Xia Luo

Immunosuppressive activity of pogostone on T cells: Blocking proliferation via S phase arrest.

Pogostone (PO) is one of the major chemical constituents of the essential oil of Pogostemon cablin (Blanco) Benth. In the present study, the effect of PO on T cell responsiveness was investigated to explore its potential in immunosuppression by a Concanavalin A (ConA)-stimulation model using splenocytes isolated from C57BL/6 mice. Cytotoxicity by PO on normal splenocytes was evaluated by MTS assays. Characteristics of apoptosis, proliferation, and cell cycle were analyzed by flow cytometry. Related expressions of cyclins and cyclin-dependent kinases (CDKs) were also determined by flow cytometry. Inflammatory cytokine profiling was performed emplying cytometric beads assays (CBA). Moreover, the T cell-mediated delayed Type hepersensity (DTH) model was applied to evaluate the immunosuppressive activity of PO. Neither viability reduction in normal splenocytes nor apoptosis in ConA-stimulated splenocytes was observed under PO treatments. Meanwhile, PO remarkably reduced the total population of ConA-stimulated T cell, blocked T cell proliferation induced by Con A, and inhibited the production of IFN-γ and IL-10. This blockade of stimulated T cell proliferation by PO was likely attributed to down-regulation of cyclin E, cyclin B and CDK1 and the subsequent S-phase arrest. Additionally, PO could inhibit the DTH reaction by alleviating ear swelling and inflammatory infiltrations in the DNCB-challenged ear. Taken together, PO exhibited an immunosuppressive property by directly blocking T cell proliferation as well as altering inflammatory cytokine profile, suggesting that PO may have clinical implications for treating autoimmune diseases and other immune-based disorders.

Effects of ceftriaxone induced intestinal dysbacteriosis on lymphocytes in different tissues in mice

The close relationship between intestinal microflora and immune system has been confirmed, stimulus from intestinal flora plays an important role in the development of the immune system and its dynamic balance. Current research is still inadequate to determine how local bacteria in gut influence the whole body. In this study, influence of ceftriaxone sodium induced intestinal dysbacteriosis on local and overall immune function was investigated. We found that the beneficial bacteria decreased significantly compared with control after oral administration of ceftriaxone; Moreover, the proportion of T cells are higher and B cells are lower in the dysbacteriosis mice, activation and proliferation of T and B cells was decreased significantly in gut-associated lymphoid tissues (GALTs), such as Peyers patches (PPs) and mesenteric lymph nodes(MLNs)with ceftriaxone treatment; The secreted sIgA in intestinal was reduced in dysbacteriosis mice than that of control as well. The similar results above are also shown on the spleen. In addition, the delayed type hypersensitivity (DTH) reaction decreased in dysbacteriosis mice. The present data suggested that intestinal microflora had impact on immune system by influencing the proportion and function of lymphocytes in PPS-MLN-spleen.

Antitumor activity of Lycium barbarum polysaccharides with different molecular weights: an in vitro and in vivo study

ABSTRACT The antitumor activity of Lycium barbarum polysaccharide (LBP) has been reported, but the structure–bioactivity relationship has still not been fully elucidated. In this study, four water-soluble LBP fractions with serial different molecular weights (MWs) were separated from LBP, designated LBP-2, LBP-3, LBP-4, and LBP-5. After a characteristic analysis, the relationship between MW and antitumor activity of LBP was investigated both in vitro using murine hepatoma H22 cells and in vivo using H22 tumor-bearing mice. In vitro, the results showed that all the LBP fractions had significant inhibition on H22 cells, in which LBP-3 had the best activity. LBP-3 could induce apoptosis, mitochondrial membrane potential destruction, and S phase arrest in H22 cells. In vivo, the results showed that LBP-2, LBP-3, LBP-4, and LBP-5 could inhibit the tumor growth in H22 tumor-bearing mice by 18.18%, 37.97%, 9.09%, and 14.44%, respectively. However, only LBP-3 was able to decrease the tumor weight significantly in H22 tumor-bearing mice. Meanwhile, all the LBP fractions did not show significant toxicity to murine splenocytes, thymus, and spleen. Taken together, these results demonstrated that the antitumor activity of LBP was closely related to its MW, and LBP-3 with medium MW (40–350 kDa) was the main active fraction.

