mimmunМедицинская иммунологияMedical Immunology (Russia)1563-06252313-741XSPb RAACI10.15789/1563-0625-MCA-2525mimmun-2525Research ArticleОРИГИНАЛЬНЫЕ СТАТЬИORIGINAL ARTICLESM-CSF и GM-CSF детерминируют фибромодулирующую активность поляризованных макрофагов человекаM-CSF and GM-CSF determinate fibromodulatory activity of polarized human macrophageshttps://orcid.org/0000-0003-0884-0226МаксимоваА. А.MaksimovaA. A.

Максимова Александра Александровна, к.м.н., младший научный сотрудник лаборатории клеточной иммунотерапии

630099, г. Новосибирск, ул. Ядринцевская, 14Тел.: 8 (383) 222-26-74Факс: 8 (383) 222-70-28

Maksimova Aleksandra A. PhD (Mecicine), Junior Research Associate, Laboratory of Cellular Immunotherapy

630099, Novosibirsk, Yadrintsevskaya str., 14Phone: 7 (383) 222-26-74Fax: 7 (383) 222-70-28

parkinson.dses@gmail.comhttps://orcid.org/0000-0001-8997-3586ШевелаЕ. Я.ShevelaE. Ya.

д.м.н., ведущий научный сотрудник лаборатории клеточной иммунотерапии

г. Новосибирск

PhD, MD (Mecicine), Leading Research Associate, Laboratory of Cellular Immunotherapy

Novosibirsk

shevelak@mail.ruhttps://orcid.org/0000-0003-3290-7910СахноЛ. В.SakhnoL. V.

к.б.н., старший научный сотрудник лаборатории клеточной иммунотерапии

г. Новосибирск

PhD (Biology), Senior Research Associate, Laboratory of Cellular Immunotherapy

Novosibirsk

lsahno53@mail.ruhttps://orcid.org/0000-0002-2366-1667ТихоноваМ. А.TikhonovaM. A.

к.б.н., старший научный сотрудник лаборатории клеточной иммунотерапии

г. Новосибирск

PhD (Biology), Senior Research Associate, Laboratory of Cellular Immunotherapy

Novosibirsk

martix-59@mail.ruhttps://orcid.org/0000-0001-6895-938XОстанинА. А.OstaninA. A.

д.м.н., профессор, главный научный сотрудник лаборатории клеточной иммунотерапии

г. Новосибирск

PhD, MD (Mecicine), Professor, Chief Research Associate, Laboratory of Cellular Immunotherapy

Novosibirsk

ostanin62@mail.ruhttps://orcid.org/0000-0003-2346-6279ЧерныхЕ. Р.ChernykhE. R.

д.м.н., профессор, член-корр. РАН, заведующая лабораторией клеточной иммунотерапии

г. Новосибирск

PhD, MD (Mecicine), Professor, Corresponding Member, Russian Academy of Sciences, Head, Laboratory of Cellular Immunotherapy

Novosibirsk

ct_lab@mail.ruФГБНУ «Научно-исследовательский институт фундаментальной и клинической иммунологии»РоссияResearch Institute of Fundamental and Clinical ImmunologyRussian Federation202231102022245931942Copyright © Максимова А.А., Шевела Е.Я., Сахно Л.В., Тихонова М.А., Останин А.А., Черных Е.Р., 20222022Максимова А.А., Шевела Е.Я., Сахно Л.В., Тихонова М.А., Останин А.А., Черных Е.Р.Maksimova A.A., Shevela E.Y., Sakhno L.V., Tikhonova M.A., Ostanin A.A., Chernykh E.R.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.mimmun.ru/mimmun/article/view/2525

