Methodological approaches to the comparative assessment of cellular reactions of innate immunity in mollusk Pomacea sp. (Caenogastropoda, Ampullariidae) in experimental aseptic inflammation
- Authors: Khrushchev K.A.1, Shilov S.Y.2,3, Barkov S.Y.2,3, Shilov Y.I.1,2,3
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Affiliations:
- Perm State University
- Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences
- E. Wagner Perm State Medical University
- Issue: Vol 27, No 4 (2024)
- Pages: 749-756
- Section: SHORT COMMUNICATIONS
- URL: https://journal-vniispk.ru/1028-7221/article/view/267846
- DOI: https://doi.org/10.46235/1028-7221-16939-MAT
- ID: 267846
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Abstract
One of the significant methodological problems that arise in the study of cells of the innate immunity of invertebrates is the unsuitability of culture media and even salt solutions intended for mammals and humans. Another important methodological point is the significantly pronounced reactions of coagulation, which play an important protective role in all invertebrates, but manifest themselves in vitro in cellular suspensions prepared even with optimal saline solutions for this animal. To solve this problem, researchers widely use an anticoagulation buffer with ethylenediaminetetraacetic acid (EDTA) even in functional tests. From the standpoint of traditional approaches of mammalian immunology, the use of EDTA is possible only for the study of cell morphology, but not their functions. The aim of the work is to adapt methodological approaches to the comparative assessment of cellular reactions of innate immunity in Pomacea sp mollusks in experimental aseptic inflammation. To evaluate the functions of phagocytic cells in mollusk Pomacea sp., we used a complete salt solution (CSS) with optimal concentrations of glucose, calcium ions, magnesium with addition to prevent coagulation, cellular aggregation and degranulation instead of EDTA sodium salt of heparin at a concentration of 100 IU/mL (CSS-hep). As a result of a specially conducted experiment, it was shown that during incubation of hemolymph cells in this solution, their viability is preserved, evaluated by flow laser cytometry during incubation with annexin V-PE (PE Annexin V) and 7-aminoactinomycin D (7- AAD), as well as in a test with trypan blue. The possibility of studying the phagocytic activity and a phagosome acidification of leukocytes from hemolymph, hematopoiesis organ (kidney) and the focus of inflammation induced by intramuscular injection of sterile suspension of zymosan using CSS-hep is shown. Unlike the Limnaea stagnalis and Biomphalaria glabrata mollusks, in which the production of reactive oxygen species by phagocytes in the luminol-dependent chemiluminescence reaction is well documented, we were unable to induce this reaction in Pomacea sp. mollusks. In a specially conducted experiment, we did not reveal the effect of the sodium salt of heparin at a concentration of 100 IU/mL on the production of reactive oxygen species by rat phagocytes.
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##article.viewOnOriginalSite##About the authors
K. A. Khrushchev
Perm State University
Email: jshilov@mail.ru
4th year Student, Biological Faculty
Russian Federation, PermS. Yu. Shilov
Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; E. Wagner Perm State Medical University
Author for correspondence.
Email: jshilov@mail.ru
PhD (Medicine), Junior Research Associate, Laboratory of Ecological Immunology, Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; Associate Professor, Department of Normal Physiology, E. Wagner Perm State Medical University
Russian Federation, 3 Golev St, Perm, 614081; PermS. Yu. Barkov
Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; E. Wagner Perm State Medical University
Email: jshilov@mail.ru
Postgraduate Student, Laboratory of Ecological Immunology, Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; Senior Laboratory Assistant, Department of Immunology, E. Wagner Perm State Medical University
Russian Federation, 13 Golev St, Perm, 614081; PermYu. I. Shilov
Perm State University; Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; E. Wagner Perm State Medical University
Email: jshilov@mail.ru
PhD (Medicine), Associate Professor, Senior Research Associate, Laboratory of Ecological Immunology, Institute of Ecology and Genetics of Microorganisms, Perm Federal Research Center, Ural Branch, Russian Academy of Sciences; Associate Professor, Department of Immunology, E. Wagner Perm State Medical University, Perm, Russian Federation; Associate Professor, Department of Microbiology and Immunology, Perm State University
Russian Federation, Perm; 13 Golev St, Perm, 614081; PermReferences
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