Laboratory of Cellular Immunology

Head of the Laboratory: Dotsika Eleni, PhD
Tel: +30-210-6478875, e-mail: e.dotsika@mail.pasteur.gr

About

Parasitic protozoa of the genus Leishmania cause a broad spectrum of diseases in humans from localized cutaneous lesions, mucosal infections of varying severity, or subclinical to acute visceral disease. Leishmaniasis are spanning throughout tropical and subtropical regions worldwide including the Mediterranean region. They are considered a major public health problem. In all forms of the disease, healing or recovery of leishmaniasis has been depended on specific Th1 cells. Inevitably, due to the importance of cellular responsiveness for the generation and maintenance of effective host immunity, extensive studies have been performed in animal models. Moreover the well-defined immune response generated upon experimental infection with Leishmania ssp in inbred mice, has made this parasitic infection an ideal experimental system for the investigation of Th1/Th2 polarization and regulation.

The objective of our work is to analyse the cellular and molecular immune mechanisms concerning the interplay between host cells and Leishmania spp. in order to define specific molecules for immune intervention.

Particularly our work is focus on the:

  • Role of CD8+ T cells in experimental L. infantum infection
  • Impact of L. infantum infection οn macrophage function: role of MAP kinases
  • DCs and their role in the induction of protective immunity in L. major infection
Members

Head of the Laboratory:

Dotsika Eleni, Veterinarian, Researcher Grade B’ (email: e.dotsika@pasteur.gr)

Researchers:

Karagouni Evdokia, Biologist, Researcher Grade B’ (email: ekaragouni@pasteur.gr)

Scientific and Technical Operatives:

Koutsoni Olga, Biologist (email: okoutsoni@pasteur.gr)

Postdoctoral Research Fellows:

Agallou Maria, Biologist (email: mariaagallou@pasteur.gr)

Toubanaki Dimitra, Chemist (email: dtouban@pasteur.gr)

PhD students:

Athanasiou Evita, Biologist (email: eathanasiou@pasteur.gr)

Kyriazis Ioannis, Biologist (email: g.kyriazis@pasteur.gr)

Margaroni Maritsa, Biologist (email: mmargaroni@pasteur.gr)

Postgraduate students

Voukelatou Golfo, Biologist (email: voukelatoug@pasteur.gr)

Research programs

Research programs

Role of CD8+ T cells in experimental L. infantum infection

We focused on the mode of action of CD8+ T cells in murine visceral leishmaniasis and their contribution to the clearance of the parasite. Genetically curative C57BL/6 and non-curative ΒΑLB/c mice were infected with L. infantum, the most common cause of visceral leishmaniasis in the Mediterranean basin. It has been showed that the spleen of infected BALB/c mice contained parasite-specific CD8+ lymphocytes shared a TC1 pattern of differentiation in an environment of compromised TH1 response. These cells had up regulated the CD25 and CD69 surface antigens and were capable to lyse target cells pulsed with Leishmania peptides and Leishmania-infected macrophages through the perforin pathway of cytotoxicity. TC lymphocytes had also the ability to kill target cells by apoptosis, as they over expressed FasL. An equally important function of this cell subset in visceral leishmaniasis was the production of massive amount of cytokines (IFN-γ and TNFα) and C-C chemokines (MIP-1α and RANTES).
On the other hand, in cure C57BL/6 mice the role of TC lymphocytes was of markedly lesser importance, as the number of parasite-specific CD8+ T cells remained very low, cytotoxicity was limited to the first weeks of infection, TNF production was virtually absent and chemokine (especially RANTES) production was limited. However, they retained the ability to secrete IFN-γ, and were thus characterized as TC1 cells. Experiments in TNF-/- mice showed that the major mediator of apoptosis was TNF. Infection of β2-microglobulin-/- mice allowed us to assess the in vivo contribution of CD8+ T cells in the resolution of visceral leishmaniasis in the curative experimental model.

Impact of L. infantum infection οn macrophage function: role of MAP kinases

MAP kinases – ERK1/2, JNK and p38 – differentially regulate iNOS and IL-12 gene expression in LPS-stimulated macrophages. In vitro infection of LPS-stimulated macrophages obtained from BALB/c mice, with L. infantum, led to the phosphorylation of MAP kinases and the iNOS and IL-12 genes transcription resulting in the reduction of parasite load. Use of MAP kinase inhibitors demonstrated that JNK and p38 MAP kinases play an important role in the induction of iNOS, while ERK1/2 does not. In addition, p38 MAP kinase induces IL-12 gene transcription, in contrast to ERK1/2 and JNK activation which abrogate IL-12 production. These findings indicate that NO and IL-12 inhibition by Leishmania parasites is JNK- and p38 MAP kinase-mediated, suggesting these molecules as candidate targets for immune intervation.

