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1.
J Control Release ; 293: 158-171, 2019 01 10.
Article in English | MEDLINE | ID: mdl-30496771

ABSTRACT

The study of capture and processing of antigens (Ags) by intestinal epithelial cells is very important for development of new oral administration systems. Efficient oral Ag delivery systems must resist enzymatic degradation by gastric and intestinal proteases and deliver the Ag across biological barriers. The recombinant unlipidated outer membrane protein from Brucella spp. (U-Omp19) is a protease inhibitor with immunostimulatory properties used as adjuvant in oral vaccine formulations. In the present work we further characterized its mechanism of action and studied the interaction and effect of U-Omp19 on the intestinal epithelium. We found that U-Omp19 inhibited protease activity from murine intestinal brush-border membranes and cysteine proteases from human intestinal epithelial cells (IECs) promoting co-administered Ag accumulation within lysosomal compartments of IECs. In addition, we have shown that co-administration of U-Omp19 facilitated the transcellular passage of Ag through epithelial cell monolayers in vitro and in vivo while did not affect epithelial cell barrier permeability. Finally, oral co-delivery of U-Omp19 in mice induced the production of Ag-specific IgA in feces and the increment of CD103+ CD11b- CD8α+ dendritic cells subset at Peyer's patches. Taken together, these data describe a new mechanism of action of a mucosal adjuvant and support the use of this rationale/strategy in new oral delivery systems for vaccines.


Subject(s)
Adjuvants, Immunologic/administration & dosage , Antigens, Bacterial/administration & dosage , Bacterial Outer Membrane Proteins/administration & dosage , Intestinal Mucosa/metabolism , Lipoproteins/administration & dosage , Protease Inhibitors/administration & dosage , Vaccines/administration & dosage , Administration, Oral , Animals , Caco-2 Cells , Epithelial Cells/metabolism , Female , HT29 Cells , Humans , Mice, Inbred BALB C
2.
Front Immunol ; 8: 171, 2017.
Article in English | MEDLINE | ID: mdl-28261222

ABSTRACT

Most pathogens infect through mucosal surfaces, and parenteral immunization typically fails to induce effective immune responses at these sites. Development of oral-administered vaccines capable of inducing mucosal as well as systemic immunity while bypassing the issues of antigen degradation and immune tolerance could be crucial for the control of enteropathogens. This study demonstrates that U-Omp19, a bacterial protease inhibitor with immunostimulatory features, coadministered with Salmonella antigens by the oral route, enhances mucosal and systemic immune responses in mice. U-Omp19 was able to increase antigen-specific production of IFN-γ and IL-17 and mucosal (IgA) antibody response. Finally, oral vaccination with U-Omp19 plus Salmonella antigens conferred protection against virulent challenge with Salmonella Typhimurium, with a significant reduction in bacterial loads. These findings prove the efficacy of this novel adjuvant in the Salmonella infection model and support the potential of U-Omp19 as a suitable adjuvant in oral vaccine formulations against mucosal pathogens requiring T helper (Th)1-Th17 protective immune responses.

3.
J Control Release ; 220(Pt A): 18-28, 2015 Dec 28.
Article in English | MEDLINE | ID: mdl-26456256

ABSTRACT

We report here that a bacterial protease inhibitor from Brucella spp. called U-Omp19 behaves as an ideal constituent for a vaccine formulation against infectious diseases. When co-administered orally with an antigen (Ag), U-Omp19: i) can bypass the harsh environment of the gastrointestinal tract by inhibiting stomach and intestine proteases and consequently increases the half-life of the co-administered Ag at immune inductive sites: Peyer's patches and mesenteric lymph nodes while ii) it induces the recruitment and activation of antigen presenting cells (APCs) and increases the amount of intracellular Ag inside APCs. Therefore, mucosal as well as systemic Ag-specific immune responses, antibodies, Th1, Th17 and CD8(+) T cells are enhanced when U-Omp19 is co-administered with the Ag orally. Finally, this bacterial protease inhibitor in an oral vaccine formulation confers mucosal protection and reduces parasite loads after oral challenge with virulent Toxoplasma gondii.


Subject(s)
Antigens/metabolism , Bacterial Proteins/pharmacology , Brucella/chemistry , Immunity, Mucosal , Protease Inhibitors/pharmacology , Vaccines/immunology , Administration, Oral , Amino Acid Sequence , Animals , Female , Mice , Mice, Inbred Strains , Molecular Sequence Data
4.
PLoS One ; 8(7): e69438, 2013.
Article in English | MEDLINE | ID: mdl-23861971

ABSTRACT

The discovery of novel mucosal adjuvants will help to develop new formulations to control infectious and allergic diseases. In this work we demonstrate that U-Omp16 from Brucella spp. delivered by the nasal route (i.n.) induced an inflammatory immune response in bronchoalveolar lavage (BAL) and lung tissues. Nasal co-administration of U-Omp16 with the model antigen (Ag) ovalbumin (OVA) increased the amount of Ag in lung tissues and induced OVA-specific systemic IgG and T helper (Th) 1 immune responses. The usefulness of U-Omp16 was also assessed in a mouse model of food allergy. U-Omp16 i.n. administration during sensitization ameliorated the hypersensitivity responses of sensitized mice upon oral exposure to Cow's Milk Protein (CMP), decreased clinical signs, reduced anti-CMP IgE serum antibodies and modulated the Th2 response in favor of Th1 immunity. Thus, U-Omp16 could be used as a broad Th1 mucosal adjuvant for different Ag formulations.


