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1.
Mol Immunol ; 47(2-3): 551-9, 2009 Dec.
Article in English | MEDLINE | ID: mdl-19850344

ABSTRACT

Chemokines are key molecules that drive migration of lymphoid and myeloid cells toward organs in basal as well as inflammatory conditions. By recruiting immature dendritic cells to the mucosal surfaces, CCL20 acts in the very early events leading to the development of a specific immune response. In order to characterize dendritic cells in birds and better understand their role in the initiation of immune responses against pathogens of economic as well as human health relevance, we have cloned and expressed chicken CCL20 (chCCL20) and its specific receptor chCCR6. chCCL20 has 51% identity (60% similarity) with human CCL20, while the chicken receptor and its human counterpart display nearly 55% identity (and up to 70% similarity). chCCL20 and its specific receptor chCCR6 mRNAs are mainly expressed in bone marrow, secondary lymphoid organs and in the mucosal surfaces, in particular lungs and intestine. Both receptor and chemokine are functionally active when expressed as genuine or tagged proteins in mammalian expression systems, that is chCCR6 is mainly located at the cell surface within lipid rafts like its human counterpart. And secondly, both human and chicken chemokines were able to drive the migration of either chicken or human CCR6-transfected cells.


Subject(s)
Chemokine CCL20/genetics , Chickens/genetics , Receptors, CCR6/genetics , Amino Acid Sequence , Animals , Blotting, Western , Cell Line , Cell Movement , Chemokine CCL20/metabolism , Cloning, Molecular , Gene Expression Regulation , Green Fluorescent Proteins/metabolism , Humans , Ligands , Membrane Microdomains/metabolism , Molecular Sequence Data , Protein Transport , Receptors, CCR6/chemistry , Receptors, CCR6/metabolism , Recombinant Fusion Proteins/metabolism , Sequence Alignment , Transfection
2.
Vaccine ; 27(38): 5284-91, 2009 Aug 20.
Article in English | MEDLINE | ID: mdl-19450633

ABSTRACT

Biodegradable nanoparticles with surface adsorbed antigens represent a promising method for in vivo delivery of vaccines targeting a wide range of infectious diseases or cancers. We investigated the feasibility of loading dendritic cells with a vaccine antigen, HIV p24 protein, on the surface of surfactant-free anionic (d,l-lactic acid, PLA) nanoparticles. The p24 protein had a high affinity for the nanoparticles and the antigenicity and immunogenicity of the p24 protein on the nanoparticle was well preserved after immunization. p24-coated nanoparticles were efficiently taken up by mouse dendritic cells (DCs), inducing DC maturation by increasing MHC-I, MHC-II, CD40, CD80 and CD86 surface expression and secreting IL-12 (p70) and IL-4. We evaluated the ability of DCs pulsed with p24-coated nanoparticles to elicit an optimal humoral and cellular immune response in the blood and intestine. DCs pulsed with p24-nanoparticles induced high seric and mucosal antibody production and elicited strong systemic and local lymproliferative responses, correlated with a Th1/Th2-type response, and systemic CTL responses in mice. Thus, DCs pulsed with antigen-loaded PLA nanoparticles may provide a novel delivery tool for cell therapy vaccination against chronic infectious diseases.


Subject(s)
AIDS Vaccines/immunology , Dendritic Cells/immunology , HIV Core Protein p24/immunology , Immunity, Cellular , Nanoparticles , Adjuvants, Immunologic , Animals , Cell Line , Cell Proliferation , Cytokines/immunology , Female , HIV Antibodies/blood , HIV Antibodies/immunology , HIV Core Protein p24/administration & dosage , HIV-1/immunology , Lactic Acid/immunology , Mice , Mice, Inbred CBA , Polyesters , Polymers , Th1 Cells/immunology , Th2 Cells/immunology
3.
Microbes Infect ; 9(4): 536-43, 2007 Apr.
Article in English | MEDLINE | ID: mdl-17350307

ABSTRACT

Immunization with antigen-pulsed dendritic cells (DCs) can be used to elicit optimal immune responses. We developed the SRDC cell line, with a morphology, phenotype and activity similar to mouse splenic CD4(-)CD8alpha(+)CD205(+)CD11b(-) dendritic cells, which induce a polarized Th1 immune response. We evaluated the ability of SRDCs pulsed with HIV-1 viral lysate, oligomeric soluble gp140 or capsid p24 to induce specific antibody and T-cell responses in CBA/J mice. Immunization with all loaded SRDCs elicited antibody responses against the antigens tested. However, only HIV-1 viral lysate and gp140-pulsed SRDCs elicited specific CD4(+) and CD8(+) T-cell responses. These findings demonstrate the value of well characterized DC lines for optimizing the antigen-loading mixture, according to the DC population targeted. Our data suggest that splenic DCs pulsed with complex antigens, such as HIV-1 viral lysate or oligomeric soluble gp140, could be used as vaccines, eliciting strong primary Th1-polarized and humoral immune responses against HIV proteins in vivo.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Dendritic Cells/immunology , HIV-1/immunology , Immunotherapy, Adoptive/methods , Th1 Cells/immunology , Viral Envelope Proteins/immunology , Animals , Antibody Formation , Female , Gene Products, env/immunology , HIV Core Protein p24/immunology , Interferon-gamma/biosynthesis , Interferon-gamma/immunology , Interleukin-12/biosynthesis , Interleukin-12/immunology , Interleukin-4/biosynthesis , Interleukin-4/immunology , Mice , Mice, Inbred CBA , Spleen/cytology , Spleen/immunology , T-Lymphocytes, Cytotoxic/immunology , env Gene Products, Human Immunodeficiency Virus
4.
Vaccine ; 24(10): 1705-9, 2006 Mar 06.
Article in English | MEDLINE | ID: mdl-16256251

