Glycyrrhiza uralensis Fisch. alleviates dextran sulfate sodium-induced colitis in mice through inhibiting of NF-κB signaling pathways and modulating intestinal microbiota.

Licorice is widely used in traditional Chinese Medicine (TCM) for compound compatibility, which could reduce toxicity and increase efficacy of certain herbal medicine, and its active components prominently effects of inhibit of inflammation and regulate of immunity. The study probed into the mechanism of the anti-inflammatory and immunomodulatory effects of licorice based on the domination of the T helper type 17/regulatory T cells (Th17/Treg) differentiation balance and the composition and structure of the intestinal flora through the nuclear factor kappa B (NF-κB) signaling pathway. BALB/c mice were inoculated with dextran sulfate sodium (DSS) to establish animal models of ulcerative colitis (UC). For the pharmacodynamic study, UC mice were observed for the anti-inflammatory effect of licorice water extraction (LWE) in vivo, including clinical observation and measurement of colon length. Hematoxylin-eosin (HE) staining was used to evaluate pathological conditions. Immunohistochemistry (IHC) and transmission electron microscopy (TEM) were performed to observe the intestinal barrier of the colons. Inflammatory cytokine levels were measured using with enzyme-linked immunosorbent assay (ELISA) kits. The proportions of T helper (Th) cells in the colons was assessed using flow cytometry. Gut microbiota diversity was detected using 16S ribosomal (r)DNA sequencing. In addition, Western blot (WB) assays were used to verify ROR-γt, Foxp3, TLR4, MyD88 and NF-κB expression according to a standard protocol. LWE exerted a pharmacological anti-inflammatory effect by attenuating inflammation in the colonic tissues through affecting the protein expression of TLR4/MyD88/NF-κB, and increasing the expression of tight junction (TJ) protein in the colons, improving the integrity of the intestinal mucosal barrier in vivo. Moreover, LWE reversed the imbalance in Th17/Treg cells differentiation and influenced the protein expression of ROR-γt and Foxp3 in UC mouse colons. In particular, LWE significantly affected the diversity of the gut microbiota in UC mice, ameliorated the composition of dominant species, and significantly increased the type and quantity of probiotics. Licorice tends to reduce inflammation and enhance the protective action of the intestinal mucosal barrier via the TLR4/MyD88/NF-κB signal transduction pathway and alter the imbalance of Th-cell differentiation. Notably, licorice may affect the diversity of intestinal microbiota and the content of beneficial bacteria in the colon, which is a potential mechanism for understanding anti-inflammatory and immunomodulatory effects in UC mice in vivo.

Shi G ，Kong J ，Wang Y ，Xuan Z ，Xu F ... -  《-》

Sanhuang Xiexin decoction ameliorates DSS-induced colitis in mice by regulating intestinal inflammation, intestinal barrier, and intestinal flora.

Sanhuang Xiexin decoction (SXD) is a widely applicated traditional Chinese medicine (TCM) with a significant intestinal anti-inflammatory effect. To evaluate the therapeutic effect and elucidate the possible underlying mechanisms of SXD on dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) in mice. To model UC, 3% DSS was added to the drinking water for 7 days. The UC mice were grouped and treated with three doses of SXD (1.3, 2.6, and 6 g/kg) orally for 7 days. Mice body weight and disease activity index (DAI) scores were recorded daily. After treatment with SXD, the colon was removed, and the colon length and histopathological changes were recorded. Blood cells were counted and colonic inflammatory cytokines and oxidative stress indicators were examined. The key proteins in TLR4-MyD88-NF-κB signaling and the colonic barrier were determined by Western blot analysis. The restorative effect of SXD on intestinal flora was determined. Treatment with SXD reduced DAI scores, increased body weight, improved colon shortening, and decreased colonic damage. SXD decreased the numbers of white blood cells (WBCs), increased the numbers of red blood cells (RBCs), and inhibited the expression of inflammatory cytokines and oxidative stress indicators. In addition, SXD displayed an effective anti-inflammatory effect by inhibiting the expression levels of p-IκBα, TLR4, MyD88, and p65. Furthermore, SXD significantly restored the integrity of the colonic barrier and the abundance of beneficial flora. SXD significantly reduced DSS-induced colon damage when the dose was higher than 1.3 g/kg, and the middle dose group (2.6 g/kg) indicated the best effect. SXD effectively ameliorated DSS-induced UC in mice, possibly by inhibiting oxidative stress, protecting the mucosal barrier, inhibiting the TLR4-MyD88-NF-κB signaling pathway, and regulating the intestinal flora.

