Immunostimulatory effects on THP-1 cells by peptide or protein pharmaceuticals associated with injection site reactions.

Injection site reaction (ISR) is a common side-effect associated with the use of peptide or protein pharmaceuticals. These types of pharmaceuticals-induced activation of antigen-presenting cells is assumed to be a key step in the pathogenesis of immune-mediated ISR. The present study was designed to evaluate the immunostimulatory properties of peptide or protein pharmaceuticals using human monocytic THP-1 cells. Here, THP-1 cells, with or without phorbol-12-myristate-13-acetate (PMA) pretreatment, were exposed to enfuvirtide and glatiramer acetate (positive controls) or evolocumab (negative control) for 6 or 24 h. PMA treatment differentiated non-adherent monocytic THP-1 (nTHP-1) cells into adherent macrophagic THP-1 (pTHP-1) cells that highly express CD11b and CD36. Enfuvirtide increased the release of cytokines, e.g. TNFα, MIP-1ß, and MCP-1, and expression of CD86 and CD54 on nTHP-1 cells at 24 h. Similar immunostimulatory properties of glatiramer acetate were observed both in the nTHP-1 and pTHP-1 cells at 6 h, but the responses were very weak in the pTHP-1 cells. Evolocumab did not affect cytokine secretion or cell surface marker expression in either cell type. Taken together, these in vitro THP-1 cell assays revealed the immunostimulatory properties of enfuvirtide and glatiramer acetate. This assay platform thus could serve as a powerful tool in evaluating potential immune-related ISR risks of peptide or protein pharmaceuticals in humans.

Dendritic cells mediate the anti-inflammatory action of omega-3 long-chain polyunsaturated fatty acids in experimental autoimmune uveitis.

We previously showed that dietary omega (ω)-3 long-chain polyunsaturated fatty acids (LCPUFAs) suppress inflammation in mice with experimental autoimmune uveitis (EAU). We have now investigated the role of antigen presenting cells (APCs) in this action of ω-3 LCPUFAs. C57BL/6 mice were fed a diet supplemented with ω-3 or ω-6 LCPUFAs for 2 weeks, after which splenocytes were isolated from the mice and cocultured with CD4+ T cells isolated from mice with EAU induced by injection of a human interphotoreceptor retinoid-binding protein peptide together with complete Freund's adjuvant. The proliferation of and production of interferon-γ and interleukin-17 by T cells from EAU mice in vitro were attenuated in the presence of splenocytes from ω-3 LCPUFA-fed mice as compared with those from mice fed ω-6 LCPUFAs. Splenocyte fractionation by magnetic-activated cell sorting revealed that, among APCs, dendritic cells (DCs) were the target of ω-3 LCPUFAs. Adoptive transfer of DCs from mice fed ω-3 LCPUFAs attenuated disease progression in EAU mice as well as the production of pro-inflammatory cytokines by T cells isolated from these latter animals. The proliferation of T cells from control Balb/c mice was also attenuated in the presence of DCs from ω-3 LCPUFA-fed mice as compared with those from ω-6 LCPUFA-fed mice. Furthermore, T cell proliferation in such a mixed lymphocyte reaction was inhibited by prior exposure of DCs from mice fed an ω-6 LCPUFA diet to ω-3 LCPUFAs in vitro. Our results thus suggest that DCs mediate the anti-inflammatory action of dietary ω-3 LCPUFAs in EAU.

Regional therapies for locoregionally advanced and unresectable melanoma.

Locoregionally advanced and unresectable disease can be seen in up to 10% of melanoma patients. Treatment options for these patients have been evolving most notably over the past few decades and have demonstrated efficacy through multiple intra-arterial as well as intralesional therapies. Isolated limb perfusions and isolated limb infusions have been utilized to treat locoregionally advanced melanoma of the extremity with overall response rates up to 90% in some reports. Intralesional therapies, for in transit metastatic melanoma, such as Bacille Calmette-Guerin, talimogene laherparepvec, and PV-10 (Rose Bengal) have all demonstrated efficacy in the treatment of unresectable cutaneous melanoma. The treatment effect due to intralesional injection has been identified in directly injected lesions as well as in distant uninjected "bystander lesions" with some injectables. This bystander effect is likely an immunologic reaction due to tumor antigen release, antigen-presenting cell uptake, T cell activation and subsequent bystander tumor destruction in uninjected lesions. Treatment options for unresectable melanoma metastases limited to the liver include isolated hepatic perfusion, which can now be performed through a minimally invasive approach known as percutaneous hepatic perfusion. These intra-arterial and intralesional regional therapies offer a variety of effective treatment modalities for unresectable disease and may potentially be combined with systemic treatments, such as immunotherapy, in the future treatment of locoregionally advanced melanoma.

