Uncovering the mechanisms of Zhubi decoction against rheumatoid arthritis through an integrated study of network pharmacology, metabolomics, and intestinal flora.

ETHNOPHARMACOLOGICAL RELEVANCE: Zhubi Decoction (ZBD) is a modified formulation derived from the classic traditional Chinese medicine prescription "Er-Xian Decoction" documented in the esteemed "Clinical Manual of Chinese Medical Prescription". While the utilization of ZBD has exhibited promising clinical outcomes in treating rheumatoid arthritis (RA), the precise bioactive chemical constituents and the underlying mechanisms involved in its therapeutic efficacy remain to be comprehensively determined. AIM OF THE STUDY: This study aims to systematically examine ZBD's pharmacological effects and molecular mechanisms for RA alleviation. MATERIALS AND METHODS: Utilizing the collagen-induced arthritis (CIA) rat model, we comprehensively evaluated the anti-rheumatoid arthritis effects of ZBD in vivo through various indices, such as paw edema, arthritis index, ankle diameter, inflammatory cytokine levels, pathological conditions, and micro-CT analysis. The UPLC-MS/MS technique was utilized to analyze the compounds of ZBD. The potential therapeutic targets and signaling pathways of ZBD in the management of RA were predicted using network pharmacology. To analyze comprehensive metabolic profiles and identify underlying metabolic pathways, we conducted a serum-based widely targeted metabolomics analysis utilizing LC-MS technology. Key targets and predicted pathways were further validated using immunofluorescent staining, which integrated findings from serum metabolomics and network pharmacology analysis. Additionally, we analyzed the gut microbiota composition in rats employing 16 S rDNA sequencing and investigated the effects of ZBD on the microbiota of CIA rats through bioinformatics and statistical methods. RESULTS: ZBD exhibited remarkable efficacy in alleviating RA symptoms in CIA rats without notable side effects. This included reduced paw redness and swelling, minimized joint damage, improved the histopathology of cartilage and synovium, mitigated the inflammatory state, and lowered serum concentrations of cytokines TNF-α, IL-1ß and IL-6. Notably, the effectiveness of ZBD was comparable to MTX. Network pharmacology analysis revealed inflammation and immunity-related signaling pathways, such as PI3K/AKT, MAPK, IL-17, and TNF signaling pathways, as vital mediators in the effectual mechanisms of ZBD. Immunofluorescence analysis validated ZBD's ability to inhibit PI3K/AKT pathway proteins. Serum metabolomics studies revealed that ZBD modulates 170 differential metabolites, partially restored disrupted metabolic profiles in CIA rats. With a notable impact on amino acids and their metabolites, and lipids and lipid-like molecules. Integrated analysis of metabolomics and network pharmacology identified 6 pivotal metabolite pathways and 3 crucial targets: PTGS2, GSTP1, and ALDH2. Additionally, 16 S rDNA sequencing illuminated that ZBD mitigated gut microbiota dysbiosis in the CIA group, highlighting key genera such as Ligilactobacillus, Prevotella_9, unclassified_Bacilli, and unclassified_rumen_bacterium_JW32. Correlation analysis disclosed a significant link between 47 distinct metabolites and specific bacterial species. CONCLUSION: ZBD is a safe and efficacious TCM formulation, demonstrates efficacy in treating RA through its multi-component, multi-target, and multi-pathway mechanisms. The regulation of inflammation and immunity-related signaling pathways constitutes a crucial mechanism of ZBD's efficacy. Furthermore, ZBD modulates host metabolism and intestinal flora. The integrated analysis presents experimental evidence of ZBD for the management of RA.

Probiotics as a targeted intervention in anti-aging: A review.

The age-induced disruption of gut flora, termed gut dysbiosis, is intimately tied to compromised immune function, augmented oxidative stress, and a spectrum of age-linked disorders. This review examines the fundamental mechanisms employed by probiotic strains to modulate gut microbiota composition and metabolic profiles, mitigate cognitive decline via the gut-brain axis, modulate gene transcription, and alleviate inflammatory responses and oxidative stress. We elucidate the capacity of probiotics as a precision intervention to restore gut microbiome homeostasis and alleviate age-related conditions, thereby offering a theoretical framework for probiotics to decelerate aging, manage age-related diseases, and elevate quality of life.

The gut microbiome in patients with Cushing's disease affects depression- and anxiety-like behavior in mice.

BACKGROUND: Depression and anxiety significantly impact the quality of life in individuals with Cushing's disease (CD), which originates from pituitary neuroendocrine tumors (PitNETs), yet our understanding of the underlying mechanisms is limited. There is substantial evidence linking gut microbes to depression, anxiety, and endocrinology. RESULTS: The gut bacterial phenotype of patients with Cushing's disease was significantly different from that of the control group, and when the mice were treated with fecal bacteria from these patients, both anxiety- and depression-like behavior were significantly increased. However, this effect can be alleviated by supplementing with 2-(14, 15-epoxyeicosatrienoyl) glycerol (2-14,15-EG) which was found at reduced levels in the peripheral blood of mice treated with coprofecal bacteria from Cushing's disease. In this process, the effects of hormone levels and immune factors were not significant. In addition, in an animal model, corticosterone has been observed to affect behavioral changes in mice through gut microbiota composition, clarifying the cause-and-effect relationship between hormones, microbiota, and behavior. Finally, there was no significant difference in gut microbiome composition and its effects on mouse behavior in patients with Cushing's disease with different levels of depression and anxiety. CONCLUSIONS: In summary, this research enhances our current understanding of how gut microbes in patients with Cushing's disease contribute to depression and anxiety, offering novel insights for clinical treatment approaches. Video Abstract.

Intestinal flora and inflammatory bowel disease: Causal relationships and predictive models.

Background: Inflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, is significantly influenced by intestinal flora. Understanding the genetic and microbiotic interplay is crucial for IBD prediction and treatment. Methods: We used Mendelian randomization (MR), transcriptomic analysis, and machine learning techniques, integrating data from the MiBioGen Consortium and various GWAS datasets. SNPs associated with intestinal flora were mapped to genes, with LASSO regression refining gene selection. Differentially expressed genes (DEGs) and immune infiltration patterns were identified through transcriptomic analysis. Six machine learning models were used for predictive modeling. Findings: MR analysis identified 25 gut microbiota classifications causally related to IBD. SNP mapping and gene expression analysis highlighted 24 significant genes. Drug target MR and colocalization validated these genes' causal relationships with IBD. Key pathways identified included the PI3K-Akt signaling pathway and epithelial-mesenchymal transition. Immune infiltration analysis revealed distinct patterns between high and low LASSO score groups. Machine learning models demonstrated high predictive value, with soft voting enhancing reliability. Interpretation: By integrating MR, transcriptomic analysis, and sophisticated machine learning approaches, this study elucidates the causal relationships between intestinal flora and IBD. The application of machine learning not only enhanced predictive modeling but also offered new insights into IBD pathogenesis, highlighted potential therapeutic targets, and established a robust framework for predicting IBD onset.

Rosa roxburghii fermented juice mitigates LPS-induced acute lung injury by modulation of intestinal flora and metabolites.

Acute lung injury (ALI) is a severe pulmonary condition with high mortality and morbidity, lacking effective pharmacotherapeutic options. Rosa roxburghii Tratt, a unique fruit from southwestern China, is valued for its rich nutritional content and functional properties. Fermentation is known to enhance the nutritional value, flavor, and shelf life of foods. In this study, we investigated the effects of fermented Rosa roxburghii juice (RRFJ) on gut microbiota, metabolites, and the levels of short-chain fatty acids in the intestines, as well as its impact on lung tissue and intestine tissue injury, inflammation, and oxidative stress in murine models. The results showed that RRFJ modulated gut microbiota and metabolites, increased short-chain fatty acid levels, and consequently reduced lung tissue injury, inflammation, and oxidative stress in mice with ALI. These findings suggest that RRFJ has the potential to serve as a functional dietary adjunct in the management of acute lung injury, providing a scientific basis for its therapeutic role.

Study on the Antidepressant Effect of Zhizichi Decoction by Regulating Metabolism and Intestinal Flora.

BACKGROUND: The incidence of depression is increasing year by year, and Zhizichi Decoction（ZZCD）has shown significant efficiency in the clinical treatment of mild depression, but its mechanism of action is still unclear. In this research, network pharmacology and metagenomics combined and metabolomics were used as research methods to explain the scientific connotation of the antidepressant effect of ZZCD from the aspects of the overall effect of organisms and microbial structure and function. METHODS: The rat model of depression was established by chronic unpredictable mild stress (CUMS), and the improvement of depressive symptoms was evaluated by behavioral and histopathological methods. Network pharmacology predicted possible targets and important pathways of ZZCD. Metabolomics revealed its possible related biological pathways. Metagenomics showed the disturbance of ZZCD on intestinal microbial diversity structure and associated biological functions in depressed rats. RESULTS: ZZCD can improve the behavioral performance of CUMS rats, and can significantly regulate the content of 5-HT, NE and other neurotransmitters in serum and brain tissue, and improve the damaged state of neurons in the hippocampus. Network pharmacology predicts that it mainly acts on biological processes such as inflammatory response and oxidative stress response. Metabolomics found that it affected metabolic pathways such as amino acid metabolism and lipid metabolism. The results of metagenomics showed that it significantly regulated the abundance of Firmicutes and Bacteroidetes. The above results predicted that it may affect signaling pathways such as the nervous system, inflammatory diseases and cell processing. CONCLUSION: ZZCD may play an antidepressant role by regulating intestinal probiotics, energy metabolism, and inflammation reduction. This provides a scientific basis for the clinical application of ZZCD in traditional Chinese medicine and also makes it an optional alternative for the treatment of depression.

Corrigendum: Study on the effects of intestinal flora on gouty arthritis.

[This corrects the article DOI: 10.3389/fcimb.2024.1341953.].

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OBJECTIVES: To observe the effect of electroacupuncture (EA) on the expressions of high mobility group protein 1(HMGB1) and myeloid differentiation factor 88 (MyD88) in the small intestine and intestinal flora of obese rats, so as to explore the potential mechanism of EA to improve obesity in rats. METHODS: After 1 week of acclimatization, 10 rats were randomly selected from 50 Wistar male rats as the normal group, and the rest rats were fed with high-fat diet for 8 weeks to establish the obese model. The successfully modeling rats were randomly divided into model group, EA group and sham EA group, with 10 rats in each group. Rats in the EA group were given EA (2 Hz, 1 mA) at "Zhongwan"(CV12), "Guanyuan"(CV4), "Zusanli" (ST36)and "Fenglong"(ST40). Rats in the sham EA group were given shallow stabs at acupoints of the EA group about 5 mm outwardly and the electrodes were clamped without being energized. Both groups were intervened for 10 min each time, 3 times (Monday, Wednesday and Friday) a week for 8 weeks. The body weights of the rats were measured before and after 8 weeks of intervention, respectively. The contents of serum lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α were measured by ELISA, and the protein and mRNA expressions of HMGB1 and MyD88 in the small intestine were detected by Western blot and real-time quantitative PCR, respectively. 16S rRNA sequencing was performed to determine the relative abundance and diversity of the bacterial flora in the fresh feces of rats. RESULTS: Compared with the normal group, the body weight, serum LPS and TNF-α contents, small intestinal HMGB1 and MyD88 protein and mRNA expression levels of rats in the model group were significantly increased (P<0.01), while the relative abundance of Lactobacillus, Muri and Bifidobacterium was decreased (p<0.01), Collinsella, Prevotella and Ruminococcus was increased (P<0.01). Compared with model group, the body weight, serum LPS and TNF-α contents, protein and mRNA expression levels of HMGB1 and MyD88 in both EA and sham EA groups were decreased (P<0.01, P<0.05), while the relative abundance of Lactobacillus, Muri and Bifidobacterium was increased (P<0.01) and Collinsella, Prevotella and Ruminococcus decreased (P<0.01). Comparison between EA group and sham EA group showed that, the contents of LPS and TNF-α in serum of rats in sham EA group were increased (P<0.01, P<0.05), the relative abundance of Lactobacillus, Muri and Bifidobacterium was lower (P<0.05, P<0.01), and Collinsella, Prevotella and Ruminococcus was higher (P<0.01). CONCLUSIONS: EA can reduce the body weight of obese rats, which may be related to the regulation of the structure of intestinal flora and the reduction of inflammatory reactions in the small intestine.

