Hemorrhoids are commonly associated with inflammatory manifestations. Puji Zhichuang suppositories (PJZC) have been widely applied in the treatment of hemorrhoids; however, their underlying therapeutic mechanisms remain insufficiently understood. Therefore, this study investigated the mechanisms of PJZC in inflammatory hemorrhoids using network pharmacology, 16 S rDNA sequencing, and fecal metabolomics analyses. First, the anti-inflammatory effects of PJZC were validated by evaluating perianal diameter, anorectal coefficient, TNF-α and IL-6 levels, intestinal barrier integrity, and damage-associated molecular patterns (DAMPs). Subsequent multi-omics analyses demonstrated that PJZC exert anti-inflammatory effects may involve reduced PI3K-Akt signaling pathway, gut microbiota composition and metabolic profiles. Finally, molecular docking analysis predicted that the core targets and active ingredients may be closely related to ITGB1, EGFR, and bile acids.

Objective:To investigate the anti-inflammatory mechanisms of Hudi enteric-coated capsule (HDEC) and its major bioactive constituent, polydatin, in ulcerative colitis (UC). Methods:Mouse models of colitis were established by transplantation adoptive transfer of CD45RBhighCD4+ T cells and treated with or without HDEC/polydatin. Therapeutic efficacy was evaluated by assessing disease activity, colon length, and histopathological damage. The differentiation of Th1, Th17, and Treg cells was analyzed using quantitative real-time polymerase chain reaction and flow cytometry. In vitro cultures of mouse and human CD4+ T cells were utilized to assess the immunomodulatory activity. RNA sequencing, Western blotting, and immunofluorescence were used to explore the underlying mechanism. Molecular docking, molecular dynamics, and surface plasmon resonance (SPR) assays were employed to confirm the interaction between polydatin and KEAP1. Results:Treatment with HDEC and polydatin significantly ameliorated murine colitis and mucosal damage. Mechanistically, polydatin directly binds to KEAP1 to promote NFE2L2 nuclear translocation. This NFE2L2 activation reduces intracellular oxidative stress, thereby inhibiting pathogenic Th1/Th17 differentiation and enhancing Treg generation. Importantly, these effects were consistently validated in human CD4+ T cells and UC mucosal tissues. Conclusion:HDEC and polydatin alleviate UC by targeting the KEAP1-NFE2L2 axis to reduce oxidative stress, thereby restoring the Th1/Th17/Treg balance. This highlights polydatin as a promising KEAP1-targeting agent with strong translational potential.

BackgroundAcute pharyngitis and tonsillitis are common in children. Xiao’er Fengreqing Oral Liquid (XFQOL) shows potential benefits in previous studies of pediatric acute upper respiratory tract infections and influenza, but high-quality clinical evidence for pediatric acute pharyngitis/tonsillitis remains limited.MethodsThis multicenter, block-randomized, double-blind, double-dummy, active-controlled trial enrolled 120 children from three tertiary hospitals. Participants were randomly assigned in a 1:1 ratio to receive either XFQOL plus Xiao’er Yanbian Granules (XYG) placebo (XFQOL group, n = 60) or XYG plus XFQOL placebo (control group, n = 60) for 5 days. Primary efficacy endpoints were the day-5 response and disappearance rates of sore throat, assessed using the Wong-Baker Faces Pain Rating Scale (WBS). Secondary endpoints included the time to WBS response and disappearance, the day-3 WBS response rate, and changes in traditional Chinese medicine (TCM) syndrome scores and individual symptoms and signs. Safety was assessed via adverse events and adverse drug reactions.ResultsA total of 120 participants were included in the Full Analysis Set. Baseline characteristics were comparable between the groups. Response rate for the WBS sore throat score after 5 days of treatment was 100% in both groups. The disappearance rate was 98.33% in the experimental group and 90% in the control group, with a hazard ratio and its 95% confidence interval (experimental group/control group) of 0.0833 (97.5% CI: 0.0008–0.1659). Non-inferiority testing demonstrated that the experimental group was not inferior to the control group. In addition, statistically significant intergroup differences were observed in TCM syndrome sore throat onset time and TCM syndrome sore throat disappearance time, no statistically significant differences were detected in any other indicators. No related adverse events were attributed to XFQOL.ConclusionXFQOL is non-inferior to XYG for the 5-day sore throat disappearance rate in pediatric acute pharyngitis/tonsillitis. Secondary endpoint analyses suggest potential advantages in the speed of symptom relief, including a shorter time to sore throat disappearance and a higher 3-day response rate, while the effects on TCM syndrome total scores and most individual symptom/sign scores are comparable between groups. Furthermore, XFQOL demonstrates a favorable safety profile, with no treatment-related adverse events identified, suggesting its potential for clinical application.Clinical Trial Registrationclinicaltrials.gov, ITMCTR2024000569.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide, and its global prevalence has shown a significant upward trend in recent years. Oxidative stress and inflammatory responses are key processes in the initiation and progression of NAFLD. This study aimed to investigate the effects of aerobic exercise combined with apigenin on hepatic oxidative stress and inflammation in mice with NAFLD. To establish the NAFLD model, 52 mice were fed a high-fat diet (HFD) with a fat content of 60%. After eight weeks, HE staining was performed to examine the pathological status of the mouse livers for confirmation of successful NAFLD model establishment. The intervention strategies included aerobic exercise (AE), apigenin (API), and their combination. The amounts of pro-inflammatory substances (tumor necrosis factor-α, TNF-α; interleukin-6, IL-6; interleukin-1β, IL-1β), and parameters of lipid peroxidation (malondialdehyde, MDA) and antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT) in the liver, as well as the levels of gene expression (Kelch-like ECH-associated protein 1, Keap1; nuclear factor erythroid 2-related factor 2, Nrf2; heme oxygenase-1, HO-1; SOD; NAD(P)H quinone dehydrogenase 1, NQO1; TNF-α; IL-6; IL-1β) and protein expression (Keap1; phosphorylated nuclear factor erythroid 2-related factor 2, P-Nrf2; Nrf2; HO-1; SOD; TNF-α; IL-6), were examined using biochemical techniques. Lipid metabolism (triglyceride, TG; total cholesterol, TC; high-density lipoprotein cholesterol, HDL-C; low-density lipoprotein cholesterol, LDL-C) was assessed as well. In mice with NAFLD, aerobic exercise combined with apigenin significantly attenuated oxidative damage and inflammatory responses, ameliorated hepatic steatosis, and reduced hepatic total cholesterol and triglyceride levels, thereby improving overall metabolic homeostasis. Both aerobic exercise and apigenin were able to activate the Keap1/Nrf2/ARE pathway, as evidenced by enhanced phosphorylation of Nrf2, which further promoted the transcription of downstream antioxidant genes and negatively modulated the expression of pro-inflammatory proteins. All treatment groups exhibited significantly downregulated expression of Keap1 and downstream pro-inflammatory factors, with the HFD + API+AE group showing the most pronounced reduction. Similarly, all treatment groups displayed markedly upregulated expression of Nrf2 and downstream antioxidant genes, with the HFD + API+AE group demonstrating the greatest increase. Furthermore, all treatment groups exhibited significantly lower MDA levels compared with the HFD group, and the HFD + API+AE group showed the most prominent reduction. Collectively, apigenin and aerobic exercise exert a favorable synergistic effect against NAFLD.

Ethnopharmacological relevance The Hewei Jiangni Prescription (HP) is a traditional Chinese prescription, which has shown clinical benefits for many years in non-erosive reflux disease (NERD)-associated esophageal hypersensitivity. However, its mechanism of effects remains unclear. Aim of the study To investigate the mechanism of HP in treating NERD-associated esophageal hypersensitivity and identify its pharmacologically active components. Materials and methods NERD-associated esophageal hypersensitivity model mice were treated with HP. Proteomics and protein interaction networks were employed to identify differentially regulated targets. Effects on key targets were validated in cell models and model mice using enzyme-linked immunosorbent assay, western blotting, reverse transcription‒quantitative polymerase chain reaction, and immunofluorescence. Bio-layer interference technology was used to identify HP components binding to targets, followed by liquid chromatography‒mass spectrometry identification and functional validation in cell models. Results HP mitigated esophageal pathology and visceral hypersensitivity in NERD-associated esophageal hypersensitivity model mice. Additionally, HP may attenuate neuroimmune dysregulation in the experimental model by downregulating MrgprX2/B2 overexpression in mast cells and dorsal root ganglion (DRG) neurons, lowering intracellular Ca2+ levels, and decreasing the release of mast cell tryptase and calcitonin gene-related peptide. Vitexin was identified as one of the bioactive components of HP, reproducing its core pharmacological effects in vitro and exhibiting high affinity for the MRGX2 target. Conclusions This study suggests that HP may attenuate neuroimmune dysregulation between mast cells and DRG neurons by regulating MrgprX2/B2, thereby mitigating NERD-associated esophageal hypersensitivity. Additionally, the study findings suggest that vitexin may be one of the bioactive components of HP.

