20 research outputs found
Heterologous production of human papillomavirus type-16 L1 protein by a lactic acid bacterium
<p>Abstract</p> <p>Background</p> <p>The expression of vaccine antigens in lactic acid bacteria (LAB) is a safe and cost-effective alternative to traditional expression systems. In this study, we investigated i) the expression of Human papillomavirus type 16 (HPV-16) L1 major capsid protein in the model LAB <it>Lactococcus lactis </it>and ii) the ability of the resulting recombinant strain to produce either capsomer-or virus-like particles (VLPs).</p> <p>Results and conclusion</p> <p>HPV-16 L1 gene was cloned into two vectors, pCYT and pSEC, designed for controlled intra- or extracellular heterologous expression in <it>L. lactis</it>, respectively. The capacity of <it>L. lactis </it>harboring either pCYT:L1 or pSEC:L1 plasmid to accumulate L1 in the cytoplasm and supernatant samples was confirmed by Western blot assays. Electron microscopy analysis suggests that, L1 protein produced by recombinant lactococci can self-assemble into structures morphologically similar to VLPs intracellularly. The presence of conformational epitopes on the <it>L. lactis</it>-derived VLPs was confirmed by ELISA using an anti-HPV16 L1 capsid antigen antibody. Our results support the feasibility of using recombinant food-grade LAB, such as <it>L. lactis</it>, for the production of L1-based VLPs and open the possibility for the development of a new safe mucosal vector for HPV-16 prophylactic vaccination.</p
Rotavirus-Like Particles: A Novel Nanocarrier for the Gut
The delivery of bioactive molecules directly to damaged tissues represents a technological challenge. We propose here a new system based on virus-like particles (VLP) from rotavirus, with a marked tropism for the gut to deliver bio-active molecules to intestinal cells. For this, nonreplicative VLP nanoparticles were constructed using a baculovirus expression system and used to deliver an exogenous biomolecule, the green fluorescent protein (GFP), into either MA104 cells or intestinal cells from healthy and 2,4,6-trinitrobenzene sulfonic acid (TNBS)-treated mice. Our results show that expression of rotavirus capsid proteins in baculovirus led to the auto assembly of VLP that display similar properties to rotavirus. In vitro experiments showed that VLP were able to enter into MA104 cells and deliver the reporter protein. Intragastric administration of fluorescent VLP in healthy and TNBS-treated mice resulted in the detection of GFP and viral proteins in intestinal samples. Our results demonstrate an efficient entry of non-replicative rotavirus VLP into the epithelial cell line MA104 and provide the first in vivo evidence of the potential of these nanoparticles as a promising safe candidate for drug delivery to intestinal cells
Uso de bacterias lácticas como vehículos de moléculas terapéuticas
La administración de moléculas terapéuticas a través de las mucosas presenta varias ventajas importantes sobre la administración sistémica clásica, tales como: reducción de efectos secundarios, fácil administración y la posibilidad de modular la respuesta inmune tanto sistémica como local. Las superficies de mucosas son los principales sitios de interacción entre un organismo y su medio ambiente y por lo tanto representan la principal puerta de entrada de patógenos. En los últimos 15 años, una gran cantidad de trabajos científicos ha reportado una variedad de vehículos de moléculas terapéuticas, vacunas principalmente, para una inmunización efectiva a nivel de mucosas. Este tipo de administración puede además, inducir una eficiente respuesta inmune sistémica con un menor riesgo de provocar efectos colaterales secundarios como lo hacen las vacunas sistémicas clásicas. Adicionalmente, la inmunización por vía mucosal se realiza de una manera más simple sin la necesidad de jeringas y agujas, eliminando así la necesidad de personal capacitado, lo cual representa una clara ventaja en programas de vacunación masiva. Sin embargo, una desventaja importante de este tipo de administración de moléculas terapéuticas, es que una gran cantidad de proteína debe ser administrada, debido al hecho de que la mayoría de la proteína será degradada en las superficies de mucosas tales como el tracto gastro-intestinal por las enzimas digestivas y las condiciones hostiles de este sitio. Por lo tanto, el desarrollo de nuevos vectores, capaces de entregar y/o expresar eficientemente moléculas terapéuticas in situ representa un reto biotecnológico.Fil: Cortes Perez, Naima G.. Institut National de la Recherche Agronomique; FranciaFil: Leblanc, Jean Guy Joseph. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Langella, Philippe. Institut National de la Recherche Agronomique; FranciaFil: Bermudez Humaran, Luis. Institut National de la Recherche Agronomique; Franci
Modification of the porcine immature dendritic cell transcriptome and functions upon interaction with pseudorabies virus
International audienc
Intranasal Coadministration of Live Lactococci Producing Interleukin-12 and a Major Cow's Milk Allergen Inhibits Allergic Reaction in Mice
The Th1/Th2 balance deregulation toward a Th2 immune response plays a central role in allergy. We previously demonstrated that administration of recombinant Lactococcus lactis strains expressing bovine β-lactoglobulin (BLG), a major cow's milk allergen, partially prevents mice from sensitization. In the present study, we aimed to improve this preventive effect by coadministration of L. lactis BLG and a second recombinant L. lactis strain producing biologically active interleukin-12 (IL-12). This L. lactis strain producing IL-12 was previously used to enhance the Th1 immune response in a tumoral murine model (L. G. Bermúdez-Humarán et al., J. Immunol. 175:7297-7302, 2005). A comparison of the administration of either BLG alone or BLG in the presence of IL-12 was conducted. A BLG-specific primary Th1 immune response was observed only after intranasal coadministration of both L. lactis BLG and IL-12-producing L. lactis, as demonstrated by the induction of serum-specific immunoglobulin G2a (IgG2a) concomitant with gamma interferon secretion by splenocytes, confirming the adjuvanticity of IL-12-producing L. lactis. Immunized mice were further sensitized by intraperitoneal administration of purified BLG, and the allergic reaction was elicited by intranasal challenge with purified BLG. Mice pretreated with BLG in either the presence or the absence of IL-12 were rendered completely tolerant to further allergic sensitization and elicitation. Pretreatment with either L. lactis BLG or L. lactis BLG and IL-12-producing L. lactis induces specific anti-BLG IgG2a production in serum and bronchoalveolar lavage (BAL) fluid. Although specific serum IgE was not affected by these pretreatments, the levels of eosinophilia and IL-5 secretion in BAL fluid were significantly reduced after BLG challenge in the groups pretreated with L. lactis BLG and L. lactis BLG-IL-12-producing L. lactis, demonstrating a decreased allergic reaction. Our data demonstrate for the first time (i) the induction of a protective Th1 response by the association of L. lactis BLG and IL-12-producing L. lactis which inhibits the elicitation of the allergic reaction to BLG in mice and (ii) the efficiency of intranasal administration of BLG for the induction of tolerance
Exploring the host-virus miRNAs regulation layer: pig dendritic cells (DCs) infected by PrV (Pseudorabies virus)
International audienc
Mice immunization with live<em> lactococci</em> displaying a surface anchored HPV-16 E7 oncoprotein.
International audienceE7 oncoprotein of human papillomavirus-16 (HPV-16) is constitutively produced in cervical cancer (CxCa) and is a good candidate for the design of therapeutic vaccines. In this work, the nisin-controlled expression system was used to display the E7 protein at the cell surface of the food-grade Gram-positive bacterium Lactococcus lactis. An efficient cell wall anchoring of E7 was obtained. Intranasal administration of these recombinant lactococci in mice induced an HPV-16 E7-specific immune response. This is the first report of E7 cell wall anchoring in L. lactis and represents one more step towards the use of live food-grade bacteria to fight against CxCa