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  Cytoinf


  Director : CAVAILLON JEAN-MARC (jmcavail@pasteur.fr)


  abstract

 

Our group studies the cytokines involved in severe inflammatory processes in humans (sepsis, trauma, ischemia/reperfusion, and cystic fibrosis) and in murine models. These pathologies are associated with an exacerbated production of cytokines as well as an immune depression that we study at the intra-cellular signaling level. In addition, we study the regulation of the cytokine production by serotonin and interleukin-10.



  report

cale

1. MOLECULAR BASIS OF IMMUNE-DEPRESSION DURING SYSTEMIC INFLAMMATION

Sepsis syndrome affects 2.26 people per 100 hospitalized patients in the USA. Altogether, 751,000 cases a year occur in USA with a 28.6% mortality. With 9.3% of the annual deaths in this country, severe sepsis leads to as many deaths as cardiac stroke. Since we established (Carlos Muñoz et al. J. Clin. Invest. 1991, 88, 1747) the in vitro hyporeactivity of circulating monocytes in sepsis patients, we have further analyzed the immune-depression associated to this pathology. We have extended our observation to circulating neutrophils (Marie et al. Blood 1998, 91, 3439) and lymphocytes (Muret et al. Shock 2000, 13, 169) of sepsis patients and of patients suffering of non-infectious systemic inflammatory response syndrome (SIRS). Minou Adib-Conquy, in collaboration with Dr. C. Adrie (Hôpital de Saint-Denis), Prof. J-F. Dhainaut (Hôpital Cochin) and Drs P. Moine et K. Asehnoune (Hôpital du Kremlin Bicêtre), studies the intracellular molecular mechanisms responsible of the observed immune-depression in sepsis patients as well as in patients with SIRS (trauma, ischemia/reperfusion). A global decrease of nuclear factor-kB (NF-kB), a unbalance between its active (p65p50) and inactive (p50p50) forms and a weak cytoplasmic expression of its inhibitor (IkBa) have been observed within mononuclear cells of sepsis patients (Adib-Conquy et al. Am. J. Respir. Crit Care Med. 2000, 162, 1877). This observation is a reminiscence of the events occurring in in vitro endotoxin-tolerized monocytes/macrophages described by numerous authors. A similar study undertaken in trauma patients revealed that the defect in NF-kB expression lasts for more than 10 days (Adib-Conquy et al. J. Leuk. Biol. 2001, 70, 30). However, the immune depression assessed by the dramatically decrease of in vitro TNF production in response to Gram negative endotoxin (lipopolysaccharide, LPS) is not a universal event since leukocytes remain responsive to other stimuli (Staphylococcus aureus, Streptococcus pyogenes) and do produce interleukin-10 (IL-10) and interleukin-1 receptor antagonist (IL-1ra) upon stimulation by LPS. We are further analyzing various intracellular signaling pathways involved in the differential cytokine production within the cells of SIRS patients.

2. IMMUNOLOGICAL STATUS OF CARDIAC ARREST AND RESCUCITATED PATIENTS

In collaboration with Dr. C. Adrie (Hôpital de St Denis) we have studied patients after cardiac arrest and rescucitation (CAR). Prognosis of these patients is poor due to neurological sequel, hemodynamic shock and possible multiple organ failure in 50% patients. This clinical situation may be linked to a systemic ischemia-reperfusion phenomenon. Accordingly, it was of interest to investigate these patients who, despite a non-infectious stress, harbor numerous parameters found in sepsis patients. We demonstrated the presence of circulating endotoxin in 46% of patients within the first two days following admission in intensive care units. (n=35). We showed that at admission (around 3h after cardiac arrest) high levels of plasma sTNFRII, IL-6, IL-8, and IL-10 were significantly higher among non-surviving patients than in survivors. On day 1, IL-1ra was also a marker associated with prognosis. A whole blood culture showed that leukocytes of CAR patients had a hyporeactivity similar to that found in sepsis patients. Production of TNF and IL-6 in response to LPS was lower than that observed in healthy controls. The use of T-cell mitogens also revealed the altered production of IFNg and IL-10 by their T-lymphocytes. Altogether these observations show that a sepsis-like phenomenon occurs in CAR patients. Similarly, we showed the complexity of the phenomenon since some signaling pathways were altered whereas others were still functional.

