Homepage » Research » Research Units » Immunopharmacology

Immunopharmacology Research Unit

 

History and Achievements

The Immunopharmacology group pursues research on the immune processes undelying the inflammatory responses and the hypersensitivity reactions. For more than 25 years, the group seeks to identify and evaluate diagnostic, prognostic and therapeutic tools for the treatment of inflammatory disorders, focusing on the role of histamine and its receptors in the regulation of immune components under physiological and pathological conditions. Interestingly, although histamine has been one of the most exploited substances in medicine for more than 75 years, the identification of the H4 receptor in 2000 and is localization primarily on immune cells revived the interest on the role of this biogenic amine in immunomodulation and inflammation.

The development of in vivo and ex vivo experimental models and techniques in the Laboratory of Pharmacology allows the preclinical investigation of the mechanisms underlying the inflammatory response in allergies, ocular pathologies and arthritis. In addition, clinical laboratory studies are performed in the ‘Laboratory of Drug Allergy & Pharmacological Modification of Hypersensitivity Reactions' that was established in 2011 in the Allergy Unit 'D. Kalogeromitros', 2nd Department of Venerology and Dermatology, Univetsity Hospital 'Attikon'. Overall, the studies are directed towards the evaluation of new pharmacological targets and interventions aiming to improve the existing diagnostic and therapeutic approaches in inflammatory disorders.

The expertise of the group in the rapidly expanding field of Immunopharmacology contributed to the exposure of novel immunomodulatory properties of histamine and to the evaluation of innovative pharmacologically active agents, mainly targeting the histamine H3 and H4 receptors. The significant scientific effort in this research area resulted in the participation of the group in the development of the Immunopharmacology section (GtoImmuPdb) of the IUPHAR/BPS Guide to PHARMACOLOGY database (GtoPdb), and in the coordination and management of the competitive European program EU FP7 COST BM0806 : Recent advances in histamine receptor H4R research. This Action aimed to foster a multidisciplinary approach to the basic understanding and the huge therapeutic potential of the histamine H4 receptor, and boasted a number of successes: It enabled Europe to achieve a leading position in basic and translational histamine research and results exploitation; it promoted sustainable cooperation with the international pharmaceutical industry; and it supported the rapid entry of H4 receptor targeting compounds into advanced clinical development for poorly treatable chronic inflammatory diseases.

 

Research Interests and Goals

  • The role of histamine and/or histamine receptors in:

           -  the orchestration of acute and chronic inflammation 

           -  allergen tolerance in allergic individuals

           -  the host-microbe communication governing symbiosis

           -  the adaptive and protective cellular stress response and developmental processes

  • Preclinical evaluation of the anti-inflammatory and/or immunomodulatory action of old and new pharmacologically active agents mainly targeting the histamine H1-H4 receptors
  • Investigation of the molecular and cellular mechanisms underlying drug hypersensitivity reactions (drug allergy)

 

 

Research Methodologies and Infrastructure

In vivo animal models of disease

Wistar rats are used as experimental models for the investigation of the (patho)physiological mechanisms and for the evaluation of the effects of various agents in acute and chronic inflammation.

-       Experimental conjunctivitis (acute topical inflammation) is induced by applying a drop of the histamine liberator C48/80 into the lower conjunctival fornix, thus causing non-immunological mast cell degranulation. The early and/or late phase reaction is evaluated by determining biochemical and histological parameters in the isolated conjunctiva following sacrifice of the animal at various time points after challenge.

-       Experimental arthritis (chronic systemic inflammation) is induced by the intradermal administration of complete Freund's adjuvant (CFA) at the base of tail. The animals are sacrificed 21 days later, and the appropriate biochemical and histological parameters are determined in blood samples and in tissue homogenates.

 

Primary tissue and immune cell cultures

-       Tissue cultures of the conjunctiva are used to determine the released inflammatory mediators in the culture medium in response to various stimuli.

-       Ex vivo lymphocyte activation is evaluated by determining blastogenesis in cultured peripheral blood lymphocytes following stimulation with phytohemagglutinin (PHA). Blastogenesis is assessed by a morphometric assay based on the relative proportions of unactivated lymphocytes, activated lymphoblasts and cells with aberrant morphology. Depending on the research hypothesis, lymphocytes may be obtained from patients  and/or from healthy volunteer .

 

 Yeast cultures

The unicellular eukaryote Saccharomyces cerevisiae is a reliable experimental model for the study of the adaptive and protective cellular stress response (heat shock, oxidative stresspharmacological stress), as well as for the elicidation of the effects of chemicals,  drugs and environmental stimuli in normal or/and modified conditions including aging.

The available facilities comprise a yeast culture room, a media preparation, sterilization and storage area, an aseptic transfer area and an observation area. The laboratory is equipped with autoclaves, hot air ovens and membrane filtration apparatus for sterilization, vertical and horizontal laminar flow chambers for aseptic transfers and for inoculation of microbes, a drying oven, controlled microbiological incubators and shaking water baths for yeast and bacteria cultures, cell staining equipment and microscopes (Οlympus ΒΗ Japan with camera, Zeiss Germany) for visual observations.

