Dissolvable microneedles based on Panax notoginseng polysaccharide for transdermal drug delivery and skin dendritic cell activation.

This work explored the feasibility of using biological polysaccharide to fabricate dissolvable microneedles (MNs) for the purpose of transdermal drug delivery and skin dendritic cell (DC) activation. Panax notoginseng polysaccharide (PNPS), a naturally derived immunoactive macromolecule, was used to fabricate dissolvable MNs. The prepared PNPS MNs showed a satisfactory mechanical strength and a skin penetration depth. By Franz diffusion cell assay, the PNPS MNs demonstrated a high transdermal delivery amount of model drugs. Furthermore, with the assistance of MNs, PNPS easily penetrated across the stratum corneum and target ear skin DCs, activating the maturation and migration of immunocytes by increasing the expressions of CD40, CD80, CD86, and MHC II of skin DCs. Consequently, the matured DCs migrated to the auricular draining lymph nodes and increased the proportions of CD4+ T and CD8+ T cells. Thus, PNPS might be a promising biomaterial for transdermal drug delivery, with adjuvant potential.

Adjuvant herbal therapy for targeting susceptibility genes to Kawasaki disease: An overview of epidemiology, pathogenesis, diagnosis and pharmacological treatment of Kawasaki disease.

BACKGROUND: Kawasaki disease (KD) is a self-limiting acute systemic vasculitis occur mainly in infants and young children under 5 years old. Although the use of acetylsalicylic acid (AAS) in combination with intravenous immunoglobulin (IVIG) remains the standard therapy to KD, the etiology, genetic susceptibility genes and pathogenic factors of KD are still un-elucidated. PURPOSE: Current obstacles in the treatment of KD include the lack of standard clinical and genetic markers for early diagnosis, possible severe side effect of AAS (Reye's syndrome), and the refractory KD cases with resistance to IVIG therapy, therefore, this review has focused on introducing the current advances in the identification of genetic susceptibility genes, environmental factors, diagnostic markers and adjuvant pharmacological intervention for KD. RESULTS: With an overall update in the development of KD from different aspects, our current bioinformatics data has suggested CASP3, CD40 and TLR4 as the possible pathogenic factors or diagnostic markers of KD. Besides, a list of herbal medicines which may work as the adjunct therapy for KD via targeting different proposed molecular targets of KD have also been summarized. CONCLUSION: With the aid of modern pharmacological research and technology, it is anticipated that novel therapeutic remedies, especially active herbal chemicals targeting precise clinical markers of KD could be developed for accurate diagnosis and treatment of the disease.

Yin Yang-1 suppresses CD40 ligand-CD40 signaling-mediated anti-inflammatory cytokine interleukin-10 expression in pulmonary adventitial fibroblasts by promoting histone H3 tri-methylation at lysine 27 modification on interleukin-10 promoter.

During the pathogenesis of early pulmonary arterial hypertension (PAH), pulmonary arterial adventitial fibroblast act as an initiator and mediator of inflammatory processes that predispose vessel walls to excessive vasoconstriction and pathogenic vascular remodeling. Emerging studies report that Yin Yang-1 (YY-1) plays important roles in inflammatory response and vascular injury. Our recent study finds that activation of CD40 ligand (CD40L)-CD40 signaling promotes pro-inflammatory phenotype of pulmonary adventitial fibroblasts. However, whether YY-1 is involved in CD40L-CD40 signaling-triggered inflammatory response in pulmonary adventitial fibroblasts and its underlying mechanism is still unclear. Here, we show that soluble CD40L (sCD40L) stimulation promotes YY-1 protein expression and suppresses anti-inflammatory cytokine, interleukin 10 (IL-10) expression in pulmonary adventitial fibroblasts, while YY-1 knockdown prevents sCD40L-mediated reduction of IL-10 expression via enhancing IL-10 gene transactivation. Further, we find that sCD40L stimulation significantly increases histone H3 tri-methylation at lysine 27 (H3K27me3) modification on IL-10 promoter in pulmonary adventitial fibroblasts, and YY-1 knockdown prevents the effect of sCD40L on IL-10 promoter by reducing the interaction with enhancer of zeste homolog 2 (EZH2), a histone methyltransferase, binding to IL-10 promoter. Moreover, we find that sCD40L stimulation promotes YY-1 protein, but not messenger RNA (mRNA) expression, via decreasing N6-methyladenosine methylation on YY-1 mRNA to suppress YTHDF2-medicated mRNA decay. Overall, this in-depth study shows that the activation of CD40L-CD40 signaling upregulates YY-1 protein expression in pulmonary adventitial fibroblasts, which results in increasing YY-1 and EZH2 binding to the IL-10 promoter region to enhance H3K27me3 modification, eventually leading to suppression of IL-10 transactivation. This study first uncovers the roles of YY-1 on CD40L-CD40 signaling-triggered inflammatory response in pulmonary adventitial fibroblasts.

