Abstract:

Secondary infections due to post-sepsis immunosuppression is a major cause of death in septic patients. Strategies aimed at restoring immune functions offer a new perspective. We used LPS-immunosuppressed (IS) mice to analyze the effects of all-trans Retinoic Acid (ATRA) on different immune parameters. IS mice showed decreased lymphocyte and increased myeloid-derived suppressor cells (MDSC) counts in lymph nodes. They also showed an impaired in vitro T cell proliferation,mediated by MDSC. ATRA administration restored T cell proliferation, associated to a decreased number of live MDSC. IS mice treated with ATRA showed an increased number of CD4+ and CD8+ T cells. ATRA partially improved the primary humoral immune response, even when the immunossuppression was first established, and ATRA administered afterwards. Our results demonstrate that ATRA restores immunocompetence, modulating the number of leukocytes and the survival of MDSC,representing a supplementary potential strategy in the treatment of the immunosuppressive state of sepsis.

Author affiliation: Martire Greco, Daiana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Landoni, Verónica Inés. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Chiarella, Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Rodriguez Rodrigues, Nahuel Emiliano. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Schierloh, Luis Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Rearte, María Bárbara. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Isturiz, Martín Amadeo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Fernández, Gabriela Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Abstract:

Gram-negative infections can result in endotoxic shock, which is the most common cause of death in intensive care units. Most of the undesirable effects in sepsis and septic shock have been ascribed to lipopolysaccharide (LPS), a normal constituent of the bacterial wall. The response to LPS involves rapid secretion of proinflammatory cytokines [tumour necrosis factor-α, interleukin (IL)-1, IL-6, IL-8, interferon-γ] and the concomitant induction of anti-inflammatory mediators such as IL-10 and transforming growth factor-β and glucocorticoids (GC), which render the host temporarily refractory to subsequent lethal doses of LPS challenge in a process known as LPS or endotoxin tolerance. Although protective from the development of sepsis or systemic inflammation, endotoxin tolerance has also been pointed out as the principal cause of the non-specific immunosuppression described in these patients. In this report we demonstrate, using a mouse model, that while the maintenance of tolerance is dependent upon GC, the establishment of tolerance by LPS could be inhibited by dexamethasone (Dex), a synthetic GC. Conversely, we demonstrated that mifepristone (RU486), a known GC receptor antagonist, was capable of inducing a transient and reversible disruption of endotoxin tolerance, also permitting partial restoration of the humoral immune response in LPS tolerant/immunosuppressed mice. These results are encouraging for the management of immunosuppression in sepsis and/or non-infectious shock, and deserve further investigation in the future. © 2009 British Society for Immunology.

Author affiliation: Rearte, María Bárbara. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Leucemia Experimental; Argentina

Author affiliation: Landoni, Verónica Inés. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Leucemia Experimental; Argentina

Author affiliation: Laborde, Evangelina Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Leucemia Experimental; Argentina

Author affiliation: Fernández, Gabriela Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Invest. Hematológicas ; Argentina

Author affiliation: Isturiz, Martín Amadeo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Invest. Hematológicas ; Argentina. Academia Nacional de Medicina de Buenos Aires. Instituto de Leucemia Experimental; Argentina

Abstract:

Los fenómenos de sepsis y shock séptico pueden ser causados por bacterias Gram- positivas y negativas como también por otros microorganismos. En el caso de las bacterias Gram-negativas, las endotoxinas, componentes normales de la pared bacteriana, también conocidas como lipopolisacáridos (LPS), han sido consideradas como uno de los principales agentes causales de los efectos indeseables de estas enfermedades. La respuesta a LPS implica, en principio, la generación de un estado inflamatorio, caracterizado por la rápida secreción de citoquinas proinflamatorias como el factor de necrosis tumoral (TNF)-α, interleuquina (IL) -1, IL-6, e interferón (IFN)-γ, seguido por un estado anti-inflamatorio en donde predominan mediadores como la IL-10, el factor de transformación del crecimiento-β (TGF)-β o glucocorticoides (GC), los cuales conducen al huésped a un estado de refractariedad temporal frente a una nueva exposición al LPS, un proceso conocido como tolerancia a LPS o tolerancia a endotoxina. Si bien se ha considerado como un mecanismo de protección frente al desarrollo de una sepsis o una inflamación sistémica, la tolerancia a endotoxina ha sido también señalada como una de las causas principales de la inmunosupresión inespecífica tanto humoral como celular descrita en dichos pacientes. En este trabajo demostramos, mediante un modelo murino, que si bien el mantenimiento de la tolerancia depende de los GC, el establecimiento del fenómeno puede ser inhibido por un GC sintético como la dexametasona (Dex). Por el contrario, demostramos que el mifepristone (RU486), un conocido antagonista de receptores para GC, fue capaz de inducir una desarticulación transitoria y reversible de la tolerancia a endotoxina, permitiendo también una restauración parcial de tanto la respuesta inmune adaptativa humoral como celular en ratones inmunosuprimidos inducido por LPS, sugiriendo el involucramiento de los GC endógenos en este fenómeno. Por otro lado, mediante el uso de ciclofosfamida y gemcitabina, demostramos que las células supresoras GR-1^+/CD11b^+ y las células T regulatorias CD4^+CD25^+FoxP3^+ no juegan un rol principal ni en el establecimiento ni en el mantenimiento de la tolerancia a endotoxina, un mecanismo central en la inducción de un estado de inmunosupresión. Estos resultados permiten considerar una nueva perspectiva para el manejo de la inmunosupresión en sepsis y en shock no infecciosos y merecen de mayores estudios e investigaciones en el futuro.

