Группа биомедицинских материалов

Группа разрабатывает и изучает различные биодеградируемые  материалы  (скаффолды) на основе природных и синтетических полимеров для тканевой инженерии и регенеративной медицины При этом такие биоматериалы  представлены в различных формах, в частности в виде пленок, нано- и микроволокон, гидрогелей, микрочастиц (микроносителей для роста клеток)  Для изучения цитотоксичности и других важных свойств полученных матриксов в модели  in vitro используются   различные культуры клеток (фибробласты, остеобласты, стволовые мезенхимальные  клетки и др). Кроме того, в группе  разрабатываются как различные  системы доставки противоопухолевых лекарств (наночастицы, мицеллы, липосомы, полиэлектролитные наноконтейнеры  и др), так и новые 3D  in vitro модели на основе мультиклеточных опухолевых сфероидов для тестирования этих систем.  Такие 3D  in vitro модели перспективны для исследования механизмов действия противораковой терапии (химиотерапия, фотодинамическая терапия и др) и скрининга новых лекарств,  а также средств их доставки  непосредственно перед клиническими испытаниями. Их использование позволяет  удешевить клинические испытания   и минимизировать  количество животных, необходимых  для этих тестов.  

Группа сотрудничает с  различными лабораториями ИБХ, а также с Национальным политехническим институтом Лотарингии (Нанси, Франция), Центром биоматериалов Льежского университета (Льеж, Бельгия), Страсбургским университетом (Страсбург, Франция), Институтом  Oniris (Нант, Франция), Королевским университетом (Кингстон, Канада) и др.

Группа основана как независимое подразделение в 2017 г., выделившись из Лаборатории полимеров для биологии.

Группа занимается разработкой биодеградируемых матриксов (микроносители, волокна, гидрогели, пленки) для регенеративной медицины (рис. 1.), систем доставки противоопухолевых лекарств (рис. 2), различных 3D-моделей in vitro на основе мультиклеточных опухолевых сфероидов (опухолевые сфероиды в микрокапсулах (рис. 3), на основе сфероидов, полученных из монослойной культуры клеток с помощью  RGD- пептидов (рис. 4), сфероидов из опухолевых  и нормальных клеток, полученных с помощью RGD-пептидов (рис. 5)).

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Рис.1. Рост мышиных фибробластов L929 на биодеградируемых полилактидных микроносителях (A), микроволокнах (B), в макропористых гидрогелях из хитозана и гиалуроновой кислоты (С), а также мезенхимальных стволовых клеток человека на хитозановых пленках, обработанных низкотемпературной плазмой в разряде постоянного тока (D). Клетки окрашены витальным красителем Сalcein AM (зеленый цвет), а структура матриксов визуализирована с помощью красителя DAPI (синий цвет). СЭМ (A,B) и конфокальная лазерная микроскопия (С,D).

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Рис. 2 Полисахаридные микроконтейнеры для доставки противораковых лекарств и их накопление в клетках M-3 (мышиная меланома) в 2D (монослойная культура) и в 3D (сфероиды) моделях.

 

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Рис.3. Опухолевые сфероиды из клеток рака молочной железы человека MCF-7, полученные культивированием в биосовместимых альгинат-хитозановых микрокапсулах.

 

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Рис. 4. Получение опухолевых сфероидов с помощью самопроизвольной агрегации клеток, индуцированной добавлением RGD-пептидов непосредственно к монослойным клеточным культурам.

 

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Рис.5. Два подхода для получения сфероидов из раковых и нормальных клеток c помощью RGD- пептидов.

Избранные публикации

  1. Attia M.F., Anton N., Akasov R., Chiper M., Markvicheva E., Vandamme T.F. (2016). Biodistribution and Toxicity of X-Ray Iodinated Contrast Agent in Nano-emulsions in Function of Their Size. Pharm. Res. 33 (3), 603–14 [+]

    This study aimed to investigate the impact of the size of X-ray iodinated contrast agent in nano-emulsions, on their toxicity and fate in vivo.

    ID:1435
  2. Demina T.S., Akopova T.A., Vladimirov L.V., Zelenetskii A.N., Markvicheva E.A., Grandfils C.h. (2016). Polylactide-based microspheres prepared using solid-state copolymerized chitosan and d,l-lactide. Mater Sci Eng C Mater Biol Appl 59, 333–8 [+]

    Amphiphilic chitosan-g-poly(d,l-lactide) copolymers have been manufactured via solid-state mechanochemical copolymerization and tailored to design polyester-based microspheres for tissue engineering. A single-step solid-state reactive blending (SSRB) using low-temperature co-extrusion has been used to prepare these copolymers. These materials have been valorized to stabilize microspheres processed by an oil/water emulsion evaporation technique. Introduction of the copolymers either in water or in the oil phase of the emulsion allowed to replace a non-degradable emulsifier typically used for microparticle preparation. To enhance cell adhesion, these copolymers were also tailored to bring amino-saccharide positively charged segments to the microbead surface. Size distribution, surface morphology, and total microparticle yield have been studied and optimized as a function of the copolymer composition.

    ID:1434
  3. Stetciura I.Y., Yashchenok A., Masic A., Lyubin E.V., Inozemtseva O.A., Drozdova M.G., Markvichova E.A., Khlebtsov B.N., Fedyanin A.A., Sukhorukov G.B., Gorin D.A., Volodkin D. (2015). Composite SERS-based satellites navigated by optical tweezers for single cell analysis. Analyst 140 (15), 4981–6 [+]

    Herein, we have designed composite SERS-active micro-satellites, which exhibit a dual role: (i) effective probes for determining cellular composition and (ii) optically movable and easily detectable markers. The satellites were synthesized by the layer-by-layer assisted decoration of silica microparticles with metal (gold or silver) nanoparticles and astralen in order to ensure satellite SERS-based microenvironment probing and satellite recognition, respectively. A combination of optical tweezers and Raman spectroscopy can be used to navigate the satellites to a certain cellular compartment and probe the intracellular composition following cellular uptake. In the future, this developed approach may serve as a tool for single cell analysis with nanometer precision due to the multilayer surface design, focusing on both extracellular and intracellular studies.

    ID:1432
  4. Akasov R., Borodina T., Zaytseva E., Sumina A., Bukreeva T.V., Burov S., Markvicheva E. (2015). Ultrasonically Assisted Polysaccharide Microcontainers for Delivery of Lipophilic Antitumor Drugs: Preparation and in vitro Evaluation. ACS Appl Mater Interfaces 7 (30), 16581–9 [+]

    High toxicity, poor selectivity, severe side effects are major drawbacks of anticancer drugs. Various drug delivery systems could be proposed to overcome these limitations. The aim of the study was to fabricate polysaccharide microcontainers (MC) loaded with thymoquinone (TQ) by one-step ultrasonication technique and to study their cellular uptake and cytotoxicity in vitro. Two MC fractions with a mean size of 500 nm (MC-0.5) and 2 µM (MC-2) were prepared and characterized. Uptake of the MC by mouse melanoma M-3 cells was evaluated in both 2D (monolayer culture) and 3D (multicellular tumor spheroids) models by confocal microscopy, flow cytometry and fluorimetry. The higher cytotoxicity of the TQ-MC-0.5 sample than that of the TQ-MC-2 fraction was in a good correlation with higher MC-0.5 accumulation in the cells. The MC-0.5 beads were more promising than the MC-2 particles because of a higher cellular uptake in both 2D and 3D models, an enhanced antitumor effect and a lower non-specific toxicity.

    ID:1304
  5. Privalova A., Markvicheva E., Sevrin C.h., Drozdova M., Kottgen C., Gilbert B., Ortiz M., Grandfils C.h. (2015). Biodegradable polyester-based microcarriers with modified surface tailored for tissue engineering. J Biomed Mater Res A 103 (3), 939–48 [+]

    Microcarriers have been proposed in tissue engineering, namely for bone, cartilage, skin, vascular, and central nervous system. Although polyester-based microcarriers have been already used for this purpose, their surface properties should be improved to provide better cell growth. The goal of this study was to prepare microbeads based on poly(D,L-lactide) acid, poly(L-lactide) acid, and to study cell behavior (adhesion, spreading, growth, and proliferation) in function of microbead topography and surface chemistry. To improve L-929 fibroblasts adhesion, microbead surface has been modified with three polycations: chitosan, poly(2-dimethylamino ethylmethacrylate) (PDMAEMA), or chitosan-g-oligolactide copolymer (chit-g-OLA). Although modification of the microbead surface with chitosan and PDMAEMA was performed through physical adsorption on the previously prepared microbeads, chit-g-OLA copolymer was introduced directly during microbead processing. This simple approach (1) bypass the use of an emulsifier (polyvinyl alcohol, PVA); (2) avoid surface "contamination" with PVA molecules limiting a control of the surface characteristics. In vitro study of the growth of mouse fibroblasts on the microbeads showed that both surface topography and chemistry affected cell attachment, spreading, and proliferation. Cultivation of L-929 fibroblasts for 7 days resulted in the formation of a 3D cell-scaffold network.

    ID:1433
  6. Privalova A.M., Uglanova S.V., Kuznetsova N.R., Klyachko N.L., Golovin Yu.I., Korenkov V.V., Vodovozova E.L., Markvicheva E.A. (2015). Microencapsulated Multicellular Tumor Spheroids as a Tool to Test Novel Anticancer Nanosized Drug Delivery Systems In Vitro. J. Nanosci. Nanotechnol. 15 (7), 4806–4814 [+]

    In the study, MCF-7 human breast adenocarcinoma cells were used to study cytotoxicity of novel anticancer nanosized formulations, such as docetaxel-loaded nanoemulsion and liposomal formulation of a lipophilic methotrexate (MTX) prodrug. In Vitro study of cytotoxicity was carried out in 2 models, namely using 3D In Vitro model based on multicellular tumor spheroids (MTS) and 2D monolayer culture. MTS were generated by tumor cell cultivation within alginate-oligochitosanmicro-capsules. In the case of the monolayer culture, cell viability was found to be 25, 18 and 12% for the samples containing nanoemulsion at concentrations 20, 300 and 1000 nM of docetaxel, respectively, after 48 hs incubation. For MTS these values were higher, namely 33, 23 and 18%, respectively. Cytotoxicity of liposomal MTX prodrug-based formulation with final concentration of 1, 2, 10, 50, 100 and 1000 nM in both models was also studied. MTX liposomal formulation demonstrated lower cytotoxicity on MTS compared to intact MTX. Moreover, MTS were also more resistant to both liposomal formulation and intact MTX than the monolayer culture. Thus, at 1000 nM MTX in the liposomal form, cell viability in MTS was 1.4-fold higher than that in the monolayer culture. MTS could be proposed as a promising tool to test novel anticancer nanosized formulations In Vitro.

    ID:1142
  7. Марквичева Е.А., Дроздова М.Г., Акасов Р.А., ЗайцеваЗотова Д.С. (2011). Биосовместимые материалы в тканевой инженерии, В кн: Клеточные технологии для регенеративной медицины / под ред.: Г.П.Пинаева, М.С.Богдановой, А.М.Кольцовой. – СПБ.: Изд-во Политехн.ун-та. , 103–126 ID:735
  8. Балабашин Д., ЗайцеваЗотова Д., Топорова В., Панина А., Марквичева Е., Свирщевская Е., Алиев Т. (2011). Способы увеличения продукции рекомбинантных антител в клеточных линиях CHO DG44. Современные проблемы науки и образования  (5), [+]

    The cell line CHO DG44 producing recombinant antibodies(Abs) to human tumor necrosis factor-alpha has been obtained. The influence of cell inoculation density and cultivation protocols on the level of Ab biosynthesis has been studied. The highest Ab yields have been observed at the inoculation density 3×106 cells/ml. The alternative method to cells-in-suspension cultivation has been proposed, which is the cell cultivation in calcium alginate hydrogel microgranules or alginate chitosan semipermeable microcapsules. It has been shown that the Ab production level by CHO DG44 cells entrapped into polymer microcapsules exceeds that of the cells-in-suspension cultivation regime.

    ID:540
  9. ZaytsevaZotova D., Balysheva V., Tsoy A., Drozdova M., Akopova T., Vladimirov L., Chevalot I., Marc A., Goergen J.L., Markvicheva E. (2011). Biocompatible Smart Microcapsules Based on Chitosan‐Poly (vinyl alcohol) Copolymers for Cultivation of Animal Cells. Advanced Biomaterials , [+]

    In this study, two novel chitosan-graft-poly(vinyl alcohol) copolymers are synthesized and used as water-soluble at physiological conditions polycations for preparation of smart microcapsules. The microcapsules provide growth and proliferation of eight mammalian cell lines, including hybridoma and tumor cells, at long-term cell cultivation in vitro. The microcapsules are stable in cell culture medium but can be dissolved by changing pH value of the medium (up to 8.0–8.2), thus making possible a simple release of the entrapped cells. Monoclonal antibody production by encapsulated hybridoma cells is demonstrated. Cultivation of tumor cells within the microcapsules allows the formation of 3D multicellular spheroids, which can be proposed as an in vitro model for anticancer drug screening.

    ID:454
  10. Borodina T., Grigoriev D., Markvicheva E., Mohwald H., Shchukin D. (2011). Vitamin E Microspheres Embedded Within a Biocompatible Film for Planar Delivery. Advanced Engineering Materials 13 (3), B123–B130 [+]

    We demonstrate a new one-batch approach to the fabrication of a biocompatible Ca-alginate film with embedded vitamin E-loaded microspheres that could be used for planar dermal drug delivery. Stable vitamin E microspheres, coated with gum acacia, are produced by ultrasonic treatment of a two-phase liquid system. The Fourier transform infrared spectroscopy indicates an interaction between biopolymer functional groups induced by ultrasonication. Confocal laser fluorescence microscopy, scanning electron microscopy (SEM) and atomic force microscopy (AFM) demonstrate a homogeneous microsphere distribution within the Ca-alginate polymer film. The kinetics of in vitro vitamin E release found for the polymer film with entrapped microspheres was much more sustained (100% in 96 h) compared to the polymer film with vitamin E embedded in the free state (100% in 5 h). The novelty of the proposed research involves the ultrasonic fabrication of loaded microspheres and formation of biodegradable coating directly doped with microspheres.

    ID:428
  11. Tsoy A., ZaytsevaZotova D., Edelweiss E., Bartkowiak A., Goergen J.L., Vodovozova E., Markvicheva E. (2010). Microencapsulated multicellular tumor spheroids as a novel in vitro model for drug screening. Biochemistry (Moscow) Supplement Series B: Biomedical Chemistry 4 (3), 243–250 [+]

    В работе использован метод микрокапсулирования  клеток аденокарциномы молочной железы человека MCF-7 в биосовместимые альгинат-хитозановые микрокапсулы с целью генерирования на основе этих клеток  мультиклеточных опухолевых сфероидов (МОС)  и их дальнейшего исследования в качестве модели in vitro для тестирования противораковых лекарственных средств. На МОС, полученных на основе клеточной линии MCF-7 (аденокарцинома молочной железы человека), было исследовано цитотоксическое действие метотрексата. В зависимости от времени культивирования клеток в микрокапсулах были  получены МОС со средним размером 150, 200 и 300 мкм. После инкубирования МОС с метотрексатом различных концентраций (1, 2, 10, 50 и 100 нМ) в течение 48 часов оценивали количество жизнеспособных клеток. Показано, что МОС гораздо устойчивее к метотрексату, чем монослойная культура. Так, при концентрации метотрексата 100 нМ в МОС размером 300 мкм доля жизнеспособных клеток в 2,5 раза превышала количество живых клеток в монослойной культуре. Таким образом, было показано, что микрокапсулированные  МОС могут более адекватно отражать состояние клеток  в малых солидных опухолях, чем монослойная культура,  и могут в дальнейшем быть предложены в качестве новой модели in vitro для тестирования противораковых лекарств.

    ID:372
  12. Бовин Н.В., Марквичева Е.А., Селина О.Е. (2009). Сорбент для удаления антител из цельной крови и способ его получения. Патент RU 2360707. , ID:400
  13. Borodina T.N., Rumsh L.D., Kunizhev S.M., Sukhorukov G.B., Vorozhtsov G.N., Feldman B.M., