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<article article-type="research-article" dtd-version="1.3" xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance" xml:lang="ru"><front><journal-meta><journal-id journal-id-type="publisher-id">mimmun</journal-id><journal-title-group><journal-title xml:lang="ru">Медицинская иммунология</journal-title><trans-title-group xml:lang="en"><trans-title>Medical Immunology (Russia)</trans-title></trans-title-group></journal-title-group><issn pub-type="ppub">1563-0625</issn><issn pub-type="epub">2313-741X</issn><publisher><publisher-name>SPb RAACI</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="doi">10.15789/1563-0625-FCD-2124</article-id><article-id custom-type="elpub" pub-id-type="custom">mimmun-2124</article-id><article-categories><subj-group subj-group-type="heading"><subject>Research Article</subject></subj-group><subj-group subj-group-type="section-heading" xml:lang="ru"><subject>ИММУНОЛОГИЧЕСКИЕ МЕТОДЫ</subject></subj-group><subj-group subj-group-type="section-heading" xml:lang="en"><subject>IMMUNOLOGICAL METHODS</subject></subj-group></article-categories><title-group><article-title>Определение и имунофенотипирование тромбоцитарно-моноцитарных комплексов в периферической крови с помощью проточной цитофлуорометрии</article-title><trans-title-group xml:lang="en"><trans-title>Flow cytofluorimetric detection and immunophenotyping of platelet-monocyte complexes in peripheral blood</trans-title></trans-title-group></title-group><contrib-group><contrib contrib-type="author" corresp="yes"><contrib-id contrib-id-type="orcid">https://orcid.org/0000-0003-4873-4081</contrib-id><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Павлов</surname><given-names>О. В.</given-names></name><name name-style="western" xml:lang="en"><surname>Pavlov</surname><given-names>O. V.</given-names></name></name-alternatives><bio xml:lang="ru"><p>д.б.н., ведущий научный сотрудник отдела иммунологии и межклеточных взаимодействий,</p><p>199034, Санкт-Петербург, Менделеевская линия, 3</p></bio><bio xml:lang="en"><p>PhD, MD (Biology), Leading Research Associate, Department of Immunology and Cell Interactions, </p><p>199034, St. Petersburg, Mendeleev line, 3</p></bio><email xlink:type="simple">ovpavlov@hotmail.com</email><xref ref-type="aff" rid="aff-1"/></contrib><contrib contrib-type="author" corresp="yes"><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Чепанов</surname><given-names>С В.</given-names></name><name name-style="western" xml:lang="en"><surname>Chepanov</surname><given-names>S. V.</given-names></name></name-alternatives><bio xml:lang="ru"><p>к.м.н., старший научный сотрудник отдела иммунологии и межклеточных взаимодействий,</p><p>Санкт-Петербург</p></bio><bio xml:lang="en"><p>PhD (Medicine), Senior Research Associate, Department of Immunology and Cell Interaction,</p><p>St. Petersburg</p></bio><email xlink:type="simple">chepanovsv@gmail.com</email><xref ref-type="aff" rid="aff-1"/></contrib><contrib contrib-type="author" corresp="yes"><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Селютин</surname><given-names>А. В.</given-names></name><name name-style="western" xml:lang="en"><surname>Selutin</surname><given-names>A. V.</given-names></name></name-alternatives><bio xml:lang="ru"><p>к.б.н., старший научный сотрудник отдела иммунологии и межклеточных взаимодействий,</p><p>Санкт-Петербург</p></bio><bio xml:lang="en"><p>PhD (Biology), Senior Research Associate, Department of Immunology and Cell Interactions,</p><p>St. Petersburg</p></bio><email xlink:type="simple">a_selutin@yahoo.com</email><xref ref-type="aff" rid="aff-1"/></contrib><contrib contrib-type="author" corresp="yes"><contrib-id contrib-id-type="orcid">https://orcid.org/0000-0002-2622-5000</contrib-id><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Зайнулина</surname><given-names>М. С.</given-names></name><name name-style="western" xml:lang="en"><surname>Zainulina</surname><given-names>M. S.</given-names></name></name-alternatives><bio xml:lang="ru"><p>д.м.н., главный врач;</p><p>профессор кафедры акушерства, гинекологии и репродуктологии,</p><p>Санкт-Петербург</p></bio><bio xml:lang="en"><p>PhD, MD (Medicine), Head Physician;</p><p>Professor, Department of Obstetrics, Gynecology and Reproductive Medicine,</p><p>St. Petersburg</p></bio><email xlink:type="simple">marzainulina@gmail.com</email><xref ref-type="aff" rid="aff-2"/></contrib><contrib contrib-type="author" corresp="yes"><contrib-id contrib-id-type="orcid">https://orcid.org/0000-0003-4512-9599</contrib-id><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Еремеева</surname><given-names>Д. Р.</given-names></name><name name-style="western" xml:lang="en"><surname>Eremeeva</surname><given-names>D. R.</given-names></name></name-alternatives><bio xml:lang="ru"><p>к.м.н., заместитель главного врача по качеству;</p><p>ассистент кафедры акушерства, гинекологии и репродуктологии,</p><p>Санкт-Петербург</p></bio><bio xml:lang="en"><p>PhD (Medicine), Deputy Head Physician for Qualityж</p><p>Assistant Professor, Department of Obstetrics, Gynecology and Reproductive Medicine,</p><p>St. Petersburg</p></bio><email xlink:type="simple">dina-bikmullina@yandex.ru</email><xref ref-type="aff" rid="aff-2"/></contrib><contrib contrib-type="author" corresp="yes"><contrib-id contrib-id-type="orcid">https://orcid.org/0000-0003-1560-7529</contrib-id><name-alternatives><name name-style="eastern" xml:lang="ru"><surname>Сельков</surname><given-names>С. А.</given-names></name><name name-style="western" xml:lang="en"><surname>Selkov</surname><given-names>S. A.</given-names></name></name-alternatives><bio xml:lang="ru"><p>д.м.н., профессор, заслуженный деятель науки РФ, заведующий отделом иммунологии и межклеточных взаимодействий;</p><p>профессор кафедры иммунологии,</p><p>Санкт-Петербург</p></bio><bio xml:lang="en"><p>PhD, MD (Medicine), Professor, Honored Scientist of the Russian Federation, Head, Department of Immunology and Cell Interactions;</p><p>Professor, Department of Immunology,</p><p>St. Petersburg</p></bio><email xlink:type="simple">selkovsa@mail.ru</email><xref ref-type="aff" rid="aff-3"/></contrib></contrib-group><aff-alternatives id="aff-1"><aff xml:lang="ru">ФГБНУ «Научно-исследовательский институт акушерства, гинекологии и репродуктологии имени Д.О. Отта»<country>Россия</country></aff><aff xml:lang="en">D. Ott Research Institute of Obstetrics, Gynecology and Reproductive Medicine<country>Russian Federation</country></aff></aff-alternatives><aff-alternatives id="aff-2"><aff xml:lang="ru">СПб ГБУЗ «Родильный дом № 6 имени профессора В.Ф. Снегирева»;&#13;
ФГБОУ ВО «Первый Санкт-Петербургский государственный медицинский университет имени академика И.П. Павлова» Министерства здравоохранения РФ<country>Россия</country></aff><aff xml:lang="en">V. Snegirev Maternity Hospital No. 6;&#13;
First St. Petersburg I. Pavlov State Medical University<country>Russian Federation</country></aff></aff-alternatives><aff-alternatives id="aff-3"><aff xml:lang="ru">ФГБНУ «Научно-исследовательский институт акушерства, гинекологии и репродуктологии имени Д.О. Отта»;&#13;
ФГБОУ ВО «Первый Санкт-Петербургский государственный медицинский университет имени академика И.П. Павлова» Министерства здравоохранения РФ<country>Россия</country></aff><aff xml:lang="en">D. Ott Research Institute of Obstetrics, Gynecology and Reproductive Medicine;&#13;
First St. Petersburg I. Pavlov State Medical University<country>Russian Federation</country></aff></aff-alternatives><pub-date pub-type="collection"><year>2021</year></pub-date><pub-date pub-type="epub"><day>24</day><month>04</month><year>2021</year></pub-date><volume>23</volume><issue>2</issue><fpage>401</fpage><lpage>410</lpage><permissions><copyright-statement>Copyright &amp;#x00A9; Павлов О.В., Чепанов С.В., Селютин А.В., Зайнулина М.С., Еремеева Д.Р., Сельков С.А., 2021</copyright-statement><copyright-year>2021</copyright-year><copyright-holder xml:lang="ru">Павлов О.В., Чепанов С.В., Селютин А.В., Зайнулина М.С., Еремеева Д.Р., Сельков С.А.</copyright-holder><copyright-holder xml:lang="en">Pavlov O.V., Chepanov S.V., Selutin A.V., Zainulina M.S., Eremeeva D.R., Selkov S.A.</copyright-holder><license license-type="creative-commons-attribution" xlink:href="https://creativecommons.org/licenses/by/4.0/" xlink:type="simple"><license-p>This work is licensed under a Creative Commons Attribution 4.0 License.</license-p></license></permissions><self-uri xlink:href="https://www.mimmun.ru/mimmun/article/view/2124">https://www.mimmun.ru/mimmun/article/view/2124</self-uri><abstract><p>Активированные тромбоциты образуют комплексы с циркулирующими лейкоцитами посредством мембраносвязанных молекул. Взаимодействие тромбоцитов с моноцитами лежат в основе патофизиологических механизмов, связывающих процессы тромбообразования и воспаления. Определение и анализ тромбоцитарно-моноцитарных комплексов (ТМК) позволяет выявить их физиологическую и патогенетическую роль, а также имеет важное диагностическое значение при изучении различных патологических состояний, в том числе, акушерских. Цель исследования состояла в разработке метода определения количественного содержания ТМК в периферической крови, позволяющего сохранить основные фенотипические характеристики тромбоцитов и моноцитов, и выявить их изменения при анализе биоматериала ex vivo. Предлагаемый метод сочетает в себе немедленную фиксацию образцов крови, иммуноцитохимическое флуоресцентное окрашивание дифференцировочных и активационных маркеров тромбоцитов и моноцитов с последующим лизисом эритроцитов и анализ с помощью проточной цитофлуориметрии. Было исследовано 14 образцов периферической крови, полученных от пациенток с отягощенным акушерским анамнезом, в первом триместре текущей беременности. Показано, что процедура фиксации позволяет сохранить количественные и качественные характеристики