Extracts of Tectona grandis and Vernonia amygdalina have anti-Toxoplasma and pro-inflammatory properties in vitro.

Tectona grandis (teak) and Vernonia amygdalina (bitter leaf) are plants used in traditional medicine in West Africa. In this study, we tested ethanolic and hydro-ethanolic extracts of bark and leaves of T. grandis and ethanolic extract of leaves of V. amygdalina for their inhibitory effect on Toxoplasma gondii, a protozoan parasite responsible for toxoplasmosis. Ethanolic extract of V. amygdalina leaves had proportional contents of phenols, tannins, flavonoids, and polysaccharides. This extract presented the highest efficacy against T. gondii, the lowest cytotoxicity to mammalian cells, but moderate anti-oxidant activity compared to other plant extracts. Ethanolic extract of T. grandis bark also had elevated anti-T. gondii activity, low cytotoxicity on mammalian cells, and one of the highest anti-oxidant activities. However, the phytochemical content of this extract was not very different from the hydro-ethanolic extract, which had no anti-T. gondii activity. In addition, ethanolic extract of V. amygdalina leaves, but not of T. grandis bark, significantly increased the production of TNF-α and NO by antigen-presenting cells. Both extracts had the tendency to decrease expression of major histocompatibility complex molecules at the surface of antigen-presenting cells, while they did not modulate the percentage of apoptotic cells. A study of signalling pathways would help to determine the mechanisms of action of these plant extracts.

TITLE: Les extraits de Tectona grandis et de Vernonia amygdalina ont des propriétés anti-Toxoplasma et pro-inflammatoires in vitro. ABSTRACT: Tectona grandis (teck) et Vernonia amygdalina sont des plantes utilisées dans la médecine traditionnelle en Afrique de l'Ouest. Dans cette étude, l'effet inhibiteur d'extraits éthanoliques et hydro-éthanoliques d'écorce et de feuilles de T. grandis et de l'extrait éthanolique des feuilles de V. amygdalina a été étudié sur Toxoplasma gondii, un parasite protozoaire responsable de la toxoplasmose. L'extrait éthanolique des feuilles de V. amygdalina avait des quantités équivalentes de phénols, tanins, flavonoïdes et polysaccharides. Cet extrait présentait la plus grande efficacité contre T. gondii, la plus faible cytotoxicité vis-à-vis de cellules de mammifères, mais une activité anti-oxydante moyenne comparée aux autres extraits de plantes. L'extrait éthanolique d'écorce de T. grandis avait aussi une activité anti-T. gondii élevée, une faible cytotoxicité vis-à-vis des cellules de mammifères et l'une des activités anti-oxydantes les plus élevées. Cependant, le contenu phytochimique de cet extrait n'était pas très différent de l'extrait hydro-éthanolique qui n'avait pas d'activité anti-T. gondii. De plus, l'extrait éthanolique des feuilles de V. amygdalina, mais pas de l'écorce de T. grandis, augmentait significativement la production de TNF-α et de NO par les cellules présentatrices d'antigènes. Les deux extraits avaient tendance à diminuer l'expression des molécules du complexe majeur d'histocompatibilité à la surface des cellules présentatrices d'antigènes alors qu'ils ne modulaient pas le pourcentage de cellules apoptotiques. L'étude des voies de signalisation permettrait de comprendre les mécanismes d'action de ces extraits de plantes.

MERS-CoV pathogenesis and antiviral efficacy of licensed drugs in human monocyte-derived antigen-presenting cells.