Wu X ，Fu S ，Jiang M ，Wang J ，Tang H ，Fang C ，Li W ，Fu C ... -  《-》

Ganluyin ameliorates DSS-induced ulcerative colitis by inhibiting the enteric-origin LPS/TLR4/NF-κB pathway.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD), that is associated with a significantly increased risk of colon cancer. As a classic traditional Chinese medicine, Ganluyin (GLY) has a long history as an anti-inflammatory medication, but its impacts on UC has not been established. This study aims to evaluate the protective effect and mechanism of GLY on a pathway involving enteric-origin lipopolysaccharide (LPS), toll-like receptor (TLR)4, and NF-κB in mice with dextran sulfate sodium (DSS)-induced UC. After three weeks of intragastric administration of GLY, a UC model was induced in mice by administration of 4% DSS in drinking water for one week. The disease activity index (DAI) was measured, and histological staining was used to detect histopathological changes of colon. LPS content of the serum was measured by ELISA, and the expression of tight junction proteins and proteins related to TLR4/NF-κB pathway in colon were analyzed by immunohistochemistry or Western Blotting. The intestinal flora was analyzed by 16S rRNA sequencing. GLY improved the histological pathological changes of DSS-induced UC, as assessed by DAI, colonic mucosal damage, inflammatory cell infiltration, and goblet cell and mucus reduction. GLY also protected the intestinal mucosal barrier by increasing the expression of the tight junction proteins, occludin, claudin-1, and ZO-1 and by reducing the serum LPS content and decreasing the expression of TLR4, MyD88, NF-κB, IL-6, IL-1β, and TNF-α proteins in colon. Analyses of the intestinal flora showed that GLY restored the homeostasis of the intestinal flora through increases in the abundance of Firmicutes and decreases in the abundance of Proteobacteria and Bacteroidetes, which is associated with the production of LPS. GLY might exert an anti-UC effect by improving the colonic mucosal barrier and inhibiting the enteric-origin LPS/TLR4/NF-κB inflammatory pathway, and restoring the homeostasis of the intestinal flora in UC mice. These discoveries lay a strong foundation for GLY as a UC treatment.

Xiong T ，Zheng X ，Zhang K ，Wu H ，Dong Y ，Zhou F ，Cheng B ，Li L ，Xu W ，Su J ，Huang J ，Jiang Z ，Li B ，Zhang B ，Lv G ，Chen S ... -  《-》

Xianglian Pill attenuates ulcerative colitis through TLR4/MyD88/NF-κB signaling pathway.

Xianglian Pill (XLP) is a classical Chinese medicine prescription applied for controlling ulcerative colitis (UC). Whereas, the underlying mechanism remains unclear. The present work was aimed to investigate the mechanism of XLP in dextran sulfate sodium (DSS)-induced UC via the Toll Like Receptor 4 (TLR4)/Myeloid Differentiation factor 88 (MyD88)/Nuclear Factor kappa-B (NF-κB) signaling in mice. The major components of XLP were detected by high-performance liquid chromatography-diode array detection (HPLC-DAD). The ulcerative colitis model was induced by DSS in mice. 5-Amino Salicylic Acid (5-ASA) group and XLP group were intragastrically treated. Disease activity index (DAI) and colon length were monitored and hematoxylin-eosin (HE) staining was conducted. Gasdermin D (GSDMD)-N and TLR4 expressions in colon tissues were visualized by immunofluorescence. TLR4 mRNA was measured by Real Time Quantitative PCR (RT-qPCR). The expressions of NOD-like receptor thermal protein domain associated protein 3 (NLRP3), active-caspase-1, GSDMD-N, TLR4, MYD88, NF-κB, p-NF-κB, and the ubiquitination of TLR4 in colon tissues were detected by Western blot. Myeloperoxidase (MPO) enzyme activity was examined and serum inflammatory factors Interleukin (IL)-1β, IL-6, Tumor Necrosis Factor-α (TNF-α), and IL-18 were determined by Enzyme-linked Immunosorbent Assay (ELISA). TLR4-/- mice were applied for verifying the mechanism of XLP attenuated DSS symptoms. The XLP treatment extended colon length, reduced DAI, and attenuated histopathological alteration in DSS-induced mice. XLP administration suppressed MPO activity and reduced the content of IL-1β, IL-6, TNF-α and IL-18 in serum. XLP also inhibited the expression levels of GSDMD-N, TLR4, NLRP3, active-caspase-1, MyD88, p-NF-κB/NF-κB in colon tissues of DSS-induced mice. TLR4-/- mice proved that TLR4 was involved in XLP-mediated beneficial effect on DSS-induced ulcerative colitis. XLP might treat ulcerative colitis by regulating the TLR4/MyD88/NF-κB signaling pathway.

