Wutou decoction alleviates arthritis inflammation in CIA mice by regulating Treg cell stability and Treg/Th17 balance via the JAK2/STAT3 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Wutou Decoction (WTD) is a classic traditional Chinese medicine formula, which has shown clinical efficacy in treating rheumatoid arthritis (RA). The Treg stability and Th17/Treg imbalance is an important immunological mechanism in RA progression. Whether WTD regulates CD4+ T cell subsets has not been thoroughly investigated yet. AIM OF THE STUDY: This study aimed to explore the potential role and mechanisms of WTD in regulating the diminished stability of Treg cells and the imbalance of CD4+ T cell subsets via in vivo and in vitro experiments. MATERIALS AND METHODS: Firstly, the therapeutic effects of WTD on the collagen-induced arthritis (CIA) mouse and its potential regulatory function on CD4+ T cell subsets were evaluated in vivo. Animal specimens were collected after 31 days of treatment with WTD. The anti-arthritic and anti-inflammatory effects of WTD were assessed through arthritis scoring, body weight, spleen index, serum IL-6 levels, and micro-PET/CT imaging. Gene enrichment analysis was performed to evaluate the activation T cell-related signaling pathway. Flow cytometry was used to determine the proportions of CD4+ T cell subsets in vitro and in vitro. Additionally, ELISA was used to assess the secretion of IL-10 and TGF-ß by Treg cells under inflammatory conditions. The suppressive function of Treg cells on cell proliferation under inflammatory conditions was examined using CFSE labeling. Immunofluorescence staining was performed to detect the phosphorylation levels of STAT3 in CD4+ T cells from mouse spleen tissues. Western blotting was used to evaluate the phosphorylation levels of JAK2/STAT3 in Treg cells. RESULTS: WTD significantly alleviated joint inflammation in CIA mice. WTD reduced serum IL-6 levels in CIA mice, improved their body weight and spleen index. WTD treatment inhibited the activation of CD4+ T cell subgroup-related signaling in the joint tissues of CIA mice. In vitro and in vitro experiments showed that WTD increased the proportion of Treg cells and decreased the proportion of Th17 cells in CIA mice spleen. Furthermore, WTD promoted the secretion of IL-10 and TGF-ß by Treg cells and enhanced the inhibitory capacity of Treg cells on cell proliferation under inflammatory conditions. Immunofluorescence detected decreased STAT3 phosphorylation levels in CD4+ T cells from CIA mice spleen, while western blotting revealed a decrease in JAK2/STAT3 phosphorylation levels in Treg cells in vitro. CONCLUSIONS: Inhibiting JAK2/STAT3 phosphorylation is a potential mechanism through which WTD improves Treg cell stability, balances CD4+ T cell subsets, and attenuates RA joint inflammation.

Stachyose Exerts Anticolitis Efficacy by Re-balancing Treg/Th17 and Activating the Butyrate-Derived PPARγ Signaling Pathway.

Ulcerative colitis (UC) is a complex chronic inflammatory disease closely associated with gut homeostasis dysfunction. The previous studies have shown that stachyose, a functional food additive, has the potential to enhance gut health and alleviate UC symptoms. However, the underlying mechanism of its effects remains unknown. In this study, our findings showed that dietary supplements of stachyose had a significant dose-dependent protective effect on colitis symptoms, regulation of gut microbiota, and restoration of the Treg/Th17 cell balance in dextran sulfate sodium (DSS) induced colitis mice. To further validate these findings, we conducted fecal microbiota transplantation (FMT) to treat DSS-induced colitis in mice. The results showed that microbiota from stachyose-treated mice exhibited a superior therapeutic effect against colitis and effectively regulated the Treg/Th17 cell balance in comparison to the control group. Moreover, both stachyose supplementation and FMT resulted in an increase in butyrate production and the activation of PPARγ. However, this effect was partially attenuated by PPARγ antagonist GW9662. These results suggested that stachyose alleviates UC symptoms by modulating gut microbiota and activating PPARγ. In conclusion, our work offers new insights into the benefical effects of stachyose on UC and its potential role in modulating gut microbiota.

BMSC-derived exosomes regulate the Treg/Th17 balance through the miR-21-5p/TLR4/MyD88/NF-κB pathway to alleviate dry eye symptoms in mice.

Bone marrow mesenchymal stem cell (BMSC)-derived exosomes (BMSC-Exos) have a variety of biological functions and are extensively involved in the regulation of inflammatory diseases, as well as tissue repair and regeneration. However, the mechanism of action of these compounds in dry eye disease (DED) in mice is still unclear. This study demonstrated that the Treg/Th17 ratio was strongly imbalanced in DED clinical samples. BMSC-Exos can modulate the Treg/Th17 balance, improve the integrity of the corneal epithelial layer, and ameliorate DED progression in mice. Mechanistically, BMSC-Exos dramatically decreased the levels of IL-17 and IL-22; increased the levels of IL-4, IL-10, and TGF-ß1; and increased tear secretion and the number of goblet cells in the conjunctiva in mice, thus alleviating the progression of DED. This effect is achieved by BMSC-Exos through the delivery of miR-21-5p to target and restrain TLR4, thereby restraining the MyD88/NF-κB pathway. Our study showed that the upregulation of miR-21-5p in BMSC-Exos may be a therapeutic target for DED. These findings support new ideas and a basis for treating DED, as well as for further study of the application value of exosomes in alleviating DED.

Regulatory role of PI16 in autoimmune arthritis and intestinal inflammation: implications for Treg cell differentiation and function.

BACKGROUND: Regulatory T cells (Tregs) are crucial in maintaining immune homeostasis and preventing autoimmunity and inflammation. A proportion of Treg cells can lose Foxp3 expression and become unstable under inflammation conditions. The precise mechanisms underlying this phenomenon remain unclear. METHODS: The PI16 gene knockout mice (PI16fl/flFoxp3Cre) in Treg were constructed, and the genotypes were identified. The proportion and phenotypic differences of immune cells in 8-week-old mice were detected by cell counter and flow cytometry. Two groups of mouse Naïve CD4+T cells were induced to differentiate into iTreg cells to observe the effect of PI16 on the differentiation and proliferation of iTreg cells, CD4+CD25+Treg and CD4+CD25- effector T cells (Teff) were selected and co-cultured with antigen presenting cells (APC) to observe the effect of PI16 on the inhibitory ability of Treg cells in vitro. The effects of directed knockout of PI16 in Treg cells on inflammatory symptoms, histopathological changes and immune cell expression in mice with enteritis and autoimmune arthritis were observed by constructing the model of antigen-induced arthritis (AIA) and colitis induced by dextran sulfate sodium salt (DSS). RESULTS: We identified peptidase inhibitor 16 (PI16) as a negative regulator of Treg cells. Our findings demonstrate that conditional knock-out of PI16 in Tregs significantly enhances their differentiation and suppressive functions. The conditional knockout of the PI16 gene resulted in a significantly higher abundance of Foxp3 expression (35.12 ± 5.71% vs. 20.00 ± 1.61%, p = 0.034) in iTreg cells induced in vitro compared to wild-type mice. Mice with Treg cell-specific PI16 ablation are protected from autoimmune arthritis (AIA) and dextran sulfate sodium (DSS)-induced colitis development. The AIA model of PI16CKO is characterized by the reduction of joint structure and the attenuation of synovial inflammation and in DSS-induced colitis model, conditional knockout of the PI16 reduce intestinal structural damage. Additionally, we found that the deletion of the PI16 gene in Treg can increase the proportion of Treg (1.46 ± 0.14% vs. 0.64 ± 0.07%, p < 0.0001) and decrease the proportion of Th17 (1.00 ± 0.12% vs. 3.84 ± 0.64%, p = 0.001). This change will enhance the shift of Th17/Treg toward Treg cells in AIA arthritis model (0.71 ± 0.06% vs. 8.07 ± 1.98%, p = 0.003). In DSS-induced colitis model of PI16CKO, the proportion of Treg in spleen was significantly increased (1.40 ± 0.15% vs. 0.50 ± 0.11%, p = 0.003), Th17 (2.18 ± 0.55% vs. 6.42 ± 1.47%, p = 0.017), Th1 (3.42 ± 0.19% vs. 6.59 ± 1.28%, p = 0.028) and Th2 (1.52 ± 0.27% vs. 2.76 ± 0.38%, p = 0.018) in spleen was significantly decreased and the Th17/Treg balance swift toward Treg cells (1.44 ± 0.50% vs. 24.09 ± 7.18%, p = 0.012). CONCLUSION: PI16 plays an essential role in inhibiting Treg cell differentiation and function. Conditional knock out PI16 gene in Treg can promote the Treg/Th17 balance towards Treg dominance, thereby alleviating the condition. Targeting PI16 may facilitate Treg cell-based therapies for preventing autoimmune diseases and inflammatory diseases. The research provides us with novel insights and future research avenues for the treatment of autoimmune diseases, particularly arthritis and colitis.

Differential Immune Response Patterns Induced by Anionic and Cationic Lipid Adjuvants in Intranasal Anti-Influenza Immunization.

