INFLUENCE OF EXOMETABOLYTES OF PALEOBACTERIA OF THE GENUS BACILLUS FROM PERMAFROST ROCKS ON THE MECHANISMS OF FORMATION OF THE IMMUNE RESPONSE IN VITRO

Abstract

It has been established that exometabolites of paleobacteria Bacillus cereus strain 875 TS significantly activate the differentiation of monocytes in the subpopulation of intermediate (CD14⁺CD16⁺) and non-classical (CD14^loCD16⁺) monocytes, effector CD4⁺ and CD8⁺ T-lymphocytes with a change in markers of early (CD69), mid (CD25) and late (HLA- DR) activation, differentiation of Treg (CD3⁺CD4⁺CD25^hiCD127^-), and also stimulate the synthesis of cytokines IFNγ and IL-4 relative to control levels. The peculiarities of the influence of exometabolites of paleobacteria include the dependence of immunomodulatory activity on the method of their preparation - “cold” (obtained from bacteria when they are cultivated at 5ºC), “mid temperature” (22ºC) and “heat” (37ºC) metabolites.  “Cold” metabolites stimulate mainly the mechanisms of the immune response with pro-inflammatory activity, in particular, the differentiation of intermediate CD14⁺CD16⁺ monocytes, an increase in the activity of differentiation of CD8⁺ T-lymphocytes and the synthesis of IFNγ. “Heat” metabolites stimulate predominantly immune response mechanisms with anti-inflammatory activity, namely the differentiation of non-classical CD14^loCD16⁺ monocytes, increased differentiation activity of CD4⁺ T-lymphocytes and IL-4 secretion. Also a distinctive feature is the relationship between pro- and anti-inflammatory mechanisms, which do not depend on the type of exometabolites. Thus, during the first three days of cell culture, the differentiation activity of CD8⁺ T-lymphocytes prevails over the differentiation of CD4⁺ T-lymphocytes, and the level of IFNγ secretion exceeds the level of IL-4. On the third day, there is a significant increase in Treg levels, which is accompanied by a tendency to normalize the balance between IFNγ(Th1) and IL-4(Th2) by the seventh day. There is a clear influence of Tregs (CD3⁺CD4⁺CD25^hiCD127^-) on the strength and duration of the immune response. The increase in Treg levels occurs moderately and briefly, which, on the one hand, prevents the excessive development of pro-inflammatory mechanisms, and on the other hand, does not lead to the development of long-term immunosuppression. An increase in Treg levels on days 1-3 is accompanied by a decrease in the activity of monocyte differentiation in the subpopulation and the synthesis of the proinflammatory cytokine IFNγ.

1. Vakhitov T.Ya., Petrov L.N. Regulatory functions of bacterial exometabolites // Microbiology, 2006, Vol. 75, No. 4, P. 483-488.

2. Vorobyova L.I. Stressors, stress and survival of bacteria // Applied biochemistry and microbiology, 2004, Vol. 40, No. 3, P. 261-269.

3. Kalenova L.F., Petrov S.A., Sukhovey Yu.G. Repair and immunomodulatory potential of low molecular weight fractions of secondary metabolites of Bacillus sp.// BEBIM. – 2021. – Vol. 172, No. 9. – P. 323-327.

4. Provorov N.A., Tikhonovich I.A. Genetic and molecular basis of symbiotic adaptations // Advances in modern biology, 2014, Vol. 134, No. 3, P. 211–226.

5. Nikolaev Yu.A., Mulyukin A.L., Stepanenko I.Yu., El-Registan G.I. Autoregulation of the stress response of microorganisms // Microbiology, 2008, Vol. 75, No. 4, P. 489-496.

6. Filippova S.N., Surgucheva N.A., Sorokin V.V. and others. Bacteriophages of low-temperature systems of the Arctic and Antarctic // Microbiology, 2016, Vol. 85, No. 3, P. 337-346.

7. Freidlin I.S. Regulatory T cells: origin and functions / Medical immunology. – 2005. – Vol. 7, No. 4. – P. 347-354.

8. Khaidukov S.V., Zurochka A.V. Cytometric analysis of T-helper subpopulations (Th1, Th2, Treg, Th17, activated T-helpers) // Medical Immunology. – 2011. – Vol.11, No. 1. – P.7-16.

9. Athanassakis I., Vassiliadis S. T-regulatory cells: are we rediscovering T suppressor? // Immunology Letters. – 2002. – Vol.84. – P.179-183.

10. Azeredo E.L., Neves-Souza P.C., Alvarenga A.R. et al. Differential regulation of toll-like receptor-2, toll-like receptor-4, CD16 and human leucocyte antigen-DR on peripheral blood monocytes during mild and severe dengue fever // Immunology. – 2010. – Vol. 130(2). – P. 202‐216.

11. Caramalho, I., Lopes Carvalho, T., Ostler, D., Zelenay, S., Haury, M., and Demengeot, J. (2003) Regulatory T cells selectively express toll like receptors and are activated by lipopolysaccharide, J. Exp. Med., 197, 403–411.