Dai Y ，Lu Q ，Li P ，Zhu J ，Jiang J ，Zhao T ，Hu Y ，Ding K ，Zhao M ... -  《-》

Fufangxiaopi formula alleviates DSS-induced colitis in mice by inhibiting inflammatory reaction, protecting intestinal barrier and regulating intestinal microecology.

Fufangxiaopi Formula (FF) is a modified form of Sishen Wan, traditionally used for treating diarrhea. The application of FF for treating ulcerative colitis (UC) has achieved desirable outcomes in clinical settings. However, the underlying mechanism of the effect of FF on UC is yet to be determined. This study aimed to evaluate the protective effect and underlying mechanism of FF on mice with dextran sodium sulfate (DSS)-induced colitis. In vivo, the efficacy of FF on the symptoms associated with DSS-induced colitis in mice was clarified by observing the body weight change, colon length, DAI score, and H&E staining. The release of inflammatory mediators in mouse colon tissues was detected by ELISA and MPO, and the contents of TLR4/NF-κB signaling pathway and MAPK signaling pathway-related proteins, as well as intestinal barrier-related proteins, were detected in mouse colon tissues by western blot method. Changes in the content of barrier proteins in mouse colon tissues were detected by immunofluorescence. 16S rRNA sequencing and FMT were performed to clarify the effects of FF on intestinal flora. In vitro, the effect of FF-containing serum on LPS-induced inflammatory mediator release from RAW264.7 cells were detected by qRT-PCR. The contents of TLR4/NF The effects of FF-containing serum on B signaling pathway and MAPK signaling pathway related proteins in RAW264.7 cells and intestinal barrier related proteins in Caco-2 cells were detected by western blot. The effects of FF-containing serum on LPS-induced nuclear translocation of p65 protein in RAW264.7 cells and barrier-associated protein in Caco-2 cells were detected by immunofluorescence. In vivo studies showed that FF could significantly alleviate the symptoms of UC, including reducing colon length, weight loss, clinical score, and colon tissue injury in mice. FF could significantly reduce the secretion of proinflammatory cytokines by suppressing the activation of the TLR4/NF-κB and MAPK signaling pathways. Moreover, FF could protect the integrity of intestinal barriers by significantly increasing claudin-3, occludin, and ZO-1 expression levels. 16S rRNA sequencing and FMT elucidate that FF can alleviate symptoms associated with colitis in mice by interfering with intestinal flora. In vitro studies showed that FF drug-containing serum could significantly inhibit proinflammatory responses and attenuate the secretion of iNOS, IL-1β, TNF-α, IL-6, and COX-2 by suppressing the activation of TLR4/NF-κB and MAPK signaling pathways in RAW264.7 cells. Furthermore, FF could protect the Caco-2 cell epithelial barrier. FF could alleviate DSS-induced colitis in mice by maintaining the intestinal barrier, inhibiting the activation of TLR4/NF-κB and MAPK signaling pathways, reducing the release of proinflammatory factors, and regulating intestinal microecology.

Liu K ，Shi C ，Yan C ，Yin Y ，Qiu L ，He S ，Chen W ，Li G ... -  《-》