Neobavaisoflavone-mediated TH9 cell differentiation ameliorates bowel inflammation.

Neobavaisoflavone (Neo), is the active constituent of the herb Psoralea corylifolial, used in the traditional Chinese medicine, and has anti-inflammatory activity, but whether Neo could regulate colitis remains unclear. T helper 9 (TH9) cells, a subset of CD4+ T helper cells characterized by secretion of IL-9, have been reported to be involved in the pathogenesis of many autoimmune and inflammatory diseases, but whether Neo could control TH9 cell differentiation also remains unclear. Here, we found that Neo could decrease IL-9 production of CD4+ T cells by targeting PU.1 in vitro. Importantly, Neo had therapeutic effects on DSS-induced colitis. Furthermore, we identified TH9 cells as the direct target of Neo for attenuating bowel inflammation. Therefore, Neo could serve as a lead for developing new therapeutics against inflammatory bowel disease.

Sophoricoside from Styphnolobium japonicum improves experimental atopic dermatitis in mice.

BACKGROUND: Abnormal immune responses, specifically excessive differentiation of Th2 cells, are associated with the development of atopic dermatitis (AD). Sophoricoside, the genistein-4'-ß-D-glucoside isolated from Styphnolobium japonicum, has previously demonstrated anti-inflammatory and immunosuppressive effects along with IL-3 and IL-5 inhibitory activities. Therefore, we speculated that sophoricoside could regulate AD by regulating abnormal immune responses. PURPOSE: To investigate the role of sophoricoside on AD-like allergic skin inflammation induced by ovalbumin (OVA) or 2,4,6-trinitrochlorobenzene (TNCB) in mouse models. METHODS: Sophoricoside was isolated from the 70% ethanol extract of S. japonicum dried mature seeds. After being submitted to a purification process, its purity was assessed by high-performance liquid chromatography (HPLC). The effects of sophoricoside were determined in vivo by OVA- and TNCB-induced AD-like allergic skin inflammation in mice. Skin tissues were subjected with hematoxylin-eosin (H&E), Giemsa and toluidine blue staining. In vitro CD4+ T cell differentiation was performed and the levels of serum immunoglobulins, cytokines, and genes related to CD4+ T cell differentiation were determined by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time PCR. Cytokine bioassay, mixed lymphocytes reaction and cell viability assay were performed. RESULTS: Topical application of sophoricoside decreased the symptoms of AD-like allergic skin inflammation, including elevated hypertrophic scars with spongiotic epidermis, epidermal hyperplasia, hyperkeratosis, infiltration of immune, and mast cells, dermal thickness, amounts of immunoglobulins, and pro-inflammatory cytokines, and the mast cell population in the skin. Sophoricoside also decreased T cell antigen receptor (TCR)-mediated immune responses. In particular, sophoricoside suppressed the differentiation of naïve CD4+ T cells into Th cell subsets, including Th1, Th2, and Th17, by inhibiting the expression of their subset-specific master transcription factors, leading to suppression of the expression and production of these cell subset-specific cytokines. CONCLUSION: Sophoricoside can improve AD-like allergic skin diseases mainly by inhibiting pathogenic CD4+ T cell differentiation and immune responses.

T Helper Cell Lineage-Defining Transcription Factors: Potent Targets for Specific GVHD Therapy?

Allogenic hematopoietic stem cell transplantation (allo-HSCT) represents a potent and potentially curative treatment for many hematopoietic malignancies and hematologic disorders in adults and children. The donor-derived immunity, elicited by the stem cell transplant, can prevent disease relapse but is also responsible for the induction of graft-versus-host disease (GVHD). The pathophysiology of acute GVHD is not completely understood yet. In general, acute GVHD is driven by the inflammatory and cytotoxic effect of alloreactive donor T cells. Since several experimental approaches indicate that CD4 T cells play an important role in initiation and progression of acute GVHD, the contribution of the different CD4 T helper (Th) cell subtypes in the pathomechanism and regulation of the disease is a central point of current research. Th lineages derive from naïve CD4 T cell progenitors and lineage commitment is initiated by the surrounding cytokine milieu and subsequent changes in the transcription factor (TF) profile. Each T cell subtype has its own effector characteristics, immunologic function, and lineage specific cytokine profile, leading to the association with different immune responses and diseases. Acute GVHD is thought to be mainly driven by the Th1/Th17 axis, whereas Treg cells are attributed to attenuate GVHD effects. As the differentiation of each Th subset highly depends on the specific composition of activating and repressing TFs, these present a potent target to alter the Th cell landscape towards a GVHD-ameliorating direction, e.g. by inhibiting Th1 and Th17 differentiation. The finding, that targeting of Th1 and Th17 differentiation appears more effective for GVHD-prevention than a strategy to inhibit Th1 and Th17 cytokines supports this concept. In this review, we shed light on the current advances of potent TF inhibitors to alter Th cell differentiation and consecutively attenuate GVHD. We will focus especially on preclinical studies and outcomes of TF inhibition in murine GVHD models. Finally, we will point out the possible impact of a Th cell subset-specific immune modulation in context of GVHD.