BACKGROUND:Traditional Chinese medicines, as a burgeoning field of medication, significantly alleviate ulcerative colitis (UC) by improving intestinal microbiota-metabolism. Our previous studies demonstrated the significant efficacy of Hudi Enteric-coated capsules (HDEC), Qingchang Wenzhong decoction (QCWZ), and Modified Wumei pill (MWMP) using a mouse model of colitis. However, the mechanism of these therapies through the modulation of microbiota-metabolism remains uncertain. OBJECTIVE:Three multicenter randomized controlled trials were designed to explore the effects of three therapies on the microbiota-metabolism of UC patients with different severity. METHODS:A total of 143 patients with different severities of UC were recruited from 10 hospitals. The clinical efficacy of HDEC for mild UC, QCWZ for moderate UC, and MWMP for severe UC (SUCs) was evaluated by colorectal Mayo scores and systemic inflammatory indicators. The 16S rRNA sequencing and metabolomics were used to analyze intestinal microbiota and metabolite profiles. RESULTS:Three therapies used alone or combined with mesalazine (MS) were comparable to MS alone in improving Mayo scores and hematic inflammatory parameters. Microbial diversities and architectures of SUCs showed the greatest response to MWMP+MS than other medications, as reflected by the enriched Ruminococcus and Anaerostipes together with the reduced Enterococcus, Streptococcus, and Streptococcus anginosus. Furthermore, MWMP+MS boosted the production of the microbiota-derived short-chain fatty acids (SCFAs) of SUCs. These differential microbes and metabolites further displayed significant statistical relationships with clinical parameters. CONCLUSION:Herbal therapies, especially MWMP+MS, effectively improve microbiota composition and SCFA metabolism, which correlates with the improvements of serum inflammatory markers and endoscopic findings in patients.

ETHNOPHARMACOLOGICAL RELEVANCE:Shufeng Jiedu Capsules (SFJDC) is a traditional Chinese patent medicine comprising eight traditional Chinese medicines (TCM). SFJDC is known for its anti-inflammatory and antipyretic effects and is mainly used in clinics to treat upper respiratory tract infections. Currently, studies on the active ingredients of the SFJDC all focus on small-molecule compounds. In contrast, bio-macromolecules, such as the anti-inflammatory activities of polysaccharides in SFJDC, have not been studied, and the composition of the polysaccharides in SFJDC is also unclear. AIM OF THE STUDY:This study aimed to isolate active polysaccharides from Shufeng Jiedu capsules and determine their structural properties and anti-inflammatory activities. MATERIALS AND METHODS:The polysaccharides with different molecular weights were prepared by organic solvent extraction, alcohol precipitation, dialysis, and cross-flow ultrafiltration. The structural characterization of polysaccharides was clarified by high-performance size exclusion chromatography (HPGPC), ion chromatography (IC), and Fourier transform infrared spectroscopy (FT-IR). Enzyme-linked immunosorbent assay (ELISA) and quantitative real-time PCR (qRT-PCR) assay were used to investigate the anti-inflammatory effects of polysaccharides on Lipopolysaccharides (LPS)-stimulated RAW264.7 cells. The in vivo study was employed on the CuSO4-induced and LPS-stimulated zebrafish inflammatory models, and the survival analysis, observation of neutrophil migration, hematoxylin-eosin (H&E) staining, and qRT-PCR assays were used to investigate the in vivo anti-inflammatory effect of polysaccharides. RESULTS:The crude polysaccharides SFJDC-CP were obtained from the mixed aqueous extract of SFJDC with a yield of 38.72 %. Both SFJDC and SFJDC-CP dose-dependently inhibited the secretion of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1β in LPS-stimulated RAW264.7 cells, and SFJDC-CP was more effective at a lower dosage. SFJDC-CP was further separated into three fractions, SFJDC-CP1-CP3, by cross-flow ultrafiltration apparatus with nominal molecular weight cut-offs of 100 kDa, 50 kDa, and 10 kDa membrane cassettes, and the yields were approximately 58.19 %, 10.88 %, and 30.94 %, respectively. The MWs of the SFJDC-CP and its SFJDC-CP1-CP3 were 35.7 kDa, 149.1 kDa, 34.5 kDa, and 15.1 kDa, respectively. The four polysaccharides were composed of rhamnose, arabinose, galactose, glucose, and galacturonic acid in different molar ratios. Non-toxic concentrations of the four polysaccharides ranged from 12.5 to 200 μg/mL. The four polysaccharides significantly reduced the mRNA expression levels and release of IL-1β, IL-6, and TNF-α (P < 0.0001) in LPS-stimulated RAW264.7 cells. Polysaccharides also decreased inflammatory cell infiltration and neutrophil migration (P < 0.05 or P < 0.001) in both CuSO4-induced and LPS-microinjected zebrafish inflammatory models. Additionally, they effectively inhibited the mRNA levels of IL-6 and TNF-α in LPS-infected zebrafish (P < 0.01 or P < 0.001). CONCLUSIONS:Polysaccharides isolated from Shufeng Jiedu capsules have demonstrated anti-inflammatory effects on LPS-stimulated RAW264.7 cells and zebrafish inflammatory models. This study provided preliminary evidence that polysaccharides are one of the main anti-inflammatory ingredients of Shufeng Jiedu capsules. Additionally, it may provide valuable perspectives for investigating polysaccharides in other TCM formulations, particularly those obtained through aqueous extraction methods.

Intestinal microbiota plays a key role in ulcerative colitis (UC), and its composition may vary between the disease's acute (AUC) and chronic (CUC) stages. Using dextran sulfate sodium-induced mice models of AUC and CUC, this study characterized their distinct microbial dysbiosis patterns and evaluated the efficacy of a traditional Chinese medicine formula, Bawei Huangqin Enema (BHE). BHE treatment effectively alleviated colonic inflammation in both models. This was achieved by remodeling the gut microbiota in a stage-specific manner and by enhancing intestinal barrier integrity through the upregulation of tight junction proteins Occludin and MUC2. These findings highlight the distinct microbial features of AUC versus CUC and suggest that BHE's therapeutic potential derives from its dual regulation of the microbiome and intestinal barrier, supporting a multi-target approach for managing UC.

OBJECTIVE:To explore if Hewei Jiangni granule (, HWJNG) could regulate esophageal hypersensitivity via stromal interaction molecule 1 (STIM1)/transient receptor potential vanilloid subfamily member 1 (TRPV1) pathway. METHODS:Qualitative analysis of HWJNG was analysis by high performance of liquid and gas chromatography. In vivo, animal model of non-erosive reflux disease (NERD) was established by fructose intake and restraint stress. HWJNG and Omeprazole were administered by gavage to the drug intervention group. Reflux and visceral hypersensitivity were analyzed by pathological changes, PH value test, mechanical paw withdrawal threshold, thermal withdrawal latency and mast cells (MCs) degranulation. In vitro, substance P (SP)-induced P815 cells and dorsal root ganglion (DRG) cells were co-cultured. Expression in both mice and cells of STIM1, TRPV1, and esophageal visceral hypersensitivity-related gastrointestinal neurochemicals were validated by enzyme linked immunosorbent assays, quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Moreover, overexpression and small interfering RNA against STIM1 were utilized to verify of the role of HWJNG in DRG cells. RESULTS:HWJNG significantly suppressed intercellular space widening, injury of mitochondrial, MCs degranulation, mechanical allodynia and heat neuropathic sensory and increased pH value of esophageal mucosa in NERD mice. HWJNG inhibited expression of visceral hypersensitivity-related gastrointestinal neurochemicals in esophageal mucosa and activated P815 cells, and expression of the STIM1, TRPV1 and related neurotransmitters in DRG and DRG cells. STIM1 siRNA and HWJNG both reduced P815 cells adhesion to DRGs cells and Ca2+ flow into the cytoplasmic space of DRG cells. Furthermore, HWJNG could reversed STIM1 overexpression induced upregulation of TRPV1. CONCLUSION:HWJNG suppressed intercellular space widening in NERD mice, stabilized MCs and restored neuronal hyperexcitability by regulating visceral hypersensitivity viaSTIM1/TRPV1 pathway.

