Increased IL-10 levels during dengue virus infection. Dr. Ana B. Perez Dept. Virology Institute for Tropical Medicine Pedro Kouri Cuba
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1 Increased IL-10 levels during dengue virus infection Dr. Ana B. Perez Dept. Virology Institute for Tropical Medicine Pedro Kouri Cuba
2 IPK Dengue virus infections: Spectrum of an illness Asymptomatic Dengue Fever Dengue Hemorrhagic Fever/ Dengue Shock Syndrome Plasma leakage Hemorrhage Hemoconcentration
3 Possible role of cytokines in DHF/SSD DV-infected monocyte Cytokines plasma leakage Activated T cell
4 Cytokines in DHF Th1 Th2 IFNγ IL-10 IL-12 TGFβ Chaturvedi UC, FEMS Immunol 2000 IPK
5 Cytokines in DHF IFN-α IFN-γ IL-10 stnfr-ii sil-2r # days after peak viremia 4 Libraty et al. J Infect Dis 2002; 185: IPK
6 CUBA: Exceptional epidemiological conditions Cases vd1 vd2 Not Endemicity vd2 vd3 vd Dengue Outbreaks (Years)
7 Objectives General: To contribute to the elucidation of the role of some innate immune response soluble mediators in the development of Dengue Hemorrhagic Fever Specific: To measure the levels of RANTES, IL-12 and IL-10 in serum samples from Cuban dengue patients. To correlate these levels with illness severity. IPK
8 Material and Methods: 34 patients Dengue 2 outbreak (Santiago de Cuba, 1997) Clinical, serological diagnosis (IgM) and viral isolation. 28--FD 6--FHD Collection of serum samples in three times after clinical upset 12 serum samples from healthy donors (Negative controls) Serum RANTES, IL-12 (p70+p40) and IL-10 cuantification by ELISA kits (ENDOGEN) IPK
9 Levels of RANTES in sera from patients with primary and secondary infections 130 RANTES (pg/ml) DF1 DF2 DHF DF1 DF2 DHF DF1 DF2 DHF Contro st 2nd 3rd Days Determinations after fever upset IPK
10 Levels of IL-12 in sera from patients with DF and DHF IL-12 (ug/ml) DHF DF DHF DF DHF DF Cont Days after fever upset
11 IPK Levels of IL-10 in sera from patients with DF and DHF 250 IL-10 IL-10 (ug/ml) DHF DF DHF DF DHF DF Controls Days Determinations after fever upset
12 Levels of IL-10 in sera from patients with primary or secondary infections IL-10 (pg/ml) DF1 DF2 DHF DF1 DF2 DHF DF1 DF2 DHF Contro Days after fever onset IPK
13 IPK Summarizing: Serum levels of RANTES were lower in DHF patients than controls. Controls and FD patients did not show significatively differences. IL-12 levels in dengue patients were no significatively higher than negative controls. IL-10 levels were much higher in dengue patients comparing to controls, and DHF patients show the highest serum levels of this cytokine, reached maximal values on the 4th and 5th days of the disease. When we compared primary and secondary cases DF cases, only secondary cases show high levels.
14 IPK Are pre-existing heterotypic antibodies involved in the increasing of IL-10 serum levels in patients with dengue secondary infection? To quantify the levels of IL-12 and IL-10 in supernatants of mononuclear cells from individual with antecedent of primary dengue 1 infection cultured with dengue virus in presence of autologous serum.
15 Healthy donors with antecedent of primary dengue 1 infection (neutralización assay) Bleeding Isolation of Peripheral Mononuclear cells Collection of serum samples. Culture of PBMC with DV 1 or DV 2 in RPMI medium suplemented with: Fetal Bovine Serum. autologous serum(abs anti-d1) Incubation for 48 hs (37 o C, 5%CO 2 ) Determination of IL-10 and IL-12 in culture supernatants (ELISA kits) IPK
16 30 IL-12 levels in culture supernatants of mononuclear cells of dengue 1 immune donors incubated with dengue virus 20 IL-12 (pg/ml) 10 0 D1 D1+Ac D2 D2+Ac Virus in culture Ab: autologous serum IPK
17 Levels of IL-10 in culture supernatants of mononuclear cells from dengue 1 immune individuals incubated with dengue virus IL-10 (pg/ml) D1 D1+Ab D2 D2+Ab Virus in culture Ab: autologous serum IPK
18 IPK? Which cells produce IL-10? Infected Monocytes or B lymphocytes? Memory T cells? Are higher levels of IL-10 involved in T cell inhibition and apoptosis and viral dissemination in dengue disease? What happens in asymptomatic cases?
19 Th1 IFNγ IL-2 Th2 IL-4 IL-13 IL-10 Th3 TGFβ Tr1 IL-10
20 Evidences that suggest the role of Th2 cytokines in the DHF pathogenesis Higher levels of IL-13 and IL-4 in DHF cases (Mustafa et al., 2001; Gagnon et al., 2002) Higher levels of total and dengue virus-specific IgE in patients with DHF (Pavri et al., 1979; AbuBakar et al., 1997; Miguez-Burbano et al., 1999) Bronchial asthma as a Risk factor for developing DHF (Bravo et al., 1987; Guzman et al., 1992; Orosco et al., 2001) IPK
21 IPK Dengue virus infections: Spectrum of an illness Asymptomatic Dengue Fever Dengue Hemorrhagic Fever/ Dengue Shock Syndrome Th1 Th2/Tr1? Low affinity antibodies:?ade Differential viral cell tropism Genetic factors
22 In secondary infection: Elevated titers of high affinity neutralizing Abs Protection (Asymptomatic) Lower titers of neutralizing Abs or less affinity Abs Virus Escape of neutralizing, Infection of new target cells, memory T cells with Th2 or Tr1 type-response Susceptible (Symptomatic: DF or DHF/DSS)
23 IPK Acknowledgments Instituto de Medicina CPHE Santiago de Tropical Pedro Kourí Cuba Lic. Gissel García a Dr. Maria V. Cabrera Dr. Beatriz Sierra MINSAP Lic. Rayner Rodríguez Dr. Eric Martínez Dr. Susana VázquezV Lic. Mayling Alvarez UMD New Jersey, Dr. María a G. Guzmán USA Dr. Thomas Denny
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