Currently, vaccine development against different respiratory diseases is at its peak. It is of utmost importance to find suitajble adjuvants that can increase the potency of the vaccine candidates. This study aimed to determine the systemic and splenic immune mechanisms in mice models induced by anionic and cationic lipid adjuvants in the presence of the vaccine-candidate influenza antigen hemagglutinin (HA). In the presence of the HA antigen, the cationic adjuvant (N3) increased conventional dendritic cell 1 (cDC1) abundance with enhanced MHCI and CD80-CD86 costimulatory marker expression, and significantly higher CD8T and Th17 populations with enhanced interferon-gamma (IFNγ) expression in CD8T and CD4T populations. Conversely, the anionic adjuvant (L3) increased the cDC2 population percentage with significantly higher MHCII and DEC205 expression, along with an increase in the CD4T and regulatory T cell populations. The L3-treated group also exhibited higher percentages of activated B and plasma cell populations with significantly higher antigen-specific IgG and IgA titer and virus neutralization potential. While the anionic adjuvant induced significantly higher humoral responses than the cationic adjuvant, the latter influenced a significantly higher Th1/Th17 response. For customized vaccine development, it is beneficial to have alternative adjuvants that can generate differential immune responses with the same vaccine candidate antigen. This study will aid the selection of adjuvants based on their charges to improve specific immune response arms in the future development of vaccine formulation.

Schisandra chinensis Bee Pollen Ameliorates Colitis in Mice by Modulating Gut Microbiota and Regulating Treg/Th17 Balance.

Colitis is a chronic disease associated with alterations in the composition of gut microbiota. Schisandra chinensis bee pollen extract (SCPE) has been proved to be rich in phenolic compounds and effective in modulating gut microbiota, but its effect on colitis and the underlying mechanism remains unclear. This study investigates the relationship between colitis amelioration and the gut microbiota regulation of SCPE via fecal microbial transplantation (FMT). The results showed that administration of 20.4 g/kg BW of SCPE could primely ameliorate colitis induced by dextran sulfate sodium (DSS) in mice, showing as more integration of colon tissue structure and the colonic epithelial barrier, as well as lower oxidative stress and inflammation levels compared with colitis mice. Moreover, SCPE supplement restored the balance of T regulatory (Treg) cells and T helper 17 (Th17) cells. Gut microbiota analysis showed SCPE treatment could reshape the gut microbiota balance and improve the abundance of gut microbiota, especially the beneficial bacteria (Akkermansia and Lactobacillus) related to the production of short-chain fatty acids and the regulation of immunity. Most importantly, the protection of 20.4 g/kg BW of SCPE on colitis can be perfectly transmitted by fecal microbiota. Therefore, the gut microbiota-SCFAS-Treg/Th17 axis can be the main mechanism for SCPE to ameliorate colitis. This study suggests that SCPE can be a new promising functional food for prevention and treatment of colitis by reshaping gut microbiota and regulating gut immunity.

Bergapten ameliorates combined allergic rhinitis and asthma syndrome after PM2.5 exposure by balancing Treg/Th17 expression and suppressing STAT3 and MAPK activation in a mouse model.

Combined allergic rhinitis and asthma syndrome (CARAS) causes chronic respiratory inflammation in allergic individuals. Long-term exposure to particulate matter 2.5 (PM2.5; particles 2.5 µm or less in diameter) can aggravate respiratory damage. Bergapten (5-methoxysporalen) is a furocoumarin mostly found in bergamot essential oil and has significant antioxidant, anticancer, and anti-inflammatory activity. This study created a model in which CARAS was exacerbated by PM2.5 exposure, in BALB/c mice and explored the potential of bergapten as a therapeutic agent. The bergapten medication increased ovalbumin (OVA)-specific immunoglobulin (Ig) G2a level in serum and decreased OVA-specific IgE and IgG1 expression. Clinical nasal symptoms diminished significantly, with weakened inflammatory reaction in both the nasal mucosa and lungs. Furthermore, bergapten controlled the T helper (Th)1 to Th2 ratio by increasing cytokines associated with Th1-like interleukin (IL)-12 and interferon gamma and decreasing the Th2 cytokines IL-4, IL-5, and IL-13. Factors closely related to the balance between regulatory T cells and Th17 (such as IL-10, IL-17, Forkhead box protein P3, and retinoic-related orphan receptor gamma) were also regulated. Notably, pro-inflammatory cytokines IL-6, IL-1ß, and tumor necrosis factor-alpha were reduced by bergapten, which suppressed the activation of both the signal transducer and activator of transcription 3 signaling pathway and the mitogen-activated protein kinase signaling pathway. Therefore, bergapten might have potential as a therapeutic agent for CARAS.

Paris polyphylla extract attenuates colitis in mice by regulating PPAR-γ mediated Treg/Th17 balance.