DCs and their role in the induction of protective immunity in L. major infection

In the context of DC-based vaccination strategy, we investigated the capability of bone marrow derived DCs-pulsed with Leishmania gp63 peptides to induce protective immunity against L. major infection. To this end, susceptible BALB/c mice were vaccinated with DCs-pulsed with gp63 epitopes 154-169aa and 467-482aa and subsequently challenged with L. major promastigotes.  DCs pulsed with gp63 epitope 154-169aa triggered antigen-specific immune responses able to reduce lesion formation and parasite load in BALB/c mice. This effect was accompanied by a modulation of the immune response towards Th1 type as confirmed by the cytokine secretion (IFN-γ vs IL 4) and parasite-specific antibodies profile (IgG2a vs IgG1). DCs-pulsed with gp63 epitopes 467-482aa failed to instruct Th1 response.
In contrast, vaccination of resistant CBA mice with DCs-pulsed with gp63 epitopes 467-482aa resulted in a shift towards a Th2 cellular immune response that justified the exacerbation of lesions. These findings suggest that vaccination with DCs pulsed with defined peptides could be a strategy against infectious diseases. Peptide selection is a prerequisite as they can differentially regulate the type of immune response in susceptible or resistant hosts.

Other attempts to explore novel vaccination approaches included vaccination of BALB/c mice with a plasmid carrying a truncated gp63 cDNA resulted in the exacerbation of the local inflammatory reaction. In contrast, the co-administration of this plasmid with another encoding CD40L induced protection in 65% of the infected mice since CD40-CD40L interaction play key role in IL-12 secretion from antigen-presenting cells.

