Role of interleukin-1 in the pathogenesis of the infection caused by Streptococcus suis serotype 2

Streptococcus suis sérotype 2 est un pathogène important du porc et un agent zoonotique en émergence causant des morts subites (porcs), des chocs septiques (humains) et des méningites (chez les deux espèces), où une réaction inflammatoire sévère est caractéristique de l’infection. Une réponse rapide et efficace du système immunitaire inné contre S. suis est critique pour contrôler la croissance bactérienne et pour limiter la propagation du pathogène sans occasionner une inflammation excessive. Bien que l’interleukine (IL)-1 soit considérée comme l’un des médiateurs pro-inflammatoires les plus efficaces et produit le plus rapidement, son rôle dans la pathogénèse de l’infection par S. suis n’a pas encore été étudié. En utilisant un modèle murin d’infection systémique bien standardisé, nous avons démontré que la souche nord-américaine de virulence intermédiaire de « sequence type » (ST) 25, la souche européenne hautement virulente ST1 ainsi que la souche épidémique chinoise ST7, induisent toutes de hauts niveaux d’IL-1 dans des organes de filtration, tels que le foie et la rate. De plus, les cellules dendritiques et les macrophages, deux types de cellules jouant un rôle central dans la pathogénèse de S. suis, sont des sources importantes de cette cytokine. Les études des mécanismes impliqués dans la production de cette cytokine ont démontré que la production d’IL-1, indépendamment de la souche bactérienne utilisée, dépendait de MyD88 et impliquait les récepteurs TLR2 et possiblement TLR7 et TLR9. Cela suggère que les composantes bactériennes responsables de l’activation cellulaire sont similaires et conservées entre les différentes souches. Cependant, seuls de très hauts niveaux de suilysine, produite par la souche ST7, provoquaient une maturation importante de proIL 1β. Cette maturation implique l’activation des inflammasomes NLRP3, NLRP1, AIM2 et NLRC4, qui est due à la formation de pores et à un efflux d’ions. De surcroît, nous avons évalué le rôle global de cette cytokine chez des souris IL-1R-/-, démontrant que l’IL-1 pourrait jouer un rôle bénéfique lors d’une infection systémique par S. suis en modulant l’inflammation requise pour contrôler et éliminer la charge bactérienne, ce qui favorise la survie de l’hôte. Toutefois, au-delà d’un certain seuil, l’inflammation causée par S. suis ne peut plus être contrebalancée par cette signalisation, ce qui complique la détermination exacte du rôle de l’IL-1. Une meilleure compréhension des mécanismes sous-jacents impliqués dans le contrôle de l’inflammation et de la charge bactérienne aiderait à développer de meilleures mesures de contrôle pour ce pathogène important à la fois chez le porc et chez l’Homme.Streptococcus suis serotype 2 is an important porcine bacterial pathogen and an emerging zoonotic agent causing sudden death (pigs), septic shock (humans), and meningitis (both species), with exacerbated inflammation being a hallmark of the infection. A rapid, effective, and balanced innate immune response against S. suis is critical to control bacterial growth and limit the spread of the pathogen without causing excessive inflammation. Even though interleukin (IL)-1 is regarded as one of the most potent and earliest pro-inflammatory mediators produced, its role in the S. suis pathogenesis has not been studied. Using a well-standardized mouse model of systemic infection, we showed that an intermediately pathogenic sequence type (ST) 25 North American strain, a highly pathogenic ST1 European strain, and the epidemic ST7 Chinese strain induce high levels of IL-1 in important filter organs such as liver and spleen. Moreover, dendritic cells and macrophages, which are two cell types centrally involved in the S. suis pathogenesis, are important sources of this cytokine, with the ST7 strain secreting the highest levels. The study of the underlying mechanisms involved in this production showed that, independently of the strain, IL-1β production required MyD88 and involved recognition via TLR2 and possibly TLR7 and TLR9. This suggests that recognized bacterial components are similar and conserved between S. suis strains. However, very high levels of the pore-forming toxin suilysin produced by the ST7 strain only, are required for efficient maturation of proIL 1β. Such maturation involved the activation of the NLRP3, NLRP1, AIM2, and NLRC4 inflammasomes via pore formation and ion efflux. Using IL-1R-/- mice, we demonstrated that IL-1 signaling may play a beneficial role during S. suis systemic infection by modulating the inflammation required to control and clear bacterial burden, thus, promoting host survival. Beyond a certain threshold, however, S. suis-induced inflammation cannot be counter-balanced by this signaling, making it difficult to discriminate its role. A better understanding of the underlying mechanisms involved in the control of inflammation could help to develop control measures for this important porcine and zoonotic agent

Interleukin-1 signaling induced by Streptococcus suis serotype 2 is strain-dependent and contributes to bacterial clearance and inflammation during systemic disease in a mouse model of infection

International audienceAbstractStreptococcus suis serotype 2 is an important porcine pathogen and zoonotic agent causing sudden death, septic shock and meningitis, with exacerbated inflammation being a hallmark of the infection. A rapid, effective and balanced innate immune response against S. suis is critical to control bacterial growth without causing excessive inflammation. Even though interleukin (IL)-1 is one of the most potent and earliest pro-inflammatory mediators produced, its role in the S. suis pathogenesis has not been studied. We demonstrated that a classical virulent European sequence type (ST) 1 strain and the highly virulent ST7 strain induce important levels of IL-1 in systemic organs. Moreover, bone marrow-derived dendritic cells and macrophages contribute to its production, with the ST7 strain inducing higher levels. To better understand the underlying mechanisms involved, different cellular pathways were studied. Independently of the strain, IL-1β production required MyD88 and involved recognition via TLR2 and possibly TLR7 and TLR9. This suggests that the recognized bacterial components are similar and conserved between strains. However, very high levels of the pore-forming toxin suilysin, produced only by the ST7 strain, are required for efficient maturation of pro-IL-1β via activation of different inflammasomes resulting from pore formation and ion efflux. Using IL-1R−/− mice, we demonstrated that IL-1 signaling plays a beneficial role during S. suis systemic infection by modulating the inflammation required to control and clear bacterial burden, thus promoting host survival. Beyond a certain threshold, however, S. suis-induced inflammation cannot be counterbalanced by this signaling, making it difficult to discriminate its role

Recognition of Lipoproteins by Toll-like Receptor 2 and DNA by the AIM2 Inflammasome Is Responsible for Production of Interleukin-1β by Virulent Suilysin-Negative Streptococcus suis Serotype 2

Streptococcus suis serotype 2 is an important porcine bacterial pathogen and zoonotic agent causing sudden death, septic shock and meningitis. These pathologies are the consequence of an exacerbated inflammatory response composed of various mediators including interleukin (IL)-1β. Elevated levels of the toxin suilysin (SLY) were demonstrated to play a key role in S. suis-induced IL-1β production. However, 95% of serotype 2 strains isolated from diseased pigs in North America, many of which are virulent, do not produce SLY. In this study, we demonstrated that SLY-negative S. suis induces elevated levels of IL-1β in systemic organs, with dendritic cells contributing to this production. SLY-negative S. suis-induced IL-1β production requires MyD88 and TLR2 following recognition of lipoproteins. However, the higher internalization rate of the SLY-negative strain results in intracellularly located DNA being recognized by the AIM2 inflammasome, which promotes IL-1β production. Finally, the role of IL-1 in host survival during the S. suis systemic infection is beneficial and conserved, regardless of SLY production, via modulation of the inflammation required to control bacterial burden. In conclusion, this study demonstrates that SLY is not required for S. suis-induced IL-1β production

Renal response to L-arginine in diabetic rats. A possible link between nitric oxide system and aquaporin-2.

The aim of this study was to evaluate whether L-Arginine (L-Arg) supplementation modifies nitric oxide (NO) system and consequently aquaporin-2 (AQP2) expression in the renal outer medulla of streptozotocin-diabetic rats at an early time point after induction of diabetes. Male Wistar rats were divided in four groups: Control, Diabetic, Diabetic treated with L-Arginine and Control treated with L-Arginine. Nitric oxide synthase (NOS) activity was estimated by [14C] L-citrulline production in homogenates of the renal outer medulla and by NADPH-diaphorase staining in renal outer medullary tubules. Western blot was used to detect the expression of AQP2 and NOS types I and III; real time PCR was used to quantify AQP2 mRNA. The expression of both NOS isoforms, NOS I and NOS III, was decreased in the renal outer medulla of diabetic rats and L-Arg failed to prevent these decreases. However, L-Arg improved NO production, NADPH-diaphorase activity in collecting ducts and other tubular structures, and NOS activity in renal homogenates from diabetic rats. AQP2 protein and mRNA were decreased in the renal outer medulla of diabetic rats and L-Arg administration prevented these decreases. These results suggest that the decreased NOS activity in collecting ducts of the renal outer medulla may cause, at least in part, the decreased expression of AQP2 in this model of diabetes and constitute additional evidence supporting a role for NO in contributing to renal water reabsorption through the modulation of AQP2 expression in this pathological condition. However, we cannot discard that another pathway different from NOS also exists that links L-Arg to AQP2 expression

Mitochondrial Dysfunction in Brain Cortex Mitochondria of STZ-Diabetic Rats: Effect of L-Arginine

Mitochondrial dysfunction has been implicated in many diseases, including diabetes. It is well known that oxygen free radical species are produced endogenously by mitochondria, and also nitric oxide (NO) by nitric oxide synthases (NOS) associated to mitochondrial membranes, in consequence these organelles constitute main targets for oxidative damage. The aim of this study was to analyze mitochondrial physiology and NO production in brain cortex mitochondria of streptozotocin (STZ) diabetic rats in an early stage of diabetes and the potential effect of L-arginine administration. The diabetic condition was characterized by a clear hyperglycaemic state with loose of body weight after 4 days of STZ injection. This hyperglycaemic state was associated with mitochondrial dysfunction that was evident by an impairment of the respiratory activity, increased production of superoxide anion and a clear mitochondrial depolarization. In addition, the alteration in mitochondrial physiology was associated with a significant decrease in both NO production and nitric oxide synthase type I (NOS I) expression associated to the mitochondrial membranes. An increased level of thiobarbituric acid-reactive substances (TBARS) in brain cortex homogenates from STZ-diabetic rats indicated the presence of lipid peroxidation. L-arginine treatment to diabetic rats did not change blood glucose levels but significantly ameliorated the oxidative stress evidenced by lower TBARS and a lower level of superoxide anion. This effect was paralleled by improvement of mitochondrial respiratory function and a partial mitochondrial repolarization.In addition, the administration of L-arginine to diabetic rats prevented the decrease in NO production and NOSI expression. These results could indicate that exogenously administered L-arginine may have beneficial effects on mitochondrial function, oxidative stress and NO production in brain cortex mitochondria of STZ-diabetic rats.Fil: Ortiz, María del Carmen. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Lores Arnaiz, Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular; Argentina;Fil: Albertoni Borghese, Maria Florencia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Balonga, Sabrina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Lavagna, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Farmacológicas (i); Argentina; Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Filipuzzi, Ana Laura. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Cicerchia, Daniela Griselda. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Química Analítica y Fisicoquímica. Cátedra de Fisicoquímica; Argentina; Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Bioquimica y Medicina Molecular; Argentina;Fil: Majowicz, Mónica Patricia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina;Fil: Bustamante, Juanita. Universidad Abierta Interamericana. Facultad de Medicina. Centro de Altos Estudios en Ciencias de la Salud; Argentina; Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas. Cátedra de Biología Celular y Molecular; Argentina

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AQP2 mRNA levels in the renal outer medulla.

<p>AQP2 mRNA levels are expressed as relative values from control untreated rats. The following formula was applied: ΔΔCT = (<i>CTAQP</i>2−<i>CTGAPDH</i>) <i>experimental</i>−(<i>CT AQP</i>2−<i>CT GAPDH</i>)<i>control untreated rats</i>. Two-way ANOVA showed a statistically significant (p<0.001) interaction between the effects of Diabetes and L-Arg treatment on AQP2 mRNA expression. Bonferroni’s post- tests: *p<0.05 vs. control untreated rats; ***p<0.001 vs. control untreated rats; &&&p<0.001 vs. diabetic untreated rats. Data are mean ± SEM (n = 6).</p

Summary of some general and renal parameters in control and diabetic rats.

<p>C: control rats; D: diabetic untreated rats; D+A: diabetic rats treated with L-Arg; C+A: control rats treated with L-Arg. Results are expressed as the mean ± SEM (n = 6).</p><p>Two-way ANOVA showed a statistically significant interaction (p<0.05) between the effects of Diabetes and L-Arg treatment on NOx urinary excretion. There was no interaction between the effects of Diabetes and L-Arg treatment on the other parameters. Diabetes had a significant overall effect on Final body weight (p<0.001), Kidney weight/body weight (p<0.001), Blood glucose (p<0.001), Urinary volume (p<0.05), Osmolar excretion rate (p<0.01), Urinary glucose (p<0.001) and Urinary ketones (p<0.001). L-Arg had no significant effect on any of the parameters tested. *p<0.05 vs. C; **p<0.01 vs. C; ***p<0.001 vs. C; ^#p<0.05 vs. C+A; ^##p<0.01 vs. C+A ^###p<0.001 vs. C+A; ^&&&p<0.001 vs. D.</p

NOS activity measured as pmol [¹⁴C] L-citrulline/g tissue/min.

<p>Two-way ANOVA showed no statistically significant interaction between the effects of Diabetes and L-Arg treatment on NOS activity. The effects of Diabetes and L-Arg were considered extremely significant (p<0.001). ***p<0.001 vs. control untreated rats; ^###p<0.001 vs. control rats treated with L-Arg; ^&&&p<0.001 vs. diabetic untreated rats. Data are mean ± SEM (n = 8).</p