12. Cibrián D., Sánchez-Madrid F. CD69: from activation marker to metabolic gatekeeper // Eur.J.Immunol. – 2017. – Vol.47, №6. – P. 946–953.

13. During M., Cabanillas Stanchi K.M., Haufe S., et al. Patterns of monocyte subpopulations and their surface expression of HLA-DR during adverse events after hematopoietic stem cell transplantation // Ann. Hematol. – 2015. – Vol. 94(5). – P.825-836.

14. Fontenot J., Rudensky A. A well adapted regulatory contrivance: regulatory T cell development and the forkhead family transcription factor Foxp3 // Nature Immunol. – 2005. – Vol.6. – P.331-337.

15. Guilliams M., Mildner A., Yona S. Developmental and functional heterogeneity of monocytes // Immunity. – 2018. – Vol. 49(4). – P. 595-613.

16. Hijdra D., Vorselaars A.D.M., Grutters J.C. et al. Phenotypic characterization of human intermediate monocytes // Front Immunol. – 2013. – №.4. – P. 4-6.

17. Jenkins S.J., Ruckerl D., Thomas G.D. et al. IL‐4 directly signals tissue‐resident macrophages to proliferate beyond homeostatic levels controlled by CSF‐1 // J. Exp. Med. – 2013. – Vol. 210(11). – P. 2477-2491.

18. Ka M.B., Olive D., Mege J.L. Modulation of monocyte subsets in infectious diseases // World J. Immunol. – 2017. – Vol. 4(3). – P.185.

19. Kalenova L.F., Novikova M.A., Kostolomova E.G. Effects of Low-Doses of Bacillus Spp. from Permafrost on Differentiation of Bone Marrow Cells // Bulletin of Experimental Biology and Medicine, 2015б, v.158, № 3, p. 364-367.

20. Kalyonova L.F., Novikova M.A., Subbotin A.M., Bazhin A.S. Effects of Temperature on Biological Activity of Permafrost Microorganisms // Bulletin of Experimental Biology and Medicine, 2015в, v. 158, № 6, р.772-775. 10.1007/s10517-015-2859-z.

21. Kalenova L.F., Kolyvanova S.S., Bazhin A.S. et al. Effects of Secondary Metabolites of Permafrost Bacillus sp. on Cytokine Synthesis by Human Peripheral Blood Mononuclear Cells // Bulletin of Experimental Biology and Medicine, 2017, vol. 163, No. 2, p.235-238.

22. Ritchie A.J., Jansson A.Р., Stallberg J.Р. et al. The Pseudomonas aeruginosa Quorum-Sensing Molecule N-3-(Oxododecanoyl)-L-Homoserine Lactone Inhibits T-Cell Differentiation and Cytokine Production by a Mechanism Involving an Early Step in T-Cell Activation // Infection and immunity, 2005, v. 45, р. 1648-1655.

23. Rodriguez-Muсoz Y., Martin-Vilchez S., Lуpez-Rodriguez R. et al. Peripheral blood monocyte subsets predict antiviral response in chronic hepatitis C // Aliment Pharmacol. Ther. – 2011. – Vol. 34(8). – P. 960-971.

24. Shipkova M., Wieland E. Surface markers of lymphocyte activation and markers of cell proliferation // Clin.Chim. Acta. – 2012. – Vol. 413, № 17-18. – Р. 1338–1349.

25. Sing, A., Rost, D., Tvardovskaia, N., Roggenkamp, A., Wiedemann, A., Kirschning, C.J., Aepfelbacher, M., and Heesemann, J. (2002) Yersinia V antigen exploits toll like receptor 2 and CD14 for interleukin 10 mediated immunosuppression, J. Exp. Med., 196, 1017–1024.

26. Smeekens S.P., van de Veerdonk F.L., Joosten L.A.B. et al. The classical CD14++CD16− monocytes, but not the patrolling CD14+CD16+ monocytes, promote Th17 responses to Candida albicans // Eur. J. Immunol. – 2011. – Vol. 41(10). – P. 2915-2924.

27. Sutmuller, R.P.M., Morgan, M.E., Netea, M.G., Grauer, O., and Adema, G.J. (2006) Toll like receptors on regulatory T cells: expanding immune regulation, Trends Immunol., 27, 387–393.

28. Wieland E., Shipkova M. Lymphocyte surface molecules as immune activation biomarkers // Clin. Biochem. – 2016. – Vol. 49, № 4-5. – P. 347–354.

29. Wong K.L., Yeap W.H., Tai J. et al. The three human monocyte subsets: implications for health and disease // Immunol. Res. – 2012. – Vol. 53(1-3). – P.41-57.

30. Wong K.L., Jing Yi, Tai J. et al. Gene expression profiling reveals the defining features of the classical, intermediate, and non-classical human monocyte subsets // Blood. – 2011. – Vol. 118(5). – P.16-32.