Cystatins of helminths: characterization of molecules with immunomodulatory potential
| Autoři | |
|---|---|
| Rok publikování | 2016 |
| Druh | Konferenční abstrakty |
| Fakulta / Pracoviště MU | |
| Citace | |
| Popis | Cystatins cysteine peptidase inhibitors are produced by a wide range of organisms, including parasites, where they act as modulators of the host immune system. In the frame of our research we focused on characterization of cystatins produced by four helminth species representing most important groups of helminth parasites, namely Eudiplozoon nipponicum (Monogenea), Fascioloides magna (Trematoda), Hymenolepis diminuta (Cestoda) and Trichinella spiralis (Nematoda). We performed a bacterial expression of optimized cystatin genes in recombinant forms (E. coli BL21 cells, pET32a and/or pET19b plasmid vector). The recombinant cystatins were purified via Histag using metal affinity chromatography (NiNTA) and successful production of cystatin was confirmed by mass spectrometric methods (MALDI TOF/TOF). Selected candidates were used for immunization of mice and cystatins were immunolocalized on cryosections of parasite tissues using positive mice sera, FITClabeled antimouse antibodies and DAPI counterstaining. The inhibitory effect of particular cystatins was fluorometrically measured by adoption of recombinant cysteine peptidases (mouse cathepsin L, cathepsin L3 from E. nipponicum) and its fluorogenic peptide substrate (FRAMC). The immunomodulatory potential of presented cystatins (modulation of antigen presentation, influence on cytokine production and proliferation of lymphocytes, macrophages, etc.) will be studied in detail. |
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