                                                                                                                                            

Biochemical tests

Various parameters are determined in human biological fluids (saliva, tears, peripheral blood, serum, plasma), culture media and primary immune cell cultures (lymphocytes, mast cells) and tissue homogenates (conjunctiva, hypothalamus, optic nerve, cartilage, blood vessels, whole blood). In addition to the strandard laboratory equipment, the following instruments are available:

   Microplate Reader (SpectraMax PLUS 384, Molecular Devices, Sunnyvale, CA, USA) with monochromator wavelength selection (190-1000nm), supporting both standard spectrophotometer and microplate reader assays, including cell viability, proliferation and cytotoxicity, ELISA based assays and immunoassays, enzyme assays, micro volume applications, microbial growth, DNA and RNA quantification and reporter gene assays. It is routinely used for cytokine, glycogen, glucose and trehalose determination.

   Spectrophotofluorometer (AMINCO-Bowman, American Instrument Co, Maryland, USA) with monochromator, used to quantify histamine and ΝΟ in biological samples.

   Spectrophotometer (Spectro 23RS, Labomed, CA USA) with continuous wavelength 320-1100nm, able to analyze and record four sample results. It is used to measure cell proliferation, protein content, glycogen, glucose, and trehalose in biological samples, as well as ΝΟhttps://www.ncbi.nlm.nih.gov/pubmed/21500983 following reduction of nitrate to nitrite ions.

Fully automated analyser Phadia 250 (Thermo Fisher Scientific Inc, Uppsala, Sweden) for allergy and autoimmunity testing. The allergen panel detects sensitization to 650 allergens and 90 allergen components. The analyser is housed at the Allergy Unit 'D. Kalogeromitros', Univetsity Hospital 'Attikon'.

 

 

Research and Scientific Programmes

International programmes

  • ‘COST FA1404: Improving current understanding and research for sustainable control of the poultry red mite Dermanyssus gallinae (COREMI),  EU Framework Programme Horizon 2020, Chair: O. Sparagano (UK), 2014-2018
  • ‘Implementation of the basophil histamine release (HR) technique’, non-profit external investigation in collaboration with RefLab ApS, Copenhagen, Denmark, 2012-2015
  • ‘COST BM1007: Mast cells and basophils - Targets for innovative therapies’, EU Framework Programme FP7, Chair: M. Maurer (DE), 2011-2015
  • ‘COST BM0806: Recent advances in histamine receptor H4R research’, EU Framework Programme FP7, Chair: E. Tiligada (EL), 2009-2013. More than >160 experts from 26 countries participated in the Action, 35 activities were organised in various European countries and >60 young investigators were trained.
  • ‘Effects of the histamine H3 receptor on the histamine content of peripheral and central tissues in a rat model of experimental arthritis’, non-profit external investigation GlaxoSmithKline R&D Ltd (UK), 2009-2013
  • ‘Pharmacological investigatιon of the H4 receptor antagonist JNJ7777120’ , non-profit external investigation Johnson & Johnson Pharmaceutical R&D (San Diego, USA), 2004-2012

 

National programmes

  • ‘Inflammatory mediators in the GI tract under pathological conditions’, State Scholarships Foundation (ΙΚΥ), Greece, 2008-2009
  • ‘Development of methodology for the determination of cytokines in tissues and biological fluids under physiological and pathological conditions’, Hellenic Ministry of Heath, Greece, 2005-2013
  • ‘The role of salicylates in preconditioning during oxidative stress ’, State Scholarships Foundation (ΙΚΥ), Greece, 2005-2007       
  • ‘The neuroimmunopharmacological role of histamine in the hypothalamus ’  ‘Herakleitos’, Hellenic Ministry of Education, Greece, 2004-2007   
  • ‘Determination of interleukins in type I hypersensitivity reactions’, Empeirikeion Foundation, Greece, 2004  
  • ‘Effects of pharmacologically  active agents  in  the  stress response  in  S. cerevisiae, NKUA, 1999-2005
  • ‘Effect of immunoregulatory and antineoplastic agents in lymphocyte blastogenesis’, NKUA, 1999
  • ‘Allergic reactions in the rat eye, Central Health Council (KESY), 1999\

 

 

Congress organization

  • Training School ‘One health: Potential risks for human health associated with Dermanyssus gallinae and dissemination strategies to communicate recommendations to professionals’, EU COST FA1404, Itea (EL), 31.7-3.8.2017
  • ‘Advances in histamine H4 receptor research’ EU COST BM0806, Sounio (EL), 21-23.3.2013
  • Training School ‘Methodological strategies for histamine receptor characterisation’ EU COST BM0806, Itea (EL), 26-28.7.2012
  • Open Forum ‘Scientific networking in the service of biomedicine and society’, EU COST BM0806, Athens (EL), 6.11.2009
  • 35th Annual Meeting  of the European Histamine Research Society (EHRS), Delphi, Greece, 10-13.5.2006