ETHNOPHARMACOLOGICAL RELEVANCE: Paris polyphylla Sm. (P.P), is a widely-used traditional Chinese medicine (TCM) in the treatment of wound, throat sores and snakebites. Furthermore, P.P was recorded as an anti-inflammatory drug by the Chinese Pharmacopoeia. AIM OF THE STUDY: We sought to decipher the anti-inflammatory effect of P.P on ulcerative colitis (UC); specifically, to explore whether P.P attenuates colitis by restoring the regulatory T cells (Tregs) and T helper 17 (Th17) cells balance and its mechanism. MATERIAL AND METHODS: We treated experimental colitis mice with extracts of Paris polyphylla (EPP). The percentage of Tregs and Th17 cells were measured using flow cytometry, and their secreted cytokines levels were evaluated employing ELISA. The expression of peroxisome proliferator-activated receptor gamma (PPAR-γ) in colon tissues was detected using immunofluorescence. Furthermore, GW9662, a PPAR-γ antagonist, was used to validate the mechanism of EPP in restoring the Treg/Th17 balance. RESULTS: The EPP effectively alleviated the clinical symptoms and inflammatory cytokine levels in mice with colitis. EPP treatment also restored the impaired Treg/Th17 balance in mice. Furthermore, EPP treatment promoted PPAR-γ expression and reduced HIF-1α and p-STAT3 expression in colon tissues, whereas PPAR-γ inhibition blocked the effects of EPP in mice models. CONCLUSION: Our study indicates that EPP exhibit excellent anti-inflammatory properties via restoring PPAR-γ/STAT3/HIF-1α axis-mediated Treg/Th17 balance in colitis mice. Hence, P. polyphylla is a promising medicinal plant-based alternative for managing colitis that requires further clinical validation.

Fecal microbiota transplantation and replenishment of short-chain fatty acids protect against chronic cerebral hypoperfusion-induced colonic dysfunction by regulating gut microbiota, differentiation of Th17 cells, and mitochondrial energy metabolism.

BACKGROUND: Little is known about the association between gut microbiota and intestinal injury under a state of chronic cerebral hypoperfusion (CCH). Here, the effects of gut microbiota and short-chain fatty acids (SCFAs), as important metabolic products, on intestinal function and potential mechanisms after CCH were investigated. METHODS: Rats were subjected to bilateral common carotid artery occlusion (BCCAo) to induce CCH. The gut microbiota and metabolites of SCFAs were assessed by 16S rRNA sequencing and targeted metabolomics, respectively. Transcriptomic analysis of colon tissues was also conducted. Subsequently, potential molecular pathways and differentially expressed genes were verified by western blot, immunoprecipitation, and immunofluorescence analyses. Furthermore, the integrity of the colonic barrier was evaluated by hematoxylin and eosin and mucin 2 staining and expression levels of tight junction proteins. Besides, colonic inflammation was further assessed by flow cytometry and expression levels of inflammatory cytokines. In addition, colonic mitochondrial dysfunction was analyzed via membrane potential, reactive oxygen species, electron transport chain (ETC) activities, adenosine triphosphate content, and mitochondrial ultrastructure. RESULTS: CCH modified gut microbial composition and microbial metabolism of SCFAs, which may be associated with inhibition of mitochondrial ETC activities and oxidative phosphorylation, leading to dysregulation of mitochondrial energy metabolism. Furthermore, CCH induced differentiation of pathogenic Th17 cells, promoted the formation of complexes of interferon regulatory factor 4 and signal transducer and activator of transcription 3 (STAT3), and increased the phosphorylation of STAT3. This was associated with an impairment of colonic barrier function and chronic colonic inflammation. In contrast, FMT and SCFA replenishment ameliorated CCH-induced gut microbial dysbiosis by increasing the intestinal content of Ruminococcus_sp_N15_MGS_57 and modulating microbial metabolism of SCFAs by increasing acetic acid contents associated with an improvment of the balance between Tregs and Th17 cells, mitochondrial ETC activities, and oxidative phosphorylation to prevent colonic inflammation and dysregulation of mitochondrial energy metabolism. CONCLUSION: These findings indicate that FMT and SCFA replenishment present a promising therapeutic strategy against colonic dysfunction under a state of chronic cerebral ischemia.

Research Advances in the Treatment of Allergic Rhinitis by Probiotics.

Allergic rhinitis (AR) impairs the quality of life of patients and reduces the efficiency of social work, it is an increasingly serious public medical and economic problem in the world. Conventional anti-allergic drugs for the treatment of allergic rhinitis (AR) can cause certain side effects, which limit the quality of life of patients. Therefore, it makes sense to look for other forms of treatment. Several studies in recent years have shown that probiotics have shown anti-allergic effects in various mouse and human studies. For example, the application of certain probiotic strains can effectively relieve the typical nasal and ocular symptoms of allergic rhinitis in children and adults, thereby improving the quality of life and work efficiency. At the same time, previous studies in humans and mice have found that probiotics can produce multiple effects, such as reduction of Th2 cell inflammatory factors and/or increase of Th1 cell inflammatory factors, changes in allergy-related immunoglobulins and cell migration, regulate Th1/Th2 balance or restore intestinal microbiota disturbance. For patients with limited activity or allergic rhinitis with more attacks and longer attack duration, oral probiotics have positive effects. The efficacy of probiotics in the prevention and treatment of allergic rhinitis is remarkable, but its specific mechanism needs further study. This review summarizes the research progress of probiotics in the treatment of allergic rhinitis in recent years.