ABSTRACT

Toxoplasma gondii enters the mucosal surfaces of the host, and so immunity at these sites is of major interest. Due to the compartmentalization of the immune response, systemic immunization does not induce high levels of immunity at mucosal surfaces. Intranasal immunization has been shown to be very effective in inducing both systemic and mucosal immune responses. Immunization with mRNA can induce both humoral and cell-mediated immune responses, both of which are important in conferring immunity to T. gondii. The efficacy of RNA vaccination by the nasal route with T. gondii RNA was evaluated. We assessed the percentage of cumulative survival after an oral challenge with a lethal dose of T. gondii cysts (40 cysts), and the number of brain cysts following a challenge with a sublethal dose of T. gondii 76 K cysts (15 cysts). Vaccinated mice were found to be significantly better protected than non-immunized mice after a challenge with a lethal dose of cysts; and a challenge with a sublethal dose also resulted in fewer brain cysts than in non-immunized mice. Sera and intestinal secretions of immunized mice recognized T. gondii antigens, suggesting that a specific humoral immune response may occur. Moreover, a specific lymphoproliferative response observed in cervical lymph nodes may confer protection. These preliminary findings suggest that RNA vaccination by a mucosal route could be feasible.


Subject(s)
Protozoan Vaccines/immunology , RNA, Protozoan/immunology , Toxoplasma/immunology , Toxoplasmosis, Animal/prevention & control , Animals , Female , Immunization , Lymphocyte Activation , Mice , Mice, Inbred C57BL , RNA, Ribosomal/immunology , Toxoplasma/genetics
5.
Infect Immun ; 72(7): 4127-37, 2004 Jul.
Article in English | MEDLINE | ID: mdl-15213158

ABSTRACT

It was previously demonstrated that immunizing mice with spleen dendritic cells (DCs) that had been pulsed ex vivo with Toxoplasma gondii antigens triggers a systemic Th1-biased specific immune response and induces protection against infection. T. gondii can cause severe sequelae in the fetuses of mothers who acquire the infection during pregnancy, as well as life-threatening neuropathy in immunocompromised patients, in particular those with AIDS. Here, we investigate the efficacy of a novel cell-free vaccine composed of DC exosomes, which are secreted antigen-presenting vesicles that express functional major histocompatibility complex class I and II and T-cell-costimulatory molecules. They have already been shown to induce potent antitumor immune responses. We investigated the potential of DC2.4 cell line-derived exosomes to induce protective immunity against toxoplasmosis. Our data show that most adoptively transferred T. gondii-pulsed DC-derived exosomes were transferred to the spleen, elicited a strong systemic Th1-modulated Toxoplasma-specific immune response in vivo, and conferred good protection against infection. These findings support the possibility that DC-derived exosomes can be used for T. gondii immunoprophylaxis and for immunoprophylaxis against many other pathogens.


Subject(s)
Adjuvants, Immunologic/pharmacology , Antigens/pharmacology , Dendritic Cells/immunology , Toxoplasma/immunology , Toxoplasmosis/immunology , Animals , Antigens/immunology , Antigens, Surface/immunology , Cell Division/immunology , Cytokines/drug effects , Cytokines/metabolism , Dendritic Cells/drug effects , Female , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Mice , Mice, Inbred C57BL , Spleen/drug effects , Spleen/immunology , Toxoplasmosis/prevention & control
6.
Infect Immun ; 71(9): 5254-65, 2003 Sep.
Article in English | MEDLINE | ID: mdl-12933872

ABSTRACT

Toxoplasma gondii, an obligate intracellular parasite pathogen which initially invades the intestinal epithelium before disseminating throughout the body, may cause severe sequelae in fetuses and life-threatening neuropathy in immunocompromised patients. Immune protection is usually thought to be performed through a systemic Th1 response; considering the route of parasite entry it is important to study and characterize the local mucosal immune response to T. gondii. Despite considerable effort, Toxoplasma-targeted vaccines have proven to be elusive using conventional strategies. We report the use of mesenteric lymph node dendritic cells (MLNDCs) pulsed ex vivo with T. gondii antigens (TAg) as a novel investigation approach to vaccination against T. gondii-driven pathogenic processes. Using a murine model, we demonstrate in two genetically distinct mouse strains (C57BL/6 and CBA/J) that adoptively transferred TAg-pulsed MLNDCs elicit a mucosal Toxoplasma-specific Th2-biased immune response in vivo and confer strong protection against infection. We also observe that MLNDCs mostly traffic to the intestine where they enhance resistance by reduction in the mortality and in the number of brain cysts. Thus, ex vivo TAg-pulsed MLNDCs represent a powerful tool for the study of protective immunity to T. gondii, delivered through its natural route of entry. These findings might impact the design of vaccine strategies against other invasive microorganisms known to be delivered through digestive tract.


Subject(s)
Dendritic Cells/immunology , Immunity, Mucosal , Th2 Cells/immunology , Toxoplasma/immunology , Adoptive Transfer , Animals , Antibodies, Protozoan/blood , Antibodies, Protozoan/metabolism , Antigens, Protozoan/administration & dosage , Cytokines/biosynthesis , Dendritic Cells/classification , Female , Humans , Immunoglobulin A, Secretory/metabolism , Immunoglobulin G/blood , Immunophenotyping , In Vitro Techniques , Intestines/immunology , Lymph Nodes/immunology , Lymphocyte Activation , Mesentery , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , Spleen/immunology , Th1 Cells/immunology , Th2 Cells/classification , Toxoplasma/pathogenicity , Toxoplasmosis, Animal/immunology
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