As respostas termorregulatórias ao lipopolissacarídeo (LPS) são influenciadas por moduladores que aumentam (febrigênicos) ou diminuem (criogênicos) a temperatura corporal (Tb). Entre eles, o neurotransmissor gasoso sulfeto de hidrogênio (H2S) modula a inflamação sistêmica induzida por endotoxina em ratos, agindo como uma molécula anti-inflamatória e criogênica, embora os mecanismos subjacentes ainda sejam pouco compreendidos. Considerando que a endotoxina é um ligando para o Toll-like receptor 4 (TLR4) e que evidências recentes revelam um cross-talk entre a via de sinalização TLR e fosfo-Akt (p-Akt), o objetivo do presente estudo foi investigar se o H2S atua como um mediador antiinflamatório e antipirético durante as fases termorregulatórias que ocorrem no choque endotoxêmico (hipotermia e febre) induzido por lipopolissacarídeo bacteriano (LPS, 2,5 mg / kg intraperitoneal (ip)) através da modulação sobre a produção de prostaglandina D2 (PGD2) e a ativação de Akt na área pré-óptica ântero-ventral do hipotálamo (AVPO). A Tb profunda de ratos mantidos a uma temperatura ambiente de 25 °C foi registrada antes e depois da inibição farmacológica da enzima cistationina ?-sintase (CBS - responsável pela produção endógena de H2S no cérebro) usando aminooxiacetato (AOA, 100 pmol, intracerebroventricular (icv)), combinado ou não com administração de LPS. Para esclarecer os mecanismos responsáveis por esses ajustes da resposta imune, foram determinados na AVPO os níveis de H2S, a produção de PGD2 e o perfil de expressão das proteínas CBS, p-Akt e p-CREB. Além disso, foi analisada a concentração de citocinas plasmáticas (IL-1?, IL-6, IL-10, TNF?, IFN-? , E IL-4). A injeção ip de LPS causou hipotermia típica seguida de febre. Os níveis de AVPO H2S aumentaram significativamente durante a hipotermia quando comparado com ratos eutérmicos e febris. A microinjeção icv de AOA não causou nenhuma alteração na Tb nem na produção basal de PGD2 durante a eutermia. Em ratos tratados com LPS, o AOA causou uma atenuação na queda da Tb durante a fase de hipotermia e uma febre exacerbada, simultaneamente com o aumento na produção de PGD2 e abolição do aumento induzido pela endotoxina na atividade de Akt. Durante a fase de febre, a expressão relativa de CBS esteve significativamente diminuída enquanto a expressão relativa de p-Akt esteve aumentada, quando comparado com ratos eutérmicos e hipotérmicos. As citocinas plasmáticas aumentaram durante a inflamação sistêmica, mas apenas a IL-4 mostrou um padrão semelhante em relação à Akt. Estes dados são consistentes com a noção de que o neurotransmissor gasoso H2S modula as fases de hipotermia e febre durante o choque endotoxêmico, atuando como uma molécula criogênica. Este papel anti-inflamatório durante a inflamação sistémica envolve uma regulação positiva da PGD2, de Akt e da IL-4 plasmática. / Thermoregulatory responses to lipopolysaccharide (LPS) are affected by modulators that increase (pro-pyretic) or decrease (cryogenic) body temperature (Tb). Among them, the gaseous messenger hydrogen sulfide (H2S) modulates endotoxin-induced systemic inflammation being an anti-inflammatory and cryogenic molecule, although the underlying mechanisms are still poorly understood. Since endotoxin is a Toll-like receptor 4 (TLR4) ligand and recent evidence indicates that there is a possible a cross-talk between the TLR and phospho-Akt (p-Akt) signaling pathway, the current study aimed to investigate whether H2S acts as an anti-inflammatory and anti-pyretic mediator during thermoregulatory phases of endotoxic shock (hypothermia and fever) induced by bacterial lipopolysaccharide (LPS, 2.5 mg/kg intraperitoneal (ip)) through the modulation of prostaglandin D2 (PGD2) production and activation of Akt in the anteroventral preoptic region of the hypothalamus (AVPO). Deep Tb in rats kept at an ambient temperature of 25 °C, was recorded before and after pharmacological inhibition of the enzyme cystathionine ?-synthase (CBS - responsible for H 2S endogenous production in the brain) using aminooxyacetate (AOA; 100 pmol/1 ?l intracerebroventricular (icv)) combined or not with endotoxin administration. To clarify the mechanisms responsible for these adjustments on immune response were verified in the AVPO H 2S levels, PGD2 production and expression profiles of CBS, p-Akt and p-CREB. In addition, plasma cytokines concentration (IL-1?, IL-6, IL-10, TNF?, IFN-?, and IL-4) was analyzed. Intraperitoneal injection of LPS caused typical hypothermia followed by fever. Intracerebroventricular microinjection of AOA neither affected Tb nor basal PGD2 production during euthermia. Levels of AVPO H2S were significantly increased during hypothermia when compared to both euthermic and febrile rats. In LPS-treated rats, AOA increased Tb values during hypothermia and fever, along with enhanced PGD2 production and abolition of endotoxin-induced increase in Akt activity. During fever, CBS relative expression was significantly decreased whereas p-Akt was significantly increased when compared to both euthermic and hypothermic rats. Plasma cytokines were increased during systemic inflammation, but only IL-4 showed a similar pattern in relation to Akt. These data are consistent with the notion that the gaseous messenger H2S modulates hypothermia and fever during endotoxic shock, acting as a cryogenic molecule. This anti-inflammatory role during systemic inflammation involves a H2S-induced up-modulation of PGD2, Akt and plasma IL-4.