Objective To evaluate the effectiveness and safety of Xiao'er Fengreqing Oral Liquid (XFOL) for pediatric acute pharyngitis/tonsillitis (external wind heat syndrome) through a multi-center, randomized, double-blind, positive-controlled, non-inferiority clinical trial.Method A total of 120 participants (60 per group) will be randomized to receive either XFOL or Xiao'er Yanbian Granules (positive control) for 5 days. The primary outcome is the throat pain resolution rate and overall effective rate at Day 5, assessed via the Wong-Baker Faces Pain Rating Scale (WBS). Secondary outcomes include time to symptom onset/resolution, fever resolution time, and traditional Chinese medicine (TCM) syndrome scores. Safety assessments will monitor adverse events, vital signs, and laboratory parameters. Statistical analyses will follow a pre-specified plan, employing non-inferiority testing, survival analysis for time-to-event endpoints, and generalized estimating equations for repeated measures. Missing data will be handled using the last observation carried forward (LOCF) method for effectiveness endpoints, while safety analyses will rely on observed cases.Conclusion This trial will provide rigorous evidence on the non-inferiority and safety profile of Fengreqing Oral Liquid, supporting its integration into pediatric care for acute upper respiratory infections. Adherence to a predefined statistical analysis plan ensures transparency and minimizes bias, ultimately guiding evidence-based clinical practice for TCM interventions.

This study explored the effect of plant-derived indigo supplementation on intestinal inflammation using in vivo, in vitro, and clinical sample analyses. Our results showed that indigo decreased mucosal inflammation by regulating CD4+ T cell differentiation in a gut microbiota-dependent manner. Microbes transferred from indigo-treated mice, indigo-induced enrichment of Roseburia intestinalis, and its metabolite butyrate played a role in Th17/Treg immunity similar to that of indigo in intestinal inflammation, which was involved in mTORC1/HIF-1α signal-mediated reprogrammed glucose metabolism. We further showed that patients with ulcerative colitis exhibited significant gut dysbiosis and CD4+ T cell differentiation abnormalities. Our findings provide new insights into the gut-immune axis in ulcerative colitis, offering a novel microbial-based immunotherapy for the treatment of inflammatory bowel disease.

Background Non-alcoholic steatohepatitis (NASH), an escalating global health concern, is a primary factor behind cirrhosis, liver transplantation, and hepatocellular carcinoma. Effective treatments remain elusive. Danggui-Shaoyao-San (DGSY), a classic famous prescription employed in treating NASH, could hold promise, although its molecular underpinnings are still under investigation. This study undertakes an exploration of the impacts of DGSY on NASH and seeks to illuminate the mechanisms at play. Methods UHPLC-Q-Orbitrap HRMS was employed to identify compounds within DGSY. Mice underwent a 25-week regimen of HFHC diet and high-sugar water, with 4 weeks of DGSY treatment for efficacy and pathogenic mechanism exploration in vivo. L02 cells were cultured with 0.2 mM FFA for 24 h, exposed to DGSY at 1 mg/ml and 2 mg/ml for efficacy and pathogenic mechanism exploration in vitro. Using online databases, we sought potential targets for NASH treatment, and through PPI networks, identified key targets. Expression levels of genes and proteins were examined by western blotting, RT-PCR, and immunofluorescence staining. Results Thirty-four compounds were identified within DGSY. DGSY brought about marked reductions in biochemical indicators and yielded significant improvements in NASH mice histological features. Additionally, it mitigated hepatic steatosis and inflammation both in vivo and in vitro. The top 10 targets from two network pharmacology analyses, one focusing on structural prediction and the other on literature mining, identified APOE and APP as potential therapeutic targets for DGSY in NASH treatment. PCR validation confirmed that DGSY reduced APP expression after treatment, and further investigation revealed that DGSY significantly suppressed hepatic APP and Aβ expression, indicating its effectiveness in treating NASH. Furthermore, it inhibited Aβ-induced Cathepsin B lysosomal release, reducing hepatic inflammation. Conclusion Danggui-Shaoyao-San has anti-steatohepatitis effects in ameliorating hepatic APP protein expression, reducing hepatic lysosomal CTSB release, and suppressing hepatic NF-κB activation. The study provided a more theoretical basis for the future clinical application of DGSY.

Abstract Background The most common progressive form of non-alcoholic fatty liver disease (NAFLD) is non-alcoholic steatohepatitis (NASH), which is characterized by the development of cirrhosis, and requires liver transplantation. We screened for the differentially expressed necroptosis-related genes in NASH in this study, and analyzed immune infiltration through microarray and bioinformatics analysis to identify potential biomarkers, and explore the molecular mechanisms involved in NASH. Methods The GSE24807 microarray dataset of NASH patients and healthy controls was downloaded, and we identified the differentially expressed genes (DEGs). Necroptosis-related differential genes (NRDEGs) were extracted from these DEGs, and functionally annotated by enrichment analyses. The core genes were obtained by constructing gene co-expression networks using weighted gene co-expression network analysis (WGCNA). Finally, the transcription factor (TF) regulatory network and the mRNA-miRNA network were constructed, and the infiltrating immune cell populations were analyzed with CIBERSORT. Results We identified six necroptosis-related genes (CASP1, GLUL, PYCARD, IL33, SHARPIN, and IRF9), and they are potential diagnostic biomarkers for NASH. In particular, PYCARD is a potential biomarker for NAFLD progression. Analyses of immune infiltration showed that M2 macrophages, γδ T cells, and T follicular helper cells were associated with the immune microenvironment of NASH, which is possibly regulated by CASP1, IL33, and IRF9. Conclusions We identified six necroptosis-related genes in NASH, which are also potential diagnostic biomarkers. Our study provides new insights into the molecular mechanisms and immune microenvironment of NASH.

目前心身疾病治疗思路多从情志病人手,较为局限.中医学广郁证理论对心身疾病辨治具有重要的临床指导价值.明末医家易思兰所著《易氏医按》体现了其辨治广郁证的学术特色,即以气血为纲,强调审脉求因,参合五运六气、天时之郁,重视顾护脾胃,以"畅"立法,创制多首新方,对后世颇有启发.总结并传承易氏辨治广郁证的学术思想,对拓宽当今心身疾病临床诊疗思路具有借鉴作用.

Damage to the intestinal mucosal barrier play an important role in the pathogenesis of ulcerative colitis (UC). Discovering the key regulators and repairing the disturbed barrier are crucial for preventing and treating UC. Traditional Chinese medicine (TCM) has been proved to be effective on treating UC and has exhibited its role in repairing the intestinal mucosal barrier. We summarized the evidence of TCM against UC by protecting and repairing the physical barrier, chemical barrier, immune barrier, and biological barrier. Mechanisms of increasing intestinal epithelial cells, tight junction proteins, and mucins, promoting intestinal stem cell proliferation, restoring the abundance of the intestinal microbiota, and modulating the innate and adaptive immunity in gut, were all involved in. Some upstream proteins and signaling pathways have been elucidated. Based on the existing problems, we suggested future studies paying attention to patients’ samples and animal models of UC and TCM syndromes, conducting rescue experiments, exploring more upstream regulators, and adopting new technical methods. We hope this review can provide a theoretical basis and novel ideas for clarifying the mechanisms of TCM against UC via repairing the intestinal mucosal barrier.

目的 研究清肠温中方对葡聚糖硫酸钠(DSS)诱导的溃疡性结肠炎(UC)小鼠NLRP6/IL-18/MUC2轴及肠道黏液屏障的影响及作用机制.方法 24只SPF级雄性C57BL/6小鼠随机分为空白组、模型组、清肠温中方组和美沙拉嗪组,每组6只.空白组自由饮用去离子水,模型组、清肠温中方组和美沙拉嗪组自由饮用3%DSS溶液制备UC模型.造模7 d后,清肠温中方组、美沙拉嗪组予相应药液灌胃,空白组、模型组予去离子水灌胃,连续7 d.每日称量小鼠体质量,记录小鼠疾病活动指数(DAI)评分,HE染色观察结肠组织病理变化,AB-PAS染色观察结肠组织杯状细胞情况,Western blot检测结肠组织核苷酸结合寡聚化结构域样受体蛋白6(NLRP6)、白细胞介素(IL)-18蛋白表达,免疫荧光染色检测结肠组织黏蛋白2(MUC2)蛋白表达.结果 与空白组比较,模型组小鼠体质量下降,DAI评分显著升高(P＜0.01),结肠长度显著缩短(P＜0.01),结肠组织黏膜屏障破坏,杯状细胞数量减少,黏液分泌减少;结肠组织NLRP6、MUC2蛋白表达显著降低(P＜0.01),IL-18蛋白表达显著升高(P＜0.01).与模型组比较,清肠温中方组小鼠体质量增加,DAI评分显著降低(P＜0.01),结肠长度显著增加(P＜0.05);结肠黏膜损伤减轻,炎性细胞浸润减少,杯状细胞数量增多;结肠组织NLRP6、MUC2蛋白表达显著升高(P＜0.01),IL-18蛋白表达显著降低(P＜0.01).结论 清肠温中方能明显缓解UC小鼠疾病活动度,降低DAI评分,修复结肠组织病理损伤,增加杯状细胞数量,促进黏液分泌,其作用机制可能与调节NLRP6/IL-18/MUC2轴,修复肠道黏液屏障有关.