Publications

Publications

  1. Karagouni, E.,  S. Frydas, E. Dotsika, C. Himonas, P. Conti and A. Trakatellis (1995). Inhibition of TNF- and IL-6, but not IFN-, by 4-deoxypyridoxine (4-DPD) in Trichinella-infected mouse serum. Int. J. Immunopath. Pharmacol., 8(1), 9-14.
  2. Haralabidis, S., E. Karagouni, S. Frydas, and E. Dotsika (1995). Immunoglobulin and cytokine profile in murine secondary hydatidosis. Parasite Immunol., 17, 625-530.
  3. Frydas, S., E. Karagouni, E. Dotsika, M. Reale, R.C. Barbacane, I. Vlemmas, G. Anogianakis, A. Trakatellis, and P. Conti (1996). Generation of TNF, IFN-, IL-6, IL-4 and IL-10 in mouse serum from trichinellosis: effect of the anti-inflammatory compound 4-deoxypyridoxine (4-DPD). Immunol. Let., 49, 179-184.
  4. Sideris, V., E. Karagouni, G. Papadopoulou, A. Garyfallou and E. Dotsika (1996). Canine visceral leishmaniasis in the great Athens area, Greece. Parasite, 3, 125-130.
  5. Chryssikopoulos A., Th. Μantzavinos, N. Kanakas, E. Karagouni, E. Dotsika, and P.A. Zourlas (1996). Correlation of serum and follicular fluid concentrations of placenta protein 14 and CA-125 in in vitro fertilization-embryo transfer patients. Fertil. Steril., 66(4), 599-603.
  6. Dotsika, E., E. Karagouni, B. Sundquist, B. Morein, A. Morgan, and M. Villacres-Eriksson (1997). Ιnfluence of Quillaja saponaria triterpenoid content on the immunomodulatory capacity of Epstein-Barr virus (EBV) iscoms. Scand. J. Immunol., 45, 261-268.
  7. Papadopoulou, G., E. Karagouni and E. Dotsika (1998). ISCOMs vaccine against experimental Leishmaniasis. Vaccine, 16, 885-892.
  8. Karagouni, E., A. Chryssikopoulos, T. Mantzavinos, N. Kanakas and E. Dotsika (1998). Ιnterleukin-1 and interleukin-1 may affect the implantation rate of patients undergoing in vitro fertilization-embryo transfer. Fertil. Steril, 70, 553-559.
  9. Doukas, V., F. Athanasopoulou, E. Karagouni and E. Dotsika (1998). Aeromonas hydrophila infection in cultured see bass, Dicentrarchus labrax L. and Puntazzo puntazzo Cuvier from the Aegean sea. J. Fish Dis., 21, 317-320.
  10. Eliades, P., E. Karagouni, I. Stergiatou and K. Miras (1998). A simple method for the serodiagnosis of human hydatid disease based on a protein A/colloidal dye conjugate. J. Immunol. Meth., 218, 123-132.
  11. Frydas, S., M. Reale, D. Vacalis, R.C. Barbacane, F.C. Placido, I. Cataldo, M. Gioacchino, E. Karagouni, E. Dotsika, G. Anogiannakis, A. Trakatellis and P. Conti (1999). IgG, IgG1 and IgM response in Trichinella spiralis-infected mice treated with 4-deoxypirydoxine or fed a Vitamin B6-deficient diet. Mol. Cell. Biochem., 194, 47-52.
  12. Sideris, V., G. Papadopoulou, E. Dotsika and E. Karagouni (1999). Asymtomatic canine leishmaniasis in greater Athens area, Greece. Eur. J. Epidemiol., 15, 271-276.
  13. Hemmerlin C., A. Phan Chan Du, Z. Elhilali, A. Moulia, V. Tsikaris, M. Sakarellos-Daitsiotis, C. Sakarellos, E.N. Dotsika, A.G. Tzioufas, H.M. Moutsopoulos, Manh-Thong Cung (2001). Conformational study of the complementary peptide to a B-cell epitope of the La/SSB autoantigen. Chimie/Chemistry, 4, 729-733.
  14. Routsias G. J., E. Touloupi, E.N. Dotsika, V. Tsikaris, C. Sakarellos, M. Sakarellos-Daitsiotis, H.M. Moutsopoulos, A.G. Tzioufas,  (2002). Unmasking the ant-La/SSB response by specific blocking of ant-idiotypic antibodies with sense-complementary peptides to La/SSB major antigenic determinants. Mol. Med., 8, 293-305.
  15. Frydas, S., M. Hatzistilianou, E. Karagouni, B. Madhappan, N. D’Orazio, G. Riccioni, F. Conti, G. Carratelli and D. Kempuraj (2003). Chemokines and parasites. Int. J. Immunopathol. Pharmacol., 16(3), 221-224.
  16. Routsias G. J., E.N. Dotsika, Touloupi E., M. Papamattheou, C. Sakarellos, M. Sakarellos-Daitsiotis, H.M. Moutsopoulos, A.G. Tzioufas, (2003). Idiotypic-antiidiotype circuit in non-autoimmune mice after immunization with the epitope and complementary epitope 289-308aa of La/SSB response. J. Autoimmun., 2, 17-26.
  17. Tsagozis, P., E. Karagouni and E. Dotsika (2003). CD8+ T cells with parasite cytotoxic activity and the TC1 profile of cytokine and chemokine secretion develop in experimental visceral leishmaniasis. Parasite Immunol., 25, 569-579.
  18. Papamattheou, M.G., J.G. Routsias, E.E. Karagouni, C. Sakarellos, M. Sakarellos-Daitsiotis, H.M. Moutsopoulos, A.G. Tzioufas and E.N. Dotsika (2004). T cell help is required to induce idiotypic-anti-idiotypic autoantibody network after immunization with complementary epitope 289-308aa of La/SSB autoantigen in non-autoimmune mice. Clin. Exp. Immunol., 135, 416-426.
  19. Frydas, S., E. Karagouni, E. Papadopoulos, M. Hatzistiliannou, S. Pappas and N. Papaioannou (2004). Chemokines and their role in parasitic diseases. Eur. J. Inflamm. 2(2), 53-56.