Safety and efficacy of a feed additive consisting of Saccharomyces cerevisiae NCYC R618 (Benesacc®) for chickens for fattening and other poultry species for fattening and reared for laying (Global Nutritech Biyoteknoloji Ltd. Sti).

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of Saccharomyces cerevisiae NCYC R618 as a zootechnical feed additive (functional group: gut flora stabiliser) for chickens for fattening and other poultry species for fattening and reared for laying. The additive has never been authorised. The strain under assessment S. cerevisiae (NCYC R618) qualifies for the qualified presumption of safety (QPS) and, consequently, the FEEDAP Panel concluded that the product is safe for the target species, the consumers and the environment. The FEEDAP Panel could not conclude on the skin and eye irritation or skin sensitisation potential of the additive. The additive should be considered a respiratory sensitiser. Due to the lack of sufficient data, the Panel cannot conclude on the efficacy of the additive under the proposed conditions of use.

Angelica sinensis Polysaccharide Alleviates Staphylococcus aureus-Induced Mastitis by Regulating The Intestinal Flora and Gut Metabolites.

The modulation of intestinal flora by various polysaccharides has been shown to mitigate disease progression. Recent research reveals a significant link between intestinal flora and the progression of mastitis. This study demonstrates that the oral administration of Angelica sinensis polysaccharide (ASP) reduces mammary inflammation and blood-milk barrier (BMB) damage induced by Staphylococcus aureus in mice, primarily through the modulation of intestinal flora. The beneficial effects of ASP were negated when antibiotics disrupted the gut microbiota in mice. Furthermore, fecal microbiota transplantation (FMT) from ASP-treated mice to recipients markedly alleviated symptoms of S. aureus-induced mastitis. Oral ASP not only enhances gut microbial diversity but also shifts its composition, increasing the abundance of Lachnospiraceae_NK4A136 while reducing Erysipelatoclostridium. Metabolomic analysis revealed that ASP alters intestinal metabolic pathways, elevating levels of metabolites, such as tabersonine and riboflavin. Notably, tabersonine was found to ameliorate S. aureus-induced mastitis. These results suggest that targeting intestinal flora and metabolism through polysaccharides could serve as a promising strategy for mastitis intervention and potentially for other infectious diseases, as well.

In Situ Expression of Yak IL-22 in Mammary Glands as a Treatment for Bovine Staphylococcus aureus-Induced Mastitis in Mice.

Since the development of dairy farming, bovine mastitis has been a problem plaguing the whole industry, which has led to a decrease in milk production, a reduction in dairy product quality, and an increase in costs. The use of antibiotics to treat mastitis can cause a series of problems, which can bring a series of harm to the animal itself, such as the development of bacterial resistance and dramatic changes in the gut flora. However, the in vivo and in vitro antibacterial activity of yak Interleukin-22 (IL-22) and its application in mastitis caused by Staphylococcus aureus have not been reported. In this study, the mammary gland-specific expression plasmid pLF-IL22 of the yak IL-22 gene was constructed and expressed in MAC-T cells and mammary tissue of postpartum female mice. The coding region of the IL-22 gene in yaks is 573 bp, which can encode 190 amino acids, and the homology difference in the IL-22 gene in yaks is less than 30%, which indicates certain conservation. IL-22 is a hydrophilic protein with a total positive charge of four, the presence of a signal peptide, and the absence of a transmembrane domain. Sufficient expression of IL-22 effectively inhibited the high expression of inflammatory factors caused by Staphylococcus aureus, reduced the symptoms of mammary gland histopathology, and alleviated mastitis. Under the action of IL-22, the intestinal flora of mastitis mice also changed, the abundance of intestinal Bacilli, Prevotellaceae, and Alloprevotella in mice increased after treatment, and the pathogenic bacteria decreased. These findings provide new insights into the potential application of the yak IL-22 gene in the treatment of bovine mastitis in the future.

