Bibenzyl-Phenylpropane Hybrids with Immunosuppressive Activities from Dendrobium Nobile Lindl.

Fifteen bibenyls and four fluorenones, including five new bibenzyl-phenylpropane hybrids, were isolated from the aerial part of Dendrobium nobile Lindl. Their structures were determined by spectroscopic methods. Bioassay on the LPS-induced proliferations of mouse splenic B lymphocytes, and Con A-induced T lymphocytes showed that compounds 1, 2, and 14 showed excellent immunosuppressive activities with IC50 values of 1.23, 1.01, and 3.87 µM, respectively, while compounds 3-4, 7, 10, 13, and 15 exhibited moderate immunosuppressive activities with IC50 values ranging from 6.89 to 14.2 µM.

Structurally diverse biflavonoids from the fruits of Citrus medica L. var. sarcodactylis Swingle and their hypolipidemic and immunosuppressive activities.

The fruit of Citrus medica L. var. sarcodactylis Swingle is not only used as a traditional medicinal plant, but also served as a delicious food. Six new (3'→7″)-biflavonoids (1-6), and twelve known biflavonoid derivatives (7-18) were isolated and characterized from the fruits of C. medica L. var. sarcodactylis Swingle for the first time. Their structures were determined by extensive and comprehensive analyzing NMR, HR-ESI-MS, UV, and IR spectral data coupled with the data described in the literature. Compounds (1-18) were evaluated for their hypolipidemic activities with Orlistat as the positive control, and assayed for their immunosuppressive activities with Dexamethasone as the positive control, respectively. Among them, compounds (1-3) exhibited moderate inhibition of pancreatic lipase activity by inhibiting 68.56 ± 1.40%, 56.18 ± 1.57%, 53.51 ± 1.59% of pancreatic lipase activities at the concentration of 100 µM, respectively. Compounds (4-6) and 8 showed potent immunosuppressive activities with the IC50 values from 16.83 ± 1.32 to 50.90 ± 1.79 µM. The plausible biogenetic pathway and preliminary structure activity relationship of the selected compounds were scientifically summarized and discussed in this study.

Formononetin protects against concanavalin-A-induced autoimmune hepatitis in mice through its anti-apoptotic and anti-inflammatory properties.

Autoimmune hepatitis (AIH) is a chronic inflammatory liver disease that seriously threatens the health of humans globally. Formononetin (FMN) is a natural herb extract with multiple biological functions. In this study, an experimental model of AIH was established in mice through the use of concanavalin A (ConA). To investigate the effects of FMN on ConA-induced hepatitis, the mice were pretreated with 50 or 100 mg/kg body mass of FMN. The results show that FMN alleviated ConA-induced liver injury of mice in a dose-dependent manner. Moreover, pretreatment with FMN inhibited the apoptosis of hepatocytes in the ConA-treated mice through downregulating the expression of pro-apoptotic proteins (Bax, cleaved caspase 9, and cleaved caspase 3) and upregulating the expression of anti-apoptotic protein (Bcl-2). It was also found that the levels of proinflammatory cytokines were greatly reduced in the serum and liver tissues of mice pretreated with FMN. Further studies showed that FMN reduced the level of phosphorylated nuclear factor kappa B (p-NF-κB) p65 and enhanced the level of IκBα (inhibitor of NF-κB), suggesting that FMN inhibits the activation of the NF-κB signaling pathway. In addition, FMN inhibited activation of the NOD-like receptor protein 3 (NLRP3) inflammasome. Therefore, FMN could be a promising agent for the treatment of AIH.

4,21-Secovincanol, a Novel Immunosuppressive Monoterpenoid Indole Alkaloid from Kopsia Hainanensis.