Purpose Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by mucosal inflammation, which may develop into ulcerative colitis-associated carcinogenesis (UCAC) with disease progression. Qingchang Wenzhong Decoction (QCWZD) is a classic and effective prescription for the clinical treatment of UC. QCWZD has been shown to alleviate intestinal mucosal injury in acute and chronic UC models. This study aimed to explore and then verify the pharmacological mechanisms of QCWZD in UC and UCAC therapy. Methods In this study, approaches including microarray analysis, network pharmacology, and biological verification are employed to clarify the mechanism of QCWZD in the treatment of UC and UCAC. TCMSP, Swiss Target Prediction, and Similarity Ensemble Approach were used to investigate the active ingredients and targets of QCWZD. UC and UCAC valid targets were identified by the microarray data in the GEO database (GSE38713 and GSE47908). The core targets were obtained by PPI network and enriched by GO and KEGG. DSS and AOM/DSS mouse models were adopted to verify the above analysis results. Results The enrichment analysis showed that the therapeutic targets of QCWZD enriched in blood circulation, cell adhesion molecules, and pathways of inflammation and cancer such as IL-17 signaling pathway and toll-like receptor signaling pathway were involved in the multiple synergies of QCWZD on UC and UCAC treatment. The results of experiments demonstrated that QCWZD can exert its effects on protecting the intestinal mucosal barrier, regulating inflammation and improving intestinal fibrosis in UC and UCAC and the main mechanism of QCWZD in treatment of UC and UCAC may be related to the activation of the IL-17, NF-κB and TLR4 signaling pathways. Conclusion Our results indicated that QCWZD treated UC and UCAC via multiple targets and pathways and the IL-17, NF-κB and TLR4 signaling pathways may be highly involved in this process.

ETHNOPHARMACOLOGICAL RELEVANCE:Ulcerative colitis (UC) is an idiopathic, chronic inflammatory disorder of the colonic mucosa, accompanied with abdominal pain, and bloody diarrhea. Currently, clinical treatment options for UC are limited. Qingchang Wenzhong Decoction (QCWZD) is an effective prescription of traditional Chinese medicine for the treatment of UC. However, the mechanism of QCWZD in alleviating intestinal barrier dysfunction in UC has not been clearly explained. AIM OF THE STUDY:To determine the mechanism whereby QCWZD promotes the recovery of intestinal barrier dysfunction in UC. MATERIALS AND METHODS:A secondary analysis of colonic mucosa from UC patients acquired from a prior RCT clinical trial was performed. The effects of QCWZD on intestinal mucus and mechanical barriers in UC patients were evaluated using colon tissue paraffin-embedded sections from UC patients. The mechanism was further investigated by in vivo and in vitro experiments. UC mice were established in sterile water with 3.0% dextran sodium sulfate (DSS). Meanwhile, mice in the treatment group were dosed with QCWZD or mesalazine. In vitro, an intestinal barrier model was constructed using Caco-2 and HT29 cells in co-culture. GC-C plasmid was used to overexpress/knock down GC-C to clarify the target of QCWZD. HE, AB-PAS, ELISA, immunohistochemistry and immunofluorescence assays were used to assess the level of colonic inflammation and intestinal barrier integrity. Rt-qPCR, Western Blot were used to detect the expression of genes and proteins related to GC-C signaling pathway. Molecular docking was used to simulate the binding sites of major components of QCWZD to GC-C. RESULTS:In UC patients, QCWZD increased mucus secretion, goblet cell number, and promoted MUC2 and ZO-1 expression. QCWZD accelerated the recovery of UC mice from DSS-induced inflammation, including weight gain, reduced disease activity index (DAI) scores, colon length recovery, and histological healing. QCWZD promoted mucus secretion and increased ZO-1 expression in in vivo and in vitro experiments, thereby repairing mucus mechanical barrier damage. The effects of QCWZD are mediated through regulation of the GC-C signaling pathway, which in turn affects CFTR phosphorylation and MUC2 expression to promote mucus secretion, while inhibiting the over-activation of MLCK and repairing tight junctions to maintain the integrity of the mechanical barrier. Molecular docking results demonstrate the binding of the main components of QCWZD to GC-C. CONCLUSION:Our study demonstrated that QCWZD modulates the GC-C signaling pathway to promote remission of mucus-mechanical barrier damage in the UC. The clarification of the mechanism of QCWZD holds promise for the development of new therapies for UC.