Ethyl pyruvate ameliorates experimental colitis in mice by inhibiting the HMGB1-Th17 and Th1/Tc1 responses

Ethyl pyruvate (EP), a simple lipophilic pyruvate ester, has demonstrated protective effects against murine colitis through inhibition the release of inflammatory factor high-mobility group protein box 1 (HMGB1). HMGB1 has been implicated in several autoimmune diseases by inducing Thl and Thl7 cells activation. This study was designed to investigate whether EP amelioration of murine colitis is related to the blocking of the HMGB1-Th17/Thl pathway. We induced murine colitis by intrarectal administration of 2, 4, 6-trinitrobenzene sulfonic acid (TNBS). Ethyl pyruvate was injected intraperitoneally once a day for 7days. One week after intrarectal challenge with TNBS, HMGB1, IL-17 and IFN-γ protein levels were remarkably increased following severe colon inflammation. Meanwhile, excessive infiltration of Th17 cells in colonic tissues, and an upregulated proportion of Th17 and Th1/Tc1 cells in the spleen and mesenteric lymph nodes (MLN) were found in the TNBS-treated group compared to the control group. Treatment with the HMGB1 inhibitor EP not only remarkably improved colon pathological damage, but also significantly reduced the number of Th17 cells in the local tissues of the colitis-induced mice. Furthermore, the percentage of Th1/Tc1 and Th17 cells in the spleen and MLN, as well as levels of serum IFN-γ and IL-17A, were all markedly decreased in the EP-treated group. Moreover, in vitro, our results showed that EP in a dose dependent manner inhibited HMGB1 release induced by LPS from CT26 cells (murine colon adenocarcinoma cell line). These results suggest that HMGB1 contributes to the development of murine colitis by promoting the Th17 and Th1/Tc1 responses, and that EP can significantly inhibit HMGB1-Th17 and Thl/Tc1 pathway activation, which may provide better protection to mice with TNBS-induced colitis.

Fraction From Lycium barbarum Polysaccharides Reduces Immunotoxicity and Enhances Antitumor Activity of Doxorubicin in Mice

The aim of the present study was to investigate whether fraction from Lycium barbarum polysaccharide (LBP) could reduce immunotoxicity and enhance antitumor activity of doxorubicin (Dox) in mice. A water-soluble LBP fraction, designated LBP3, was isolated from edible Chinese herbal Lycium barbarum and used in this study. To investigate the effect of LBP3 on Dox-induced immunotoxicity, tumor-free mice were used and treated with either normal saline, Dox, or Dox plus LBP3. To investigate the effect of LBP3 on antitumor activity of Dox, H22 tumor-bearing mice were used and treated with either normal saline, Dox, LBP3, or Dox plus LBP3. The results showed that LBP3 did not protect against the body weight loss caused by Dox, but it promoted the recovery of body weight starting at day 5 after Dox treatment in tumor-free mice. LBP3 also improved peripheral blood lymphocyte counts, promoted cell cycle recovery in bone marrow cells, and restored the cytotoxicity of natural killer cells. Furthermore, in H22 tumor-bearing mice, LBP3 enhanced antitumor activity of Dox and improved peripheral blood lymphocyte counts and the cytotoxicity of splenocytes. In brief, our results demonstrated that LBP3 could reduce the immunotoxicity and enhance antitumor activity of Dox.

Identification of genes underlying the enhancement of immunity by a formula of lentinan, pachymaran and tremelia polysaccharides in immunosuppressive mice

The efficacy of polysaccharides is widespread, especially in immune regulation. However, the genetic basis of the changes in polysaccharides regulating immunity is unclear. To obtain genome-wide insights into transcriptome changes and regulatory networks, we designed a polysaccharide formula, comprising lentinan, pachymaran and tremelia, to increase the availability of their optimized active sites. In this case, we focused on a model of immunosuppression to investigate genes by digital gene expression (DGE) tag profiling in T and B cells. These genes were further validated by qRT-PCR and Western blot experiments. Consequently, polysaccharide formula treatment helped to recover the expression of immune-related genes, including CADM1, CCR2, IGLL1, LIGP1, and FCGR3, FCGR2 in B cells, as well as S100A8, S100A9, ChIL3, MMP8 and IFITM3 in T cells. These results suggest that treatment with polysaccharides improves the immunity of immunosuppressive mice by regulating genes associated with T and B cell functions.

Rhubarb Peony Decoction Ameliorates Ulcerative Colitis in Mice by Regulating Gut Microbiota to Restoring Th17/Treg Balance

ETHNOPHARMACOLOGY RELEVANCE Rhubarb Peony Decoction (RPD) is a formula of traditional Chinese medicine chronicled in Jin Gui Yao Lve, commonly used to treat ulcerative colitis (UC). However, the underlying mechanism of RPD treating UC remains elusive. In our study, we investigated the therapeutic efficacy of RPD and potential mechanism involved in inhibiting dextran sulfate sodium (DSS)-induced ulcerative colitis in mice. METHODS The colitis was induced by DSS in mice for 5 days and estimated body weight loss, disease activity index (DAI) and colon length. Histological changes were observed by H&E staining. The number and abundance of gut mircrobiota were measured with 16 S rDNA sequencing. GC-MS was used to detect the concentration of short chain fatty acids (SCFAs) in cecum. Flow cytometry analyzed the proportion of Th17 and Treg cells in mesenteric lymph nodes (MLNs). IL-17A and Foxp3 in colon were determined by immunohistochemical analyses. The level of cytokine was determined by Multi-Analyte Flow Assay Kit. RESULTS Administration of RPD significantly alleviated the pathological changes of UC mice, involving rescued the inflammation-related reduction of colon length, ameliorated body weight loss and damaged tissue. In addition, RPD altered the gut microbiota, involving restored α diversity, increased significantly the abundance of Firmicutes and Actinobacteria, decreased the Proteobacteria and Bacteroidetes. Furthermore, the number of Butyricicoccus pullicaecorum, a butyrate-producing bacterium, were augmented obviously by RPD. Besides, RPD restored the content of SCFA in intestinal tract. Additionally, the proportion of Th17 cells and Treg cells in mesenteric lymph nodes, likewise, the expression of IL-17A and Foxp3 in colon were regulated by RPD, contributing to the restoration of Th17/Treg balance. Moreover, RPD significantly decreased the level of IL-6, TNF-α, IFNγ, IL-10, IL-17A, IL-21, IL-22 in colon, simultaneously increased Treg-related cytokine TGF-β at dose-dependently. CONCLUSIONS These results demonstrated that RPD had effect on ulcerative colitis, which was related to regulating gut microbiota, especially Butyricicoccus pullicaecorum, and SCFAs to restore the gut Th17/Treg homeostasis.

Against NF-κB/thymic stromal lymphopoietin signaling pathway, catechin alleviates the inflammation in allergic rhinitis

Background: The prevalence of allergic rhinitis has risen sharply. Previous work has demonstrated the anti‐inflammatory effect of catechin, including in models of allergic disease. However, the molecular mechanisms underlying this therapeutic effect remain unclear. Thymic stromal lymphopoietin(TSLP), as a molecule from epithelial cell, has been identified that plays a significant role in the development of allergic disease, and the production of TSLP is related to activation of the NF‐&kgr;B signaling pathway. For that, we try to research the treatment of catechin for allergic rhinitis and found out possible mechanism under this effect, which was based on TSLP factor. Materials and methods: Here, the anti‐inflammatory effects of catechin were studied in an ovalbumin‐induced allergic rhinitis murine model and a vitro experiments using poly(I:C)‐stimulated human nasal epithelial cells(HNEpCs). The pharmacological effects of catechin in allergic rhinitis mice were assessed by observing the allergic symptoms, performing hematoxylin and eosin staining and Giemsa staining of the nasal tissues. Additionally, the TSLP expression in epithelial cells was tested by enzyme‐linked immunosorbent assays, immunohistochemistry, and western blots. The serum levels of interleukin‐5, interleukin‐13, and ovalbumin‐specific immunoglobulin‐E were detected by enzyme‐linked immunosorbent assays, and the balance between T helper type 1 and T helper type 2 cells was assessed by flow cytometry. The expression levels of phospho‐NF‐&kgr;Bp65, I&kgr;B&agr;, and NF‐&kgr;Bp65 proteins were further investigated by western blots or immunofluorescence. Results: Our results reveal that catechin, in doses of 75, 150, or 300 mg/kg, remitted the allergic symptoms in mice with allergic rhinitis, like sneezing and nasal rubbing. Catechin could reduce the levels of interleukin‐5, interleukin‐13, and ovalbumin‐specific immunoglobulin‐E in the serum and restored the T helper type 2/T helper type 1 cell balance and also had anti‐thymic stromal lymphopoietin effects. Moreover, as an upstream regulator of TSLP, the NF‐&kgr;B signal pathway was also suppressed after catechin treatment, which was demonstrated by the observed decrease in phospho‐NF‐&kgr;Bp65 and NF‐&kgr;Bp65 levels and the reduction of I&kgr;B&agr; degradation and NF‐&kgr;Bp65 nuclear translocation. Conclusions: Catechin effectively reduced the inflammation in allergic rhinitis. The underlying mechanism is that catechin inhibited the expression of TSLP in epithelial cells by influencing NF‐&kgr;B/TSLP pathway. HighlightsCatechin effectively remits the allergic symptom and Th2 inflammation in allergic rhinitis mice.Catechin reduces the expression of thymic stromal lymphopoietin(TSLP) and NF‐&kgr;Bp65 in nasal mucosa of allergic rhinitis mice.Catechin inhibits the expression of TSLP and the activation of NF‐&kgr;B signal pathway in human nasal epithelial cells(HNEpCs).

Emodin ameliorates ulcerative colitis by the flagellin-TLR5 dependent pathway in mice

ABSTRACT Emodin is an anthraquinone compound derived from Rheum officinale Baill. Several reports showed that emodin had efficacy on acute pancreatitis, keratitis, myocarditis, and rheumatoid arthritis. However, the potency of emodin on ulcerative colitis(UC) remains unclear. In this study, we investigated the effect of emodin on dextran sodium sulfate (DSS)‐induced ulcerative colitis in mice. Our results showed that emodin significantly alleviated the symptoms of DSS‐induced UC in mice, involving prevented the loss of body weight and colon shortening, decreased the disease activity index (DAI), intestinal damages and the count of white blood cells (WBC) in peripheral blood. In addition, emodin treatment decreased the level of anti‐flagellin antibody in serum and significantly down‐regulated the expression of TLR5 and NF‐&kgr;B p65 in colon of the UC mice. Further study in vitro showed that emodin down‐regulated the expression of TLR5 and MyD88, up‐regulated the expression of I&kgr;B, inhibited the nuclear translocation of NF‐&kgr;B p65 and decreased the release of IL‐8 in flagellin‐stimulated HT‐29 cells. These results, for the first time, demonstrated that emodin had the therapeutic potential to ameliorate UC symptoms possibly via regulating the flagellin‐TLR5 signaling pathway. Emodin may be a potential candidate ingredient for ulcerative colitis. HIGHLIGHTSWe found that emodin was a potential candidate ingredient for ulcerative colitis.Effect of emodin on UC was related to regulation of flagellin‐TLR5 signaling pathway.

Effects of ceftriaxone-induced intestinal dysbacteriosis on regulatory T cells validated by anaphylactic mice

ABSTRACT Both probiotics and pathogens in the human gut express pathogen‐associated molecular patterns (PAMPs) and die with the release of endotoxin and bacterial DNA, which can stimulate our immune system and cause immune reaction. However, its interesting and fascinating to address why the normal intestinal flora will not generate immunological rejection like the pathogen does. By investigating the changes in cells and molecules relevant to immune tolerance in mice with ceftriaxone‐induced dysbacteriosis, our study discovered that both the Evenness indexes and Shannon Wiener index of intestinal flora showed a decrease in dysbacteriosis mice. Moreover, the proportion of &agr;&bgr;+TCR+CD3+CD4−CD8− cells, CD3+&ggr;&dgr;TCR+ cells and CD4+CD25+FoxP3+ cells in the Peyers patches (PPs), mesenteric lymph nodes (MLNs) and spleen (SP) and the level of TGF‐&bgr;1, IL‐2, IL‐4 and IL‐10 in the serum also changed. Intestinal dysbacteriosis in an asthma murine model resulted in enhancement of immunologic response to the allergen ovalbumin (OVA), which was an agent that aggravates asthma symptoms. In summary, it is integral to maintain a certain amount or variety of intestinal microflora for regulatory T cells to act in averting hypersensitivity. HIGHLIGHTSIntestinal dysbacteriosis reduced the proportion and function of regulatory T cells in miceIntestinal dysbacteriosis enhances sensibility to allergen OVA in mice.Anaphylaxis is directly related to the dysfunction of regulatory T cells caused by intestinal flora dysbacteriosis.