Колониестимулирующие факторы, GM-CSF и M-CSF, вызывают различные фенотипические изменения в популяциях линии макрофагов, способствуя дифференцировке клеток в М1- и М2-подобные макрофаги соответственно. Про- и противовоспалительные свойства макрофагов, генерируемых под влиянием данных колониестимулирующих факторов, хорошо описаны в литературе, однако вклад дифференцировочных и поляризующих сигналов в фибромодулирующую активность макрофагов остается неизученным. Для выяснения различий в регуляторных механизмах, контролирующих фиброгенез, которые присущи различно активированным макрофагам, нами было проведено исследование влияния кондиционных сред макрофагов на пролиферацию и дифференцировку фибробластов дермы. В работе были исследованы макрофаги человека, генерированные из моноцитов периферической крови, которые дифференцировали M-CSF или GM-CSF и далее поляризовали в М1-направлении липополисахаридом и в М2-направлении IL-4 или дексаметазоном. Пролиферативный ответ фибробластов определяли радиометрически по включению [3H]-тимидина. Дифференцировку в миофибробласты определяли по экспрессии специфического маркера α-гладкомышечного актина (α-SMA) с помощью проточной цитометрии. Уровень продукции TGF-β1 макрофагами оценивали с помощью соответствующего ELISA kit. Полученные результаты свидетельствуют о том, что макрофаги, дифференцированные под влиянием «гомеостатического» M-CSF, характеризуются умеренным стимулирующим влиянием на пролиферацию фибробластов, причем эффекты М2 (IL-4) и М2 (Dex) макрофагов превышают эффекты М1 (LPS), при этом не различаются значимо между собой. M-CSF-индуцированные М1 (LPS) и М2 (IL-4) макрофаги, но не М2 (Dex), также усиливают дифференцировку фибробластов, демонстрируя схожий уровень стимуляции. В отличие от M-CSF, макрофаги, индуцированные «провоспалительным» GM-CSF, оказывают выраженный стимулирующий эффект на пролиферацию фибробластов, причем эффекты М2-макрофагов превышают таковые М1-клеток и наиболее ярко выражены у М2 (Dex). В то же время способность усиливать дифференцировку фибробластов проявляют только GM-CSF-индуцированные макрофаги с М2 (IL-4) фенотипом. Поляризованные дексаметазоном макрофаги – вне зависимости от используемого CSF (M-CSF или GM-CSF) – не влияют значимо на дифференцировку фибробластов. Содержание TGF-β1 в супернатантах различно активированных макрофагов не коррелирует с уровнем стимулирующего влияния кондиционных сред макрофагов на дифференцировку фибробластов. В целом, полученные данные свидетельствуют о вовлечении дифференцировочных и поляризующих сигналов в модуляцию про- и антифиброгенных свойств макрофагов.

GM-CSF and M-CSF, the hematopoietic colony-stimulating factors, induce various phenotypic changes in macrophage lineage populations and promote cell differentiation, respectively, into M1- and M2-like macrophages. The pro- and anti-inflammatory properties of macrophages generated by these colony-stimulating factors are well described, but the contribution of differentiation and polarization signals to the fibromodulatory activity of macrophages remains unexplored. To clarify the differences in the fibrogenesis regulation mechanisms inherent in differently activated macrophages, we studied the effects of macrophage-conditioned media on proliferation and differentiation of dermal fibroblasts. In this study, the human macrophages generated from peripheral blood monocytes were investigated. They were induced for differentiation by M-CSF or GM-CSF, being further polarized in the M1 direction with lipopolysaccharide and, in the M2 direction, with IL-4 or dexamethasone. Proliferative response of the fibroblasts was determined radiometrically by [3H]-thymidine incorporation. Differentiation into myofibroblasts was determined with flow cytometry technique, as expression of a specific marker α-smooth muscle actin (α-SMA). The level of macrophage TGF-β1 production was assessed using an appropriate ELISA kit. The data obtained indicate that the macrophages differentiated under the influence of “homeostatic” M-CSF are characterized by a moderate stimulating effect upon fibroblast proliferation, and the effects of M2 (IL-4) and M2 (Dex) macrophages exceed that of M1 (LPS), but do not differ significantly from each other. The M-CSF-induced M1 (LPS) and M2 (IL-4) macrophages, but not M2 (Dex), enhance the fibroblast differentiation and show similar level of stimulation. In contrast to M-CSF, the macrophages induced by “pro-inflammatory” GM-CSF exhibit a pronounced stimulatory effect on fibroblast proliferation, and the effects of M2 macrophages exceed those of M1 cells, being most pronounced for M2 (Dex). At the same time, only GM-CSF-induced M2 (IL-4) macrophages enhance fibroblast differentiation. Dexamethasone-polarized macrophages do not significantly affect fibroblast differentiation regardless of the CSF used (M-CSF or GM-CSF). The content of TGF-β1 in the supernatants of differently activated macrophages does not correlate with the level of stimulating effect of macrophage-conditioned media upon fibroblast differentiation. In general, the data obtained suggest the involvement of differentiation and polarization signals into modulation of pro- and anti-fibrogenic properties of macrophages.

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The authors declare that there are no conflicts of interest present.