Subject(s)
Adjuvants, Immunologic , Bacterial Outer Membrane Proteins/immunology , Brucella/immunology , Milk Hypersensitivity/immunology , Milk Proteins/immunology , Th1 Cells/immunology , Th2 Cells/immunology , Adjuvants, Immunologic/administration & dosage , Animals , Antigens/immunology , Antigens/metabolism , Bacterial Outer Membrane Proteins/administration & dosage , Bacterial Outer Membrane Proteins/chemistry , Bronchoalveolar Lavage Fluid/cytology , Bronchoalveolar Lavage Fluid/immunology , Cattle , Central Nervous System/immunology , Central Nervous System/pathology , Cytokines/biosynthesis , Disease Models, Animal , Female , Immunoglobulin E/immunology , Immunoglobulin G/immunology , Lung/immunology , Lung/pathology , Mice , Milk Hypersensitivity/metabolism , Nasal Mucosa/immunology , Nasal Mucosa/metabolism , Spleen/immunology , Th1 Cells/metabolism , Th2 Cells/metabolism
5.
Neuroendocrinology ; 92(4): 207-14, 2010.
Article in English | MEDLINE | ID: mdl-20975260

ABSTRACT

Dopamine D2 receptor (D2R) participation in prolactin regulation is well documented, but the role of D2Rs in the control of other hormones involved in growth, food intake and glucose metabolism has not been extensively studied. The study of D2R knockout mice (Drd2(-/-)) puts forward new insights into the role of the D2R in growth hormone (GH)-releasing hormone-GH regulation, peptides involved in food intake, glucose homeostasis, as well as in prolactinoma development. The expected phenotype of chronic hyperprolactinemia and prolactinoma development was found in the Drd2(-/-) mouse, and this model constitutes a valuable tool in the study of dopamine-resistant prolactinomas. Unexpectedly, these mice were growth retarded, and the importance of functional hypothalamic D2Rs in the neonatal period was revealed. In the Drd2(-/-) mouse there was a failure of high neonatal GH levels and therefore the expansion of pituitary somatotropes was permanently altered. These mice also had increased food intake, and a sexually dimorphic participation of the D2R in food intake regulation is suggested. The effect described is probably secondary to D2R regulation of prolactin secretion. Furthermore, the negative modulation of D2Rs on α-melanocyte-stimulating hormone release and positive action on the hypothalamic expression of orexins reveals the complex D2R regulation of food intake. Finally, pancreatic D2Rs inhibit glucose-stimulated insulin release. Lack of dopaminergic inhibition throughout development in the Drd2(-/-) mouse may exert a gradual deteriorating effect on insulin homeostasis, so that eventually glucose intolerance develops. These results highlight the complex endocrine actions of the D2Rs at different levels, hypothalamus, pituitary or pancreas, which function to improve fitness, reproductive success and survival.


Subject(s)
Endocrine System/physiology , Metabolism/genetics , Receptors, Dopamine D2/genetics , Receptors, Dopamine D2/physiology , Animals , Eating/genetics , Eating/physiology , Endocrine System/metabolism , Growth Hormone/metabolism , Growth Hormone-Releasing Hormone/metabolism , Mice , Mice, Knockout , Prolactin/metabolism , Receptors, Dopamine D2/metabolism
6.
Front Horm Res ; 38: 59-69, 2010.
Article in English | MEDLINE | ID: mdl-20616496

ABSTRACT

The role of dopaminergic receptors in the control of GH release remains controversial. The dopamine receptor 2 (D2R) knockout mouse represents a useful model to study the participation of the D2R on growth and GHRH-GH regulation. These knockout mice have hyperprolactinemia and lactotrope hyperplasia, but unexpectedly, they are also growth retarded. In D2R knockout mice there is a significant decrease in somatotrope population, which is paralleled by decreased GH content and output from pituitary cells. The sensitivity of GHRH-induced GH and cAMP release is similar between genotypes, even though the response amplitude is lower in knockouts. We point to an involvement of D2R signaling at the hypothalamic level as dopamine did not release GH acting at the pituitary level, and both somatostatin and GHRH mRNA expression are altered in knockout mice. The similarity of the pituitary defect in the D2R knockout mouse to that of GHRH deficient models suggests a probable mechanism. Loss of dopamine signaling via hypothalamic D2Rs at a critical age may cause inadequate GHRH secretion subsequently leading to inappropriate somatotrope lineage development. Furthermore, GH pulsatility, which depends on a regulated temporal balance between GHRH and somatostatin output might be compromised in D2R knockout mice, leading to lower IGF-I, and growth retardation.


Subject(s)
Dopamine/physiology , Growth Hormone-Releasing Hormone/physiology , Growth Hormone/physiology , Neurotransmitter Agents/physiology , Acromegaly/drug therapy , Animals , Growth , Humans , Mice , Receptors, Dopamine D2/physiology
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