to observe the pathological changes of lung tissue, ELISA was used to detect the protein levels of interleukin-4 (IL-4), interleukin-5 (IL-5) and interleukin-17(IL-17) in BALF, and RT-PCR was used to detect the mRNA levels of CXC motif chemokine ligand-10 (CXCL-10) and IL-17 in lung. Results: The inflammatory score (3.33±0.82 vs 0.67±0.52) and BALF total cell count [(10.13±4.83) ×105/ml vs (3.76±0.84) ×105/ml] in B group were higher than those in the A group with statistical significance (P<0.05). There was no significant difference between group C and group B in inflammatory infiltration score (2.83±0.75 vs 3.33±0.82) and BALF total cell count [(9.80±3.19) ×105/ml vs (10.13±4.83) ×105/ml] (P>0.05). Moreover the protein level of CARD9 was increased in the B group than A group (0.245±0.090 vs 0.047±0.014, P=0.004). Compared to E group and F group, more obviously inflammatory cells, neutrophils, eosinophils infiltration and tissue injury were observed in G group (P<0.05), so did the expression of IL-4 (P<0.05), IL-5 and IL-17. Meanwhile the mRNA expression levels of IL-17 and CXCL-10 also increased in lung tissue (P<0.05) of G group. Conclusion: CARD9 gene deletion may aggravate the steroid resistant of asthma by increasing neutrophil chemokines, such as IL-17 and CXCL-10, therefore increasing infiltration of neutrophils in C57BL/6 mice asthma model.

Dexmedetomidine alleviates acute lung injury by promoting Tregs differentiation via activation of AMPK/SIRT1 pathway.

To explore the anti-inflammatory effect and the potential mechanism of dexmedetomidine in ARDS/ALI. C57BL/6 mice and EL-4 cells were used in this research. The ALI model was established by CLP. The level of inflammatory cytokines in the lung and blood, the severity of lung injury, the expression of Foxp3, and the proportion of Tregs were detected before and after dexmedetomidine treatment. The expression of the AMPK/SIRT1 after dexmedetomidine treatment was detected in vivo and in vitro. After blocking the AMPK/SIRT1 pathway or depleting Tregs in vivo, the level of the inflammatory response, tissue injury, and Tregs differentiation were detected again to clarify the effect of dexmedetomidine. Dexmedetomidine significantly reduced systemic inflammation and lung injury in CLP mice. Dexmedetomidine enhanced the Foxp3 expression in the lungs and the frequency of Tregs in the spleen. Dexmedetomidine up-regulated the protein expression of p-AMPK and SIRT1 in lungs and EL-4 cells and facilitated the differentiation of naïve CD4+ T cells into Tregs in vitro. Meanwhile, DEX also increased the expression of Helios in Treg cells. DEX could improve ARDS/ALI by facilitating the differentiation of Tregs from naïve CD4+ T cells via activating the AMPK/SIRT1 pathway.

S100A1 expression is increased in spinal cord injury and promotes inflammation, oxidative stress and apoptosis of PC12 cells induced by LPS via ERK signaling.

Spinal cord injury (SCI) is a severe neurological disorder and the molecular mechanisms leading to its poor prognosis remain to be elucidated. S100A1, a mediator of Ca2+ handling of sarcoplasmic reticulum and mitochondrial function, operates as an endogenous danger signal (alarmin) associated with inflammatory response and tissue injury. The aim of the present study was to investigate the expression and biological effects of S100A1 in SCI. A rat model of SCI and a PC12 cell model of lipopolysaccharide (LPS)‑induced inflammation were established to examine S100A1 expression at the mRNA and protein levels. The inflammation level, which was mediated by S100A1, was determined based on inflammatory factor (IL‑1β, IL‑6 and TNF‑α) and anti‑inflammatory factor (IL‑10) expression. The effects of S100A1 on cellular oxidation and anti‑oxidation levels were observed by detecting the levels of reactive oxygen species, superoxide dismutase, catalase activities and nuclear factor erythroid 2‑related factor 2 expression. The protein levels of Bax, Bcl2 and cleaved caspase‑3 were used for the evaluation of the effects of S100A1 on apoptosis. Phosphorylated (p‑)ERK1/2 expression was used to evaluate the effects of S100A1 on ERK signaling. The results revealed that S100A1 expression was significantly upregulated invivo and invitro in the PC12 cell model of LPS‑inflammation. The silencing and overexpression of S100A1 helped alleviate and aggravate LPS‑induced inflammation, oxidative stress and apoptosis levels, respectively. S100A1 was found to regulate the ERK signaling pathway positively. An inhibitor of ERK signaling (MK‑8353) partially abolished the promoting effects of the overexpression of S100A1 on inflammation, oxidative stress damage and apoptosis. In conclusion, S100A1 expression was elevated in model of SCI and in the PC12 cell model of LPS‑induced inflammation. Furthermore, the overexpression/silencing S100A1 aggravated/mitigated the inflammation, oxidative stress damage and the apoptosis of LPS‑stimulated PC12 cells via the ERK signaling pathway. The present study revealed the mechanism of S100A1 in SCI, which provided a new theoretic reference for future research on SCI.

Diminished antiviral innate immune gene expression in the placenta following a maternal SARS-CoV-2 infection

BackgroundCoronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and is associated with critical illness requiring hospitalization, maternal mortality, stillbirth, and preterm birth. SARS-CoV-2 has been shown to induce placental pathology, however, substantial gaps exist in our understanding of the pathophysiology of COVID-19 disease in pregnancy and the long-term impact of SARS-CoV-2 on the placenta and fetus. To what extent a SARS-CoV-2 infection of the placenta alters the placental antiviral innate immune response is not well understood. A dysregulated innate immune response in the setting of maternal COVID-19 disease may increase risk for inflammatory tissue injury or placental compromise and contribute to deleterious pregnancy outcomes.ObjectiveWe sought to determine the impact of a maternal SARS-CoV-2 infection on placental immune response by evaluating gene expression of a panel of six antiviral innate immune mediators that act as biomarkers of the antiviral and interferon cytokine response. Our hypothesis was that a SARS-CoV-2 infection during pregnancy would result in an upregulated placental antiviral innate immune response.Study DesignWe performed a case control study on placental tissues [chorionic villous (CV) tissues and chorioamniotic membrane (CAM)] collected from pregnant patients with (N=140) and without (N=24) COVID-19 disease. We performed real-time quantitative polymerase chain reaction (qPCR) and immunohistochemistry. Placental histopathology was evaluated. Clinical data was abstracted. Fisher’s exact test, Pearson correlations, and linear regression models were used to examine proportions and continuous data between patients with active (<10 days since diagnosis) versus recovered COVID-19 (>10 days since diagnosis) at time of delivery. Secondary regression models adjusted for labor status as a covariate and evaluated potential correlation between placental innate immune gene expression and other variables.ResultsSARS-CoV-2 vRNA was detected in placental tissues from 5 women with COVID-19 and from no controls (0/24, 0%). Only 1 of 5 cases with detectable SARS-CoV-2 vRNA in placental tissues was confirmed to express SARS-CoV-2 nucleocapsid and spike proteins in syncytiotrophoblast cells. We detected a significantly lower gene expression of five critical innate immune mediators (IFNB, IFIT1, MXA, IL6, IL1B) in CV and CAM from women with active or recovered COVID-19 compared to controls, which remained significant after adjustment for labor status. There were minimal correlations between placental gene expression and other studied variables including: gestational age at diagnosis, time interval from COVID-19 diagnosis and delivery, pre-pregnancy body mass index, COVID-19 disease severity or placental pathology.ConclusionA maternal SARS-CoV-2 infection was associated with an impaired placental innate immune response in chorionic villous tissues and chorioamniotic membranes that was not correlated with gestational age at COVID-19 diagnosis, timer interval from COVID-19 diagnosis to delivery, maternal obesity, disease severity or placental pathology.

Neutrophil extracellular traps-triggered impaired autophagic flux via METTL3 underlies sepsis-associated acute lung injury

Neutrophil extracellular traps (NETs) assist pathogen clearance, while excessive NETs formation is associated with exacerbated inflammatory responses and tissue injury in acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). Autophagy is generally considered to be a protective process, but autophagy dysfunction is harmful. Whether and how NETs affect autophagic flux during sepsis-induced ALI are currently unknown. Here, we confirmed that the level of NETs was increased in ARDS patients and mice models, which led to impairment of autophagic flux and deterioration of the disease. Mechanistically, NETs activated METTL3 mediated m6A methylation of Sirt1 mRNA in alveolar epithelial cells, resulting in abnormal autophagy. These findings provide new insights into how NETs contribute to the development of sepsis-associated ALI/ARDS.

AFF3, a susceptibility factor for autoimmune diseases, is a molecular facilitator of immunoglobulin class switch recombination.

Immunoglobulin class switch recombination (CSR) plays critical roles in controlling infections and inflammatory tissue injuries. Here, we show that AFF3, a candidate gene for both rheumatoid arthritis and type 1 diabetes, is a molecular facilitator of CSR with an isotype preference. Aff3-deficient mice exhibit low serum levels of immunoglobulins, predominantly immunoglobulin G2c (IgG2c) followed by IgG1 and IgG3 but not IgM. Furthermore, Aff3-deficient mice show weak resistance to Plasmodium yoelii infection, confirming that Aff3 modulates immunity to this pathogen. Mechanistically, the AFF3 protein binds to the IgM and IgG1 switch regions via a C-terminal domain, and Aff3 deficiency reduces the binding of AID to the switch regions less efficiently. One AFF3 risk allele for rheumatoid arthritis is associated with high mRNA expression of AFF3, IGHG2, and IGHA2 in human B cells. These findings demonstrate that AFF3 directly regulates CSR by facilitating the recruitment of AID to the switch regions.

TRPV6 channel mediates alcohol-induced gut barrier dysfunction and systemic response.

SUMMARYIntestinal epithelial tight junction disruption is a primary contributing factor in alcohol-associated endotoxemia, systemic inflammation, and multiple organ damage. Ethanol and acetaldehyde disrupt tight junctions by elevating intracellular Ca2+. Here we identify TRPV6, a Ca2+-permeable channel, as responsible for alcohol-induced elevation of intracellular Ca2+, intestinal barrier dysfunction, and systemic inflammation. Ethanol and acetaldehyde elicit TRPV6 ionic currents in Caco-2 cells. Studies in Caco-2 cell monolayers and mouse intestinal organoids show that TRPV6 deficiency or inhibition attenuates ethanol- and acetaldehyde-induced Ca2+ influx, tight junction disruption, and barrier dysfunction. Moreover, Trpv6−/− mice are resistant to alcohol-induced intestinal barrier dysfunction. Photoaffinity labeling of 3-azibutanol identifies a histidine as a potential alcohol-binding site in TRPV6. The substitution of this histidine, and a nearby arginine, reduces ethanol-activated currents. Our findings reveal that TRPV6 is required for alcohol-induced gut barrier dysfunction and inflammation. Molecules that decrease TRPV6 function have the potential to attenuate alcohol-associated tissue injury.