Regulation of colon injury and improvement of exercise performance in exhausted running mice by Lactobacillus pentosus CQZC02.

In this study, strenuous forced exercise caused intestinal damage and reduced the exercise capacity of mice. However, the antioxidant and anti-inflammatory properties of Lactobacillus pentosus CQZC02 (LPCQZC02) were found to improve both the intestinal barrier and exercise function in mice. The effectiveness of LPCQZC02 was confirmed through various methods, including kit detection, pathological observation, quantitative reverse transcription polymerase chain reaction (qRT-PCR), and intestinal flora analysis. The findings demonstrated that LPCQZC02 could control colonic index, lessen colonic enlargement caused by intense exercise, and extend the running duration of mice. Serum levels of total superoxide dismutase (T-SOD), glutathione (GSH), and interleukin-10 (IL-10) were elevated, whereas those of malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α) were reduced. The findings of the mRNA expression analysis revealed that in the colons of mice who remarkably exercised, LPCQZC02 could increase the expression levels of zonula occludens-1 (ZO-1), occludin-1, and claudin-1 genes. Additionally, in skeletal muscle tissue, it could downregulate TNF-α expression level and upregulate copper/zinc superoxide dismutase (Cu/Zn-SOD) and manganese superoxide dismutase (Mn-SOD) expression levels. Furthermore, LPCQZC02 could both reduce and promote beneficial bacteria in the intestines of mice undergoing intense exercise. In conclusion, LPCQZC02 emerged as a functional probiotic and demonstrated a notable advantage over sulfasalazine, a medication for intestinal conditions, in mitigating oxidative inflammation, repairing intestinal barrier damage, and enhancing motor function in mice subjected to strenuous exercise.

Intestinal flora and bile acid interactions impact the progression of diabetic kidney disease.

In recent years, with the rapid development of omics technologies, researchers have shown that interactions between the intestinal flora and bile acids are closely related to the progression of diabetic kidney disease (DKD). By regulating bile acid metabolism and receptor expression, the intestinal flora affects host metabolism, impacts the immune system, and exacerbates kidney injury in DKD patients. To explore interactions among the gut flora, bile acids and DKD, as well as the related mechanisms, in depth, in this paper, we review the existing literature on correlations among the gut flora, bile acids and DKD. This review also summarizes the efficacy of bile acids and their receptors as well as traditional Chinese medicines in the treatment of DKD and highlights the unique advantages of bile acid receptors in DKD treatment. This paper is expected to reveal a new and important potential strategy for the clinical treatment of DKD.

Regulation effect of the intestinal flora and intervention strategies targeting the intestinal flora in alleviation of pulmonary fibrosis development.

Pulmonary fibrosis is an end-stage respiratory disease characterized by fibroblast proliferation and accumulation of extracellular matrix and collagen, which is accompanied by inflammatory damage. The disease is mainly based on pulmonary dysfunction and respiratory failure, the incidence of it is increasing year by year, and the current treatment methods for it are limited. In recent years, it has been found that gut microbes play a crucial role in the pathogenesis and development of pulmonary fibrosis. The microecological disturbance caused by changes in the composition of the intestinal flora can affect the course of pulmonary fibrosis. The regulatory network or information exchange system for gut-lung crosstalk is called the "gut-lung axis". This review focuses on the frontier research on entero-pulmonary regulation in pulmonary fibrosis and on intervention strategies for changing the gut microbiota to improve pulmonary fibrosis, including fecal microbiota transplantation, traditional Chinese medicine interventions, and supplementation with probiotics. In addition, the present problems in this field are also raised in order to provide strong theoretical and strategic support for the future exploration of regulatory mechanisms and therapeutic drug development. This paper reviews the interaction of the intestinal flora with pulmonary fibrosis, introduces the research progress for improving pulmonary fibrosis through interventions targeted at the intestinal flora, and provides new ideas for the treatment of pulmonary fibrosis.

Effect of Morchella esculenta polysaccharides on the rectal microbiota of mice challenged with lipopolysaccharides.

Introduction: Intestinal dysfunction poses a severe problem by preventing the digestion and absorption of nutrients. The gut, being the most vital organ for these processes, plays a crucial role in ensuring our body receives the nutrients it needs. We explored the mitigating effect of Morchella esculenta polysaccharides (MEP) on intestinal injury induced by lipopolysaccharides (LPS) through the modulation of intestinal flora. Methods: For this purpose, Kunming mice (KM) were divided into three groups, namely, PC, PM, and PY. Group PY was treated with MEP, while groups PM and PY were induced with LPS. Results: The results showed that weight loss in the PM group was significantly greater than that in the PY group (P < 0.05), and the organ indexes of the lung and spleen in the PM group were significantly higher than those in the PC (P < 0.01) and PY (P < 0.05) groups. LPS caused severe injuries in KM mice in the PM group, characterized by broken villi. However, MEP treatment could alleviate this damage in the PY group, resulting in relatively intact villi. The serum analysis showed that tumor necrosis factor alpha (TNF-ɑ) (P < 0.01), interleukin 6 (IL-6) (P < 0.01), and 3,4-methylenedioxyamphetamine (MDA) (P < 0.05) levels were significantly higher in the PM group, while IL-10 (P < 0.001), superoxide dismutase (SOD) (P < 0.01) and glutathione peroxidase (GSH-Px) (P < 0.01) were significantly lower in that group. Interestingly, supplementation with MEP could lower the levels of TNF-ɑ, IL-10, IL-6, MDA while increasing the levels of superoxide dismutase (SOD) (P < 0.01) and GSH-Px. The gut microbiota analysis yielded 630,323 raw reads and 554,062 clean reads, identifying 3,390 amplicon sequencing variants (ASVs). One phylum and five genera were notably different among animal groups, including Escherichia_Shigella, Limosilactobacillus, unclassified_Geminicoccaceae, unclassified_Rhodobacteraceae, and Parabacteroides (P. distasonis). Discussion: In conclusion, we found that MEP could mitigate the intestinal damage caused by LPS by modulating the inflammatory response, oxidative resistance, and intestinal flora of KM mice. Our results may provide insights into novel treatment options for intestine-related diseases.

Ulcerative colitis: molecular insights and intervention therapy.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by abdominal pain, diarrhea, rectal bleeding, and weight loss. The pathogenesis and treatment of UC remain key areas of research interest. Various factors, including genetic predisposition, immune dysregulation, and alterations in the gut microbiota, are believed to contribute to the pathogenesis of UC. Current treatments for UC include 5-aminosalicylic acids, corticosteroids, immunosuppressants, and biologics. However, study reported that the one-year clinical remission rate is only around 40%. It is necessary to prompt the exploration of new treatment modalities. Biologic therapies, such as anti-TNF-α monoclonal antibody and JAK inhibitor, primarily consist of small molecules targeting specific pathways, effectively inducing and maintaining remission. Given the significant role of the gut microbiota, research into intestinal microecologics, such as probiotics and prebiotics, and fecal microbiota transplantation (FMT) shows promising potential in UC treatment. Additionally, medicinal herbs, such as chili pepper and turmeric, used in complementary therapy have shown promising results in UC management. This article reviews recent findings on the mechanisms of UC, including genetic susceptibility, immune cell dynamics and cytokine regulation, and gut microbiota alterations. It also discusses current applications of biologic therapy, herbal therapy, microecologics, and FMT, along with their prospects and challenges.

Alleviating effect of Lactobacillus rhamnosus SDSP202418 on exercise-induced fatigue in mice.