Sepsis and septic shock can be caused by Gram-positive and -negative bacteria and other microorganisms. In the case of Gram-negative bacteria, endotoxin, a normal constituent of the bacterial wall, also known as lipopolysaccharide (LPS), has been considered as one of the principal agents causing the undesirable effects in this critical illness. The response to LPS involves a rapid secretion of proinflammatory cytokines such as tumour necrosis factor (TNF)-a, interleukin (IL)-1, IL-6, interferon (IFN)-g and the concomitant induction of anti-inflammatory mediators such as IL-10, transforming growth factor (TGF)-b or glucocorticoids (GC), which render the host temporarily refractory to subsequent lethal doses of LPS challenge in a process known as LPS or endotoxin tolerance. Although protective from the development of sepsis or systemic inflammation, endotoxin tolerance has also been pointed out as the main cause of the non-specific humoral and cellular immunosuppression described in these patients. In this report we demonstrate, using a mouse model, that while the maintenance of tolerance is dependent upon GC, the establishment of tolerance by LPS could be inhibited by dexamethasone (Dex), a synthetic GC. Conversely, we demonstrated that mifepristone (RU486), a known GC receptor antagonist, was capable of inducing a transient and reversible disruption of endotoxin tolerance, also permitting partial restoration of both adaptive humoral and cellular immune response in LPS immunosuppressed mice, suggesting the involvement of endogenous GC in this phenomenon. On the other hand, using cyclophosphamide and gemcitabine, we demonstrated that regulatory/suppressor CD4^CD25^+FoxP3^+ and GR-1+CD11b^+ cells do not play a major role in the establishment or the maintenance of endotoxin tolerance, a central mechanism for inducing an immunosuppression state. These results are encouraging for the management of immunosuppression in sepsis and/or noninfectious shock, and deserve further investigation in the future.

Author affiliation: Rearte, María Bárbara. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

Abstract:

Large amounts of anti-inflammatory mediators, such as interleukin (IL)-10, are produced and found early in the course of sepsis. We explore the role of IL-10 on neutrophil (PMN) activation/function using an in vitro model. Isolated human PMN were pre-incubated with polysaccharide (LPS) and/or IL-10 for 18 h. Subsequently, a second LPS exposure was performed and CD11b and CD66b up-regulation, and the reactive oxygen species (ROS) generation were measured 2 h later. We found that IL-10 prevented PMN activation and the secretion of TNF-a and IL-8 induced by the first LPS contact. In the absence of IL-10, a second LPS exposure induced additive effects that were prevented by IL-10. Only ROS generation was highly affected by the blockade of PMN-secreted TNF-a or IL-8. Additionally, IL-10 prevented other possible mechanisms of LPS priming. Therefore, IL-10 modulates PMN activation preventing autocrine activating loops and priming mechanisms, rendering PMN less responsive to a second LPS exposure.

Author affiliation: Martire Greco, Daiana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Rodriguez Rodrigues, Nahuel Emiliano. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Landoni, Verónica Inés. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Rearte, María Bárbara. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Isturiz, Martín Amadeo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Author affiliation: Fernández, Gabriela Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

Abstract:

Hemolytic-uremic syndrome (HUS) is generally caused by Shiga toxin (Stx)-producing Escherichia coli. Endothelial dysfunction mediated by Stx is a central aspect in HUS development. However, inflammatory mediators such as bacterial lipopolysaccharide (LPS) and polymorphonuclear neutrophils (PMN) contribute to HUS pathophysiology by potentiating Stx effects. Acute renal failure is the main feature of HUS, but in severe cases, patients can develop neurological complications, which are usually associated with death. Although the mechanisms of neurological damage remain uncertain, alterations of the blood-brain barrier associated with brain endothelial injury is clear. Astrocytes (ASTs) are the most abundant inflammatory cells of the brain that modulate the normal function of brain endothelium and neurons. The aim of this study was to evaluate the effects of Stx type 1 (Stx1) alone or in combination with LPS in ASTs. Although Stx1 induced a weak inflammatory response, pretreatment with LPS sensitized ASTs to Stx1-mediated effects. Moreover, LPS increased the level of expression of the Stx receptor and its internalization. An early inflammatory response, characterized by the release of tumor necrosis factor alpha (TNF-α) and nitric oxide and PMN-chemoattractant activity, was induced by Stx1 in LPS-sensitized ASTs, whereas activation, evidenced by higher levels of glial fibrillary acid protein and cell death, was induced later. Furthermore, increased adhesion and PMN-mediated cytotoxicity were observed after Stx1 treatment in LPS-sensitized ASTs. These effects were dependent on NF-κB activation or AST-derived TNF-α. Our results suggest that TNF-α is a pivotal effector molecule that amplifies Stx1 effects on LPS-sensitized ASTs, contributing to brain inflammation and leading to endothelial and neuronal injury. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Author affiliation: Landoni, Verónica Inés. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: de Campos Nebel, Ildefonso Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: Schierloh, Luis Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: Calatayud, Cecilia Alicia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires; Argentina

Author affiliation: Fernández, Gabriela Cristina. Academia Nacional de Medicina de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Author affiliation: Ramos, Maria Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: Rearte, María Bárbara. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: Palermo, Marina Sandra. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina

Author affiliation: Isturiz, Martín Amadeo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina