biopreparatБИОпрепараты. Профилактика, диагностика, лечениеBiological Products. Prevention, Diagnosis, Treatment2221-996X2619-1156Scientific Centre for Expert Evaluation of Medicinal Products10.30895/2221-996X-2026-761biopreparat-761Research ArticleМЕХАНИЗМЫ ИММУНОПАТОЛОГИЧЕСКИХ ПРОЦЕССОВMECHANISMS OF IMMUNOPATHOLOGICAL PROCESSESПрименение метода детекции антителозависимого усиления инфекции in vitro на примере вируса ЧикунгуньяApplication of an in vitro method for detecting antibody-dependent enhancement of Chikungunya virus infectionhttps://orcid.org/0000-0002-2491-4072ОтрашевскаяЕ. В.OtrashevskajaE. V.

Отрашевская Елена Викторовна

Малый Казенный пер., д. 5а, Москва, 105064

Еlena V. Otrashevskaja

5A Maly Kazenny Ln, Moscow, 105064

https://orcid.org/0000-0003-3264-6722СамарцеваТ. Г.SamartsevaT. G.

Самарцева Татьяна Геннадьевна

Малый Казенный пер., д. 5а, Москва, 105064

Tatyana G. Samartseva

5A Maly Kazenny Ln, Moscow, 105064

https://orcid.org/0000-0002-8600-7347ОксаничА. С.OksanichA. S.

Оксанич Алексей Сергеевич, канд. биол. наук

Малый Казенный пер., д. 5а, Москва, 105064

Aleksey S. Oksanich, Cand. Sci. (Biol.)

5A Maly Kazenny Ln, Moscow, 105064

https://orcid.org/0009-0009-2889-9680ГогинаС. С.GoginaS. S.

Гогина Софья Сергеевна

Малый Казенный пер., д. 5а, Москва, 105064

Sofya S. Gogina

5A Maly Kazenny Ln, Moscow, 105064

https://orcid.org/0000-0001-5808-2246ЗверевВ. В.ZverevV. V.

Зверев Виталий Васильевич, д-р биол. наук, проф., академик РАН

Малый Казенный пер., д. 5а, Москва, 105064

Vitaly V. Zverev, Dr. Sci. (Biol.), Prof., Acad. RAS

5A Maly Kazenny Ln, Moscow, 105064

https://orcid.org/0000-0002-9731-3681ИгнатьевГ. М.IgnatyevG. M.

Игнатьев Георгий Михайлович, д-р мед. наук, проф.

Малый Казенный пер., д. 5а, Москва, 105064

George M. Ignatyev, Dr. Sci. (Med.), Prof.

5A Maly Kazenny Ln, Moscow, 105064

Marburgman@mail.ruФедеральное государственное бюджетное научное учреждение «Научно-исследовательский институт вакцин и сывороток им. И.И. Мечникова»РоссияI. Mechnikov Research Institute for Vaccines and SeraRussian Federation202604072026262183195Copyright © Отрашевская Е.В., Самарцева Т.Г., Оксанич А.С., Гогина С.С., Зверев В.В., Игнатьев Г.М., 20262026Отрашевская Е.В., Самарцева Т.Г., Оксанич А.С., Гогина С.С., Зверев В.В., Игнатьев Г.М.Otrashevskaja E.V., Samartseva T.G., Oksanich A.S., Gogina S.S., Zverev V.V., Ignatyev G.M.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.biopreparations.ru/jour/article/view/761ВВЕДЕНИЕ

ВВЕДЕНИЕ. Механизм антителозависимого усиления инфекции (antibody-dependent enhacement, ADE), опосредуемый через Fcγ-рецептор иммунных клеток, принято считать основным. Комплекс вируса со специфическим иммуноглобулином G (IgG) взаимодействует с данным рецептором, обеспечивая вирусу лучшее проникновение в клетку. Развитие ADE возможно и у вакцинированных, что предполагает его изучение при оценке безопасности разрабатываемых вакцин.

ЦЕЛЬ

ЦЕЛЬ. Применение метода детекции антителозависимого усиления in vitro с использованием клеток линий К562 и Vero, а также штамма вируса Чикунгунья Nika21.