Protective effect of Schistosoma japonicum eggs on TNBS-induced colitis is associated with regulating Treg/Th17 balance and reprogramming glycolipid metabolism in mice.

Inflammatory bowel diseases (IBDs) have been classified as modern refractory diseases. However, safe, well-tolerated, and effective treatments for IBDs are still lacking. Therefore, there is an urgent need to develop novel therapeutic targets with fewer undesirable adverse reactions. A growing body of research has shown that infection with live helminths or exposure to defined helminth-derived components can downregulate pathogenic inflammation due to their immunoregulatory ability. Here we were to explore the protective role of Schistosoma japonicum eggs on murine experimental colitis caused by trinitrobenzene sulfonic acid (TNBS) and the underlying mechanism. Frequencies of splenic Treg and Th17 cells were detected by flow cytometry. Protein and mRNA expressions of Foxp3 and RORγt were investigated by Western Blot and quantitative real-time polymerase chain reaction (qPCR), respectively. Concentrations of transforming growth factor-beta1 (TGF-ß1), interleukin-10 (IL-10) and IL-17A were assessed with ELISA. Expression levels of genes related to glycolipid metabolism were measured with qPCR. The results showed that pre-exposure to S. japonicum eggs contributed to the relief of colitis in the TNBS model, evidenced by improved body weight loss, reversing spleen enlargement and colon shortening, and decreased histology scores. Compared with the TNBS group, the TNBS+Egg group had increased Treg immune response, accompanied by decreased Th17 immune response, leading to the reconstruction of Treg/Th17 balance. In addition, a ratio of Treg/Th17 was correlated negatively with the histological scores in the experiment groups. Furthermore, the regulation of Treg/Th17 balance by S. japonicum eggs was associated with inhibiting the glycolysis pathway and lipogenesis, along with promoting fatty acid oxidation in the TNBS+Egg group. These data indicate that S. japonicum eggs have a protective effect against TNBS-induced colitis, which is related to restoring Treg/Th17 balance and regulating glucose and lipid metabolism.

Gut Microbiota Protected Against pseudomonas aeruginosa Pneumonia via Restoring Treg/Th17 Balance and Metabolism.

Backgrounds and Purpose: The theory of "entero-pulmonary axis" proves that pneumonia leads to gut microbiota disturbance and Treg/Th17 immune imbalance. This study is aimed to explore the potential mechanism of fecal microbiota transplantation (FMT) in the treatment of Pseudomonas aeruginosa pneumonia, in order to provide new insights into the treatment of pneumonia. Methods: Pseudomonas aeruginosa and C57/BL6 mice were used to construct the acute pneumonia mouse model, and FMT was treated. Histopathological changes in lung and spleen were observed by HE staining. The expression of CD25, Foxp3 and IL-17 was observed by immunofluorescence. The proportion of Treg and Th17 cells was analyzed by flow cytometry. Serum IL-6, LPS, and IFN-γ levels were detected by ELISA. The expression of TNF-α, IFN-γ, IL-6, IL-2, Foxp3, IL-17, IL-10, and TGFß1 in lung tissue homogenate was detected by qRT-PCR. 16S rRNA sequencing and non-targeted metabolomics were used to analyze gut microbiota and metabolism. Results: Pseudomonas aeruginosa caused the decrease of body weight, food and water intake, lung tissue, and spleen injury in mice with pneumonia. Meanwhile, it caused lung tissue and serum inflammation, and Treg/Th17 cell imbalance in mice with pneumonia. Pseudomonas aeruginosa reduced the diversity and number of gut microbiota in pneumonia mice, resulting in metabolic disorders, superpathway of quinolone and alkylquinolone biosynthesis. It also led to the decrease of 2-heptyl-3-hydroxy-4(1H)-quinolone biosynthesis, and the enrichment of Amino sugar and nucleotide sugar metabolism. FMT with or without antibiotic intervention restored gut microbiota abundance and diversity, suppressed inflammation and tissue damage, and promoted an immunological balance of Treg/Th17 cells in mice with pneumonia. In addition, FMT inhibited the aerobactin biosynthesis, 4-hydroxyphenylacetate degradation, superpathway of lipopolysaccharide biosynthesis and L-arabinose degradation IV function of microbiota, and improved amino sugar and nucleotide sugar metabolism. Conclusions: FMT restored the Treg/Th17 cells' balance and improved inflammation and lung injury in mice with Pseudomonas aeruginosa pneumonia by regulating gut microbiota disturbance and metabolic disorder.

Losartan Attenuates Atherosclerosis in Uremic Mice by Regulating Treg/Th17 Balance via Mediating PTEN/PI3K/Akt Pathway.

