Activity Of Lymph Node Cells Research Articles

Modulation of antigen delivery and lymph node activation in non-human primates by saponin adjuvant SMNP

Abstract Saponin-based vaccine adjuvants are potent in preclinical animal models and humans, but their mechanisms of action remain poorly understood. Here, using a stabilized HIV envelope trimer immunogen, we carried out studies in non-human primates (NHPs) comparing the most common clinical adjuvant alum with Saponin/MPLA Nanoparticles (SMNP), a novel ISCOMs-like adjuvant. SMNP elicited substantially stronger humoral immune responses than alum, including 7-fold higher peak antigen-specific germinal center B cell responses, 18-fold higher autologous neutralizing antibody titers, and higher levels of antigen-specific plasma and memory B cells. PET-CT imaging in live NHPs showed that, unlike alum, SMNP promoted rapid antigen accumulation in both proximal and distal lymph nodes (LNs). SMNP also induced strong type I interferon transcriptional signatures, expansion of innate immune cells, and increased antigen presenting cell activation in LNs. These findings indicate that SMNP promotes multiple facets of the early immune response relevant for enhanced immunity to vaccination.

Modulation of antigen delivery and lymph node activation in non-human primates by saponin adjuvant SMNP.

Saponin-based vaccine adjuvants are potent in preclinical animal models and humans, but their mechanisms of action remain poorly understood. Here, using a stabilized HIV envelope trimer immunogen, we carried out studies in non-human primates (NHPs) comparing the most common clinical adjuvant alum with Saponin/MPLA Nanoparticles (SMNP), a novel ISCOMs-like adjuvant. SMNP elicited substantially stronger humoral immune responses than alum, including 7-fold higher peak antigen-specific germinal center B cell responses, 18-fold higher autologous neutralizing antibody titers, and higher levels of antigen-specific plasma and memory B cells. PET-CT imaging in live NHPs showed that, unlike alum, SMNP promoted rapid antigen accumulation in both proximal and distal lymph nodes (LNs). SMNP also induced strong type I interferon transcriptional signatures, expansion of innate immune cells, and increased antigen presenting cell activation in LNs. These findings indicate that SMNP promotes multiple facets of the early immune response relevant for enhanced immunity to vaccination.

Focused Ultrasound-Responsive Nanocomposite with Near-Infrared II Mechanoluminescence for Spatiotemporally Selective Immune Activation in Lymph Nodes.

The immune regulation of the lymphatic system, especially the lymph node (LN), is of great significance for the treatment of diseases and the inhibition of pathogenic organisms spreading in the body. However, achieving precise spatiotemporal control of immune cell activation in LN in vivo remains a challenge due to tissue depth and off-target effects. Furthermore, minimally invasive and real-time feedback methods to monitor the regulation of the immune system in LN are lacking. Here, focused ultrasound responsive immunomodulator loaded nanoplatform (FURIN) with near-infrared II (NIR-II) luminescence is designed to achieve spatiotemporally controllable immune activation in LN in vivo. The NIR-II persistent luminescence of FURIN can track its delivery in LN through bioimaging. Under focused ultrasound (FUS) stimulation, the immunomodulator encapsulated in FURIN can be released locally in the LN to activate immune cells such as dendritic cells and the NIR-II mechanoluminescence of FURIN provides real-time optical feedback signals for immune activation. This work points to a FUS mediated, spatiotemporal selective immune activation strategy in vivo with the feedback control of luminescence signals via ultrasound responsive nanocomposite, which is of great significance in improving the efficacy and reducing the side effect of immune regulation for the development of potential immunotherapeutic methods in the future.

Oligodendrocyte progenitor cells differentiation induction with MAPK/ERK inhibitor fails to support repair processes in the chronically demyelinated CNS.

Remyelination failure is considered a major obstacle in treating chronic-progressive multiple sclerosis (MS). Studies have shown blockage in the differentiation of resident oligodendrocyte progenitor cells (OPC) into myelin-forming cells, suggesting that pushing OPC into a differentiation program might be sufficient to overcome remyelination failure. Others stressed the need for a permissive environment to allow proper activation, migration, and differentiation of OPC. PD0325901, a MAPK/ERK inhibitor, was previously shown to induce OPC differentiation, non-specific immunosuppression, and a significant therapeutic effect in acute demyelinating MS models. We examined PD0325901 effects in the chronically inflamed central nervous system. Treatment with PD0325901 induced OPC differentiation into mature oligodendrocytes with high morphological complexity. However, treatment of Biozzi mice with chronic-progressive experimental autoimmune encephalomyelitis with PD0325901 showed no clinical improvement in comparison to the control group, no reduction in demyelination, nor induction of OPC migration into foci of demyelination. PD0325901 induced a direct general immunosuppressive effect on various cell populations, leading to a diminished phagocytic capability of microglia and less activation of lymph-node cells. It also significantly impaired the immune-modulatory functions of OPC. Our findings suggest OPC regenerative function depends on a permissive environment, which may include pro-regenerative inflammatory elements. Furthermore, they indicate that maintaining a delicate balance between the pro-myelinating and immune functions of OPC is of importance. Thus, the highly complex mission of creating a pro-regenerative environment depends upon an appropriate immune response controlled in time, place, and intensity. We suggest the need to employ a multi-systematic therapeutic approach, which cannot be achieved through a single molecule-based therapy.

Stromal structure remodeling by B lymphocytes limits T cell activation in lymph nodes of Mycobacterium tuberculosis-infected mice.

An effective adaptive immune response depends on the organized architecture of secondary lymphoid organs, including the lymph nodes (LNs). While the cellular composition and microanatomy of LNs under steady state are well defined, the impact of chronic tissue inflammation on the structure and function of draining LNs is incompletely understood. Here we showed that Mycobacterium tuberculosis infection remodeled LN architecture by increasing the number and paracortical translocation of B cells. The formation of paracortical B lymphocyte and CD35+ follicular dendritic cell clusters dispersed CCL21-producing fibroblastic reticular cells and segregated pathogen-containing myeloid cells from antigen-specific CD4+ T cells. Depletion of B cells restored the chemokine and lymphoid structure and reduced bacterial burdens in LNs of the chronically infected mice. Importantly, this remodeling process impaired activation of naive CD4+ T cells in response to mycobacterial and unrelated antigens during chronic tuberculosis infection. Our studies reveal a mechanism in the regulation of LN microanatomy during inflammation and identify B cells as a critical element limiting the T cell response to persistent intracellular infection in LNs.

Tumor-educated Tregs drive organ-specific metastasis in breast cancer by impairing NK cells in the lymph node niche.

Breast cancer is accompanied by systemic immunosuppression, which facilitates metastasis formation, but how this shapes organotropism of metastasis is poorly understood. Here, we investigate the impact of mammary tumorigenesis on regulatory Tcells (Tregs) in distant organs and how this affects multi-organ metastatic disease. Using a preclinical mouse mammary tumor model that recapitulates human metastatic breast cancer, we observe systemic accumulation of activated, highly immunosuppressive Tregs during primary tumor growth. Tumor-educated Tregs show tissue-specific transcriptional rewiring in response to mammary tumorigenesis. This has functional consequences for organotropism of metastasis, as Treg depletion reduces metastasis to tumor-draining lymph nodes, but not to lungs. Mechanistically, we find that Tregs control natural killer (NK) cell activation in lymph nodes, thereby facilitating lymph node metastasis. In line, an increased Treg/NK cell ratio is observed in sentinel lymph nodes of breast cancer patients compared with healthy controls. This study highlights that immune regulation of metastatic disease is highly organ dependent.

PI3-Kinase p110α Deficiency Modulates T Cell Homeostasis and Function and Attenuates Experimental Allergic Encephalitis in Mature Mice.

Class I phosphoinositide 3-kinases (PI3K) are involved in the development of normal and autoimmune responses, including Experimental Autoimmune Encephalomyelitis (EAE), a mouse model for human multiple sclerosis (MS). Here, the role of the ubiquitously expressed class IA PI3K p110α catalytic subunits in EAE has been analyzed using a model of Cre/flox mediated T cell specific deletion of p110α catalytic chain (p110αΔT). Comparison of two month-old (young) and six month-old (mature) p110αΔT mice and their wild type (WT) counterparts indicated loss of spleen CD4+ T cells that increased with age, indicating a role of p110α in their homeostasis. In contrast, CD4+ T regulatory (Treg) cells were enhanced in mature p110αΔT mice when compared to WT mice. Since Myelin Oligodendrocyte Glycoprotein (MOG) peptide-induced EAE is dependent on, or mediated by CD4+ T cells and CD4+ T cell-derived cytokines and controlled by Treg cells, development of EAE in young and mature WT or p110αΔT mice was analyzed. EAE clinical symptoms and disease scores in six month p110αΔT mice were significantly lower than those of mature WT, or young WT and p110αΔT mice. Furthermore, ex vivo antigen activation of lymph node cells from MOG immunized mature p110αΔT mice induced significantly lower levels of IFN-γ and IL-17A than young p110αΔT or young and mature WT mice. Other cytokines including IL-2, IL-10 or TNF-α showed no significant differences between p110αΔT and WT mature mice. Our data show a lower incidence of MOG-induced EAE in mature p110αΔT mice linked to altered T cell homeostasis and lower secretion of inflammatory cytokines.

Nano-Immune-Engineering Approaches to Advance Cancer Immunotherapy: Lessons from Ultra-pH-Sensitive Nanoparticles.

Immunotherapy has transformed the field of oncology and patient care. By leveraging the immune system of the host, immunostimulatory compounds exert a durable, personalized response against the patient's own tumor. Despite the clinical success, the overall response rate from current therapies (e.g., immune checkpoint inhibitors) remains low (∼20%) because tumors develop multiple resistance pathways at molecular, cellular, and microenvironmental levels. Unlike other oncologic therapies, harnessing antitumor immunity requires precise activation of a complex immunological system with multiple levels of regulation over its function. This requires the ability to exert control over immune cells in both intracellular compartments and various extracellular sites, such as the tumor microenvironment, in a spatiotemporally coordinated fashion.The immune system has evolved to sense and respond to nano- and microparticulates (e.g., viruses, bacteria) as foreign pathogens. Through the versatile control of composition, size, shape, and surface properties of nanoparticles, nano-immune-engineering approaches are uniquely positioned to mount appropriate immune responses against cancer. This Account highlights the development and implementation of ultra-pH-sensitive (UPS) nanoparticles in cancer immunotherapy with an emphasis on nanoscale cooperativity. Nanocooperativity has been manifested in many biological systems and processes (e.g., protein allostery, biomolecular condensation), where the system can acquire emergent properties distinct from the sum of individual parts acting in isolation.Using UPS nanoparticles as an example, we illustrate how all-or-nothing protonation cooperativity during micelle assembly/disassembly can be leveraged to augment the cancer-immunity cycle toward antitumor immunity. The cooperativity behavior enables instant and pH-triggered payload release and dose accumulation in acidic sites (e.g., endocytic organelles of antigen presenting cells, tumor microenvironment), intercepting specific immunological and tumor pathophysiological processes for therapy. These efforts include T cell activation in lymph nodes by coordinating cytosolic delivery of tumor antigens to dendritic cells with simultaneous activation of stimulator of interferon genes (STING), or tumor-targeted delivery of acidotic inhibitors to reprogram the tumor microenvironment and overcome T cell retardation. Each treatment strategy represents a nodal intervention in the cancer-immunity cycle, featuring the versatility of UPS nanoparticles. Overall, this Account aims to highlight nanoimmunology, an emerging cross field that exploits nanotechnology's unique synergy with immunology through nano-immune-engineering, for advancing cancer immunotherapy.

A directly GP130-targeting small molecule ameliorates collagen-induced arthritis (CIA) by inhibiting IL-6/GP130 signalling and Th17 differentiation.

Rheumatoid arthritis is a chronic inflammatory disease associated with joint inflammation and destruction driven by T helper 17 (Th17) cells. Interleukin-6 (IL-6) is secreted by many cell types, including macrophages and synovial fibroblasts. It induces the differentiation and function of Th17 cells that can increase lymphocytic infiltration in the joint. LMT-28 can suppress IL-6 signalling through direct binding to glycoprotein-130 and alleviate inflammatory diseases such as rheumatoid arthritis and inflammatory bowel disease. The purpose of this study was to assess whether LMT-28 could potently inhibit Th17 differentiation and to determine the mechanism involved in the attenuating effect of LMT-28 on rheumatoid arthritis through the IL-6 signalling pathway. LMT-28 reduced the arthritis score and showed protective effects against bone and cartilage destruction in collagen-induced arthritis (CIA) mice. In mice with CIA, LMT-28 markedly decreased serum levels of IL-6, TNF and IL-1β compared to vehicle control. Moreover, LMT-28 attenuated Th17 cell activation in lymph nodes of CIA mice. We demonstrated that LMT-28 suppressed differentiation of Th17 in mouse splenocytes and human peripheral blood mononuclear cells (PBMCs). Additionally, LMT-28 inhibited phosphorylation of GP130, STAT3 and ERK induced by Hyper-IL-6 in human fibroblast-like synoviocytes (FLS). Collectively, these results suggest that LMT-28 can inhibit differentiated/activated-Th17 cells in rheumatoid arthritis by blocking activation of the STAT3 pathway. LMT-28 can attenuate rheumatoid arthritis by inhibiting differentiation/activation of Th17 cells and suppressing the proliferation and signalling activation of the IL-6/solubleIL-6 receptor complex stimulated FLS.

Возрастные изменения лимфатических узлов (собственные данные и обзор литературы)

Aim. To study the morphological features of lymph nodes in people aged 65-69 years and to analyze the results with data from other researchers. Materials and methods. The study was carried out on 30 lymph nodes obtained during autopsy of people aged 65-69 years. Lymph nodes were subjected to standard histological protocol; slides were stained with hematoxylin and eosin. An immunohistochemical study was performed to determine the expression of the markers Ki-67, p53, bcl-2 and Cyclin B1. Results. The thickening of the capsule and connective tissue trabecules were determined on the sections of the lymph nodes, the cleavage of trabecules by adipocytes. The zonal structure of the lymph node is smoothed. Lymphatic nodules in the cortex have unclear contours, they do not have germinal centers. The boundaries of the internodular zone and paracortex are not determined. The marginal sinus is unevenly expanded. In the medulla, medullary cords are replaced by white adipose tissue. The medullary sinuses are dilated, adipocytes are located in the lumen of some of them. Small arteries and arterioles with thickened walls and signs of hyalinosis occur. An immunohistochemical study of the proliferative activity of lymph node cells shows a significant decrease in the number of positively stained lymphocytes in both the cortex and medulla by the expression of the Ki-67 marker. In a study of the expression of Cyclin B1 and p53 receptors, the reaction is negative in both cases, while a reaction with antibodies to bcl-2 revealed weak cytoplasmic staining of lymphocytes of the cortex and medulla. Conclusion. Own and literature data shows that with age, unidirectional degenerative and dystrophic changes occur in the lymph nodes of humans, rats and mice. This results in increase in the proportion of connective and adipose tissue and a decrease in lymphoid tissue in the lymph nodes. These changes can be considered morphological signs of inhibition of lymphocytopoiesis, humoral and cellular immunity in the lymph nodes.

A directly GP130-targeting small molecule ameliorates collagen-induced arthrites by inhibiting IL-6/GP130 signalling and Th17 differentiation

Abstract Rheumatoid arthritis is a chronic inflammatory disease that is characterized by joint destruction and inflammation by T helper 17 (Th17) cells. Interleukin-6 (IL-6) is secreted in cells such as macrophage or synovial fibroblast and induces the differentiation and function of Th17 cells that increase lymphocytic infiltration in the joint. LMT-28 suppresses IL-6 signaling through direct binding to glycoprotein-130, and alleviates inflammatory disease (such as rheumatoid arthritis, inflammatory bowel disease) in previous study. The purpose of this study is to assess whether LMT-28 would potently inhibit Th17 differentiation, and the mechanism of LMT-28 that attenuates rheumatoid arthritis through IL-6 signaling pathway. LMT-28 reduced the arthritis score and showed protective effects against bone and cartilage destruction in collagen induced arthritis (CIA) mice. In mice with CIA, LMT-28 markedly decreased the serum levels of IL-6, TNF, and IL-1β compared with each vehicle treated group. Moreover, LMT-28 attenuated Th17 cell activation in lymph nodes of CIA mice. We demonstrated that LMT-28 suppressed differentiation of Th17 in mouse splenocytes and human PBMCs. Additionally, LMT-28 inhibited the phosphorylation of GP130, STAT3, and ERK induced by Hyper IL-6 in human fibroblast-like synoviocytes(FLS). Collectively, these results suggested that LMT-28 inhibits differentiated/activated-Th17 cells in rheumatoid arthritis by blocking activation of the STAT3 pathway.

Improvement of Outcome Measures of Dry Eye by a Novel Integrin Antagonist in the Murine Desiccating Stress Model.

We investigated the effects of GW559090, a novel, competitive, and high-affinity α4 integrin antagonist, in a murine model of dry eye. Through interaction with vascular cell adhesion molecule 1 (VCAM-1) and fibronectin α4β1 integrin is involved in leukocyte trafficking and activation. Female C57BL/6 mice, aged 6 to 8 weeks, were subjected to desiccating stress (DS). Bilateral topical twice daily treatment with GW559090 was compared to vehicle-treated controls. Treatment was initiated at the time of DS induction. Treatment effects were assessed on corneal staining with Oregon Green Dextran (OGD) and expression of inflammatory markers in ocular surface tissues by real time PCR. Dendritic cell activation was measured in draining cervical lymph nodes (CLN) by flow cytometry. Separate groups of mice received GW559090 subcutaneously to evaluate the effects of systemic administration on corneal staining and cells in CLN. Topical GW559090 significantly reduced corneal uptake of OGD compared to vehicle-treated disease controls in a dose-dependent manner (1, 3, 10, and 30 mg/mL) with 30 mg/mL showing the greatest reduction in OGD staining. When administered topically, corneal expression of IL-1α, matrix metalloproteinase (MMP)-9, chemokine ligand 9 (CXCL9), and TGF-β1 was reduced in GW559090-treated eyes. Topical treatment with GW559090 decreased dendritic cell activation in lymph nodes. The effects on corneal staining and cellular composition in CLN were not reproduced by systemic administration of GW559090, suggestive of a local role for integrin antagonism in the treatment of dry eye. The novel α4 integrin antagonist, GW559090, improved outcome measures of corneal staining and ocular surface inflammation in this murine model of dry eye. These results indicate the potential of this novel agent for the treatment of dry eye disease.

The level of antigen expression in the liver predicts the fate and functional outcome of antigen-specific CD8 T cells (LYM5P.706)

Abstract A wide spectrum of CD8 T cell responses can be elicited against antigens (Ag) expressed in the liver, ranging from tolerance to full effector function. Identifying the factors involved would allow us to predict the outcome of liver transplantation and viral hepatitis. To investigate how CD8 T cells respond to liver Ags, we have developed recombinant adeno-associated viral vectors that, when injected into mice, allow de novo Ag expression in a variable number of hepatocytes. Within the first day of Ag expression by at least 70% of hepatocytes, most naïve Ag-specific CD8 T cells were activated in the liver via direct presentation by hepatocytes. CD8 T cell activation in lymph nodes and spleen via cross presentation was detected a day later, suggesting that primary activation of CD8 T cells against hepatocyte-expressed Ag is compartmentalised. These T cells proliferated, developed into CTLs in the first week but were unable to kill all Ag-expressing hepatocytes. Most effector T cells were rapidly deleted while those surviving became exhausted and unresponsive over time. In contrast, when less than 10% of hepatocytes expressed the targeted Ag, CD8 T cell were gradually activated in liver and lymphoid tissues but were not deleted. Instead, they eliminated all Ag-expressing hepatocytes and maintained their effector function overtime. In summary, this study shows that intrahepatic Ag load dictates both the fate and function of CD8 T cells recognising hepatocyte-expressed Ags.

How many dendritic cells are required to initiate a T-cell response?

AbstractT-cell activation in lymph nodes relies on encounters with antigen (Ag)–bearing dendritic cells (DCs) but the number of DCs required to initiate an immune response is unknown. Here we have used a combination of flow cytometry, 2-photon imaging, and computational modeling to quantify the probability of T cell–DC encounters. We calculated that the chance for a T cell residing 24 hours in a murine popliteal lymph nodes to interact with a DC was 87, 587, and 997 in the presence of 10, 100, and 1000 Ag-bearing DCs, respectively. Our results reveal the existence of a threshold in DC numbers below which T-cell responses fail to be elicited for probabilistic reasons. In mice and probably humans, we estimate that a minimum of 85 DCs are required to initiate a T-cell response when starting from precursor frequency of 10−6. Our results have implications for the rational design of DC-based vaccines.

Synthesis, biological evaluation and molecular docking studies of 1,3,4-oxadiazole derivatives as potential immunosuppressive agents

A series of 1,3,4-oxadiazole derivatives derived from 4-methoxysalicylic acid or 4-methylsalicylic acid (6a–6z) have been first synthesized for their potential immunosuppressive activity. Among them, compound 6z displayed the most potent biological activity against lymph node cells (inhibition=38.76% for lymph node cells and IC50=0.31μM for PI3Kγ). The preliminary mechanism of compound 6z inhibition effects was also detected by flow cytometry (FCM) and the compound exerted immunosuppressive activity via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Docking simulation was performed to position compound 6z into the PI3Kγ structure active site to determine the probable binding model.

Design, synthesis, and biological evaluation of chalcone oxime derivatives as potential immunosuppressive agents

A series of deoxybenzoin oximes were recently reported as potent immunosuppressive agents by our group. In order to continue the original research for potential immunosuppressive agents with high efficacy and low toxicity, we synthesized a series of new chalcone oximes and evaluated them for their cytotoxicities and immunosuppressive activities. Among the synthesized compounds, chalcone oximes 25 and 27 exhibited lower cytotoxicities and higher inhibitory activities on anti-CD3/anti-CD28 co-stimulated lymph node cells than other compounds. Specially, compound 27 displayed 200-fold lower cytotoxicity (CC50=2174.39μM) than cyclosporin A (CC50=10.10μM) and showed SI value (SI=176.69) close to cyclosporin A (SI=154.13). Besides, the preliminary mechanism of inhibition effect of compounds 25 and 27 was also detected by flow cytometry, and the compounds exerted immunosuppressive activities via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Also, the deep mechanism of apoptosis was detected by Western blot analysis.

Mechanisms of NK cell activation: CD4(+) T cells enter the scene.

Natural killer (NK) cells are innate lymphocytes involved in immunosurveillance through their cytotoxic activity and their capacity to secrete inflammatory cytokines. NK cell activation is necessary to initiate effector functions and results from a complex series of molecular and cellular events. We review here the signals that trigger NK cells and discuss recent findings showing that, besides antigen-presenting cells, T cells can play a central role in the initiation of NK cell activation in lymph nodes.

Loss of anti-mycobacterial efficacy in mice over time following vaccination with Mycobacterium bovis bacillus Calmette-Guérin

Mycobacterium bovis bacillus Calmette-Guérin (BCG) is the most often used vaccine worldwide and sole vaccine against tuberculosis. BCG is protective against severe form of childhood tuberculosis but less or not protective to adult pulmonary tuberculosis. Therefore, improved vaccination strategies and development of new tuberculosis vaccines are urgent demands. For those purposes, appropriate animal models that reflect human are critically useful. However, in animal models, BCG vaccination protects well against subsequent challenge of Mycobacterium tuberculosis. In this study we evaluated the duration of protective efficacy of the BCG vaccination in mice over time and found that efficacy was diminished 40 weeks after vaccination. The aged mice older than 45 weeks are protected sufficiently after the vaccination with BCG, suggesting that loss of its efficacy is not dependent on the age of mice but rather depends on the period from vaccination. The loss of protection occurred in TH1 polarized STAT6 deficient mice despite the maintenance of interferon (IFN)-gamma production activity of lymph node cells and splenic CD4 + T cells against M. tuberculosis antigens. Our data suggest that the duration from vaccination may explain the variation in BCG efficacy against adult pulmonary tuberculosis.

Administration of 2-arachidonoylglycerol ameliorates both acute and chronic experimental autoimmune encephalomyelitis

Background and purpose Experimental autoimmune encephalomyelitis (EAE) is a widely used model of multiple sclerosis (MS) and both conditions have been reported to exhibit reduced endocannabinoid activity. The purpose of this study was to address the effect of exogenously administered 2-arachidonoylglycerol (2AG), an endocannabinoid receptor ligand, on acute phase and chronic disability in EAE. Experimental approach Acute and chronic EAE models were induced in susceptible mice and 2AG-treatment was applied for 14 days from day of disease induction. Key results 2AG-treatment ameliorated acute phase of disease with delay of disease onset in both EAE models and reduced disease mortality and long-term (70 days post-induction) clinical disability in chronic EAE. Reduced axonal pathology in the chronic EAE- ( p < 0.0001) and increased activation and ramification of microglia in the 2AG-treated acute EAE- ( p < 0.05) model were noticed. The latter was accompanied by a 2- to 4-fold increase of the M2-macrophages in the perivascular infiltrations ( p < 0.001) of the 2AG-treated animals in the acute (day 22), although not the chronic (day 70), EAE model. Expression of cannabinoid receptors 1 (CB1R) and 2 (CB2R) was increased in 2AG-treated animals of acute EAE vs. controls ( p < 0.05). In addition, ex vivo viability assays exhibited reduced proliferation of activated lymph node cells when extracted from 2AG-treated EAE animals, whereas a dose-dependent response of activated lymphocytes to 2AG-treatment in vitro was noticed. Conclusion and implications Our data indicate for the first time that 2AG treatment may provide direct (via CBRs) and immune (via M2 macrophages) mediated neuroprotection in EAE.

Synthesis, biological evaluation of chrysin derivatives as potential immunosuppressive agents

A series of novel chrysin derivatives was firstly synthesized and evaluated on their immunosuppressive activity in the search for potential immunosuppressive agents. Among them, compounds 5c displayed the most potent immunosuppressive inhibitory activity with IC50 of 0.78 μM, which was comparable to that of cyclosporin A (IC50 = 0.06 μM). The preliminary mechanism of compound 5c inhibition effects was also detected by flow cytometry (FCM), and the compound exerted immunosuppressive activity via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Furthermore, the estimated LD50 (in mg/kg) in vivo of compound 5c is 738.2, which indicated that compound 5c was low toxic.