Carvacrol inhibits the excessive immune response induced by influenza virus A via suppressing viral replication and TLR/RLR pattern recognition.

ETHNOPHARMACOLOGICAL RELEVANCE: Carvacrol, a monoterpene phenol from Mosla chinensis Maxim, which is a commonly Chinese herbal medicine. The most important pharmacology of it is dispelling exogenous evils by increasing perspiration. And it is the gentleman medicine in the Chinese herbal compound prescription of Xin-Jia-Xiang-Ru-Yin, mainly for the treatment of summer colds with dampness including influenza virus A infection. AIM OF THE STUDY: Our preliminary study verified that the Xin-Jia-Xiang-Ru-Yin could inhibit acute lung injury of mice with influenza virus A infection. And there have been some reports implicating the high antimicrobial activity of carvacrol for a wide range of product preservation, but little research including the effects of it on viral infection. The aim of this study was to reveal the antiviral effects of carvacrol, the main constituent in Mosla chinensis Maxim. MATERIALS AND METHODS: Initially, C57BL/6 mice were grouped and intranasally administered FM1 virus to construct viral infection models. After treatment with ribavirin and carvacrol for 5 days, all mice were euthanized, and specimens were immediately obtained. Histology, flow cytometry and Meso Scale Discovery (MSD) analysis were used to analyze pathological changes in lung tissue, the expression levels of cytokines and the differentiation and proportion of CD4+ T cells subsets, while Western blot and qRT-PCR were used to detect the expression of related proteins and mRNA. RESULTS: Carvacrol attenuated lung tissue damage, the proportions of Th1, Th2, Th17 and Treg in CD4+ T cells and the relative proportions of Th1/Th2 and Th17/Treg cells. Carvacrol inhibited the expression of inflammation-associated cytokines including IFN-γ, IL-2, IL-4, IL-5, IL-12 and TNF-ɑ, IL-1, IL-10, IL-6. Decreased levels of TLR7, MyD88, IRAK4, TRAK6, NF-κB, RIG-I, IPS-I and IRF mRNA in carvacrol-treated mice were observed comparing to the mice in VC group. Further, the total expression of RIG-I, MyD88 and NF-κB proteins had increased significantly in the VC group but reduced obviously in the group treated with ribavirin or carvacrol. CONCLUSIONS: These results indicate that carvacrol is a potential alternative treatment for the excessive immune response induced by influenza virus A infection, the cold-fighting effect of Mosla chinensis Maxim may depend on the anti-virus of carvacrol.

Coptis chinensis Franch polysaccharides provide a dynamically regulation on intestinal microenvironment, based on the intestinal flora and mucosal immunity.

ETHNOPHARMACOLOGICAL RELEVANCE: Coptis chinensis Franch is one of the most widely used traditional Chinese herbs in China and was firstly recorded in "Shennong's Classic of Materia Medica" in the Han Dynasty. The medical records in past thousands years have fully confirmed the clinical efficacies of Coptis chinensis Franch against intestinal diseases. The polysaccharides in herbal medicines can be digested by the flora and uptaken by the Peyer's patches (PPs) in intestine. It can be reasonably presumed that the polysaccharides in Coptis chinensis Franch (CCP) should be one of the critical element in the regulation of intestinal microenvironment. AIM OF THE STUDY: This study intended to explore the dynamic regulation of CCP on intestinal microenvironment from the perspective of the intestinal mucosal immunity and the intestinal flora, in order to provide a new research perspective for the pharmacological mechanism of Coptis chinensis Franch. MATERIALS AND METHODS: The absorption and distribution of CCP in intestinal tissues were observed after the perfusion of FITC labeled CCP. The influences of CCP on intestinal flora were evaluated by the 16sRNA gene illumina-miseq sequencing after gavage. The regulations of CCP on intestinal mucosal immunity were evaluated by the immunohistochemical analysis of the interferon-γ (IFN-γ), interleukin-4 (IL-4), interleukin-17 (IL-17) and transforming growth factor-ß (TGF-ß) secretion in PPs and intestinal epithelial tissue. RESULTS: With the self-aggregation into particles morphology, CCP can be up-taken by PPs and promote the IFN-γ, IL-4, IL-17 and TGF-ß secretion in PPs in a dose-dependent manner. The CCP can also be utilized by the intestinal flora and dynamically regulate the diversity, composition and distribution of the intestinal flora. The temporal regulations of CCP on IFN-γ, IL-4, IL-17 and TGF-ß secretions in intestinal epithelial tissues are consistent with the variation tendency of intestinal flora. CONCLUSION: CCP can provide effective, dynamical and dose-dependent regulations on intestinal microenvironment, not only the intestinal flora but also the PPs and intestinal epithelium related immune response. These may be involved in the multiple biological activities of Coptis chinensis Franch.

YYFZBJS ameliorates colorectal cancer progression in ApcMin/+ mice by remodeling gut microbiota and inhibiting regulatory T-cell generation.

BACKGROUND: Progression of Colorectal cancer (CRC) is influenced by single or compounded environmental factors. Accumulating evidence shows that microbiota can influence the outcome of cancer immunotherapy. T cell, one of the main populations of effector immune cells in antitumor immunity, has been considered as a double-edged sword during the progression of CRC. Our previous studies indicate that traditional Chinese herbs (TCM) have potential anticancer effects in improving quality of life and therapeutic effect. However, little is known about the mechanism of TCM formula in cancer prevention. METHODS: Here, we used C57BL/6 J ApcMin/+ mice, an animal model of human intestinal tumorigenesis, to investigate the gut bacterial diversity and their mechanisms of action in gastrointestinal adenomas, and to evaluate the effects of Yi-Yi-Fu-Zi-Bai-Jiang-San (YYFZBJS) on of colon carcinogenesis in vivo and in vitro. Through human-into-mice fecal microbiota transplantation (FMT) experiments from YYFZBJS volunteers or control donors, we were able to differentially modulate the tumor microbiome and affect tumor growth as well as tumor immune infiltration. RESULTS: We report herein, YYFZBJS treatment blocked tumor initiation and progression in ApcMin/+ mice with less change of body weight and increased immune function. Moreover, diversity analysis of fecal samples demonstrated that YYFZBJS regulated animal's natural gut flora, including Bacteroides fragilis, Lachnospiraceae and so on. Intestinal tumors from conventional and germ-free mice fed with stool from YYFZBJS volunteers had been decreased. Some inflammation' expression also have been regulated by the gut microbiota mediated immune cells. Intestinal lymphatic, and mesenteric lymph nodes (MLN), accumulated CD4+ CD25+ Foxp3 positive Treg cells were reduced by YYFZBJS treatment in ApcMin/+ mice. Although YYFZBJS had no inhibition on CRC cell proliferation by itself, the altered Tregs mediated by YYFZBJS repressed CRC cancer cell growth, along with reduction of the phosphorylation of ß-catenin. CONCLUSIONS: In conclusion, we demonstrated that gut microbiota and Treg were involved in CRC development and progression, and we propose YYFZBJS as a new potential drug option for the treatment of CRC. Video abstract.

Coptidis alkaloids extracted from Coptis chinensis Franch attenuate IFN-γ-induced destruction of bone marrow cells.

Coptidis alkaloids are the primary active components of Coptis chinensis Franch. Clinical and pharmacodynamic studies have confirmed that Coptidis alkaloids have multiple therapeutic effects including anti-inflammatory, antioxidant and antitumor effects, and they are usually used to treat various inflammatory disorders and related diseases. Mouse bone marrow cells (BMCs) were isolated from BALB/c mice. Immune-mediated destruction of BMCs was induced by interferon (IFN) -γ. High-performance liquid chromatography-electrospray ionization/ mass spectrometry was used to analyze the ingredients of the aqueous extract from Coptis chinensis Franch. The results confirmed that Coptidis alkaloids were the predominant ingredients in the aqueous extract from Coptis chinensis. The functional mechanism of Coptidis alkaloids in inhibiting immune-mediated destruction of BMCs was studied in vitro. After Coptidis alkaloid treatment, the percentages of apoptotic BMCs and the proliferation and differentiation of helper T (Th) cells and regulatory T (Treg) cells were measured by flow cytometry. The expression and distribution of T-bet in BMCs were observed by immunofluorescence. Western blotting analysis was used to assay the expression of key molecules in the Fas apoptosis and Jak/Stats signaling pathways in BMCs. We identified five alkaloids in the aqueous extract of Coptis chinensis. The apoptotic ratios of BMCs induced by IFN-γ were decreased significantly after Coptidis alkaloid treatment. The levels of key molecules (Fas, Caspase-3, cleaved Caspase-3, Caspase-8 and Caspase-8) in Fas apoptosis signaling pathways also decreased significantly after treatment with low concentrations of Coptidis alkaloids. Coptidis alkaloids were also found to inhibit the proliferation of Th1 and Th17 cells and induce the differentiation of Th2 and Treg cells; further, the distribution of T-bet in BMCs was decreased significantly. In addition, the levels of Stat-1, phospho-Stat-1 and phospho-Stat-3 were also reduced after Coptidis alkaloid treatment. These results indicate that Coptidis alkaloids extracted by water decoction from Coptis chinensis Franch could inhibit the proliferation and differentiation of T lymphocytes, attenuate the apoptosis of BMCs, and suppress the immune-mediated destruction of the BMCs induced by pro-inflammatory cytokines.

Multi-functional nanocomplex codelivery of Trp2 and R837 to activate melanoma-specific immunity.

Antigen-adjuvant combination could induce a protective and long-lasting anti-tumor immune response. However, exploiting system which could co-deliver melanoma antigen peptide Trp2 (Tyrosinase-related protein 2) and Toll-like-receptor-7 (TLR7) agonists imiquimod (R837) both are poor aqueous solubility is still challenging. Our new nanocomplex was explored for specific delivery of Trp2 and R837 into antigen-presenting cells (APCs). R837 was loaded into mannosylated-ß-cyclodextrin (Man-CD) to target dendritic cells (DCs) by binding mannose receptors (MR) on DCs. A fusion peptide (WT) was constructed by incorporating the amino acid region of TAT (cell-penetrating peptide) into Trp2 to improve the TAT-mediated intracellular efficiency. Negatively charged sodium alginate (SA), a biocompatible polymer, which can induce adjuvant responses by affecting the functions of DCs, was complexed with Man-CD/R837 and WT via physical adsorption. The optimized nanocomplex promoted the cellular uptake and showed remarkable efficacy to enhance the secreting of Th1-cytokines. This multi-functional nanocomplex system may allow effective targeting-codelivery of multi-hydrophobic drugs and be a promising subunit vaccine candidate as a potential prevention action of tumor.

Bupi Hewei decoction ameliorates 5-fluorouracil-induced intestinal dysbiosis in rats through T helper 17/T regulatory cell signaling pathway.

OBJECTIVE: To observe the effects of the Bupi Hewei (BPHW) decoction on diarrhea and intestinal flora disorder induced by 5-fluorouracil (5-FU) and investigate the possible mechanism underlying these actions. METHODS: Thirty-five male Sprague-Dawley rats were randomly divided into four groups: normal control, 5-FU, 5-FU + BPHW decoction (10.5 g/kg for 5 consecutive days), and 5-FU + Bacillus licheniformis capsule groups (0.2 g/kg for 5 consecutive days). Animal models were established via the intraperitoneal injection of 5-FU (30 mg/kg for 5 consecutive days). At the end of the treatment period, diarrhea was assessed, and the change of the intestinal flora was examined using 16S rDNA high- throughput sequencing. Interleukin (IL)-17, IL-21, IL-6, IL-10, RAR-related orphan receptor gamma (RORγt), and forkhead box P3 (Foxp3) expression in the jejunum was detected using immunohistochemistry, quantitative real-time polymerase chain reaction (PCR), Western blotting, and enzyme- linked immuno sorbent assay. RESULTS: In this study, the BPHW decoction effectively lowered the diarrhea score, increased the proportions of Bacteroidetes and Prevotellaceae-Alloprevotella species, and reduced the proportions of Proteobacteria, Escherichia-Shigella, Ruminococcaceae NK4A214, and Ruminococcaceae UCG-005 species in the rat intestine after 5-FU chemotherapy. In addition, the BPHW decoction significantly suppressed the expression of IL-17, IL-21, IL-6, IL-10, RORγt, and Foxp3 in the jejunum. CONCLUSION: Our findings suggest that the BPHW decoction can improve the intestinal immune balance and reduce intestinal inflammation by targeting T helper cell/T regulatory cell-associated factors.

Fermented Morinda citrifolia (Noni) Alleviates DNCB-Induced Atopic Dermatitis in NC/Nga Mice through Modulating Immune Balance and Skin Barrier Function.

Morinda citrifolia, a fruit generally known as "Noni", has been traditionally used in parts of East Asia to relieve inflammatory diseases. Although several studies using noni have been reported, the effect of fermented Morinda citrifolia (F.NONI) on atopic dermatitis (AD) has not been investigated. Thus, we aimed to investigate the improving effect of F.NONI treatment on AD-like skin lesions and elucidate molecular mechanisms. F.NONI was prepared by the fermentation of noni fruit with probiotics and then extracted. F.NONI was orally administrated to NC/Nga mice to evaluate its therapeutic effect on 2,4-dinitrochlorobenzene (DNCB)-induced AD. Oral administration of F.NONI significantly alleviated AD lesions and symptoms such as dermatitis scores, ear thickness, scratching behavior, epidermal thickness, and infiltration of inflammatory cells (e.g., mast cells and eosinophils). In addition, F.NONI treatment reduced the levels of histamine, IgE and IgG1/IgG2a ratio, thymus and activation regulated chemokine (TARC), and thymic stromal lymphopoietin (TSLP) in serum and beneficially modulated the expressions of Th1, Th2, Th17, and Th22-mediated cytokines in lesioned skin and splenocytes. Furthermore, the expressions of the skin barrier-related proteins including filaggrin (FLG), loricrin (LOR), involucrin (IVL), zonula occludens-1 (ZO-1), and occludin (OCC) were restored by F.NONI treatment. Taken together, these results suggest that F.NONI could be a therapeutic agent to attenuate AD-like skin lesions through modulating the immune balance and skin barrier function.

CD13-specific ligand facilitates Xanthatin nanomedicine targeting dendritic cells for therapy of refractory allergic rhinitis.

Relapse in Allergic Rhinitis (AR) is triggered by various unclear mechanisms. Xanthium strumarium L. as a traditional folk medicine can inhibit inflammatory responses through multiple mechanisms. Xanthatin (XT) is a bioactive compound derived from Xanthium strumarium L, and we developed a polymeric micelle (PM) that is dendritic cells (DCs)-specific targeting delivery system loading XT (NGR-XT-PM) based on a cyclic peptide moiety (NGR) to render DCs maturation-resistant for therapy of refractory AR. A murine model of AR was employed to investigate the in vivo therapeutic efficiency and relapse rate compared with the commercial product Budesonide. The results showed intranasal administration of NGR-XT-PM presented significant anti-allergy effect with no recurrence, in contrast, all mice treatment with Budesonide relapsed. NGR-XT-PM could effectively reverse the Th1/Th2 imbalance by depleting the serum inflammatory levels (IgE, histamine and IL-4) and DCs surface costimulatory molecules (CD80, CD86 and I-A/I-E), and promote immune tolerance by upregulating the level of Treg cells and reducing the levels of Th2, Th9 and Th17 cells. Furthermore, we appealed to virtual screening of inflammatory targets and found XT blocking the COX-2/PGE2 signaling pathway, which is a key effector in immune responses. These indicated CD13-specific NGR could facilitate XT selectively targeting DCs for efficiently ameliorating refractory rhinitis, and NGR-XT-PM should be a potential anti-AR drug.

Ruxolitinib Cream Has Dual Efficacy on Pruritus and Inflammation in Experimental Dermatitis.

The goal of this study was to elucidate the anti-pruritic and anti-inflammatory efficacy of ruxolitinib cream in experimentally-induced dermatitis. Atopic dermatitis (AD), the most common chronic relapsing inflammatory skin disease, significantly impairs patients' quality of life, with pruritus being a common complaint. The sensation of itch results from the interplay between epidermal barrier dysfunction, upregulated immune signaling and the activation of the central nervous system. The Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway plays a central role in pro-inflammatory cytokine signaling in AD. Ruxolitinib cream is a potent and selective JAK1/2 inhibitor currently undergoing clinical evaluation in adults with mild-to-moderate AD (NCT03745638, NCT03920852 and NCT03745651). The efficacy of ruxolitinib cream was tested in murine models of acute and chronic dermatitis and was also characterized in an ex vivo human skin dermatitis model. Ruxolitinib cream was highly effective at ameliorating disease symptoms in multiple murine dermatitis models through downregulation of T helper (Th)2-driven inflammation, resulting in reduced skin thickening and decreased itch. Pathway analysis of mouse ear tissue and human skin explants underscored the role for ruxolitinib in ameliorating inflammation and reducing itch via modulation of the JAK-STAT pathway. Together, the data offer a strong rationale for the use of ruxolitinib cream as a potent therapeutic agent for the clinical management of atopic dermatitis.

Clematichinenoside AR ameliorated spontaneous colitis in Il-10-/- mice associated with improving the intestinal barrier function and abnormal immune responses.

OBJECTIVES: Clematichinenoside AR (AR) is a saponin extracted for traditional Chinese medicine with the effects of improving the expression of tight junction (TJ) proteins and mediating anti-inflammatory activities. However, its effect on Crohn's disease (CD) is still unknown. We aimed to investigate the impact of AR on CD-like colitis and determine the mechanism underlying its effects. METHODS: Interleukin-10 gene knockout (Il-10-/-) mice (male, fifteen weeks old) with spontaneous colitis were allocated to the positive control and AR-treated (32 mg/kg AR administered every other day by gavage for 4 weeks) groups. Wild-type (WT) mice (male, fifteen weeks old) composed the negative control group. The effects of AR on intestinal barrier function and structure and T cell responses as well as the potential mechanisms underlying these effects were investigated. RESULTS: AR treatment significantly improved spontaneous colitis in Il-10-/- mice as demonstrated by reductions in the inflammatory score, disease activity index (DAI) and levels of inflammatory factors. The effects of AR on colitis in Il-10-/- mice were related to protecting intestinal barrier function and maintaining immune system homeostasis (regulatory T cell (Treg)/T helper 17 (Th17) cell balance). The anticolitis effect of AR may partly act by downregulating PI3K/Akt signaling. CONCLUSIONS: AR may have therapeutic potential for treating CD in humans.

Total Glucosides of Paeony protects against collagen-induced mouse arthritis via inhibiting follicular helper T cell differentiation.

BACKGROUND: The development of rheumatoid arthritis (RA) is related to germinal center (GC) response and autoreactive T cells, which mediate adaptive immunity and play an important role in stimulating the production of autoantibodies and pro-inflammatory cytokines by B cells and macrophages. Total Glucosides of Paeony (TGP) has anti-inflammatory, immunomodulatory and analgesic effects and is widely used to treat RA. However, few studies investigated whether the therapeutic effect of TGP is associated with the inhibition of autoimmune response. PURPOSE: The aim of this study was to investigate the effects and mechanisms of TGP on RA. STUDY DESIGN: Type II collagen-induced arthritis (CIA) mouse model was used, and TGP and paeoniflorin were intragastrically treated. METHODS: DBA/1 mice were divided into 5 groups: control, model, positive drug (paeoniflorin) and high- and low-dose TGP group. After 21 days of intragastric administration, the pathological change, inflammation expression and molecular mechanism of each group of mice were detected by Micro-CT, histochemical analysis, ELLSA, Western blot, RT-qPCR and flow cytometry. RESULTS: Our study found that TGP treatment effectively improved inflammation and joint destruction in CIA mice. It reduced the production of serum IgG2a and pro-inflammatory cytokines, including serum interleukin (IL)-21, tumor necrosis factor (TNF)-α and IL-6, and the phosphorylation of NF-κB p65 and STAT3 in a dose-dependent manner. More importantly, TGP could suppress the frequency of germinal center B cells and Tfh cells in the spleen. CONCLUSION: TGP can not only improve symptoms, but also inhibit bone destruction. The therapeutic effect of TGP on CIA is mainly achieved by inhibiting spleen Tfh cell differentiation and GC formation through STAT3 signaling pathway.

Dihydroartemisinin Regulates the Th/Treg Balance by Inducing Activated CD4+ T cell Apoptosis via Heme Oxygenase-1 Induction in Mouse Models of Inflammatory Bowel Disease.

Dihydroartemisinin (DHA) is a derivative of the herb Artemisia annua L. that has prominent immunomodulatory activity; however, its underlying mechanism remains elusive. Inflammatory bowel disease (IBD) is an idiopathic inflammatory condition characterized as an autoimmune disorder that includes dysfunctions in the T helper (Th)/T regulatory cell (Treg) balance, which normally plays pivotal roles in immune homeostasis. The aim of this study was to explore the potential of DHA to ameliorate IBD by restoring the Th/Treg cell balance. To this end, we established mouse models of colitis induced by oxazolone (OXA) and 2,4,6-trinitro-benzene sulfonic acid (TNBS). We then treated mice with DHA at 4, 8, or 16 mg/kg/day. DHA treatment ameliorated colitis signs and reduced lymphocyte infiltration and tissue fibrosis. Moreover, DHA decreased the numbers of Th1 and Th17 cells and Th9 and Th22 cells in TNBS- or OXA-induced colitis, respectively, and increased Tregs in both models. DHA (0.8 mg/mL) also inhibited activated CD4+ T lymphocytes, which was accompanied by apoptosis induction. Moreover, it promoted heme oxygenase-1 (HO-1) production in vitro and in vivo, concomitant with CD4+ T cell apoptosis and restoration of the Th/Treg balance, and these effects were blocked by treatment with the HO-1 inhibitor Sn-protoporphyrin IX. Overall, these results suggest that DHA is a novel and valuable candidate for IBD therapy or Th/Treg immunoregulation.

In vitro human helper T-cell assay to screen antibody drug candidates for immunogenicity.

Monoclonal antibody (mAb) drugs offer a number of valuable treatments. Many newly developed mAb drugs include artificial modification of amino acid sequences from human origin, which may cause higher immunogenicity to induce anti-drug antibodies (ADA). If the immunogenicity of a new candidate can be understood in the nonclinical phase, clinical studies will be safer and the success rate of development improved. Empirically, in vitro immunogenicity assays with human cells have proved to be sufficiently sensitive to nonhuman proteins, but not to human/humanized mAb. To detect the weaker immunogenicity of human-based mAb, a more sensitive biomarker for in vitro assays is needed. The in vitro study here developed a proliferation assay (TH cell assay) using flow cytometry analysis that can detect a slight increase in proliferating TH cells. Samples from 218 donors treated with a low-immunogenic drug (etanercept) were measured to determine a positive threshold level. With this threshold, positive donor percentages among PBMC after treatment with higher-immunogenicity mAb drugs were noted, that is, 39.5% with humanized anti-human A33 antibody (hA33), 27.3% with abciximab, 25.9% with adalimumab, and 14.8% with infliximab. Biotherapeutics with low immunogenicity yielded values of 0% for basiliximab and 3.7% for etanercept. These data showed a good comparability with previously reported incidences of clinical ADA with the evaluated drugs. Calculations based on the data here showed that a TH cell assay with 40 donors could provide statistically significant differences when comparing low- (etanercept) versus highly immunogenic mAb (except for infliximab). Based on the outcomes here, for screening purposes, a practical cutoff point of 3/20 positives with 20 donors was proposed to alert immunogenicity of mAb drug candidates.

Structural and diffusion weighted MRI demonstrates responses to ibrutinib in a mouse model of follicular helper (Tfh) T-cell lymphoma.

Recent analyses of the genetics of peripheral T-cell lymphoma (PTCL) have shown that a large proportion of cases are derived from normal follicular helper (Tfh) T-cells. The sanroque mouse strain bears a mutation that increases Tfh cell number and heterozygous animals (Roquinsan/+) develop lymphomas similar to human Tfh lymphoma. Here we demonstrate the usefulness of Roquinsan/+ animals as a pre-clinical model of Tfh lymphoma. Long latency of development and incomplete penetrance in this strain suggests the lymphomas are genetically diverse. We carried out preliminary genetic characterisation by whole exome sequencing and detected tumor specific mutations in Hsp90ab1, Ccnb3 and RhoA. Interleukin-2-inducible kinase (ITK) is expressed in Tfh lymphoma and is a potential therapeutic agent. A preclinical study of ibrutinib, a small molecule inhibitor of mouse and human ITK, in established lymphoma was carried out and showed lymphoma regression in 8/12 (67%) mice. Using T2-weighted MRI to assess lymph node volume and diffusion weighted MRI scanning as a measure of function, we showed that treatment increased mean apparent diffusion coefficient (ADC) suggesting cell death, and that change in ADC following treatment correlated with change in lymphoma volume. We suggest that heterozygous sanroque mice are a useful model of Tfh cell derived lymphomas in an immunocompetent animal.

A novel RORγt inhibitor is a potential therapeutic agent for the topical treatment of psoriasis with low risk of thymic aberrations.

BACKGROUND: Retinoic acid receptor-related orphan receptor gamma t (RORγt) has critical roles in the development, maintenance and function of interleukin (IL)-17-producing cells and is a highly attractive target for the treatment of IL-17-mediated autoimmune disease, particularly psoriasis. On the other hand, RORγt is also critical for controlling apoptosis during thymopoiesis, and genetic RORγt ablation or systematic RORγt inhibition cause progressive thymic aberrations leading to T cell lymphomas. OBJECTIVE: We investigated whether topical administration of our novel RORγt inhibitor, S18-000003 has therapeutic potential for psoriasis with low risk of thymic aberrations. METHODS: We evaluated the effect of topical S18-000003 on psoriasis-like skin inflammation and influence on the thymus in a 12-O-tetradecanoylphorbol-13-acetate-induced K14.Stat3C mouse psoriasis model. RESULTS: S18-000003 markedly inhibited the development of psoriatic skin inflammation via suppression of the IL-17 pathway. In the skin, S18-000003 suppressed all subsets of IL-17-producing cells that we previously identified in this psoriasis model: Th17 cells, Tc17 cells, dermal γδ T cells, TCR- cells that probably included innate lymphoid cells, and CD4-CD8- double-negative αß T cells. Notably, neither reduction of CD4+CD8+ double-positive thymocytes nor dysregulation of cell cycling was observed in S18-000003-treated mice, even at a high dose. CONCLUSION: Our topically administered RORγt inhibitor is a potential therapeutic agent for psoriasis with low risk of thymic lymphoma.

Unaccounted risk of cardiovascular disease: the role of the microbiome in lipid metabolism.

PURPOSE OF REVIEW: Not all of the risk of cardiovascular disease can be explained by diet and genetics, and the human microbiome, which lies at the interface of these two factors, may help explain some of the unaccounted risk. This review examines some of the well established links between the microbiome and cardiovascular health, and proposes relatively unexplored associations. RECENT FINDINGS: Byproducts of microbial metabolism are associated with health and disease: Trimethylamine N oxide is associated with atherosclerosis; whereas short-chain fatty acids are associated with decreased inflammation and increased energy expenditure. More broadly, a large number of association studies have been conducted to explore the connections between bacterial taxa and metabolic syndrome. In contrast, the relationship between the microbiome and triglycerides levels remains poorly understood. SUMMARY: We suggest that deeper understanding of the molecular mechanisms that drive linkages between the microbiome and disease can be determined by replacing 16S rRNA gene sequencing with shotgun metagenomic sequencing or other functional approaches. Furthermore, to ensure translatability and reproducibility of research findings, a combination of multiple different complementary '-omic' approaches should be employed.

Inflammatory response under zinc deficiency is exacerbated by dysfunction of the T helper type 2 lymphocyte-M2 macrophage pathway.

Nutritional zinc deficiency leads to immune dysfunction and aggravates inflammation. However, the underlying mechanism remains unknown. In this study, the relationship between macrophage subtypes (M1 and M2) and helper T lymphocytes (Th1 and Th2) was investigated using the spleen from rats fed zinc-deficient or standard diet. In experiment I, 5-week-old male Sprague-Dawley rats were fed a zinc-deficient diet (without zinc additives) or a standard diet (containing 0·01% zinc) for 6 weeks. In experiment II, the rats were divided into four groups: one group was fed a standard diet for 6 weeks; two groups were fed zinc-deficient diets and were injected three times a week with either saline or interleukin-4 (IL-4) (zinc-deficient/IL-4 i.p.); a fourth group (zinc-deficient/standard) was fed a zinc-deficient diet for 6 weeks followed by a standard diet for 4 weeks. In experiment I; GATA-binding protein 3 (GATA-3) protein level, M2 macrophage, CD3+  CD8+ cells, and IL-4/IL-13-positive cells significantly decreased in the spleens of the zinc-deficient group. Additionally, IL-1ß and macrophage inflammatory protein-1α (MIP-1α) mRNA levels significantly increased in the splenic macrophages of the zinc-deficient group. In experiment II; M2 macrophages, CD3+  CD8+ cells, IL-4/IL-13-positive cells, and GATA-3 protein levels significantly increased in the spleens of the zinc-deficient/IL-4 i.p. and zinc-deficient/standard groups. Furthermore, IL-1ß and MIP-1α mRNA levels decreased in the splenic macrophages of the zinc-deficient/IL-4 i.p. and zinc-deficient/standard groups. Zinc deficiency-induced aggravated inflammation is related to Th2 lymphocytes and followed by the association with loss of GATA-3, IL-4 and anti-inflammatory M2 macrophages. Importantly, IL-4 injection or zinc supplementation can reverse the effects of zinc deficiency on immune function.