Transient modification of macrophages for activation their pro-inflammatory functions

Macrophages are innate immune cells that play an important role in both physiological and pathological conditions. According to the concept of binary polarization, there are two the most polar states of macrophages: pro-inflammatory and anti-inflammatory macrophages. We proposed that targeted programming of macrophages derived from the specific monocyte subpopulation is perspective strategy for activation their inflammation properties that could be used in therapy of cancer or endometriosis. This work is devoted to the study of the effect of Indoleamine 2,3-dioxygenase 1 (IDO1) and its inhibitor BIN1 on the shift in the polarization of macrophages into a pro-inflammatory phenotype. We used magnetic sorting, differentiation, and further pro-inflammatory polarization of CD14+ monocytes from the blood of healthy donors. We obtained data on the transcriptome and proteome and carried out a bioinformatic analysis to identify the most significant differences between two groups of cells - naive and pro-inflammatory macrophages. IDO1 has been identified as an enzyme with immunomodulatory functions and is also involved in the catabolism of the essential amino acid L-tryptophan. This enzyme is expressed in macrophages and its expression is induced by pro-inflammatory cytokines. It was shown that the BIN1 protein is an inhibitor of IDO1 and can inhibit the growth of cancer cells. It was shown that upon the polarization of macrophages towards the pro-inflammatory phenotype by LPS addition, the expression of IDO1 increases, while the expression of BIN1 decreases (p <0.05). To reveal the influence of the IDO1 and BIN1 genes on the shift of the polarization of macrophages towards the M1 phenotype, we decided to carry out knockdown by using siRNAs on the primary line of CD14+-derived macrophages. The efficiency of transfection was assessed at different concentrations of siRNA ranged from 30 to 80%, but high concentrations of siRNA led to non-specific activation of pro-inflammatory phenotype. Expression analysis confirmed the silencing of the genes IDO1 and BIN1 using 5 nM siRNA. It was shown that knockdown of IDO1 and BIN1 leads to increased expression of pro-inflammatory genes (CCR7 and TNFa) and decreased the level of anti-inflammatory interleukin 10. Also, co-cultivation CD14+ macrophages after knockdown of genes IDO1 and BIN1 didn't influent on macrophage's phagocytic functions. The proposed study has a scientific novelty since for the first time a transient modification of the primary line of macrophages from blood CD14 + was carried out and the influence of IDO1 and BIN1 on the activation of the pro- or anti-inflammatory phenotype was shown. Directed programming of macrophages is a new and rapidly developing field. © FASEB.

Poltavets A.1 , Vishnyakova P. 1, 2 , Zatsepin T.3, 4 , Fomina M.5 , Elchaninov A. 1, 2 , Fatkhudinov T. 2, 6
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  • 1 Laboratory of regenerative medicine, National Medical Research Center for Obstetrics, Gynecology and Perinatology named after Academician V.I. Kulakov of the Ministry of Healthcare of the Russian FederationMoscow, Russian Federation
  • 2 Peoples' Friendship University of RussiaMoscow, Russian Federation
  • 3 Skolkovo Institute of Science and TechnologyMoscow, Russian Federation
  • 4 Department of Chemistry, Lomonosov Moscow State UniversityMoscow, Russian Federation
  • 5 Lomonosov Moscow State UniversityMoscow, Russian Federation
  • 6 A.P. Avtsyn Research institute of human morphologyMoscow, Russian Federation
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