4,21-Secovincanol (1), a novel C-21/N-4 cleavage monoterpenoid indole alkaloid, along with four analogs (2-5), were obtained from the aerial parts of Kopsia hainanensis. Structurally, compound 1 might be a derivative of epivincanol (2) via C-21/N-4 cleavage. Their structures were confirmed by means of comprehensive spectroscopic data analysis and comparison with the reported data. All isolates significantly inhibited Con A-stimulated mice splenocytes proliferation at 10-40 µM in a dose-dependent manner in vitro. Especially, compound 3 exhibited potent activities comparable to positive control (Dexamethasone, DXM).

Ergocytochalasin A, a polycyclic merocytochalasan from an endophytic fungus Phoma multirostrata XJ-2-1.

Ergocytochalasin A (1), an unprecedented merocytochalasan formed via Diels-Alder cycloaddition of a cytochalasin with an ergosterol, was isolated from an endophytic fungus Phoma multirostrata XJ-2-1. Compound 1 possessed a unique 5/6/14/6/5/6/6/6 fused octacyclic ring system and its structure was established by detailed NMR and HRESIMS spectroscopic analyses. The absolute configuration of 1 was determined by single-crystal X-ray diffraction. A plausible biogenetic pathway of 1 was postulated. Compound 1 was evaluated for its cytotoxicity against six cancer cell lines and showed inhibitory effects with IC50 values ranging from 6.92 to 26.63 µM. The in vitro immunosuppressive activity of 1 against ConA-induced T cell and LPS-induced B cell proliferation, as well as its antiviral activity against Human dengue virus type 3 (DV3), influenza A virus (H1N1) and respiratory syncytial virus (RSV), was also evaluated. Ergocytochalasin A is the first example of a merocytochalasan which consists of one cytochalasin moiety and one ergosterol moiety. Containing eighteen chiral centers, ergocytochalasin A owns a folded framework, which makes it extremely compact in space.

Alopecines A-E, five chloro-containing matrine-type alkaloids with immunosuppressive activities from the seeds of Sophora alopecuroides.

Alopecines A-E (1-5), five unusual matrine-type alkaloids featuring with an additional dichlorocyclopropane (1-3) or a di/tri-chloromethyl (4/5) attached on the D ring, were isolated from the seeds of Sophora alopecuroides. Their structures and absolute configurations were elucidated by extensive spectroscopic techniques, and X-ray diffraction analyses or time-dependent density functional theory-based electronic circular dichroism (TDDFT-ECD) calculations. Alkaloid 4 exhibited potent inhibitory effects on the proliferation of ConA-induced T lymphocytes or LPS-induced B cells with IC50 value of 3.98 or 3.74 µM, respectively.

Diversiform and Transformable Glyco-Nanostructures Constructed from Amphiphilic Supramolecular Metallocarbohydrates through Hierarchical Self-Assembly: The Balance between Metallacycles and Saccharides.

During the past decade, self-assembly of saccharide-containing amphiphilic molecules toward bioinspired functional glycomaterials has attracted continuous attention due to their various applications in fundamental and practical areas. However, it still remains a great challenge to prepare hierarchical glycoassemblies with controllable and diversiform structures because of the complexity of saccharide structures and carbohydrate-carbohydrate interactions. Herein, through hierarchical self-assembly of modulated amphiphilic supramolecular metallocarbohydrates, we successfully prepared various well-defined glyco-nanostructures in aqueous solution, including vesicles, solid spheres, and opened vesicles depending on the molecular structures of metallocarbohydrates. More attractively, these glyco-nanostructures can further transform into other morphological structures in aqueous solutions such as worm-like micelles, tubules, and even tupanvirus-like vesicles (TVVs). It is worth mentioning that distinctive anisotropic structures including the opened vesicles (OVs) and TVVs were rarely reported in glycobased nano-objects. This intriguing diversity was mainly controlled by the subtle structural trade-off of the two major components of the amphiphiles, i.e., the saccharides and metallacycles. To further understand this precise structural control, molecular simulations provided deep physical insights on the morphology evolution and balancing of the contributions from saccharides and metallacycles. Moreover, the multivalency of glyco-nanostructures with different shapes and sizes was demonstrated by agglutination with a diversity of sugar-binding protein receptors such as the plant lectins Concanavalin A (ConA). This modular synthesis strategy provides access to systematic tuning of molecular structure and self-assembled architecture, which undoubtedly will broaden our horizons on the controllable fabrication of biomimetic glycomaterials such as biological membranes and supramolecular lectin inhibitors.

Epigynumgenane-type pregnane glycosides from Epigynum cochinchinensis and their immunosuppressive activity.

Five undescribed C21 pregnane glycosides, epigycosides D-H, together with four known analogues, two lignans, and a flavonoid have been isolated from the stems of Epigynum cochinchinensis. The structures of pregnane glycosides were elucidated using spectroscopic techniques and acid hydrolysis. The in vitro immunological activities were assessed against Con A-stimulated proliferation of mice splenocytes. The C21 pregnane glycosides showed immunosuppressive activity in a concentration-dependent manner. Moreover, epigycoside E exhibited a potent immunosuppressive effect, and the IC50 value on Con A-stimulated mice splenocytes was 22.1 ±â€¯6.4 µM. Epigycoside E also caused G0/G1 arrest, and inhibited TNF-α and IL-2 production.

Synthesis and biological evaluation of (1,2,4)triazole[4,3-a]pyridine derivatives as potential therapeutic agents for concanavalin A-induced hepatitis.

A series of (1,2,4)triazole[4,3-a]pyridine (TZP) derivatives have been designed and synthesized. Compound 8d was identified as having the most potent inhibitory activity on NO release in response to lipopolysaccharide (LPS) stimulation and inhibition of the migration induced by MCP-1 protein on RAW264.7 macrophages. Based on the screening data, an immunofluorescence assay and a real-time qPCR assay were conducted, indicating that compound 8d suppressed NF-κB p65 translocation and expression of inflammatory genes by concanavalin A (Con A)-induced RAW264.7 macrophages. More importantly, 8d also exhibited potent efficacy, alleviating Con A-induced hepatitis by downregulating the levels of plasma alanine transaminase (ALT), aspartate transaminase (AST) and inflammatory infiltration in a mouse autoimmune hepatitis (AIH) model. In addition, the flow cytometry (FCM) data showed that compound 8d inhibited the accumulation of MDSCs in the liver of Con A-induced mice. These findings raise the possibility that compound 8d might serve as a potential agent for the treatment of AIH.

Highly diverse cembranoids from the South China Sea soft coral Sinularia scabra as a new class of potential immunosuppressive agents.

The first and in-depth chemical investigation of the South China Sea soft coral Sinularia scabra has resulted to the isolation of a library of diverse cembrane type diterpenoids, including six new compounds, namely xiguscabrates A and B (1 and 2), xiguscabral A (3), xiguscabrols A and B (4 and 5), and 8-epi-xiguscabrol B (6), and twenty-seven known analogs (7-33). Their structures were elucidated by extensive spectroscopic analysis and by the comparison with literature data. In bioassay, several isolates exhibited inhibitory effects on the ConA-induced T lymphocytes and/or LPS-induced B lymphocytes proliferation. Among them, compound 24 showed considerable specific inhibition on B cell proliferation, with IC50 value of 4.4 µM and selectivity index (SI) of 10.9. The structure-activity relationship (SAR) of the tested metabolites was analyzed, and the further mechanism study of the specific B-cell targeted immunosuppressive compound 24 on purified CD19+ B cells was also performed to uncover the effects on the function and maturity of B cells, including cytokines production, abnormal activation, antigen presenting capacity and plasma cells formation.

Selenium-biofortified corn peptides: Attenuating concanavalin A-Induced liver injury and structure characterization.

The relationship between hepatoprotective effects of selenium-biofortified corn (Zea mays Linn) peptides (SeCPs) and its antioxidant ability was evaluated and the structure of SeCPs was identified. SeCPs and corn peptides (CPs) both had good antioxidant ability, and the effect of SeCPs was significantly higher than CPs within a certain concentration range (P < 0.05). Additionally, animal experiments indicated that SeCPs (200 mg/kg) had a significantly protective effect against concanavalin A (Con A) induced hepatic lesions, as it significantly declined glutamic-pyruvic transaminase (AST), alanine transaminase (ALT) activities, tumor necrosis factor alpha (TNF-α), interferon (IFN)-γ contents in serum, and malondialdehyde (MDA) contents in liver (P < 0.05). Superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in liver were also significantly increased by SeCPs (P < 0.05). The amino acid composition of SeCPs with Mw < 1 kDa was mainly glutamic acid (Glu, 31.18%), leucine (Leu, 21.06%) and alanine (Ala, 13.26%). According to the retention time, the amino acid sequences of 8 selenium-biofortified corn peptides and 29 selenium-free corn peptides were identified. Our results illustrated that the mechanisms of SeCPs against Con A induced hepatic injury in mice may be related to its antioxidant ability and reduction of lipid peroxidation, inhibiting the release of immune factors, such as TNF-α and IFN-γ.

Direct Evaluation of Live Uropathogenic Escherichia coli Adhesion and Efficiency of Antiadhesive Compounds Using a Simple Microarray Approach.

Many pathogens use host glycans as docking points for adhesion. Therefore, the use of compounds blocking carbohydrate-binding adhesins is a promising strategy for fighting infections. In this work, we describe a simple and rapid microarray approach for assessing the bacterial adhesion and efficiency of antiadhesive compounds targeting uropathogenic Escherichia coli UTI89, which displays mannose-specific adhesin FimH at the tip of fimbriae. The approach consisted in direct detection of live fluorescently labeled bacteria bound to mannan printed onto microarray slides. The utility of the arrays for binding/inhibition assays was first validated by comparing array-derived results for the model mannose-binding lectin concanavalin A with data obtained by isothermal titration calorimetry. Growth phase-dependent binding of UTI89 to the arrays was observed, proving the usefulness of the setup for detecting differences in FimH expression. Importantly, bacteria labeling and binding assays entailed minimal manipulation, helping to preserve the integrity of fimbriae. The efficiency of three different dodecamannosylated fullerenes as FimH-targeted antiadhesives was next evaluated in competition assays. The results revealed a superior activity of the mannofullerenes (5- to 18-fold per mannose residue) over methyl α-d-mannopyranoside. Moreover, differences in activity were detected for mannofullerenes differing in the structure/length of the spacer used for grafting mannose onto the fullerene core, further demonstrating the sensitivity of the assay. Overall, the approach combines straightforward and time-saving protocols for microarray preparation, bacteria labeling, and binding assays, and it can be easily tailored to other bacteria bearing carbohydrate-binding adhesins.

Salecan protected against concanavalin A-induced acute liver injury by modulating T cell immune responses and NMR-based metabolic profiles.

Salecan, a water-soluble extracellular ß-glucan produced by Agrobacterium sp. ZX09, has been reported to exhibit a wide range of biological effects. The aims of the present study were to investigate the protective effect of salecan against Concanavalin A (ConA)-induced hepatitis, a well-established animal model of immune-mediated liver injury, and to search for possible mechanisms. C57BL/6 mice were pretreated with salecan followed by ConA injection. Salecan treatment significantly reduced ConA-induced acute liver injury, and suppressed the expression and secretion of inflammatory cytokines including interferon (IFN)-γ, interleukin (IL)-6 and IL-1ß in ConA-induced liver injury model. The high expression levels of chemokines and adhesion molecules such as MIP-1α, MIP-1ß, ICAM-1, MCP-1 and RANTES in the liver induced by ConA were also down-regulated after salecan treatment. Salecan inhibited the infiltration and activation of inflammatory cells, especially T cells, in the liver induced by ConA. Moreover, salecan reversed the metabolic profiles of ConA-treated mice towards the control group by partly recovering the metabolic perturbations induced by ConA. Our results suggest the preventive and therapeutic potential of salecan in immune-mediated hepatitis.

Sixteen novel C-21 steroidal glycosides from the roots of Cynanchum mooreanum.

Fourteen novel 14,15-diseco-18-nor-pregnane-type steroidal glycosides, mooreanoside A-C (1-3) and mooreanoside F-P (6-16) and two new 14,15-diseco-pregnane-type steroidal glycosides, mooreanoside D-E (4-5) were isolated from the roots of Cynanchum mooreanum Hemsl. Their structures were determined on the basis of spectroscopic and chemical analysis. Compounds 1-6, 8-10, 12-14 and 16 were tested for their immunological activities in vitro against Con-A induced proliferation of mice splenocytes.

Phainanoids A-F, a new class of potent immunosuppressive triterpenoids with an unprecedented carbon skeleton from Phyllanthus hainanensis.

Phainanoids A-F (1-6), six highly modified triterpenoids with a new carbon skeleton by incorporating two unique motifs of a 4,5- and a 5,5-spirocyclic systems, were isolated from Phyllanthus hainanensis. Their structures with absolute configurations were determined by spectroscopic data, chemical methods, and X-ray crystallography. Compounds 1-6 exhibited exceptionally potent immunosuppressive activities in vitro against the proliferation of T and B lymphocytes. The most potent one, phainanoid F (6), showed activities against the proliferation of T cells with IC50 value of 2.04 ± 0.01 nM (positive control CsA = 14.21 ± 0.01 nM) and B cells with IC50 value of <1.60 ± 0.01 nM (CsA = 352.87 ± 0.01 nM), which is about 7 and 221 times as active as CsA, respectively. The structure-activity relationships of 1-6 are discussed.

Hydrogen sulfide, a potential novel drug, attenuates concanavalin A-induced hepatitis.

BACKGROUND: Hydrogen sulfide (H2S) is known to exert anti-inflammatory properties. Apoptosis and autophagy play important roles in concanavalin A (Con A)-induced acute hepatitis. The purpose of this study was to explore both the effect and mechanism of H2S on Con A-induced acute hepatitis. METHODS: BALB/c mice were randomized into sham group, Con A-injection group, and 14 µmol/kg of sodium hydrosulfide (NaHS, an H2S donor) pretreatment group. RESULTS: Aspartate aminotransferase, alanine aminotransferase, and pathological damage were significantly ameliorated by NaHS pretreatment. NaHS pretreatment significantly reduced the levels of interleukin-6 and tumor necrosis factor-α compared with those of the Con A group. The expression of Bcl-2, Bax, Beclin-1, and LC3-2, which play important roles in the apoptosis and autophagy pathways, were also clearly affected by NaHS. Furthermore, NaHS affected the p-mTOR and p-AKT. CONCLUSION: H2S attenuates Con A-induced acute hepatitis by inhibiting apoptosis and autophagy, in part, through activation of the PtdIns3K-AKT1 signaling pathway.

Discovery of structurally simplified analogs of colchicine as an immunosuppressant.

We have discovered a new class of colchicine-derived therapeutic agents for immune diseases including rejection of organ-transplantation and autoimmune disease. Compound 2, which had been developed to overcome poor pharmacokinetic properties of compound 1, a first-generation colchicine analog, turned out to show toxicity such as intestinal toxicity and loss of weight during in vivo tests. The deletion of 7-carboxamide group and middle ring-truncation in colchicine allowed us to have structurally simplified analogs with strong immunosuppressive activity. Herein, we report non-alkaloid tricyclic compound 7 and 12 as immunosuppressants which exhibited a strong immunosuppressive in vivo efficacy on the T-dependent antibody response, the Zymosan A-induced arthritis model and the Carrageenan-induced edema model. Compound 7 and 12 revealed less toxicity than the previous lead compound 2, and their minimum lethal doses (MLD) were proved to exceed 100 mg/kg.

Discovery of a new class of immunosuppressants from Trichothecium roseum co-inspired by cross-kingdom similarity in innate immunity and pharmacophore motif.

The limited selection of immunosuppressants in the clinic hampers the efficient management of immune disorders such as rejections after organ transplantations. However, the search for new immunosuppressive compounds remains random and creates inevitably financial and laborious wastes. Herein, we present an immunity-inspired discovery strategy that rationally allows an efficient identification of immunosuppressive compounds from the endophyte culture, as exemplified by the new peptide trichomide A. This compound exerts its immunosuppressive action more selectively than cyclosporin A. It was found that trichomide A decreases the expression of Bcl-2, increases the expression of Bax, and has a small or negligible effect on the expressions of p-Akt, CD25, and CD69. Our study strengthens the idea that the cross-kingdom similarity in immunity among living things could provide a shorter route towards the identification of natural products valuable for the development of new immunosuppressants.

Vγ4 γδ T cell-derived IL-17A negatively regulates NKT cell function in Con A-induced fulminant hepatitis.

Con A-induced fulminant hepatitis is a well-known animal model for acute liver failure. However, the role of γδ T cells in this model is undefined. In this report, using TCR δ(-/-) mice, we demonstrated a protective role of γδ T cells in Con A-induced hepatitis model. TCR δ(-/-) mice showed significantly decreased levels of IL-17A and IL-17F in the Con A-treated liver tissue, and reconstitution of TCR δ(-/-) mice with wild-type (Wt), but not IL-17A(-/-), γδ T cells significantly reduced hepatitis, strongly suggesting a critical role of IL-17A in mediating the protective effect of γδ T cells. Interestingly, only Vγ4, but not Vγ1, γδ T cells exerted such a protective effect. Furthermore, depletion of NKT cells in TCR δ(-/-) mice completely abolished hepatitis, and NKT cells from Con A-challenged liver tissues of TCR δ(-/-) mice expressed significantly higher amounts of proinflammatory cytokine IFN-γ than those from Wt mice, indicating that γδ T cells protected hepatitis through targeting NKT cells. Finally, abnormal capacity of IFN-γ production by NKT cells of TCR δ(-/-) mice could only be downregulated by transferring Wt, but not IL-17(-/-), Vγ4 γδ T cells, confirming an essential role of Vγ4-derived IL-17A in regulating the function of NKT cells. In summary, our report thus demonstrated a novel function of Vγ4 γδ T cells in mediating a protective effect against Con A-induced fulminant hepatitis through negatively regulating function of NKT cells in an IL-17A-dependent manner, and transferring Vγ4 γδ T cells may provide a novel therapeutic approach for this devastating liver disease.

Role of immunoregulatory transcription factors in differential immunomodulatory effects of tocotrienols.

Tocotrienols have been shown to possess antioxidant, antitumor, cardioprotective, and antiproliferative effects. This report describes novel immunomodulatory effects of tocotrienols in murine lymphocytes. γ-Tocotrienol (GT) was more effective in suppressing concanavalin A (Con A)-induced T cell proliferation and cytokine production compared to α-tocotrienol (AT) when present continuously in the culture. GT inhibited T cell activation markers and costimulatory molecule. GT modulated intracellular glutathione in lymphocytes, and the suppressive effects of GT could not be abrogated by thiol or nonthiol antioxidants, indicating a poor link between anti-inflammatory properties of tocotrienols and cellular redox status. It was also observed that GT suppressed Con A-induced activation of NF-κB, AP-1, and NF-κB-dependent gene expression. Cellular uptake studies with tocotrienols showed higher accumulation of GT compared to AT. Similar immunosuppressive effects of GT were also observed when administered to mice. In contrast, transient exposure of lymphocytes to GT (4 h) resulted in higher survival and proliferation of lymphocytes in vitro and in vivo in syngeneic and allogeneic hosts. This was attributed to the ability of GT to induce NF-κB, AP-1, and mTOR activation in lymphocytes upon transient exposure. Our results demonstrated that antioxidants such as tocotrienols may exhibit pleiotropic effects by activating multiple mechanisms in cells.