L'arthrose est une maladie multifactorielle complexe. Parmi les facteurs impliqués dans sa pathogénie, les certains prostaglandines exercent un rôle inflammatoire et d’autres un rôle protecteur. La prostaglandine D2 (PGD2) est bien connue comme une PG anti-inflammatoire, qui est régulée par l’enzyme «Lipocalin prostaglandine D-synthase». Avec l’inflammation de l'arthrose, les chondrocytes essaient de protéger le cartilage en activant certaines voies de récupération dont l'induction du gène L-PGDS. Dans cette étude, nous étudions la voie de signalisation impliquée dans la régulation de l'expression du (L-PGDS) sur les chondrocytes traités avec différents médiateurs inflammatoires. Le but de projet: Nous souhaitons étudier la régulation de la L-PGDS dans le but de concevoir des approches thérapeutiques qui peuvent activer la voie intrinsèque anti-inflammatoire. Méthode et conclusions: In vivo, l'arthrose a été suivie en fonction de l’âge chez la souris ou chirurgicalement suivant une intervention au niveau des genoux de souris. Nous avons confirmé les niveaux d’expression de L-PGDS histologiquement et par immunohistochimie. In vitro, dans les chondrocytes humains qui ont été traités avec différents médiateurs de l'inflammation, nous avons observé une augmentation de l’expression de la L-PGDS dose et temps dépendante. Nous avons montré, in vivo et in vitro que l’inflammation induit une sécrétion chondrocytaire de la L-PGDS dans le milieu extracellulaire. Enfin, nous avons observé la production de différentes isoformes de la L-PGDS en réponse à l'inflammation. / Osteoarthritis is a complex multifactorial disease; many factors are involved in its pathogenesis, among those factors prostaglandins. Some prostaglandins have inflammatory role and some have anti-inflammatory role. Prostaglandin D2 (PGD2) is a well-established anti-inflammatory PG which is synthesized by the Lipocalin prostaglandin-D synthase (L-PGDS) enzyme. Upon the initiation of the inflammatory process in osteoarthritis, chondrocytes try to save themselves by activating salvage pathways, among these pathways is the induction of L-PGDS gene. In this study we are addressing the signaling pathways involved in the regulation of (L-PGDS) gene expression, in chondrocytes treated with different inflammatory mediator. Rationale: understanding the regulation of L-PGDS will allow us to design therapeutic targets that can switch on the intrinsic anti-inflammatory pathway. Method and findings: In vivo, osteoarthritis was induced in mice knees surgically or naturally following aging, the development of osteoarthritis was then confirmed histologically. The expression levels of L-PGDS were detected by Immunohistochemistry. In vitro, human chondrocytes were treated with different inflammatory mediators (interleukin-1 beta, interleukin-17, hydrogen peroxide and tert-Butyl hydroperoxide). Interestingly, in most cases chondrocytes increased expression of L-PGDS in a dose and time dependent manner. We discovered that chondrocytes release L-PGDS into the extracellular space in response to inflammation, in vivo and in vitro. Lastly we observed different isoforms of L-PGDS generated in response to inflammation.