3. COMPARTMENTALIZATION DURING ENDOTOXIN TOLERANCE

Catherine Fitting demonstrated that the in vivo induction of endotoxin tolerance leads to diverse states of hyporeactivity (intensity, length) of the cells depending upon the compartment they derived from (spleen, bone marrow, peritoneal cavity, lungs).

4. PARADOXICAL PRO-INFLAMMATORY ACTIVITY OF INTERLEUKIN-10

We have demonstrated that adherence is a parameter which markedly affects the properties of IL-10 on monocytes/macrophages. IL-10 pre-treated monocytes in the absence of adherence have a further enhanced capacity to produce TNF in response to LPS in subsequent adhering condition while this is not the case if IL-10 pre-treatment is achieved in adhering conditions. This paradoxical event is found at the level of mRNA as well as for NF-kB activation. We consider that our model can mimic some in vivo events when circulating monocytes encounter IL-10 before adhering to endothelium and marginating towards tissues. Anne-France Petit-Bertron has shown that there is no significant difference in response to IL-10 with or without adherence in terms of cell surface marker involved in LPS signaling (CD14, TLR4, MD2), IL-10 responsiveness (IL-10R), antigen presentation (HLA DR), co-activation (CD40) and adherence (CD11a, CD62L). Only CD11b expression was more markedly inhibited by IL-10 together with adherence as compared to non-adhering conditions. IL-10 is known to increase phagocytosis and we have shown that this effect is significantly amplified when the pre-treatment occurred in the absence of adherence. Finally, kinetics of STAT3 activation and SOCS3 induction in response to IL-10 are influenced by adherence.

5. SEROTONIN AND CYTOKINE PRODUCTION

Serotonin (5-hydroxytryptamine = 5-HT) is released by activated platelets and can be present at the micromolar concentration within the inflammatory site. Isabelle Cloëz-Tayarani studies the effect of 5-HT on cytokine production and cytokine gene expression by human peripheral blood mononuclear cells (PBMC) upon stimulation by LPS. 5-HT inhibits in a dose-dependent manner TNF production whereas it increases that of IL-1b. The productions of IL-6, IL-10 and IL-1ra remain unchanged. Melatonin, which has a structure homologous to that of 5-HT, behaves as 5-HT in inhibiting TNF production but failed to modify that of IL-1b. Present studies attempt to characterize the various intracellular pathways involved in the observed phenomenon. The use of specific agents and antagonists should help to identify the specific receptors and/or transporter involved in the modulation of cytokine production by 5-HT.

6. COLLABORATIONS

Cytokines and cystic fibrosis

With Drs Jacky Jacquot and Edith Puchelle (INSERM 314, Reims) we have studied the consequences of the interaction of circulating neutrophils with glandular epithelial cells from cystic fibrosis patients in terms of up-regulation of IL-8 production. With Drs Harriet Corvol and Annick Clément (Hôpital Trousseau) we analyze the spontaneous and LPS-induced productions of IL-8 by isolated neutrophils derived from the blood compartment or from sputum.

Cytokines and human cryptococcosis

With Drs Olivier Lortholary and Françoise Dromer (Unité de Mycologie, Institut Pasteur) we study the cytokine profiles within the cerebro-spinal fluids of HIV positive and HIV negative patients suffering or not of a cryptococcal meningitis (Lortholary O et al. J. Infect. Dis. 2001, 183, 294), and within broncho-alveolar lavages of patients with lung infections.

Cytokines and Bordetella pertussis

With Drs Laurence Bassinet and Nicole Guiso (Unité des Bordetelles, Inst. Pasteur), the study of the influence of virulence factors of Bordetella pertussis in the induction of IL-6 and IL-8 production by epithelial cells has been undertaken.



  publications

puce Publications of the unit on Pasteur's references database


  personnel

  Office staff Researchers Scientific trainees Other personnel
   

Isabelle Cloëz-Tayarani, Chargée de recherche IP, icloez@pasteur.fr

Minou Adib-Conquy, Assistante IP, madib@pasteur.fr

Anne-France Petit-Bertron PhD student

Virginie Maxime MD, Master student

Karim Asehnoune MD, Ms, trainee

Catherine Fitting Technician IP cfitting@pasteur.fr


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