In this study, the effects of Lactobacillus rhamnosus SDSP202418 isolated from shrimp paste on the exercise performance of fatigued mice were analyzed, and the potential action mechanism was revealed. L. rhamnosus SDSP202418 significantly improved the exhaustion time of the mice and regulated the biochemical indices (lactate dehydrogenase, nitrogen, and uric acid) of the fatigued mice to resist fatigue. L. rhamnosus SDSP202418 also upregulated the mRNA expression of slow muscle fibers and downregulated the mRNA expression of fast muscle fibers in the exercise mice by activating the AMPK/PGC-1α pathway in the fatigued mice. It also increased the contents of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH)) in the liver and muscle. These enzymes removed and repaired oxidative free radicals to achieve antifatigue. In addition, L. rhamnosus SDSP202418 can change the gut microbial structure and modulate the abundance and balance of fatigue-related gut microbiota, which in turn exerts antifatigue effects. L. rhamnosus SDSP202418 is a functional food component that relieves fatigue after exercise.

Regulation of hippocampal miRNA expression by intestinal flora in anxiety-like mice.

This study investigated the possible interaction between gut flora and miRNAs and the effect of both on anxiety disorders. The model group was induced with chronic restraint stress (CRS) and each group was tested for anxiety-like behaviour by open field test and elevated plus maze test. Meanwhile, the gut flora was analysed by 16S rRNA high-throughput sequencing. The miRNAs in hippocampus were analysed by high-throughput sequencing, and the key miRNAs were obtained by using the method of bioinformatics analysis. PCR was used to verify the significantly related key miRNAs. Spearman correlation analysis was used to explore the correlation between behaviour, key miRNAs and differential gut microbiota. The 16S rRNA high-throughput sequencing result showed that the gut flora was dysregulated in the model group. In particular, Verrucomicrobia, Akkermansia, Anaerostipes, Ralstonia, Burkholderia and Anaeroplasma were correlated with behaviour. The results of miRNA high-throughput sequencing analysis and bioinformatics analysis showed that 7 key miRNAs influenced the pathogenesis of anxiety, and qRT-PCR results were consistent with the high-throughput sequencing results. Mmu-miR-543-3p and mmu-miR-26a-5p were positively correlated with Verrucomicrobia, Akkermansia and Anaerostipes. Therefore, we infer that chronic stress caused the decrease of Akkermansia abundance, which may aggravate the decrease of mmu-miR-543-3p and mmu-miR-26a-5p expression, leading to the increase of SLC1A2 expression. In conclusion, gut flora has played an important influence on anxiety with changes in miRNAs.

Ruhao Dashi granules exert therapeutic effects on H1N1 influenza virus infection by altering intestinal microflora composition.

Objective: Antiviral medications for influenza could be ineffective due to the emergence of resistant influenza virus strains. Ruhao Dashi (RHDS) granules possess anti-inflammatory and antibacterial effects. The present study aimed to determine the efficacy of RHDS granules in treating influenza-infected mice and the mechanism underlying this treatment as well as its effect on the intestinal flora composition of the infected mice. Methods: The HPLC-UV method was used to identify the active components of RHDS granules. ICR mice were infected with influenza A virus (IAV) H1N1 subtype through a nasal drip. After the influenza mice model was successfully established, the pathological changes in the lungs were observed for 5 days after gavage treatment with 0.9% sterile saline and low, medium, and high doses (0.07, 0.14, and 0.28 g/mL, respectively) of RHDS granules. The serum levels of the cytokines IL-6 and TNF-α and sIgA were detected by ELISA. Real-time fluorescence quantitative PCR and western blotting assay were performed to determine the expression levels of the tight junction (TJ) proteins claudin-1, occludin, and zonula occludens-1 (ZO-1) in colon tissues. Furthermore, 16S rRNA gene sequencing of feces samples was conducted to assess the effect of RHDS granules on the gut microbiota. Results: RHDS granules exerted a protective effect on the lung tissues of IAV-infected mice; moreover, the granules reduced the synthesis of proinflammatory cytokines and increased the relative expression levels of claudin-1, occludin, and ZO-1 in colon tissues. Furthermore, RHDS granule treatment increased the relative abundance of Lactobacillus, Akkermansia, and Faecalibaculum and decreased the relative abundance of Muribaculaceae; thus, RHDS granules could stabilize the intestinal microbiota to some extent. Conclusion: RHDS granules exert a therapeutic effect on IAV-infected mice probably by modifying the structural composition of their intestinal microbiota.