МАТЕРИАЛЫ И МЕТОДЫ

МАТЕРИАЛЫ И МЕТОДЫ. В исследовании использовали вирус Чикунгунья (ЧикВ) штамм Nika21 и сыворотки, содержащие cпецифические анти-ЧикВ IgG. Специфические сыворотки использовались в разных разведениях. Для оценки ADE клетки линий К562 и Vero заражали «смесью» ЧикВ со специфическими антителами (АТ). На четвертые сутки после заражения определяли биологический титр ЧикВ и содержание вирусной РНК. Титр ЧикВ определяли с помощью стандартного метода титрования вируса in vitro, а количество РНК — с помощью флуоресцентного метода ПЦР с обратной транскрипцией в режиме реального времени. Наличие ADE считали подтвержденным, если значения биологического титра ЧикВ в присутствии исследуемой сыворотки в данном разведении были >3 SD относительно среднего титра ЧикВ в контроле, который представлял собой «смесь» ЧикВ с сывороткой здорового добровольца.

РЕЗУЛЬТАТЫ

РЕЗУЛЬТАТЫ. В клетках линии Vero с помощью обоих методов было отмечено усиление репликации ЧикВ при увеличении разведения иммунных сывороток, которое, однако, не превышало значений титров ЧикВ в контроле. В клетках линии К562 исследуемые сыворотки в одном разведении продемонстрировали статистически достоверное усиление репликации ЧикВ относительно соответствующего контроля. Таким образом, в нашем исследовании ADE было обнаружено в присутствии промежуточных разведений иммунных сывороток с относительно средним или низким титром анти-ЧикВ АТ.

ВЫВОДЫ

ВЫВОДЫ. Репликация ЧикВ в FcR-негативных клетках линии Vero подтвердила высокую чувствительность данной линии клеток к вирусу и возможность использования ее исключительно для оценки нейтрализующей активности специфических АТ. Клетки FcR-экспрессирующей линии К562 позволили анализировать обе функции АТ: и нейтрализующую вирус, и активирующую инфекцию. Результаты работы подтвердили возможность проведения исследований ADE in vitro, что значительно упрощает исследование феномена ADE при оценке безопасности разрабатываемых вакцин.

INTRODUCTION

INTRODUCTION. The mechanism of antibody-dependent enhancement (ADE) caused by the Fcγ receptor on immune cells is a key factor. When the virus binds to specific immunoglobulin G

(IgG), it enhances cell penetration. ADE can also occur in vaccinated individuals, making its study crucial for evaluating vaccine safety.

AIM

AIM. This study aimed to apply the method of detection of antibody-dependent enhacement in vitro using cells of the K562 and Vero lines, as well as the Chikungunya virus strain Nika21.

MATERIALS AND METHODS

MATERIALS AND METHODS. Chikungunya virus (ChikV) Nika21 strain and different dilutions sera containing anti-ChikV IgG were used. To evaluate ADE, K562 and Vero cells were challenged with a ChikV with specific IgG mixture. On the fourth day after infection, the biological ChikV titer and viral RNA content were determined. ChikV titer was determined by standard in vitro virus titration and RNA amount was determined by real-time fluorescence reverse transcription PCR. ADE was considered confirmed if the values of the biological ChikV titer in the presence of the tested serum were >3 SD relative to the mean ChikV titer in the control, which represented ChikV with serum of a healthy volunteer.

RESULTS

RESULTS. In Vero cells, both methods showed increased ChikV replication with an increase in the dilution of immune sera, which, however, did not exceed the values of ChikV titers in the control. In K562 cells, serum with anti-ChikV IgG in one dilution demonstrated statistically significant enhancement of ChikV replication relative to the corresponding control. Thus, in our study, ADE was detected in the presence of intermediate dilutions of immune sera with a relatively medium or low titer of anti-ChikV AT.

CONCLUSION

CONCLUSION. ChikV replication in FcR-negative Vero cell line confirmed the high sensitivity to the virus and the possibility of using it exclusively for assessing the neutralizing activity of specific IgG. FcR-expressing K562 cell line allowed to analyze both virus neutralizing and activating infection functions of IgG. Our results confirmed the possibility of conducting ADE studies in vitro, which greatly simplifies the study of the ADE phenomenon when assessing the safety of vaccines under development.

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