INTRODUCTION: Uremia could accelerate atherosclerosis (AS) formation involving Treg/Th17 imbalance. Losartan regulates the imbalance between regulatory T cells (Treg cells) and T helper 17 cells (Th17 cells). However, their interactions in uremia accelerated AS (UAAS) remained poorly understood. METHODS: UAAS mice model was established, and after losartan and VO-OHpic (VO, phosphatase and tensin homolog [PTEN] inhibitor) injection, biological indexes, and inflammatory cytokines (transforming growth factor-ß1, TGF-ß1; interleukin-10 [IL-10]; IL-17 and IL-6) levels were determined using enzyme-linked immunosorbent assay. Pathological changes on aorta were observed using hematoxylin-eosin staining. Percentages of Treg cells (CD4+CD25+Foxp3+) and Th17 cells (CD4+IL-17+) in total CD4+ T cells were determined using flow cytometry. PTEN expressions were measured using Western blot, quantitative real-time polymerase chain reaction, and immunohistochemistry staining as needed. RESULTS: After UAAS mice model construction, biological indexes (urea, cholesterol, and triglycerides) levels were increased, and aortic atherosclerotic plaque was formed. In UAAS mice, in total CD4+ T cells, Treg cells percentage was decreased yet Th17 cells percentage was increased, and TGF-ß1 and IL-10 levels were downregulated yet IL-17 and IL-6 levels were upregulated. An opposite effect was found after losartan treatment. PTEN was downregulated in UAAS mice, and suppressing PTEN reversed the alleviating effects of losartan in UAAS mice. CONCLUSION: Losartan attenuated UAAS in mice by regulating Treg/Th17 cells balance via mediating PTEN/PI3K/Akt pathway, providing possible therapeutic method for UAAS in clinical practice.

Paeonol Attenuated Vascular Fibrosis Through Regulating Treg/Th17 Balance in a Gut Microbiota-Dependent Manner.

Background: Paeonol (Pae) is a natural phenolic compound isolated from Cortex Moutan, which exhibits anti-atherosclerosis (AS) effects. Our previous work demonstrated that gut microbiota plays an important role during AS treatment as it affects the efficacy of Pae. However, the mechanism of Pae in protecting against vascular fibrosis as related to gut microbiota has yet to be elucidated. Objective: To investigate the antifibrosis effect of Pae on AS mice and demonstrate the underlying gut microbiota-dependent mechanism. Methods: ApoE-/- mice were fed with high-fat diet (HFD) to replicate the AS model. H&E and Masson staining were used to observe the plaque formation and collagen deposition. Short-chain fatty acid (SCFA) production was analyzed through LC-MS/MS. The frequency of immune cells in spleen was phenotyped by flow cytometry. The mRNA expression of aortic inflammatory cytokines was detected by qRT-PCR. The protein expression of LOX and fibrosis-related indicators were examined by western blot. Results: Pae restricted the development of AS and collagen deposition. Notably, the antifibrosis effect of Pae was achieved by regulating the gut microbiota. LC-MS/MS data indicated that the level of SCFAs was increased in caecum contents. Additionally, Pae administration selectively upregulated the frequency of regulatory T (Treg) cells as well as downregulated the ratio of T helper type 17 (Th17) cells in the spleen of AS mice, improving the Treg/Th17 balance. In addition, as expected, Pae intervention can significantly downregulate the levels of proinflammatory cytokines IL-1ß, IL-6, TNF-α, and IL-17 in the aorta, and upregulate the levels of anti-inflammatory factor IL-10, a marker of Treg cells. Finally, Pae's intervention in the gut microbiota resulted in the restoration of the balance of Treg/Th17, which indirectly downregulated the protein expression level of LOX and fibrosis-related indicators (MMP-2/9 and collagen I/III). Conclusion: Pae attenuated vascular fibrosis in a gut microbiota-dependent manner. The underlying protective mechanism was associated with the improved Treg/Th17 balance in spleen mediated through the increased microbiota-derived SCFA production. Collectively, our results demonstrated the role of Pae as a potential gut microbiota modulator to prevent and treat AS.

Modified Gegen Qinlian Decoction Regulates Treg/Th17 Balance to Ameliorate DSS-Induced Acute Experimental Colitis in Mice by Altering the Gut Microbiota.

Inflammatory bowel disease (IBD) is characterized by chronic pathology associated with extensive intestinal microbial dysregulation and intestinal inflammation. Thus, efforts are underway to manipulate the gut microbiome to improve inflammatory pathology. Gegen Qinlian decoction (GQD), a traditional Chinese medicine prescription, has been widely utilized for treating diarrhea and ulcerative colitis (UC) for thousands of years. However, the underlying mechanism of its efficacy and whether its protective effect against colitis is mediated by the gut microbiota are poorly understood. In the present study, our data demonstrated that modified GQD (MGQD) administration significantly improved the pathological phenotypes and colonic inflammation challenged by DSS in mice, which were specifically manifested as reduced loss of body weight, shortening of colon length, DAI score, histological score and suppressed inflammatory response. 16S rRNA sequencing and targeted metabonomics analysis showed that MQGD altered the diversity and community landscape of the intestinal microbiota and the metabolic profiles. In particular, MQGD significantly boosted the abundance of the intestinal microbiota producing short-chain fatty acids (SCFAs), which are causally associated with promoting the development of Treg cells and suppressing the differentiation of pro-inflammatory Th17 cells. More importantly, transferring fecal microbiota from MGQD-treated or healthy controls exhibited equivalent alleviative effects on colitis mice. However, this protective effect could not be replicated in experiments of mice with depleted intestinal microbes through broad-spectrum antibiotic cocktails (ABX), further supporting the importance of SCFA-producing gut microbiota in the beneficial role of MGQD. In general, MGQD therapy has the potential to remodel the intestinal microbiome and reestablish immune homeostasis to ameliorate DSS-induced colitis.

Stigmasterol Restores the Balance of Treg/Th17 Cells by Activating the Butyrate-PPARγ Axis in Colitis.

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder with gut microbiota disequilibrium and regulatory T (Treg)/T helper 17 (Th17) immune imbalance. Stigmasterol, a plant-derived sterol, has shown anti-inflammatory effects. Our study aimed to identify the effects of stigmasterol on experimental colitis and the related mechanisms. Stigmasterol treatment restored the Treg/Th17 balance and altered the gut microbiota in a dextran sodium sulfate (DSS)-induced colitis model. Transplantation of the faecal microbiota of stigmasterol-treated mice significantly alleviated inflammation. Additionally, stigmasterol treatment enhanced the production of gut microbiota-derived short-chain fatty acids (SCFAs), particularly butyrate. Next, human naïve CD4+ T cells sorted from IBD patients were cultured under Treg- or Th17-polarizing conditions; butyrate supplementation increased the differentiation of Tregs and decreased Th17 cell differentiation. Mechanistically, butyrate activated peroxisome proliferator-activated receptor gamma (PPARγ) and reprogrammed energy metabolism, thereby promoting Treg differentiation and inhibiting Th17 differentiation. Our results demonstrate that butyrate-mediated PPARγ activation restores the balance of Treg/Th17 cells, and this may be a possible mechanism, by which stigmasterol attenuates IBD.

An engineered probiotic secreting Sj16 ameliorates colitis via Ruminococcaceae/butyrate/retinoic acid axis.

Most inflammatory bowel disease (IBD) patients are unable to maintain a lifelong remission. Developing a novel therapeutic strategy is urgently needed. In this study, we adopt a new strategy to attenuate colitis using the Escherichia coli Nissle 1917 probiotic strain to express a schistosome immunoregulatory protein (Sj16) in the gastrointestinal tract. The genetically engineered Nissle 1917 (EcN-Sj16) highly expressed Sj16 in the gastrointestinal tracts of dextran sulfate sodium-induced colitis mice and significantly attenuated the clinical activity of colitis mice. Mechanistically, EcN-Sj16 increased the intestinal microbiota diversity and selectively promoted the growth of Ruminococcaceae and therefore enhanced the butyrate production. Butyrate induced the expression of retinoic acid, which further attenuated the clinical activity of colitis mice by increasing Treg cells and decreasing Th17. Strikingly, retinoic acid inhibitor inhibited the therapeutic effects of EcN-Sj16 in colitis mice. These findings suggest that EcN-Sj16 represents a novel engineered probiotic that may be used to treat IBD.

Allicin regulates Treg/Th17 balance in mice with collagen-induced arthritis by increasing the expression of MEKK2 protein.

To study the role of Allicin in regulating Treg/Th17 ratio in splenic lymphocyte by increasing the expression of MEKK2 protein in MAPK signaling pathway, and to explore the mechanism of immune response in mice with collagen-induced arthritis (CIA). Mouse CIA model was induced by chicken collagen type II, and experimental mice were randomly divided into NC group, Model group, and Allicin group. HE staining was used to compare the degree of joint pathological damage in mice of each group, and Masson staining to observe the proliferation of collagen tissue in each group. Flow cytometry detected Treg/Th17 ratio in splenic lymphocytes. Furthermore, RT-PCR and WB were used to detect the mRNA and protein expression of related transcription factors and inflammatory factors Foxp3, ROR-γt, and IL-17A, as well as MEK2 protein expression in splenic lymphocytes. The results showed that Allicin treatment could reduce the severity of arthritis and the proliferation of collagen fibers on the surface of cartilage and bone joints in CIA mice. Compared with NC group, Treg decreased and Th17 increased in spleen lymphocyte of Model group (p < .01); after Allicin treatment, Treg increased while Th17 decreased significantly (p < .01). Meanwhile, MEKK2 protein expression in spleen lymphocyte of Model group decreased compared to that in NC group (p < .01), and MEK2 protein expression increased significantly after Allicin treatment (p < .01). To sum up, the present study suggests that MEKK2 protein plays an important role in the pathogenesis of CIA model. In terms of mechanism, Allicin may play a therapeutic role in rheumatoid arthritis (RA) by increasing the expression of MEKK2 protein and affecting Treg/Th17 ratio.

Dihydroartemisinin attenuated the symptoms of mice model of systemic lupus erythematosus by restoring the Treg/Th17 balance.

The Treg/Th17 imbalance is associated with the development of systemic lupus erythematosus (SLE). Dihydroartemisinin (DHA), a semi-synthetic derivative of artemisinin, is isolated from the traditional Chinese herb Artemisia annua Artemisia annua L. This study aims to evaluate the effects of DHA alone or in combination with prednisone in immunodeficiency of SLE. In vivo, the therapeutical effect of DHA alone or in combination with prednisone was assessed in the pristane-induced SLE mouse model. Then, the level of serum antibodies, creatinine (Cre), blood urea nitrogen (BUN), urine protein, kidney histopathology, interleukin (IL)-17, IL-6, transforming growth factor (TGF)-ß, the expression of RORγt and Foxp3, the percentages of Treg and Th17 in peripheral blood and spleen were assayed. In vitro, the mouse spleen lymphocytes were separated and treated with DHA alone or DHA in combination with prednisone. Then the percentages of Treg and Th17, the concentration of IL-17, IL-6, TGF-ß, and the expression of RORγt and Foxp3 were assayed. It was shown that DHA alone or in combination with prednisone treatment significantly alleviated the manifestations of pristane-induced SLE mice, suppressed inflammation and restored the Treg/Th17 balance. DHA alone or in combination with prednisone significantly inhibited Th17 cell differentiation while induced Treg cell differentiation in vitro. DHA alone or in combination with prednisone also reduced the transcription of RORγt and increased Foxp3 in lymphocytes, as well as IL-17 and TGF-ß levels. Our data indicated that DHA can produce synergistic effect with prednisone to attenuate the symptoms of SLE by restoring Treg/Th17 balance.

Paeonol attenuated vascular fibrosis through regulating Treg/Th17 balance in gut microbiota-dependent manner / 中国药理学与毒理学杂志

OBJECTIVE Atherosclerosis (AS) is a chronic inflammatory disease characterized by the accumulation of lipids, vascular fibrosis, and inflammation. Paeonol (Pae) is a natural phenolic compounds isolated from a traditional Chinese medicine, Cortex Moutan, which exhibits anti-AS effects. Our previous work demonstrated that gut microbiota plays an important role during AS treatment as it affects the efficacy of Pae. However, the mechanism of Pae in protect?ing against vascular fibrosis as related to gut microbiota has yet to be elucidated. To investigate the anti-fibrosis effect of Pae on AS mice and demonstrate the underlying gut microbiota-dependent mechanism. METHODS ApoE-/- mice were fed with high-fat-diet (HFD) to replicate the AS model. HE and Masson staining were used to observe the plaque forma?tion and collagen deposition. Gut microbiota alteration and short-chain fatty acids (SCFAs) production were analyzed through 16S rRNA sequencing and LC-MS/MS. The frequency of immune cells in spleen were phenotyped by flow cytometry. The mRNA expression of aortic inflammatory cytokines were detected by qRT-PCR. The protein expression of LOX and fibrosis related indicators were examined by Western blotting. RESULTS Pae restricted the development of AS and collagen deposition. Notably, the anti-fibrosis effect of Pae was achieved by regulating the gut microbiota. 16S rRNA sequencing and LC-MS/MS data indicated that the relative abundance of SCFAs-producing bacteria and SCFAs production was increased. Additionally, Pae administration selectively up-regulated the frequency of regulatory T (Treg) cells as well as down-regulated the ratio of T helper type 17 (Th17) cells in the spleen of AS mice, improving the Treg/Th17 balance. In addition, as expected, Pae intervention significantly down-regulate the mRNA expression levels of pro-inflammatory cytokines IL-1β, IL-6, TNF-αand IL-17 in the aorta tissue, up-regulate the levels of anti-inflammatory factor IL-10, a marker of Treg cells. Finally, Pae's intervention in the gut microbiota resulted in the restoration of the balance of Treg/Th17, which indirectly down-regulated the protein expression level of LOX and fibrosis-related indicators (MMP-2/9 and collagenⅠ/Ⅲ). CONCLUSION Pae attenuates vascular fibrosis in a gut microbiota-dependent manner. The under?lying protective mechanism is associated with the improved Treg/Th17 balance in spleen mediated through the increased microbiota-derived SCFAs production.