ТМК, существующие in vivo, тогда как при отсутствии фиксации наблюдается многократное увеличение количества ТМК и уровня экспрессии активационных маркеров тромбоцитов и моноцитов ex vivo. Используемая панель моноклональных антител и примененные стратегии гейтирования обеспечивают оценку относительного и абсолютного количества ТМК и их фенотипических характеристик как в общей популяции (CD45+CD14+) моноцитов, так и в субпопуляциях «классических» (CD14+CD16-), «промежуточных» (CD14+CD16+) и «неклассических» (CD14lowCD16+) моноцитов. Выработанный подход позволяет оценить вклад отдельных субпопуляций моноцитов в формирование ТМК и степень их участия в физиологических и патофизиологических процессах. В отдельных образцах было выявлено увеличение количества ТМК, сопровождавшееся существенным повышением в них экспрессии активационного маркера тромбоцитов CD62P и снижением экспрессии его моноцитарного лиганда CD162. Данные изменения могут свидетельствовать об изменении активационного статуса комплексообразующих клеток и возможном участии ТМК в патофизиологических механизмах некоторых акушерских осложнений. Иммунофенотипирование ТМК позволяет дополнительно охарактеризовать их провоспалительный, прокоагулянтный и адгезионный потенциал и может быть применено как в фундаментальных исследованиях, так и в целях диагностики. В частности, данный метод может быть использован для определения и характеристики ТМК при акушерских осложнениях, сопровождающихся воспалением и нарушениями гемостаза.</p></abstract><trans-abstract xml:lang="en"><p>Activated platelets aggregate with monocytes by binding membrane bound molecules. Platelet-monocyte interaction is considered to underlie pathophysiological mechanisms bridging thrombosis and inflammation. Detection and analysis of platelet-monocyte complexes (PMC) provide means for revealing their physiological and pathogenetic roles and are instrumental in the diagnostics of various pathological conditions including obstetric complications. The aim of the study was to develop the method of quantitative determination of peripheral blood PMC, that preserve phenotypic features of platelets and monocytes, and to reveal their changes by ex vivo analysis. The suggested procedure includes immediate fixation of blood sample, immunocytochemical staining with fluorochrome-conjugated specific antibodies against markers of activation and differentiation followed by lysis of erythrocytes, and flow cytometric analysis. Fourteen samples of peripheral blood from patients with history of pregnancy complication were obtained in first trimester of ongoing pregnancy and analyzed. It was demonstrated that quantitative and qualitative in vivo characteristics of PMC remained unchanged in fixed samples, whereas the number of PMC and expression levels of the markers of platelet and monocyte activation dramatically increased in the unfixed blood. The set of monoclonal antibodies and gating strategies, used in this study, ensure phenotyping and evaluation of percentage/absolute count of PMC in the total monocyte population (CD45+CD14+) and in the subpopulations of classical (CD14+CD16-), intermediate (CD14+CD16+), and non-classical (CD14lowCD16+) monocytes. This approach provides insight into the participation of different monocyte subsets in the formation of PMC and their roles in physiological and pathophysiological processes. In some samples, elevated PMC proportion was observed, accompanied by significant increase in the expression of platelet activation marker CD62P and decrease in the expression of its monocytic ligand CD162. These changes suggested altered activation of PMC and their participation in the pathophysiological mechanisms of some pregnancy complications. Immunophenotyping of PMC affords an opportunity to characterize their proinflammatory, procoagulant and adhesive properties; these results can be used for research and diagnostics. In particular, the method is suitable for detection and phenotyping of PMC in pregnancy complications and other pathological conditions associated with the disorders of hemostasis and thrombosis.</p></trans-abstract><kwd-group xml:lang="ru"><kwd>тромбоцитарно-моноцитарные комплексы</kwd><kwd>тромбоциты</kwd><kwd>моноциты</kwd><kwd>периферическая кровь</kwd><kwd>иммунофенотипирование</kwd><kwd>проточная цитофлуориметрия</kwd></kwd-group><kwd-group xml:lang="en"><kwd>platelet-monocyte complexes</kwd><kwd>platelets</kwd><kwd>monocytes</kwd><kwd>peripheral blood</kwd><kwd>immunophenotyping</kwd><kwd>flow cytofluorimetry</kwd></kwd-group></article-meta></front><back><ref-list><title>References</title><ref id="cit1"><label>1</label><citation-alternatives><mixed-citation xml:lang="ru">Aleva F.E., Temba G., de Mast Q., Simons S.O., de Groot P.G., Heijdra Y.F., van der Ven A. Increased plateletmonocyte interaction in stable COPD in the absence of platelet hyper-reactivity. 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