Many studies have reported obvious seasonal differences in the intestinal flora of rats, and this stable distribution of the seasonal flora helps in maintaining the normal physiological function of the host. However, the mechanism underlying these seasonal differences in intestinal flora remains unclear. To explore the correlation among seasonal factors and intestinal water metabolism and intestinal flora, 20 Sprague Dawley (SD) rats were divided into spring, summer, autumn, and winter groups. The environment for the four seasons was simulated using the Balanced Temperature and Humidity Control system. The intestinal water metabolism was evaluated by determining the intestinal transmission function, fecal water content, water content of colonic tissue, and the colonic expression levels of AQP3, AQP4, and AQP8. The composition and relative abundance of intestinal microflora in rats in each season were assessed through 16S rDNA amplifier sequencing, and the relationship between the dominant flora and intestinal water metabolism in each season was analyzed using Spearman correlation analysis. The high temperature and humidity season could lead to an increase in intestinal water metabolism and intestinal water content in rats, whereas the low temperature and humidity season could lead to a decrease, which was closely related to the change in microflora. To explore the molecular mechanism of seasonal changes in intestinal water metabolism, the concentration of colonic 5-HT, VIP, cAMP, and PKA associated with intestinal water metabolism in rats were also examined. Seasonal changes could affect the concentration of colonic 5-HT and VIP in rats, and then regulate AQPs through cAMP/PKA pathway to affect the intestinal water metabolism. These results suggest that seasonal factors affect the level of intestinal water metabolism in rats and result in seasonal differences in intestinal flora.

Objective: To describe the status of outcome indicators selected in randomized controlled trial（RCT） in the treatment of ulcerative colitis（UC） with Traditional Chinese Medicine（TCM）, analyze the existing problems, and provide the basis for the construction of UC clinical efficacy evaluation criteria based on the core outcome set of Traditional Chinese Medicine（COS-TCM）. Methods: Retrieved 2017-2022 published in CNKI Database, WanFang Database, VIP Database, SinoMed, Embase, Cochrane library, Web of Science and PubMed literature on randomized controlled trials of TCM treatment of UC. Two authors independently screened the literature and extracted the data in strict accordance with the inclusion and exclusion criteria. Any differences were resolved through discussion. Results: The study included 73 RCT, including 6 729 patients, using 79 outcome indicators. It can be divided into 8 broad categories: clinical effect（eight kinds, 101 times）, TCM syndrome effect（two kinds, 48 times）, intestinal mucosa and histopathological effect（10 kinds, 59 times）, symptoms and signs（one kind, nine times）, physical and chemical detection（49 kinds, 10 kinds, 213 times）, quality of life（seven kinds, 30 times）, long-term prognosis（one kind, seven times）, safety event（one kind, 36 times）. The outcome indexes with high frequency of use include: clinical efficacy/total effective rate, TCM syndrome score, incidence of adverse reactions, Mayo score/DAI score, Baron score, quality of life in inflammatory bowel disease（IBDQ） score, etc. Conclusion: There are many problems in the outcome indicators of clinical studies on TCM treatment of UC, such as non-standard efficacy indicators of TCM syndromes, inaccurate expression of clinical efficacy indicators, inconsistent reference standards, unclear distinction between primary outcome indicators and secondary outcome indicators, inadequate safety event reporting, and missing outcome indicators of economic evaluation. It is necessary to establish a unified UC TCM efficacy evaluation standard based on COS-TCM, so as to further improve the quality of TCM clinical research.