bloodjourГематология и трансфузиологияRussian journal of hematology and transfusiology0234-57302411-3042ООО Издательский дом «Практика»10.35754/0234-5730-2024-69-3-330-343bloodjour-579Research ArticleОРИГИНАЛЬНЫЕ СТАТЬИORIGINAL ARTICLESВлияние рекомбинантной глюкоцереброзидазы на пролиферацию и дифференцировку мультипотентных мезенхимных стромальных клеток человекаEffect of glucocerebrosidase on human bone marrow multipotent mesenchymal stromalhttps://orcid.org/0000-0001-7795-4564МамоновВ. Е.MamonovV. E.

Мамонов Василий Евгеньевич, кандидат медицинских наук, заведующий отделением травматологии и ортопедии

125167, г. Москва

Vasily E. Mamonov, Cand. Sci. (Med.), Head of the Orthopedic Department

125167, Moscow

vasily-mamonov@yandex.ruhttps://orcid.org/0000-0002-0221-9351СадовскаяА. В.SadovskayaA. V.

Садовская Александра Вячеславовна, лаборант отдела сопровождения клинических исследований 

125167, г. Москва

Aleksandra V. Sadovskaya, Laboratory Assistant of the Clinical Research Support Department

125167, Moscow

sadovskaya.sasha@gmail.comhttps://orcid.org/0000-0001-6591-3183ПетинатиН. А.PetinatiN. A.

Петинати Наталия Арнольдовна, кандидат медицинских наук, заведующая лабораторией физиологии кроветворения 

125167, г. Москва

Nataliya A. Petinati, Cand. Sci. (Med.), Head of the Laboratory of Hematopoiesis Physiology

125167, Moscow

petinati.n@blood.ruhttps://orcid.org/0000-0003-1189-0283ШипуноваИ. Н.ShipounovaI. N.

Шипунова Ирина Николаевна, доктор биологических наук, старший научный сотрудник лаборатории физиологии кроветворения 

125167, г. Москва

Irina N. Shipounova,  Dr. Sci. (Biol.), Senior Researcher of the Laboratory of Hematopoiesis Physiology

125167, Moscow

shipounova.i@blood.ruhttps://orcid.org/0000-0002-1154-5669ДорофееваА. И.DorofeevaA. I.

Дорофеева Алена Игоревна, кандидат биологических наук, научный сотрудник лаборатории физиологии кроветворения 

125167, г. Москва

Alena I. Dorofeeva, Cand. Sci. (Biol.), Researcher of the Laboratory of Hematopoiesis Physiology

125167, Moscow

dorofeeva.a@blood.ruhttps://orcid.org/0000-0002-7150-0403ДризеН. И.DrizeN. I.

Дризе Нина Иосифовна, доктор биологических наук, ведущий научный сотрудник лаборатории физиологии кроветворения 

125167, г. Москва

Nina I. Drize, Dr. Sci. (Biol.), Leading Researcher of the Laboratory of Hematopoiesis Physiology

125167, Moscow

ndrize@yandex.ruhttps://orcid.org/0000-0002-8774-850XЛукинаЕ. А.LukinaE. A.

Лукина Елена Алексеевна, доктор медицинских наук, профессор, заведующая отделением гематологии и химиотерапии орфанных заболеваний 

125167, г. Москва

Elena A. Lukina, Dr. Sci. (Med.), Professor, Head of the Department of Hematology and Chemotherapy of Orphan Diseases

125167, Moscow

lukina.e@blood.ruФГБУ «Национальный медицинский исследовательский центр гематологии» Министерства здравоохранения Российской ФедерацииРоссияNational Medical Research Center for HematologyRussian Federation202425112024693330343Copyright © Мамонов В.Е., Садовская А.В., Петинати Н.А., Шипунова И.Н., Дорофеева А.И., Дризе Н.И., Лукина Е.А., 20242024Мамонов В.Е., Садовская А.В., Петинати Н.А., Шипунова И.Н., Дорофеева А.И., Дризе Н.И., Лукина Е.А.Mamonov V.E., Sadovskaya A.V., Petinati N.A., Shipounova I.N., Dorofeeva A.I., Drize N.I., Lukina E.A.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.htjournal.ru/jour/article/view/579Введение

Введение. При проведении заместительной ферментной терапии у больных болезнью Гоше (БГ) рекомбинантной глюкоцереброзидазой (ГЦБ) наблюдаются изменения в костной ткани, которые могут быть обусловлены нарушением дифференцировки мультипотентных мезенхимных стромальных клеток (МСК).