  20. Tsagozis, P., V. Tseveleki, L. Probert, E. Dotsika and E. Karagouni (2004). Vaccination with plasmids encoding the Leishmania major gp63 glycoprotein and CD40L results in a partial suppression of the inflammatory reaction after experimental infection. Eur. J. Inflamm. 2(2), 91-96.
  21. Tsagozis, P., E. Karagouni and E. Dotsika (2004). Dendritic cells pulsed with peptides of gp63 induce differential protection against experimental cutaneous leishmaniasis. Int. J. Immunopathol. Pharmacol., 17(3), 343-352.
  22. Frydas, S., E. Karagouni, M. Hatzistilianou, D. Kempuraj, S. Comani, C. Petrarca, T. Iezzi, N. Verna, P. Conti and M.L. Castellani (2004). Cytokines and allergic disorders: Revisited study. Int. J. Immunopathol. Pharmacol. 17(3), 233-235.
  23. Frydas, S., E. Karagouni, T. Iezzi, D. Kempuraj, C.M.V. Conti, W. Boucher, N.G. Papadopoulou, J. Donelan, B. Madhappan, R. Letourneau, G. Carratelli, A. Grilli, F. Conti, G. Riccioni, D. Tripodi, F. Menegilini, I. Stellin and G. Sabatino (2004). Human genome project and parasitic infection. Eur. J. Inflamm., 2(1), 1-3.
  24. Athanassopoulou, F., E. Karagouni, E. Dotsika, V. Ragias, J. Tavla and P. Christofilloyanis (2004). Efficacy and toxicity of orally administrated anti-coccidial drugs for innovative treatments of Polysporoplasma sparis Sitja-Bobadilla & Alvarez-Pellitero 1885 infection in Sparus aurata L. J Appl. Ichthyol., 20, 345-354.
  25. Athanassopoulou, F., E. Karagouni, E. Dotsika, V. Ragias, J. Tavla, P. Christofilloyanis and I. Vatsos (2004). Efficacy and toxicity of orally administrated anti-coccidial drugs for innovative treatments of Myxobolus sp. infection in Puntazzo puntazzo C. Dis. Aquat. Org 62, 217-226.
  26. Di Giannantonio, S. Frydas, D. Kempuraj,  E. Karagouni,  M. Hatzistilianou, C.M.V. Conti, W. Boucher, N. Papadopoulou, J. Donelan, J. Cao, B. Madhappan, P. Boscolo, C. Petrarca, L. Castellani, R. Quartesan, R. Doyle and F.M. Ferro (2005). Cytokines in stress. Int. J. Immunopathol. Pharmacol., 18(1), 1-5.
  27. Frydas, S., N. Papaioannou, M. Papazahariadou, M. Hatzistilianou, E. Karagouni, M. Trakatelli, G. Brellou, C. Petrarca, M.L. Castellani, P. Conti, G. Riccioni, A. Patruno and A. Grilli (2005). Inhibition of MCP-1 and MIP-2 chemokines in murine trichinellosis: effect of the anti-inflammatory compound L-mimosine. Int. J. Immunopathol. Pharmacol., 18(1), 85-93.
  28. Karagouni, E., F. Athanassopoulou, P. Tsagozis, E. Ralli, Th. Moustakareas, K. Lytra and E. Dotsika (2005). The impact of a successful anti-myxosporean treatment on the phagocyte functions of juvenile and adult Sparus aurata L. Int. J. Immunopathol. Pharmacol. 18(1), 121-132.
  29. Tsagozis, P., E. Karagouni and E. Dotsika. Function of CD8+ T lymphocytes in a curing model of visceral leishmaniasis. Int. Parasitol. (Accepted)
  30. Karagouni, E., F. Athanassopoulou, A. Lytra, C. Comis and E. Dotsika.  Anti-parasitic and immunomodulatory effect of innovative treatments against Myxobolus sp. infection in Diplodus puntazzo. Vet. Parasitol. (Accepted)
Funding
  • 1993-1996, EC, Biotechnology grand : «An investigation into the mechanisms of adjuvanticity of immunostimulating  complexes (iscoms) of defined chemical composition using a primate model of EBV virus induced lymphoma»
  • 1994-1996, EC, Human Capital and Mobility. «The development of PCR assay for the detection of cryptic leishmaniasis and its application to the study of the disease in Greece and throughout Europe».
  • 1995-1998, Minister of Industry, Energy and Technology, EC support Framework II, Operational Program for Research and Development (EPET II): «Formulation and manufacture of fish feeds characterized by high nutritional value and low environmental impact».
  • 1999-2000, Ministry of Industry, Energy and Technology. Programme for Industrial Research Development (PAVE97) «Studies on pathology, transmission and eradication of myxosporidiasis in intensive aqua cultures of Sparus aurata L. and Puntazzo puntazzo».
  • 1999-2000, Ministry of Industry, Energy and Technology. Programme for Industrial Research Development (PAVE97). «Nutritional factors affecting the growth, health and resistance to infections in intensive cultures of Puntazzo puntazzo».
  • 2000-2001, Ministry of Industry, Energy and Technology. Programme Stregthening Young Researchers (PENED III) «Development of immune intervention techniques modulating autoimmune responses to intracellular antigens».
  • 2001-2002, HPI-Research Acount 99. «Dialogue between dendritic cells and CD8+ lymphocytes in models of experimental leishmaniasis».
  • 2003, Industrial project (ΕLPΕΝ INC), «Study of the pro-inflammatory cytokines in experimental model of obstructive jaundice».
  • 2003-2004, HPI-Research Acount 02,. «Role of MAPKs in DCs: impact of L. donovani infection in the MAPKs cascade events».