Ras isoforms: signaling specificities in CD40 pathway.

BACKGROUND: Ras are small cellular GTPases which regulate diverse cellular processes. It has three isoforms: H-Ras, K-Ras, and N-Ras. Owing to the N-terminus (1-165 residues) sequence homology these isoforms were thought to be functionally redundant. However, only K-Ras-deficient mice but not H-Ras- and N-Ras-deficient mice show embryonic lethality. Similarly, mutations in a given Ras isoform are associated with a particular type of cancer. Moreover, we have previously reported that Ras isoforms perform unique functions in Leishmania major infection. Thus, Ras isoforms are implicated to have signaling and functional specificity but the mechanism remains to be elucidated. RESULT: Using CD40 as a model receptor, we showed that depending on the strength of signaling, specific Ras isoforms are activated. Weak CD40 signal activates N-Ras, whereas strong signal activates H-Ras and K-Ras. Additionally, we showed that suppression of N-Ras expression reduced CD40-induced extracellular signal-regulated kinase-1/2 (ERK-1/2) activation and Interleukin (IL)-10 production; whereas suppression of H-Ras or K-Ras reduced CD40-induced p38 mitogen-activated protein kinase (p38MAPK) activation and IL-12 production. Furthermore, we showed that Ras isoforms have activator (GEF) specificity as weak CD40 signal-activated N-Ras requires Sos-1/2 whereas strong CD40 signal-activated H-Ras/K-Ras requires Ras-GRP as the guanine-nucleotide exchange factor (GEF) inducing ERK-1/2- or p38MAPK-mediated IL-10 or IL-12 productions, respectively, in macrophages. Silencing of syk reduced CD40-induced N-Ras activation but silencing of lyn inhibited H-Ras and K-Ras activation. In CD40 signaling, Ras isoforms also showed effector specificity; while H-Ras and K-Ras showed specificity for phosphatidyl inositol-3 kinase activation at high dose of CD40 stimulation, N-Ras primarily associated with Raf-1 at low dose of CD40 stimulation. Moreover, fractal analysis showed that functional site surface roughness for H-Ras (SurfaceFD = 2.39) and K-Ras (SurfaceFD = 2.39) are similar but significantly different from N-Ras (SurfaceFD = 2.25). CONCLUSION: The activator and effector specificities of Ras isoforms in CD40 signaling indicated their differential involvement in CD40 pathway and in maintaining the reciprocity. Our observations reveal Ras-regulated signaling outcome and its potential for developing Ras isoform-targeted immunotherapy and prophylaxis.

A CD40 targeting peptide prevents severe symptoms in experimental autoimmune encephalomyelitis.

CD40/CD154-interaction is critical in the development of Experimental Autoimmune Encephalomyelitis (EAE; mouse model of Multiple Sclerosis). Culprit CD4+CD40+ T cells drive a more severe form of EAE than conventional CD4 T cells. Blocking CD40/CD154-interaction with CD154-antibody prevents or ameliorates disease but had thrombotic complications in clinical trials. We targeted CD40 using a CD154-sequence based peptide. Peptides in human therapeutics demonstrate good safety. A small peptide, KGYY6, ameliorates EAE when given as pretreatment or at first symptoms. KGYY6 binds Th40 and memory T cells, affecting expression of CD69 and IL-10 in the CD4 T cell compartment, ultimately hampering disease development.

Cytotoxic effect caspase activation dependent of a genetically engineered fusion protein with a CD154 peptide mimetic (OmpC-CD154p) on B-NHL cell lines is mediated by the inhibition of bcl-6 and YY1 through MAPK p38 activation.

The interaction between CD40, and its ligand, CD154, is essential for the development of humoral and cellular immune responses. The selective inhibition or activation of this pathway forms the basis for the development of new therapeutics against immunologically based diseases and malignancies. We are developing a gene fusion of Salmonella typhi OmpC protein expressing the CD154 Tyr140-Ser-149 amino acid strand. This OmpC-CD154 binds CD40 and activates B cells. In this study, we demonstrate that OmpC-CD154p treatment inhibits cell growth, proliferation and induced apoptosis in the B-NHL cell lines Raji and Ramos. The Bcl-2 family proteins were regulated and the Bcl-6 and YY1 oncoproteins were inhibited. p38 MAPK activation is an important mechanism underlying the effect on proliferation and apoptosis mediated by this fusion protein. This study establishes a basis for the possible use of fusion protein OmpC-CD154 as an alternative treatment for B-NHL.

Radiation and Local Anti-CD40 Generate an Effective in situ Vaccine in Preclinical Models of Pancreatic Cancer.

Radiation therapy induces immunogenic cell death, which can theoretically stimulate T cell priming and induction of tumor-specific memory T cell responses, serving as an in situ vaccine. In practice, this abscopal effect is rarely observed. We use two mouse models of pancreatic cancer to show that a single dose of stereotactic body radiation therapy (SBRT) synergizes with intratumoral injection of agonistic anti-CD40, resulting in regression of non-treated contralateral tumors and formation of long-term immunologic memory. Long-term survival was not observed when mice received multiple fractions of SBRT, or when TGFß blockade was combined with SBRT. SBRT and anti-CD40 was so effective at augmenting T cell priming, that memory CD8 T cell responses to both tumor and self-antigens were induced, resulting in vitiligo in long-term survivors.

Nonclinical Safety Assessment of CFZ533, a Fc-Silent Anti-CD40 Antibody, in Cynomolgus Monkeys.

CFZ533 is a pathway blocking, nondepleting anti-CD40 antibody that is in clinical development for inhibition of transplant organ rejection and therapy for autoimmune diseases. A 26-week GLP toxicity study in sexually mature Cynomolgus monkeys was conducted in order to support chronic application of CFZ533. CFZ533 was subcutaneously administered at doses up to 150 mg/kg/week and was safe and generally well tolerated. CFZ533 showed no adverse effects for cardiovascular, respiratory, and neurobehavioral endpoints, and no changes were observed for blood lymphocyte and platelet counts or blood coagulation markers. In line with the nondepleting nature of CFZ533, CD20+ B cells in the blood were only marginally reduced. A complete suppression of germinal center (GC) development in lymph nodes and spleen was the most prominent result of post-mortem histological investigations. This was corroborated by an abrogated T-dependent antibody response (TDAR) to the antigen Keyhole Limpet Hemocyanin (KLH) as well as an absence of anti-drug antibodies (ADAs) in the absence of B cell depletion as seen with immunophenotyping and histology. When serum levels of CFZ533 in recovery animals dropped levels necessary for full CD40 occupancy on B cells, all animals were able to mount a TDAR to KLH. All histological changes also reverted to normal appearance after recovery. In summary, CFZ533 was shown to be well tolerated and safe in the 26-week toxicity study with a distinct pharmacodynamic profile in histology and immune function.

Tespa1 Deficiency Dampens Thymus-Dependent B-Cell Activation and Attenuates Collagen-Induced Arthritis in Mice.

Thymocyte-expressed, positive selection-associated 1 (Tespa1) plays an important role in both T cell receptor (TCR)-driven thymocyte development and in the FcεRI-mediated activation of mast cells. Herein, we show that lack of Tespa1 does not impair B cell development but dampens the in vitro activation and proliferation of B cells induced by T cell-dependent (TD) antigens, significantly reduces serum antibody concentrations in vivo, and impairs germinal center formation in both aged and TD antigen-immunized mice. We also provide evidence that dysregulated signaling in Tespa1-deficient B cells may be linked to CD40-induced TRAF6 degradation, and subsequent effects on 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase gamma-2 (PLCγ2) phosphorylation, MAPK activation, and calcium influx. Furthermore, we demonstrate that Tespa1 plays a critical role in pathogenic B cells, since Tespa1-deficient chimeric mice showed a lower incidence and clinical disease severity of collagen-induced arthritis. Overall, our study demonstrates that Tespa1 is essential for TD B cell responses, and suggests an important role for Tespa1 during the development of autoimmune arthritis.

Activation and Induction of Antigen-Specific T Follicular Helper Cells Play a Critical Role in Live-Attenuated Influenza Vaccine-Induced Human Mucosal Anti-influenza Antibody Response.

There is increasing interest recently in developing intranasal vaccines against respiratory tract infections. The antibody response is critical for vaccine-induced protection, and T follicular helper cells (TFH) are considered important for mediating the antibody response. Most data supporting the role for TFH in the antibody response are from animal studies, and direct evidence from humans is limited, apart from the presence of TFH-like cells in blood. We studied the activation and induction of TFH and their role in the anti-influenza antibody response induced by a live-attenuated influenza vaccine (LAIV) in human nasopharynx-associated lymphoid tissue (NALT). TFH activation in adenotonsillar tissues was analyzed by flow cytometry, and anti-hemagglutinin (anti-HA) antibodies were examined following LAIV stimulation of tonsillar mononuclear cells (MNC). Induction of antigen-specific TFH by LAIV was studied by flow cytometry analysis of induced TFH and CD154 expression. LAIV induced TFH proliferation, which correlated with anti-HA antibody production, and TFH were shown to be critical for the antibody response. Induction of TFH from naive T cells by LAIV was shown in newly induced TFH expressing BCL6 and CD21, followed by the detection of anti-HA antibodies. Antigen specificity of LAIV-induced TFH was demonstrated by expression of the antigen-specific T cell activation marker CD154 upon challenge by H1N1 virus antigen or HA. LAIV-induced TFH differentiation was inhibited by BCL6, interleukin-21 (IL-21), ICOS, and CD40 signaling blocking, and that diminished anti-HA antibody production. In conclusion, we demonstrated the induction by LAIV of antigen-specific TFH in human NALT that provide critical support for the anti-influenza antibody response. Promoting antigen-specific TFH in NALT by use of intranasal vaccines may provide an effective vaccination strategy against respiratory infections in humans.IMPORTANCE Airway infections, such as influenza, are common in humans. Intranasal vaccination has been considered a biologically relevant and effective way of immunization against airway infection. The vaccine-induced antibody response is crucial for protection against infection. Recent data from animal studies suggest that one type of T cells, TFH, are important for the antibody response. However, data on whether TFH-mediated help for antibody production operates in humans are limited due to the lack of access to human immune tissue containing TFH In this study, we demonstrate the induction of TFH in human immune tissue, providing critical support for the anti-influenza antibody response, by use of an intranasal influenza vaccine. Our findings provide direct evidence that TFH play a critical role in vaccine-induced immunity in humans and suggest a novel strategy for promoting such cells by use of intranasal vaccines against respiratory infections.

ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies: an infectious diseases perspective (Agents targeting lymphoid or myeloid cells surface antigens [II]: CD22, CD30, CD33, CD38, CD40, SLAMF-7 and CCR4).

BACKGROUND: The present review is part of the ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies. AIMS: To review, from an Infectious Diseases perspective, the safety profile of agents targeting CD22, CD30, CD33, CD38, CD40, SLAMF-7 and CCR4 and to suggest preventive recommendations. SOURCES: Computer-based MEDLINE searches with MeSH terms pertaining to each agent or therapeutic family. CONTENT: The risk and spectrum of infections in patients receiving CD22-targeted agents (i.e. inotuzumab ozogamicin) are similar to those observed with anti-CD20 antibodies. Anti-Pneumocystis prophylaxis and monitoring for cytomegalovirus (CMV) infection is recommended for patients receiving CD30-targeted agents (brentuximab vedotin). Due to the scarcity of data, the risk posed by CD33-targeted agents (gemtuzumab ozogamicin) cannot be assessed. Patients receiving CD38-targeted agents (i.e. daratumumab) face an increased risk of varicella-zoster virus (VZV) infection. Therapy with CD40-targeted agents (lucatumumab or dacetuzumab) is associated with opportunistic infections similar to those observed in hyper-IgM syndrome, and prevention strategies (including anti-Pneumocystis prophylaxis and pre-emptive therapy for CMV infection) are warranted. SLAMF-7 (CD319)-targeted agents (elotuzumab) induce lymphopenia and increase the risk of infection (particularly due to VZV). The impact of CCR4-targeted agents (mogamulizumab) on infection susceptibility is difficult to distinguish from the effect of underlying diseases and concomitant therapies. However, anti-Pneumocystis and anti-herpesvirus prophylaxis and screening for chronic hepatitis B virus (HBV) infection are recommended. IMPLICATIONS: Specific management strategies should be put in place to reduce the risk and/or the severity of infectious complications associated to the reviewed agents.

Isorhamnetin, the active constituent of a Chinese herb Hippophae rhamnoides L, is a potent suppressor of dendritic-cell maturation and trafficking.

Dendritic cells (DCs) have been recognized as major targets of immunosuppressive therapies for their significant roles in connecting innate and adaptive immunity. Isorhamnetin (Iso), one of the most common flavonoid compounds extracted from the Chinese herb Hippophae rhamnoides L, has been proved to have anti-inflammatory, anticarcinogenic, and antioxidant activities in many chronic inflammatory conditions, but the effects of Iso on DCs have rarely been reported before. Here we investigated the functions and the mechanisms of Iso on bone marrow-derived DCs (BMDCs) including maturation, phagocytosis, and trafficking. Our data showed that Iso effectively inhibited the maturation of lipopolysaccharide (LPS)-treated BMDCs by down regulation of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1ß and IL-12p70, up regulation of IL-10, and depression of costimulatory molecules CD40, CD80, and CD86, while had no effects on phagocytosis. Furthermore, Iso inhibited the migration of LPS-treated BMDCs, which may be due to its inhibition on chemokine receptor 7 (CCR7) expression. These findings strongly suggest that Iso is a potent immunosuppressive agent by inhibiting DC activation and trafficking, and may be used to prevent or treat chronic inflammation, autoimmune diseases, and graft rejections.

Capparis spinosa Fruit Ethanol Extracts Exert Different Effects on the Maturation of Dendritic Cells.

Capparis spinosa L. (C. spinosa) has been used as food and traditional medicine and shows anti-inflammatory and anti-oxidant activities. Here, we prepared the C. spinosa fruit ethanol extracts (CSEs) using different procedures and investigated the effects of CSE on the maturation of mouse bone marrow-derived dendritic cells (DCs) in the absence or presence of lipopolysaccharide (LPS). DC maturation and cytokine production were detected by flow cytometry and ELISA, respectively. We obtained three different CSEs and dissolved in water or DMSO, named CSE2W, CSEMW, CSE3W, CSE2D, CSEMD, and CSE3D, respectively. These CSEs showed different effects on DC maturation. CSEMW and CSEMD significantly increased the expressions of CD40, CD80, and CD86, in a dose-dependent manner. CSE2W and CSE2D also showed a modest effect on DC maturation, which enhanced the expression of CD40. CSE3W and CSE3D did not change DC maturation but suppressed LPS-induced DC maturation characterized by the decreased levels of CD40 and CD80. CSE3W and CSE3D also significantly inhibited the secretions of IL-12p40, IL-6, IL-1ß, and TNF-α induced by LPS. CSE3W further increased the level of IL-10 induced by LPS. Moreover, CSE3D suppressed LPS-induced DC maturation in vivo, which decreased the expressions of CD40 and CD80. These results suggested that CSE3W and CSE3D might be used to treat inflammatory diseases.

CD40 Negatively Regulates ATP-TLR4-Activated Inflammasome in Microglia.

During acute brain injury and/or sterile inflammation, release of danger-associated molecular patterns (DAMPs) activates pattern recognition receptors (PRRs). Microglial toll-like receptor (TLR)-4 activated by DAMPs potentiates neuroinflammation through inflammasome-induced IL-1ß and pathogenic Th17 polarization which critically influences brain injury. TLR4 activation accompanies increased CD40, a cognate costimulatory molecule, involved in microglia-mediated immune responses in the brain. During brain injury, excessive release of extracellular ATP (DAMPs) is involved in promoting the damage. However, the regulatory role of CD40 in microglia during ATP-TLR4-mediated inflammasome activation has never been explored. We report that CD40, in the absence of ATP, synergizes TLR4-induced proinflammatory cytokines but not IL-1ß, suggesting that the response is independent of inflammasome. The presence of ATP during TLR4 activation leads to NLRP3 inflammasome activation and caspase-1-mediated IL-1ß secretion which was inhibited during CD40 activation, accompanied with inhibition of ERK1/2 and reactive oxygen species (ROS), and elevation in p38 MAPK phosphorylation. Experiments using selective inhibitors prove indispensability of ERK 1/2 and ROS for inflammasome activation. The ATP-TLR4-primed macrophages polarize the immune response toward pathogenic Th17 cells, whereas CD40 activation mediates Th1 response. Exogenous supplementation of IFN-γ (a Th1 cytokine and CD40 inducer) results in decreased IL-1ß, suggesting possible feedback loop mechanism of inflammasome inhibition, whereby IFN-γ-mediated increase in CD40 expression and activation suppress neurotoxic inflammasome activation required for Th17 response. Collectively, the findings indicate that CD40 is a novel negative regulator of ATP-TLR4-mediated inflammasome activation in microglia, thus providing a checkpoint to regulate excessive inflammasome activation and Th17 response during DAMP-mediated brain injury.

New Murine Model of Early Onset Autoimmune Thyroid Disease/Hypothyroidism and Autoimmune Exocrinopathy of the Salivary Gland.

Sixty to seventy percent of IFN-γ(-/-) NOD.H-2h4 mice given sodium iodide (NaI)-supplemented water develop a slow onset autoimmune thyroid disease, characterized by thyrocyte epithelial cell (TEC) hyperplasia and proliferation (H/P). TEC H/P develops much earlier in CD28(-/-) mice and nearly 100% (both sexes) have severe TEC H/P at 4 mo of age. Without NaI supplementation, 50% of 5- to 6-mo-old CD28(-/-)IFN-γ(-/-) mice develop severe TEC H/P, and 2-3 wk of NaI is sufficient for optimal development of severe TEC H/P. Mice with severe TEC H/P are hypothyroid, and normalization of serum thyroxine levels does not reduce TEC H/P. Activated CD4(+) T cells are sufficient to transfer TEC H/P to SCID recipients. Thyroids of mice with TEC H/P have infiltrating T cells and expanded numbers of proliferating thyrocytes that highly express CD40. CD40 facilitates, but is not required for, development of severe TEC H/P, as CD40(-/-)IFN-γ(-/-)CD28(-/-) mice develop severe TEC H/P. Accelerated development of TEC H/P in IFN-γ(-/-)CD28(-/-) mice is a result of reduced regulatory T cell (Treg) numbers, as CD28(-/-) mice have significantly fewer Tregs, and transfer of CD28(+) Tregs inhibits TEC H/P. Essentially all female IFN-γ(-/-)CD28(-/-) NOD.H-2h4 mice have substantial lymphocytic infiltration of salivary glands and reduced salivary flow by 6 mo of age, thereby providing an excellent new model of autoimmune exocrinopathy of the salivary gland. This is one of very few models where autoimmune thyroid disease and hypothyroidism develop in most mice by 4 mo of age. This model will be useful for studying the effects of hypothyroidism on multiple organ systems.

In vitro inhibitory effects of thymol and carvacrol on dendritic cell activation and function.

CONTEXT: Thyme has been used in traditional medicine for medicinal purposes since ancient times. OBJECTIVE: The objective of this study was to investigate the effects of thymol and carvacrol as two major constituents of thyme on dendritic cells (DCs) maturation and T cell activation. MATERIALS AND METHODS: Splenic DCs were treated with non-cytotoxic concentrations of the components and then analyzed for MHC II, CD86, and CD40 expression by flow cytometry. The effects of compounds on mitogenic, as well as allogenic T cell responses in mixed lymphocyte culture (MLR) and the release of cytokines were investigated. RESULTS: At 0.1 µg/ml, reduced mean fluorescent intensity (MFI) of CD86 for thymol (80.3 ± 0.2% of untreated control) and CD40 for carvacrol (79.5 ± 0.14%) was observed (p < 0.001). Decreased mitogenic T cell proliferation by thymol [proliferation index (PI) from 0.93 ± 0.11 at 1 µg/ml to 0.42 ± 0.16 at 100 µg/ml (p < 0.01)] and carvacrol [PI from 1.08 ± 0.3 at 1 µg/ml to 0.28 ± 0.1 at 100 µg/ml (p < 0.001)] was seen. Ten micrograms/ml thymol (PI, 0.85 ± 0.04) and carvacrol (PI, 0.89 ± 0.03) inhibited allogenic T cell response (p < 0.05). Decreased IFN-γ level in MLR supernatant from 1441 ± 27.7 pg/ml in untreated cells to 944 ± 32.1 at 10 µg/ml of thymol and of carvacrol (886 ± 31.7 pg/ml) (p < 0.01) was found. IL-4 levels were decreased in the presence of both compounds (p < 0.01). CONCLUSION: These data showed the suppressive effects of thymol and carvacrol on DCs maturation and function, as well as T cell responses.

Nicotinic acid inhibits vascular inflammation via the SIRT1-dependent signaling pathway.

Nicotinic acid (NA) has recently been shown to inhibit inflammatory response in cardiovascular disease. Sirtuin1 (SIRT1), a NAD(+)-dependent class III histone deacetylase, participates in the regulation of cellular inflammation. We hypothesized that dietary supplementation of NA could attenuate vascular inflammation via modulation of SIRT1 pathway. New Zealand White rabbits received chow or chow supplemented with 0.6% (wt/wt) NA for 2 weeks. Acute vascular inflammation was induced in the animals by placing a non-occlusive silastic collar around the left common carotid artery. At 24 h after collar implantation, the collar-induced production of C-reactive protein and monocyte chemotactic protein-1 was significantly suppressed in the NA-supplemented animals. Meanwhile, NA also decreased the expression of cluster of differentiation 40 (CD40) and CD40 ligand, but up-regulated SIRT1 expression, both in rabbits and in lipopolysaccharide-stimulated endothelial cells. Moreover, knockdown of SIRT1 reversed the inhibitory effect of NA on CD40 expression. Further study revealed that NA also decreased the expression of CD40 partly through mammalian target of rapamycin. These results indicate that NA protects against vascular inflammation via the SIRT1/CD40-dependent signaling pathway.

Malaysian endophytic fungal extracts-induced anti-inflammation in Lipopolysaccharide-activated BV-2 microglia is associated with attenuation of NO production and, IL-6 and TNF-α expression.

BACKGROUND: Excessive production of inflammatory mediators such as nitric oxide (NO) and proinflammatory cytokines like tumour necrosis factor-alpha (TNF-α) from activated microglia contributes to uncontrolled inflammation in neurodegenerative diseases. This study investigated the protective role of five endophytic extracts (HAB16R12, HAB16R13, HAB16R14, HAB16R18 and HAB8R24) against LPS-induced inflammatory events in vitro. These endophytic extracts were previously found to exhibit potent neuroprotective effect against LPS-challenged microglial cells. METHODS: The effects of these fungal endophytic extracts against nitric oxide (NO), CD40 phenotype and, pro- and anti-inflammatory cytokine production in lipopolysaccharide (LPS)-stimulated BV2 microglia cells were examined using commercially available assay kits, immunophenotyping and flow cytometry, respectively. RESULTS: Microglia pre-treated with the five endophytic extracts (0.1 mg/mL) reduced NO production without compromising cell viability. Whilst CD40 expression in LPS-stimulated microglia was not significantly different with or without the influence of endophytic extracts, expression of the proinflammatory cytokines, IL-6 and TNF-α in LPS-stimulated microglia was significantly (P < 0.05) inhibited by these endophytic extracts. CONCLUSIONS: The outcomes suggest that the neuroprotective effect of the fungal endophytic extracts is likely mediated through supression of neuroinflammation. To our knowledge, this is the first report of the effect of a fungal endophytic extract in controlling inflammation in BV2 microglia cells.

Anti-asthmatic effects of oxymatrine in a mouse model of allergic asthma through regulating CD40 signaling.

The aim of the study was to investigate the anti-asthmatic effects of oxymatrine (OXY) and the possible underlying mechanisms. The mouse asthma model was established by ovalbumin (OVA) intraperitoneal injection. A total of fifty mice were randomly assigned to five groups: control, OVA, OVA + dexamethasone (Dex, 2 mg · kg(-1)), and OVA + OXY (40 mg · kg(-1)), and OVA + OXY (80 mg · kg(-1)), respectively. Histological studies were conducted by the hematoxylin and eosin (HE) staining, the levels of interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-13, and IgE were evaluated by enzyme-linked immunosorbent assay (ELISA), and the protein level of CD40 was analyzed by Western blotting. OXY inhibited OVA-induced increases in eosinophil count; the levels of IL-4, IL-5, IgE, and IL-13 were recovered. It also substantially inhibited OVA-induced eosinophilia in lung tissues and the expression of CD40 protein. These findings suggest that OXY may effectively ameliorate the progression of asthma and could be explored as a possible therapy for patients with allergic asthma.

[High Expression of co-stimulatory molecule CD40 in silicosis patients and intervention effect of yiqi huoxue decoction].

OBJECTIVE: To study whether co-stimulatory molecule CD40 of alveolar macrophage (AM) participated in the occurrence and development of silicosis, and to explore the intervention of Yiqi Huoxue Decoction (YHD) in the fibrosis of silicosis patients. METHODS: Totally 46 silicosis inpatients and outpatients were recruited and randomly assigned to the Western treatment group (A) and the Chinese medicine (CM) treatment group (B), 23 in each group. Patients in Group A received routine symptomatic treatment such as anti-inflammation, phlegm resolving, anti-spasm, and asthma relief, and so on. Patients in Group B additionally took YHD, one dose daily for 14 successive days. Besides, another 18 patients with chronic cough and sense of laryngeal foreign bodies were recruited as the normal control group, who had no obvious lesion confirmed by bronchofi6roscope and clinical diagnosis of the lung. They were treated by symptomatic supporting treatment. The alveolar lavage fluid was collected from all patients and isolated, and AM cells were cultured. The level of CD40 mRNA was detected by RT-PCR. The expression of CD40 protein was detected by Western blot. RESULTS: Compared with the normal control group, expression levels of CD40 mRNA and CD40 protein significantly increased in Group A (P < 0.01). Compared with Group A, expression levels of CD40 mRNA and CD40 protein significantly decreased in Group B (P < 0.01). CONCLUSIONS: Highly expressed co-stimulatory molecule CD40 of AM might participate in pulmonary fibrosis. YHD could hinder its roles, inhibit the progression of fibrosis, thereby playing an interventional role of treatment.