SVIBOR - Project code: 3-01-142

MINISTRY OF SCIENCE AND TECHNOLOGY

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tel.: +385 1 459 44 44, fax: +385 1 459 44 69
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Project code: 3-01-142


OPIOIDERGIC AND SEROTONINERGIC CONTROL OF HEMATOPOIESIS AND IMMUNITY


Main researcher: BORANIĆ, MILIVOJ (4493)


Assistants
Type of research: basic
Duration from: 06/01/91. to 12/31/93.
Papers on project (total): 56
Papers on project quoted in Current Contents: 16
Institution name: Institut "Ruđer Bošković", Zagreb (98)
Department/Institute: Experimental Biology and Medicine (EBM) Laboratory for Experimental Hematology, Immunology and Oncology
Address: Bijenička cesta 54, p.p. 1016
City: 10000 - Zagreb, Croatia
Communication
Phone: 385 (01) 45 61 011; 45 61 111
Fax: 385 (01) 425-497
E-mail: boranic@olimp.irb.hr
Summary: RESULTS 1994/5 In clonal cultures of mouse bone marrow cells, opioid antagonist naloxone partly blocked the suppressive effect of methionine-enkephalin on the generation of granulocyte-macrophage (GM) colonies. In the long-term culture, naloxone inhibited cell proliferation, particularly of the granulocyte lineage. On the other hand, thiorphan - an inhibitor of the membrane-bound enzyme cleaving the enkephalins and related neuropeptides (EC3.4.24.11, 'enkephalinase') stimulated the proliferation of the granulocytic lineage cells in the long-term cultures of mouse bone marrow. In the model system of humoral immune reaction in vitro (mouse spleen cells stimulated with sheep erythrocytes as the antigen), neurotransmitter serotonin inhibited the generation of hemolyzin-producing cells (PFC). The inhibition was partly blocked by serotonin antagonists ketanserine and propranolol. In continuous cultures of neoplastically transformed lymphoid cells (myelomas, hybridomas), serotonin in some concentrations stimulated the cell proliferation, in contrast to its inhibition of normal cells. Serotonin antagonists did not block the stimulatory effect. Comparative analysis of intraocular and intrathecal IgG response in aqueous humor and cerebrospinal fluid has been performed by means of immunochemical methods (detection of oligoclonal IgG, total IgG, IL-4, beta-2-microglobulin, C3, C4) and by numerical models of protein/IgG synthesis. The optimal parameters and models to define the intraocular and intrathecal IgG response in physiological and pathological conditions have been derived. In the long-term culture of dog bone marrow cells, monoclonal antibody S5 which recognizes the adhesion molecules (CD44) on the membranes of hematopoietic cells promoted the generation of granulocyte-macrophage precursors (GM-CFU) with concomitant decrease of nonadherent (mature) cells. The expression of the MHC class II cell surface marker was enhanced. In addition, S5 enhanced the expression of CD8 and VLA4 on small (lymphoid) cells, and the expression of CD4 and CD18 on large (blast) cells. The NK-cell activity was stimulated. Peripheral blood lymphocyte cultures of the patients with encephalomyelitis or uveitis, as well as the lymphocytes of helathy control persons, were cultured in the presence of peptide M. This peptide is an antigenic fragment isolated from the retina and the pineal gland. Lymphocyte sensitization to the peptide was observed in the patients' samples. In short-term lymphocyte cultures, peptide M decreased the number of chromosomal aberrations.

Keywords: hematopoesis, immune reaction, neuroendocrine regulation, serotonin, opioid peptides, peptide M, cell cultures, mathematical models

Research goals: Participation of opioidergic and serotoninergic signals in the control of hematopoiesis and immunity, respectively, was investigated using in vivo and in vitro model systems. Opioid peptides were known to affect lymphocyte, macrophage and granulocyte functions, as well as the proliferation of mesenchymal cells. It was postulated, therefore, that enkephalins would affect the proliferation and differentiation of cells in the hematopoietic tissue. The results were expected (a) to confirm and further the recognition of the regulatory role of signal molecules (mediators) shared by the central nervous, hematopoietic and immune systems, and (b) to support the view that hematopoiesis is not a self-contained process regulated by local mechanisms only (growth factors, cellular interactions), but also receives macroregulatory, systemic neuroendocrine signals. The specificity of opioidergic signals in hematopoiesis was tested using an opioid antagonist, naloxone; their dependence on cellular interactions, by working with purified cell populations; and the role of the opioid peptide-degrading enzyme D10, by using its specific inhibitor, thiorphan. In order to investigate the role of cell surface adherent molecules (CD44) in the control of cell proliferation and differentiation, hematopoietic cell cultures were treated with specific anti-CD44 monoclonal antibody. Experiments comparing the effects of serotonin on the immune response and on the myeloma/hybridoma cell lines were intended to show whether serotonin inhibits the response by diminishing the cell proliferation, or interferes with other functions. The use of serotonin antagonists was expected to reveal specificity of the serotonin effects, i.e. the presence of 5HT receptors on the cells involved. In patients with autoimmune encephalomyelitis or uveitis, sensitivity to peptide M (isolated from the retina and the pineal gland) was explored. Definition of optimal models for intraocular and intrathecal immune responses was attempted by mathematical modelling.

COOPERATION - PROJECTS


  1. Name of project: > Haematologische und immunologische Untersuchungen von Leukaemie--Erkrankungen verschidener Genese (Leukemia, immunotherapeutic Models)
    Name of institution: GSF-Forschungzentrum fuer Umwelt und Gesundheit GmbH, Institut fuer Immunologie
    City: D-8000 - Muenchen, SR Njemačka

  2. Name of project: Peptide M tolerization in multiple sclerosis model
    Name of institution: Institut fuer Immunologie und Thymusforschung
    City: Bad Harzburg, Njemačka / Deutschland

  3. Name of project: Peptide M tolerization in uveitis model
    Name of institution: Institut fuer Immunologie und Thymusforschung
    City: Bad Harzburg, Njemačka / Deutschland

COOPERATION - INSTITUTIONS


  1. Name of institution: Institut fuer Immunologie der Gesellschaft fuer Strahlen- und Umweltforschung GmbH
    City: D-8000 - Muenchen, SR Njemačka

  2. Name of institution: Fred Hutchinson Cancer Resarch Center, Clinical Research Division
    Type of institution: University/Faculty
    City: WA 98104 - Seattle, SAD

  3. Name of institution: Institut fiziologii Russkoj akademii nauk
    City: Novosibirsk, Rusija

  4. Name of institution: Institut fuer Immunologie und Thymusforschung
    Type of institution: Economical/Production
    Type of cooperation: Joint project
Other information about the project.
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