Xuanfu Daizhe Tang alleviates reflux esophagitis in rats by inhibiting the STAT1/TREM-1 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Reflux esophagitis (RE) is a common chronic inflammatory disease of the esophageal mucosa with a high prevalence and recurrence rate, for which a satisfactory therapeutic strategy is still lacking. Chinese medicine has its characteristics and advantages in treating RE, and the clinical application of Xuanfu Daizhe Tang (XDT) in treating RE has achieved sound therapeutic effects. However, there needs to be more research on its mechanism of action. AIM OF THE STUDY: The present work aimed to investigate the mechanism of XDT action in RE through the Signal Transducer and Activator of Transcription 1 (STAT1)/Triggering Receptor Expressed on Myeloid cells-1 (TREM-1) pathway. MATERIALS AND METHODS: The main active components of XDT were analyzed by ultra-performance liquid chromatography-mass spectrometer (UPLC-MS). The effect of XDT on RE was evaluated in a rat model of RE induced by "Cardioplasty + pyloric ligation + Roux-en-Y esophagojejunostomy". Each administration group was treated by gavage. The degree of damage to the esophageal mucosa was evaluated by visual observation, and the Potential of Hydrogen (PH) method and Hematoxylin-eosin staining (HE) staining were performed. Serum levels of Interleukin-1ß (IL-1ß), Interleukin-6 (IL-6), Tumor Necrosis Factor alpha (TNF-α), and Inducible Nitric Oxide Synthase (iNOS) were measured by ELISA. Quantitative Real-time PCR (qPCR), Western Blot (WB), and Immunofluorescence (IF) methods were used to detect Claudin-4, Claudin-5, TREM-1, and p-STAT1 in esophageal tissues for studying the mechanism of action and signaling pathway of XDT. Immunohistochemistry (IHC) analysis was used to detect the expression of TREM-1 and CD68 in esophageal tissues. Flow Cytometry (FC) was used to detect the polarization of macrophages in the blood. After conducting preliminary experiments to verify our hypothesis, we performed molecular docking between the active component of XDT and STAT1 derived from rats and parallel experiments with STAT1 inhibitor. The selective increaser of STAT1 transcription (2-NP) group was used to validate the mechanism by which XDT acts. RESULTS: XDT alleviated esophageal injury and attenuated histopathological changes in RE rats. XDT also inhibited the inflammatory response and decreased serum IL-1ß, IL-6, TNF-α, and iNOS levels in RE rats. qPCR and WB results revealed that XDT inhibited the expression of Claudin-4, Claudin-5, TREM-1, and STAT1 in the esophageal mucosa of RE rats. IHC and FC results showed that XDT reduced TREM-1 levels in esophageal tissues and polarized macrophages toward M2. The molecular docking results showed that rat-derived STAT1 can strongly bind to Isochronogenic acid A in XDT. The parallel experimental results of STAT1 inhibitor showed that XDT has anti-inflammatory effects similar to STAT1 inhibitors. The 2-NP group confirmed that XDT exerts its therapeutic effect on reflux esophagitis through the STAT1/TREM-1 pathway, with STAT1 as the upstream protein. CONCLUSIONS: This study suggests that XDT may treat reflux esophagitis by modulating the STAT1/TREM-1 pathway.

Salidroside alleviates ulcerative colitis via inhibiting macrophage pyroptosis and repairing the dysbacteriosis-associated Th17/Treg imbalance.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by flora disequilibrium and mucosal immunity disorder. Here, we report that salidroside effectively restricts experimental colitis from two aspects of intestinal macrophage pyroptosis and dysbacteriosis-derived colonic Th17/Treg imbalance. In innate immunity, the upregulated TREM1 and pyroptosis-related proteins in inflamed colons were inhibited by salidroside administration and further experiments in vitro showed that salidroside suppressed LPS/ATP-induced bone marrow-derived macrophages (BMDMs) pyroptosis evident by the decline of LDH and IL-1ß release as well as the protein level of NLRP3, caspase-1, and GSDMD p30. Moreover, the TREM1 inhibitor weakened the effect of salidroside on BMDMs pyroptosis, whereas salidroside still could downregulate TREM1 when NLRP3 was inhibited. In adaptive immunity, salidroside improved the gut microflora diversity and Th17/Treg ratio in DSS-induced mice, especially promoting the abundance of Firmicutes. Clearance of the gut flora blocked the benefit of salidroside on colonic inflammation and Th17/Treg adaptive immunity, but transplanting salidroside-treated foecal bacterium into flora-depleted wild mice reproduced the resistance of salidroside to gut inflammation. Taken together, our data demonstrated that salidroside protected experimental colitis via skewing macrophage pyroptosis and Th17/Treg balance, indicating its potential effect on UC and other immune disorders.

Shengxian decoction decreases doxorubicin­induced cardiac apoptosis by regulating the TREM1/NF­κB signaling pathway.

Shengxian decoction (SXT) is a traditional Chinese medicine that is clinically used for treating cardiovascular diseases. It is known for its beneficial effect on cardiomyocyte injuries, some of which can be induced by anticancer agents including doxorubicin (DOX). To determine the molecular mechanisms involved in the cardioprotective effects of SXT, DOX­induced H9c2 cells were analyzed for apoptosis and expression levels of apoptosis biomarkers. Cell viability and apoptosis were measured by CCK­8 and flow cytometry. Triggering receptors expressed on myeloid cells 1 (TREM1), cleaved caspase­3, survivin and NF­κBp65 expression levels were measured by reverse transcription­quantitative PCR and/or western blotting. A total of 30 adult male Sprague­Dawley rats were randomly allocated into five groups (n=6 each); control group receiving 0.9% saline, 1 DOX group receiving 2.5 mg/kg of DOX and 3 DOX + SXT groups, receiving a DOX dose equivalent to the DOX­only group and either 0.4, 0.8 or 1.6 g/kg of SXT. It was found that DOX increased apoptosis and NF­κB activation of H9c2 cells by increasing TREM1 expression and that SXT inhibited apoptosis and NF­κB activation of H9c2 cells induced by DOX or Trem1 overexpression. SXT also significantly reversed DOX­induced cardiotoxicity in rats. The results suggested that the protective effects of SXT against DOX­induced apoptosis may be attributed to its downregulation of TREM1.

Epigallocatechin-3-Gallate (EGCG), an Active Compound of Green Tea Attenuates Acute Lung Injury Regulating Macrophage Polarization and Krüpple-Like-Factor 4 (KLF4) Expression.

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) are serious clinical complications with a high frequency of morbidity and mortality. The initiation and amplification of inflammation is a well-known aspect in the pathogenesis of ALI and related disorders. Therefore, inhibition of the inflammatory mediators could be an ideal approach to prevent ALI. Epigallocatechin-3-gallate (EGCG), a major constituent of green tea, has been shown to have protective effects on oxidative damage and anti-inflammation. The goal of the present study was to determine whether EGCG improves phenotype and macrophage polarisation in LPS-induced ALI. C57BL/6 mice were given two doses of EGCG (15 mg/kg) intraperitoneally (IP) 1 h before and 3 h after LPS instillation (2 mg/kg). EGCG treatment improved histopathological lesions, Total Leucocyte count (TLC), neutrophils infiltration, wet/dry ratio, total proteins and myeloperoxidase (MPO) activity in LPS-induced lung injury. The results displayed that EGCG reduced LPS-induced ALI as it modulates macrophage polarisation towards M2 status. Furthermore, EGCG also reduced the expression of proinflammatory M1 mediators iNOS TNF-α, IL-1ß and IL-6 in the LPS administered lung microenvironment. In addition, it increased the expression of KLF4, Arg1 and ym1, known to augment the M2 phenotype of macrophages. EGCG also alleviated the expression of 8-OHdG, nitrotyrosine, showing its ability to inhibit oxidative damage. TREM1 in the lung tissue and improved lung regenerative capacity by enhancing Ki67, PCNA and Ang-1 protein expression. Together, these results proposed the protective properties of EGCG against LPS-induced ALI in may be attributed to the suppression of M1/M2 macrophages subtype ratio, KLF4 augmentation, lung cell regeneration and regulating oxidative damage in the LPS-induced murine ALI.

Inhibition of a triggering receptor expressed on myeloid cells-1 (TREM-1) with an extracellular cold-inducible RNA-binding protein (eCIRP)-derived peptide protects mice from intestinal ischemia-reperfusion injury.

BACKGROUND: Intestinal ischemia-reperfusion injury results in morbidity and mortality from both local injury and systemic inflammation and acute lung injury. Extracellular cold-inducible RNA-binding protein is a damage associated molecular pattern that fuels systemic inflammation and potentiates acute lung injury. We recently discovered a triggering receptor expressed on myeloid cells-1 serves as a novel receptor for extracellular cold-inducible RNA-binding protein. We developed a 7-aa peptide, named M3, derived from the cold-inducible RNA-binding protein, which interferes with cold-inducible RNA-binding protein's binding to a triggering receptor expressed on myeloid cells-1. Here, we hypothesized that M3 protects mice against intestinal ischemia-reperfusion injury. METHODS: Intestinal ischemia was induced in C57BL/6 mice via clamping of the superior mesenteric artery for 60 minutes. At reperfusion, mice were treated intraperitoneally with M3 (10 mg/kg body weight) or normal saline vehicle. Mice were killed 4 hours after reperfusion and blood and lungs were collected for various analysis. A 24-hours survival after intestinal ischemia-reperfusion was assessed. RESULTS: Serum levels of organ injury markers aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and lactate were increased with intestinal ischemia-reperfusion, while treatment with M3 significantly decreased their levels. Serum, intestinal, and lung levels of proinflammatory cytokines and chemokines were also increased by intestinal ischemia-reperfusion, and treatment with M3 significantly reduced these values. Intestinal ischemia-reperfusion caused significant histological intestinal and lung injuries, which were mitigated by M3. Treatment with M3 improved the survival from 40% to 80% after intestinal ischemia-reperfusion. CONCLUSION: Inhibition of triggering receptor expressed on myeloid cells-1 by an extracellular cold-inducible RNA-binding protein-derived small peptide (M3) decreased inflammation, reduced lung injury, and improved survival in intestinal ischemia-reperfusion injury. Thus, blocking the extracellular cold-inducible RNA-binding protein-triggering receptor expressed on myeloid cells-1 interaction is a promising therapeutic avenue for mitigating intestinal ischemia-reperfusion injury.

Novel TREM-1 Inhibitors Attenuate Tumor Growth and Prolong Survival in Experimental Pancreatic Cancer.

Pancreatic cancer (PC) is a highly lethal cancer with an urgent need to expand the limited treatment options for patients. Tumor-associated macrophages (TAMs) promote tumor aggressiveness and metastasis. High expression of triggering receptor expressed on myeloid cells 1 (TREM-1) on TAMs directly correlates with poor survival in patients with non-small cell lung cancer (NSCLC). We have previously hypothesized that blockade of TREM-1 could be a promising therapeutic strategy to treat cancer and shown that the novel, ligand-independent TREM-1 inhibitory peptides rationally designed using the signaling chain homooligomerization (SCHOOL) strategy suppress NSCLC growth in vivo. Here, we evaluated the therapeutic potential of these inhibitors in three human PC xenograft mouse models. Administration of SCHOOL peptides resulted in a strong antitumor effect achieving an optimal treatment/control (T/C) value of 19% depending on the xenograft and formulation used and persisting even after treatment was halted. The effect correlated significantly with increased survival and suppressed TAM infiltration. The peptides were well-tolerated when deployed either in free form or formulated into lipopeptide complexes for peptide half-life extension and targeted delivery. Finally, blockade of TREM-1 significantly reduced serum levels of interleukin (IL)-1α, IL-6, and macrophage colony-stimulating factor (M-CSF), but not vascular endothelial growth factor, suggesting M-CSF-dependent antitumor mechanisms. Collectively, these promising data suggest that SCHOOL TREM-1-specific peptide inhibitors have a cancer type independent, therapeutically beneficial antitumor activity and can be potentially used as a stand-alone therapy or as a component of combinational therapy for PC, NSCLC, and other solid tumors.

Inhibitory effects of dietary soyasaponin on 2,4-dinitrofluorobenzene-induced contact hypersensitivity in mice.

Soyasaponins (SSs) abundant in soybean have anti-inflammatory activities; however, their therapeutic effects on allergic contact dermatitis (ACD) remain unknown. To assess the effects of SS-enriched diets on ACD, we used a mouse model of contact hypersensitivity (CHS). Mice were fed low-dose or high-dose SS-containing diets for 3 weeks prior to CHS induction with 2,4-dinitrofluorobenzene (DNFB). The low-dose SS diet attenuated DNFB-induced ear swelling and tissue oedema, and reduced the number of infiltrating Gr-1-positive myeloid cells. Low-dose, but not high-dose, SSs decreased chemokine (C-X-C motif) ligand 2 (CXCL2) and triggering receptor expressed on myeloid cells (TREM)-1 production in ear tissues, compared to a control. Taxonomic 16S rRNA analysis revealed significant alterations in faecal microbiota caused by CHS, which were reversed by low-dose SSs. The low-dose SS and non-CHS groups clustered together, while the high-dose SS group split between CHS and non-CHS clusters. Our results demonstrated that low-dose SSs alleviated CHS symptoms by attenuating inflammation and improving the intestinal microbiota composition, suggesting that dietary SSs may have beneficial effects on ACD.