Middle East respiratory syndrome coronavirus (MERS-CoV) presents an emerging threat to public health worldwide by causing severe respiratory disease in humans with high virulence and case fatality rate (about 35%) since 2012. Little is known about the pathogenesis and innate antiviral response in primary human monocyte-derived macrophages (MDMs) and dendritic cells (MDDCs) upon MERS-CoV infection. In this study, we assessed MERS-CoV replication as well as induction of inflammatory cytokines and chemokines in MDMs and immature and mature MDDCs. Immature MDDCs and MDMs were permissive for MERS-CoV infection, while mature MDDCs were not, with stimulation of proinflammatory cytokine and chemokine upregulation in MDMs, but not in MDDCs. To further evaluate the antiviral activity of well-defined drugs in primary antigen presenting cells (APCs), three compounds (chloroquine, chlorpromazine and toremifine), each with broad-spectrum antiviral activity in immortalized cell lines, were evaluated in MDMs and MDDCs to determine their antiviral effect on MERS-CoV infection. While chloroquine was not active in these primary cells, chlorpromazine showed strong anti-MERS-CoV activity, but it was associated with high cytotoxicity narrowing the potential window for drug utilization. Unlike in established cells, toremifene had marginal activity when tested in antigen presenting cells, with high apparent cytotoxicity, also limiting its potential as a therapeutic option. These results demonstrate the value of testing drugs in primary cells, in addition to established cell lines, before initiating preclinical or clinical studies for MERS treatment and the importance of carefully assessing cytotoxicity in drug screen assays. Furthermore, these studies also highlight the role of APCs in stimulating a robust protective immune response to MERS-CoV infection.

Understanding the Effect of Surface Chemistry of Mesoporous Silica Nanorods on Their Vaccine Adjuvant Potency.

Mesoporous silica nanoparticles are reported as adjuvants in nanovaccines in generating robust antigen-specific immunity. However, the effect of surface chemistry in initiating and modulating the immune response remains largely unexplored. In this study, mesoporous silica nanorods (MSNRs) are modified with NH2 and C18 groups to investigate the influence of surface functional groups (OH, NH2 , and C18 ) on their adjuvant efficacy. It is found that compared to OH and NH2 groups, the hydrophobic C18 modification significantly enhances antigen uptake by antigen presenting cells and endosomal-lysosomal escape in vitro, dendritic cells, and macrophages maturation ex vivo, and elicits secretion of interferon-γ level and antibody response in immunized mice. Moreover, bare MSNR and MSNRNH2 exhibit T-helper 2 biased immune response, while MSNRC18 shows a T-helper 1 biased immune response. These findings suggest that the surface chemistry of nanostructured adjuvants has profound impact on the immune response, which provides useful guidance for the design of effective nanomaterial based vaccines.

Effect of Antidepressants on Immunological Reactivity in ASC Mice with Genetically Determined Depression-Like State.

The effect of chronic treatment with antidepressant drugs fluoxetine (20 mg/kg) and imipramine (25 mg/kg) on the number of antibody-producing cells and the main T cell subpopulations in ASC mice characterized by genetic predisposition to depression-like states was studied at the peak of the SE-induced immune response (5×10(8)). Fluoxetine produced an immunostimulatory effect manifested in an increase in the relative and absolute number of IgM antibody-producing cells in the spleen and index of immunoreactivity (CD4/CD8). Administration of fl uoxetine to parental mouse strains without depression (CBA and AKR) had no effect (CBA) or reduced the immune response. The CD4/CD8 ratio did not increase under these conditions. Imipramine was ineffective in the correction of immune reactions in a depression-like state.

Resistance of Nonmelanoma Skin Cancer to Nonsurgical Treatments. Part I: Topical Treatments. / Resistencias al tratamiento no quirúrgico en cáncer cutáneo no melanoma. Parte I: tratamientos tópicos.

A wide range of treatments is now available for nonmelanoma skin cancer (NMSC), including 5-fluorouracil, ingenol mebutate, imiquimod, diclofenac, photodynamic therapy, methotrexate, cetuximab, vismodegib, and radiotherapy. All are associated with high clinical and histologic response rates. However, some tumors do not respond due to resistance, which may be primary or acquired. Study of the resistance processes is a broad area of research that aims to increase our understanding of the nature of each tumor and the biologic features that make it resistant, as well as to facilitate the design of new therapies directed against these tumors. In this article we review resistance to the authorized topical treatments for NMSC.

New strategy for testing efficacy of immunotherapeutic compounds for diabetes in vitro.

BACKGROUND: The valuable role of immunotherapy in treating autoimmune diseases is increasingly recognized by those involved in the research and clinical application of new biopharmaceuticals products. However, many aspects related to the mechanisms of immune-modulated therapies remain to be elucidated in order to explore fully the emerging opportunities. The non-obese diabetic NOD mouse develops insulin-dependent diabetes mellitus spontaneously as a consequence of an autoimmune process in the presence of pathogenic CD4(+) T cells that typically exhibit Th17 cell phenotypes. The change of a Th17 phenotype into a pattern of regulatory T cells (Treg) is extremely important in controlling autoimmune diseases. Heat shock proteins (HSPs) are stress-induced proteins with immunoregulatory properties. In the current study, the capacity of Hsp65 and Hsp70 mycobacterial HSPs and a constructed DNA encoded Hsp65 (DNAhsp65) to transform the pattern of the immune response from Th17 into Treg cells has been studied in vitro using co-cultures of antigen presenting cells (APCs) and T cells in NOD mice. RESULTS: Cells harvested from NOD mice and cultured for 48 h (without immunoregulatory compounds) presented with Th1/Th17 patterns and secretions of IL-6, IFN-γ, IL-10 and IL-17 cytokines. The cultured cells from the non-diabetic BALB/C mice exhibited a Th1 pattern and the production of IL 6 and IFN-γ secretions. An up-regulation was observed in the supernatants from the co-cultures of NOD cells that were stimulated with DNAhsp65, Hsp65 or Hsp70 through increased levels of IL-10 secretion and the suppression of IL-6, IFN-γ and IL-17 production. In addition, immunoregulation was demonstrated through IL-17 suppression in the co-culture stimulated by the specific insulin antigen. Moreover, an increase of immunoregulatory compounds were observed in the co-culture through the expression of CD11b(+)CD86(+) activation markers on APCs, as well as the frequency of Treg cells expressing CD4(+)CD3(+) and CD4(+)CD25(hi). CONCLUSIONS: The in vitro observation of Th17 cells differentiating into Tregs in NOD mice could raise the hypothesis that the immune regulatory activity of HSPs could be an efficient strategy for diabetes prevention and treatment.

Dietary proanthocyanidins inhibit UV radiation-induced skin tumor development through functional activation of the immune system.

The incidence of skin cancer is equivalent to the incidence of malignancies in all other organs combined. The main risk factor for this disease is overexposure of the skin to solar ultraviolet (UV) radiation. UV irradiation induces inflammation, oxidative stress, DNA damage, and suppression of the immune system in the skin, which together contribute to carcinogenesis. The use of dietary phytochemicals shows great promise as a complementary and alternative strategy for skin cancer prevention. Grape seed proanthocyanidins (GSPs) have been tested extensively for their anti-skin cancer effect using in vivo animal models. Supplementation of an AIN76A control diet with GSPs (0.2 and 0.5%, w/w) significantly inhibits UV radiation-induced skin tumor development as well as malignant transformation of papillomas to carcinoma in mice. The inhibition of UVB-induced skin tumor development by GSPs is mediated through interrelated mechanisms of action including: (i) inhibition of inflammation, (ii) rapid repair of damaged DNA, and (iii) stimulation of immune system. Additionally, the chemopreventive effects of GSPs involve DNA repair-dependent functional activation of antigen-presenting cells and stimulation of CD8(+) effector T cells. These effects of GSPs could be useful in attenuation of the adverse effects of UV radiation and may have health benefits in humans.

[In vitro anti-tumor effect of human dendritic cells vaccine induced by astragalus polysacharin: an experimental study].

OBJECTIVE: To explore the in vitro anti-tumor effect and mechanism of dendritic cell (DC) tumor vaccine induced by astragalus polysacharin (APS). METHODS: Peripheral blood mononuclear cells (PBMCs) isolated from human peripheral blood. DCs obtained from human peripheral blood were cultivated and added with culture solution for in vitro inducing them to immature DCs. On the 5th day of culture, 100 microg/mL (as the final concentration) APS was added to cells in the APS group. DCs were induced to mature in the cytokine groups by adding 20 ng/mL rhTNF-alpha (as the final concentration). Changes of morphology and phenotype of DCs were observed. Mature DCs were sensitized with tumor antigen SGC-7901 and co-cultured with allogeneic T cells. The proliferative function of T lymphocytes was detected by MTT assay. Levels of IL-12 and IFN-gamma in co-cultured supernatant were detected by ELISA. Cytotoxic lymphocytes (CTL) activated by DC were co-cultured with tumor cell SGC-7901. The specific killing capacity of CTL to target cells was detected by LDH release assay. RESULTS: The morphological observation and phenotypic identification of APS induced DCs were in accordance with the characteristics of mature DCs. APS induced mature DCs could stimulate the proliferation of allogeneic T lymphocytes. The proliferation index of T cells increased with increased ratio of stimulator cells to effector cells (P < 0.05). Levels of IL-12 and IFN-gamma in co-culture supernatant significantly increased in a time-dependent manner (P < 0.05). CTL cells activated by sensitization of DCs could significantly kill tumor cells, and the killing effect increased along with increased effector-to-target ratio. CONCLUSION: APS could in vitro induce DCs to mature, promote its antigen-presenting capacity, effectively activate CTLs, and enhance anti-tumor function of the organism.

Collagen-induced arthritis: severity and immune response attenuation using multivalent N-acetyl glucosamine.

Rheumatoid arthritis is an autoimmunity leading to considerable impairment of quality of life. N-acetyl glucosamine (GlcNAc) has been described previously as a potent modulator of experimental arthritis in animal models and is used for osteoarthritis treatment in humans, praised for its lack of adverse effects. In this study we present a comprehensive immunological analysis of multivalent GlcNAc-terminated glycoconjugate (GC) application in the treatment of collagen-induced arthritis (CIA) and its clinical outcome. We used immunohistochemistry and FACS to describe conditions on the inflammation site. Systemic and clinical effects were evaluated by FACS, cytotoxicity assay, ELISA, cytometric bead array (CBA), RT-PCR and clinical scoring. We found reduced inflammatory infiltration, NKG2D expression on NK and suppression of T, B and antigen-presenting cells (APC) in the synovia. On the systemic level, GCs prevented the activation of monocyte- and B cell-derived APCs, the rise of TNF-α and IFN-γ levels, and subsequent type II collagen (CII)-specific IgG2a formation. Moreover, we detected an increase of anti-inflammatory IL-4 mRNA in the spleen. Similar to the synovia, the GCs caused a significant reduction of NKG2D-expressing NK cells in the spleen without influencing their lytic function. GCs effectively postponed the onset of arthritic symptoms, reduced their severity and in 18% (GN8P) and 31% (GN4C) of the cases completely prevented their appearance. Our data prove that GlcNAc glycoconjugates prevent the inflammatory response, involving proinflammatory cytokine rise, APC activation and NKG2D expression, leading to the attenuation of clinical symptoms. These results support the glycobiological approach to the treatment of collagen-induced arthritis/rheumatoid arthritis (CIA/RA) as a way of bringing new prospects for more effective therapeutic interventions.

Inflammasome-dependent and -independent IL-18 production mediates immunity to the ISCOMATRIX adjuvant.

Adjuvants are an essential component of modern vaccines and used for their ability to elicit immunity to coadministered Ags. Many adjuvants in clinical development are particulates, but how they drive innate and adaptive immune responses remains poorly understood. Studies have shown that a number of vaccine adjuvants activate inflammasome pathways in isolated APCs. However, the contribution of inflammasome activation to vaccine-mediated immunity in vivo remains controversial. In this study, we evaluated immune cell responses to the ISCOMATRIX adjuvant (IMX) in mice. Like other particulate vaccine adjuvants, IMX potently activated the NALP-3-ASC-Caspase-1 inflammasome in APCs, leading to IL-1ß and IL-18 production. The IL-18R pathway, but not IL-1R, was required for early innate and subsequent cellular immune responses to a model IMX vaccine. APCs directly exposed to IMX underwent an endosome-mediated cell-death response, which we propose initiates inflammatory events locally at the injection site. Importantly, both inflammasome-related and -unrelated pathways contributed to IL-18 dependence in vivo following IMX administration. TNF-α provided a physiological priming signal for inflammasome-dependent IL-18 production by APCs, which correlated with reduced vaccine-mediated immune cell responses in TNF-α- or TNFR-deficient mice. Taken together, our findings highlight an important disconnect between the mechanisms of vaccine adjuvant action in vitro versus in vivo.

Tanshinone II A inhibits dendritic cell-mediated adaptive immunity: potential role in anti-atherosclerotic activity.

OBJECTIVE: Antigen-presenting cells such as monocytes and dendritic cells (DCs) stimulate T-cell proliferation and activation during adaptive immunity. This cellular interaction plays a role in the growth of atherosclerotic plaques. Tanshinone II A (TSN) had been shown to decrease the growth of atherosclerotic lesions. We therefore investigated the ability of TSN to inhibit human monocyte-derived DCs and their T-cellstimulatory capacity. METHODS: DCs derived from human monocytes cultured with recombinant human interleukin (IL)-4 and recombinant human granulocyte-macrophage colony-stimulating factor were co-cultured with TSN and lipopolysaccharide for 48 h. Phosphate-buffered saline was used as a negative control. Activation markers and the capacity of DCs for endocytosis were measured by flow cytometry, and proinflammatory cytokines were measured by enzyme-linked immunosorbent assays. DCs were co-cultured with lymphocytes to measure T-cell proliferation and IL-2 secretion by mixed lymphocyte reactions. RESULTS: TSN dose-dependently attenuated DC expression of costimulatory molecules (CD86), and decreased expression of major histocompatibility complex class II (human loukocyte antigen-DR) and adhesion molecules (CD54). Moreover, TSN reduced secretion of the proinflammatory cytokines IL-12 and IL-1 by human DCs, and restored the capacity for endocytosis. Finally, TSN-preincubated DCs showed a reduced capacity to stimulate T-cell proliferation and cytokine secretion. CONCLUSIONS: TSN inhibits DC maturation and decreases the expression of proinflammatory cytokines, while impairing their capacity to stimulate T-cell proliferation and cytokine secretion. These effects may contribute to the influence of TSN on the progression of atherosclerotic lesions.

Ulmus davidiana var. japonica Nakai upregulates eosinophils and suppresses Th1 and Th17 cells in the small intestine.

The bark of Ulmus davidiana var. japonica Nakai (Ulmaceae) has been used in traditional Korean medicine for chronic inflammation in the gastrointestinal tract. Here we investigated the frequency and cytokine profile of the major immune cells in the small intestinal lamina propria (SI LP), spleen, and mesenteric lymph nodes (MLNs) of mice treated orally with Ulmus davidiana var. japonica Nakai bark water extract (UDE) to address the immunomodulatory role of this herb in intestinal homeostasis. B6 mice were given 5g/kg UDE once daily for 14 days. They were then sacrificed, and cells were isolated from the spleen, MLNs, and SI LP. The proportion of B versus T lymphocytes, CD4(+) versus CD8(+) T lymphocytes, Th1 and Th17 cells, and Foxp3(+) regulatory T cells in the spleen, MLNs, and SI LP were analyzed. The frequency of antigen-presenting cells (APCs), including dendritic cells, macrophages, and eosinophils in the SI LP and the expression of costimulatory molecules on APCs were also evaluated. The numbers and frequencies of Th1 and Th17 cells in the SI LP were significantly reduced in the UDE-treated mice compared with PBS controls. In addition, the proportion of IL-4-producing eosinophils in the SI LP was significantly elevated in the UDE-treated mice compared with controls. Taken together, these data indicate that UDE up-regulates the number and frequency of SI LP eosinophils, which can down-regulate the Th1 and Th17 responses via IL-4 secretion and contribute to intestinal homeostasis.

Red blood cells as innovative antigen carrier to induce specific immune tolerance.

The route of administration, the dose of antigen as well as the type of antigen-presenting cells (APCs) targeted are important factors to induce immune tolerance. Despite encouraging results obtained in animal models, intravenous injection of soluble antigen is unsuccessful in human clinical trials on autoimmune disease due to inefficient antigen delivery. To improve antigen delivery, we used mouse red blood cells (RBCs) as antigen vehicles to specifically target APCs which are responsible for removal of senescent RBCs after phagocytosis. In this study, we demonstrated that antigen-delivery by RBCs induced a strong decrease in the humoral response compared with the ovalbumin (OVA) free form in mice. In addition, OVA-loaded RBC treated with [bis(sulphosuccinimidyl)] suberate (BS3), a chemical compound known to enhance RBC phagocytosis, induced an inhibition of antigen-specific T cell responses and an increase in the percentage of regulatory T cells. The state of tolerance induced is long lasting, antigen-specific and sufficiently robust to withstand immunization with antigen mixed with cholera toxin adjuvant. This RBC strategy, which does not abolish the immune system, constitutes an attractive approach for induction of tolerance compared to systemic immunosuppressant therapies already in use.

New conjugates of tuftsin and muramyl dipeptide as stimulators of human monocyte-derived dendritic cells.

Muramyl dipeptide (MDP) and tuftsin are known biologically active compound displaying a significant influence on various cell populations of innate immune response. MDP, as a fragment of bacterial cell wall, stimulates not only macrophages and monocytes, but also dendritic cells. In contrast, little is known about tuftsin influence on these cells. Therefore it seemed vital to access whether tuftsin or its derivatives conjugated with MDP could influence the activity of this subpopulation of antigen presenting cells (APC). Immature dendritic cells (iDCs) were derived from human monocytes through eight-day tissue culture supplemented with hrIL-4 and hrGM-CSF. On the day 9 DCs were stimulated with newly synthesized conjugates of tuftsin and muramyl dipeptide. The influence of the examined compounds on the activity and maturity of monocyte-derived DCs was estimated by flow cytometry analysis. The flow cytometry analysis revealed that tuftsin and some of its analogues do stimulate maturation and activity of DCs but to a lesser extend in comparison to MDP. The obtained results suggest further development of the experiments concerning the influence of MDP and tuftsin analogues on the activity of dendritic cells.

Involvement of trypsin-digested silk peptides in the induction of RAW264.7 macrophage activation.

The activation of macrophages by trypsin-digested silk peptides was investigated by considering CD1 lb and CD40 expression in the RAW264.7 cell, a murine macrophage. Silk protein hydrolysates were digested with trypsin, following by centrifugal purification using the Centriprep 30k concentrator. Trypsin-digested total silk peptides and its centrifugal fractions were tested for macrophage activation in vitro. The functional peptide of fractionated silk peptides was examined by LC/MS/MS analysis. Trypsin-digested and fractionated silk peptides of more than 30 kDa induced an increase in the activation markers CD1 lb and CD40 in RAW264.7 cells. These results are supported by morphological changes reflecting an increase in the number of dendrites in activated cells. The fractionated silk peptides examined by LC/MS/MS contained partial peptides of Bombyx mori fibroin. These results suggest that the activation of RAW264.7 macrophages may be induced not by sericin-derived peptides but by fibroin-derived ones.

Abatacept mechanism of action: concordance with its clinical profile.

The double and simultaneous molecular interaction between antigen-presentig cells (APC) and T lymphocytes is essential for the optimal activation of the immunological response and requires the participation of two membrane receptor groups. Abatacept is a fusion protein that selectively modulates one of these two ways, by binding to CD80 and CD86 receptors on APC. In this way, the drug inhibits T cell activation, selectively blocking the specific interaction of CD80/CD86 receptors to CD28 and, therefore, inhibiting T cell proliferation and B cell immunological response. This pharmacological action results in the normalization of inflammatory mediators in rheumatoid arthritis patients and in a safe and efficacious clinical response. Abatacept in combination with methotrexate prevents the progression of joint damage and improves physical function in rheumatoid arthritis patients.

1,25-Dihydroxyvitamin D3 inhibits proliferation but not the suppressive function of regulatory T cells in the absence of antigen-presenting cells.

Vitamin D3 is known to induce regulatory T (Treg) cells by rendering antigen-presenting cells tolerogenic, its direct effect on human naturally occurring Treg cells is unclear. Here, we investigated if and how 1,25-dihydroxyvitamin D(3) [1,25(OH)2D3] can directly affect the proliferation and function of human naturally occurring Treg cells in vitro. First, we demonstrated that these Treg cells express vitamin D receptors that were up-regulated following anti-CD3/CD28-bead stimulation. 1,25(OH)2D3 inhibited proliferation of Treg cells even when exogenous interleukin-2 was provided. Treg cells were more susceptible to the inhibitory effect of 1,25(OH)2D3 than conventional T cells(.) 1,25(OH)2D3 neither affected the anergic state nor the suppressive function of Treg cells but induced a subtle increase in interleukin-10-secreting cells. The cell-division-inhibiting effect of 1,25(OH)2D3 on Treg cells was also demonstrated in vivo by supplementing vitamin D-deficient HIV-1-infected patients with 2000 IU cholecalciferol (vitamin D3). Increased serum 1,25(OH)2D3 levels were associated with a drop in the number and percentage of Treg cells, which may be attributed to a decrease in the proliferating Foxp3+ Treg cell population. In conclusion, 1,25(OH)2D3 directly affects Treg cell growth and promotes interleukin-10 production without apparent effects on activation status and suppressive phenotype whereas in vivo, high serum 1,25(OH)2D3 levels are associated with reduced Treg cell proliferation and a reduced number of Treg cells.