Цель — изучить изменения в МСК здоровых доноров и больной БГ при культивировании в присутствии ГЦБ.

Методы

Методы. МСК выделяли из костного мозга 17 здоровых доноров и 1 больной БГ стандартным методом, культивировали в присутствии различных концентраций ГЦБ после второго пассажа от 2 до 7 недель. Анализировали пролиферацию клеток, способность к дифференцировке, в том числе после индукции. Оценку проводили методами дифференциального окрашивания, элюции и экспрессии генов — маркеров дифференцировки методом ПЦР в реальном времени.

Результаты

Результаты. Низкие дозы рекомбинантной ГЦБ (0,25–1,5 ед/мл) не влияли на пролиферативную активность МСК. Длительное культивирование МСК в присутствии низких концентраций ГЦБ приводило к изменению дифференцировочного потенциала этих клеток в направлении адипогенеза. Дозы фермента 3–5 ед/мл угнетали пролиферацию МСК и вызывали существенные изменения в дифференцировке клеток. Высокие концентрации ГЦБ (7–10 ед/мл) обладали цитотоксическим эффектом и приводили к гибели клеток в течение одного пассажа. Пролиферативный и дифференцировочный потенциал МСК больной БГ существенно отличались от клеток здоровых доноров по исследованным параметрам.

Заключение

Заключение. Культивирование МСК доноров в присутствии рекомбинантной ГЦБ изменяет пролиферацию и дифференцировочный потенциал этих клеток. Эти изменения зависят от дозы фермента в среде и длительности культивирования.

Introduction

Introduction. During enzyme replacement therapy in patients with Gaucher disease (GD) with recombinant glucocerebrosidase (GCase), regression of bone manifestations is possible, but with prolonged therapy osteonecrosis may occur. These changes may be due to impaired differentiation of multipotent mesenchymal stromal cells (MSCs).

Aim

Aim: to study changes in the MSCs of healthy donors and a patient with GD when cultured in the presence of GCase.

Material and methods

Material and methods. MSCs were isolated from the bone marrow of 17 healthy donors and a female patient with GD by a standard method and cultured in the presence of various concentrations of GCase after the second passage from 2 to 7 weeks. Cell proliferation and the ability to differentiate were analyzed, including after induction. The assessment was carried out by differential staining, elution, and expression of differentiation marker genes by real-time PCR.

Results

Results. Low concentrations of recombinant GCase (0.25–1.5 U/ml) did not affect the proliferative activity of MSCs. Prolonged cultivation of MSCs in the presence of low doses of GCase led to a change in the differentiation potential of these cells in the direction of adipogenesis. Concentrations of GCase of 3–5 U/ml inhibited the proliferation of MSCs and caused significant changes in cell differentiation. High doses of the enzyme (7–10 U/ml) had a cytotoxic effect and led to cell death within one passage. The proliferative and differentiation potential of the MSCs of a patient with GD differed significantly from the cells of healthy donors in all the parameters studied.

Conclusion

Conclusion. The cultivation of donor MSCs in the presence of recombinant GCase alters the proliferation and differentiation potential of these cells. These changes depend on the dose of the enzyme in the medium and the duration of cultivation.

мультипотентные мезенхимные стромальные клеткиглюкоцереброзидазадифференцировкаэкспрессия геновmultipotent mesenchymal stromal cellsglucocerebrosidasedifferentiationexpression of genesРабота выполнена в рамках государственного задания «Оптимизация диагностики, лечения и мониторинга неопухолевых заболеваний крови у взрослых на основе молекулярно-генетических, иммунофенотипических, биологических параметров», рег. № НИОКТР 122012700239–9The study was carried out on assignment: “Optimization of diagnosis, treatment and monitoring of non-tumor blood diseases in adults based on molecular-genetic, immunophenotypic, biological parameters” № 122012700239-9ReferencesMeikle P.J., Hopwood J.J., Clague A.E., et al. Prevalence of lysosomal storage disorders. JAMA. 1999; 281(3): 249–54. DOI: 10.1001/jama.281.3.249.Meikle P.J., Hopwood J.J., Clague A.E., et al. Prevalence of lysosomal storage disorders. JAMA. 1999; 281(3): 249–54. DOI: 10.1001/jama.281.3.249.Zimran A. How I treat Gaucher disease. 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The authors declare that there are no conflicts of interest present.