Xijiao Dihuang decoction combined with Yinqiao powder promotes autophagy-dependent ROS decrease to inhibit ROS/NLRP3/pyroptosis regulation axis in influenza virus infection.

BACKGROUND: Influenza viral pneumonia is a common complication after influenza virus infection. Xijiao Dihuang Decoction combined with Yinqiao Powder (XDY) is effective on improving influenza viral pneumonia. PURPOSE: This study further explores the anti-inflammatory mechanism of XDY in the treatment of influenza viral pneumonia. STUDY DESIGN: The effects of XDY on inflammation, autophagy, NACHT-LRR-PYD-containing protein 3 (NLRP3) inflammasome and pyroptosis were assessed in the mice with influenza viral pneumonia. In addition, the mouse macrophage cell line (J774A.1) infected with influenza virus was adopted to decode the in vitro effects of XDY on autophagy, reactive oxygen species (ROS), NLRP3 inflammasome and pyroptosis. We analyzed the XDY-induced autophagy, especially the mitophagy-related ROS clearance, and the subsequent inhibition of ROS/NLRP3 inflammasome/pyroptosis signaling in the infected macrophages by different assays based on quantitative polymerase chain reaction, western blot, flow cytometry, immunofluorescence and enzyme-linked immunosorbent assay. RESULTS: In vivo, XDY could effectively improve the lung inflammatory response in the mice with influenza virus pneumonia, due to an intact autophagy flux-promoting effect and the inhibiting roles on NLRP3 inflammasome and pyroptosis. Notably, in vitro, compared with the infected macrophages treated by the NLRP3 inflammasome agonist (Monosodium urate) or the mitochondrial-targeted antioxidant agent, the XDY-dependent treating could inhibit pyroptosis by negatively regulating the signaling axis of ROS/NLRP3 inflammasome/pyroptosis in the influenza virus-infected macrophages. More interestingly, XDY could promote an intact autophagy flux, inducing mitophagy eliminating the damaged mitochondria to reduce the intracellular ROS accumulation, and thus decrease the oxidative stress in the infected macrophages. Especially, the inhibitor of autophagy inition, 3-Methyladenine, could reverse the inhibitory effect of XDY on ROS-NLRP3 inflammasome-mediated pyroptosis, indicating an XDY-promoted mitophagy-dependent ROS scavenging. CONCLUSION: XDY can promote an intact autophagy flux to eliminate damaged mitochondria, namely mitophagy, which reduces the intracellular ROS accumulation contributing to NLRP3 inflammasome activation, restricting pyroptosis and eventually alleviating the influenza virus-induced inflammatory lesions. The obtained results provide new insights into the mechanism of action of XDY in alleviating influenza virus pneumonia, especially the roles of XDY in anti-oxidation, anti-inflammation and anti-pyroptosis, with potential therapeutic targets for future application in integrative medicine.

Probiotics (Lactobacillus plantarum HNU082) Supplementation Relieves Ulcerative Colitis by Affecting Intestinal Barrier Functions, Immunity-Related Gene Expression, Gut Microbiota, and Metabolic Pathways in Mice.

Probiotics can effectively improve ulcerative colitis (UC), but the mechanism is still unclear. Here, shotgun metagenome and transcriptome analyses were performed to explore the therapeutic effect and the mechanism of the probiotic Lactobacillus plantarum HNU082 (Lp082) on UC. The results showed that Lp082 treatment significantly ameliorated dextran sulfate sodium (DSS)-induced UC in mice, which was manifested as increases in body weight, water intake, food intake, and colon length and decreases in disease activity index (DAI), immune organ index, inflammatory factors, and histopathological scores after Lp082 intake. An in-depth study discovered that Lp082 could improve the intestinal mucosal barrier and relieve inflammation by cooptimizing the biological barrier, chemical barrier, mechanical barrier, and immune barrier. Specifically, Lp082 rebuilt the biological barrier by regulating the intestinal microbiome and increasing the production of short-chain fatty acids (SCFAs). Lp082 improved the chemical barrier by reducing intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule (VCAM) and increasing goblet cells and mucin2. Lp082 ameliorated the mechanical barrier by increasing zonula occludens-1 (ZO-1), zonula occludens-2 (ZO-2), and occludin while decreasing claudin-1 and claudin-2. Lp082 optimized the immune barrier by reducing the content of interleukin-1ß (IL-1ß), IL-6, tumor necrosis factor-α (TNF-α), myeloperoxidase (MPO), and interferon-γ (IFN-γ) and increasing IL-10, transforming growth factor-ß1 (TGF-ß1), and TGF-ß2, inhibiting the NF-κB signaling pathway. Taken together, probiotic Lp082 can play a protective role in a DSS-induced colitis mouse model by protecting the intestinal mucosal barrier, attenuating the inflammatory response, and regulating microbial imbalance. This study provides support for the development of probiotic-based microbial products as an alternative treatment strategy for UC. IMPORTANCE Many studies have focused on the therapeutic effect of probiotics on ulcerative colitis (UC), but few studies have paid attention to the mechanism of probiotics, especially the therapeutic effect. This study suggests that Lp082 has a therapeutic effect on colitis in mice. Its mechanisms of action include protecting the mucosal barrier and actively modulating the gut microbiome, modulating inflammatory pathways, and reducing neutrophil infiltration. Our study enriches the mechanism and provides a new prospect for probiotics in the treatment of colitis, helps to deepen the understanding of the intestinal mucosal barrier, and provides guidance for the future probiotic treatment of human colitis.

Berberine ameliorates pulmonary inflammation in mice with influenza viral pneumonia by inhibiting NLRP3 inflammasome activation and gasdermin D-mediated pyroptosis.

Viral pneumonia is a common complication caused by Influenza A virus infection and is characterized by severe pulmonary inflammation. A previous study showed that berberine (BBR) significantly ameliorated the pulmonary inflammation in mice with influenza viral pneumonia but its underlying mechanism is not entirely understood. In this study, we reproduced the mouse model of influenza viral pneumonia through intranasal infection of A/Puerto Rico/8/34 (H1N1), to further investigate the anti-inflammatory mechanism of BBR based on nucleotide-binding oligomerization domain-like receptor protein (NLRP) 3 inflammasome activation and Gasdermin D (GSDMD)-mediated pyroptosis. Consistent with MCC950 (10 mg/kg, a specific NLRP3 inflammasome inhibitor), BBR (10 mg/kg) obviously improved the weight loss and survival rate of infected mice, alleviated their pulmonary inflammation, and suppressed the accumulation of tumor necrosis factor and interleukin (IL)-6 in lungs without obvious inhibition on viral multiplication (hemagglutinin titer and nucleoprotein messenger RNA). Moreover, BBR (10 mg/kg) reduced the expressions of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), and cysteinyl aspartate-specific proteinase (Caspase)1 (Caspase1 precursor [Pro-caspase1] + Caspase1p20 subunit) and the ratio of Caspase1p20 subunit to Caspase1, thus inhibiting the NLRP3 inflammasome activation and resulting in the decreased contents of mature IL-1ß and IL-18 in lungs. The GSDMD expression (GSDMD precursor [Pro-GSDMD] + GSDMD-N terminal [NT]) and the ratio of GSDMD-NT to GSDMD were also declined by BBR (10 mg/kg). These evidence indicate that BBR may ameliorate pulmonary inflammation in mice with influenza viral pneumonia through inhibiting NLRP3 inflammasome activation, as well as depressing GSDMD-mediated pyroptosis via declining GSDMD expression and restraining NLRP3 inflammasome-mediated GSDMD activation.

Lactobacillus plantarum HNU082 alleviates dextran sulfate sodium-induced ulcerative colitis in mice through regulating gut microbiome.

Probiotics have shown good efficacy in the prevention of ulcerative colitis (UC), but the specific mechanism remains unclear. Therefore, shotgun metagenomic and transcriptome analyses were performed to explore the preventive effect of a potential probiotic Lactobacillus plantarum HNU082 (Lp082) on UC and its specific mechanism. The results showed that Lp082 intervention ameliorated dextran sulfate sodium (DSS)-induced UC in mice, which was manifested in the increase in body weight, water intake, food intake, and colon length and the decrease in the DAI index, immune organ index, inflammatory factors and histopathological scores after Lp082 intake. The mechanism is deeply studied and it is discovered that Lp082 improves the intestinal mucosal barrier by co-optimizing biological barriers, chemical barriers, mechanical barriers, and immune barriers. Specifically, Lp082 improved the biological barrier by increasing the diversity, optimizing the species composition and the structure of the gut microbiota, increasing bacteria producing short chain fatty acids (SCFAs), and activating microbial metabolic pathways producing SCFAs so as to enhance the content of SCFAs. Lp082 optimized the chemical barrier by decreasing the mRNA expression of ICAM-1 and VCAM and by increasing the content of goblet cells and the mRNA expression and immunofluorescent protein content of mucin2. Lp082 ameliorated the mechanical barrier by decreasing the mRNA expression of claudin-1 and claudin-2, and by increasing the mRNA expression of ZO-1 and ZO-2 and the immunofluorescent protein content of ZO-1. Lp082 also optimized the immune barrier by increasing the mRNA expression of IL-10, TGF-ß1, and TGF-ß2 and by decreasing the mRNA expression and protein contents of IL-6, tumour necrosis factor-alpha (TNF-α) and myeloperoxidase (MPO). In addition, Lp082 can also regulate the metabolic pathways of inflammation and disease in mice, and notably, Lp082 inhibits the NF-κB signaling pathway by inhibiting NF-κB signaling molecules to alleviate UC. In conclusion, improving gut microbiota dysbiosis, protecting the intestinal mucosal barrier, regulating inflammatory and disease pathways, and affecting neutrophil infiltration are the potential mechanisms of probiotic Lp082 in alleviating UC. Our study enriches the mechanism and provides a new prospect for Lactobacillus plantarum HNU082 in the prevention of colitis, provides support for the development of probiotic-based microbial products as an alternative prevention strategy for UC, and provides guidance for the future probiotic prevention of human colitis.

Establishing a novel inflammatory bowel disease prediction model based on gene markers identified from single nucleotide variants of the intestinal microbiota.

The intestinal microbiota is a crucial environmental factor in the development of inflammatory bowel disease (IBD). The abundance of Faecalibacterium prausnitzii is significantly decreased in IBD patients, which is used as a biomarker for IBD diagnosis. However, this can be observed in both IBD and colorectal cancer, which would confound the diagnostic results. Thus, we first established a new model for predicting Crohn's disease (CD) with high precision according to gene characteristics based on single nucleotide variants (SNVs). Next, five gene markers belonging to two species, F. prausnitzii and Eubacterium rectale, that were enriched in the CD group were obtained to build a CD prediction model, and high accuracy in distinguishing the CD and control groups was observed in the discovery (area under curve [AUC] = 91.13%) and validation cohorts (AUC = 79.55%). The model still maintained high accuracy after expanding the healthy cohort (AUC = 89.75%). High disease specificity in distinguishing CD and CRC groups (AUC = 95.74%) was also proven. This study establishes a novel diagnostic method for predicting IBD that also provides unprecedented insight for the early, painless diagnosis of other non-communicable diseases.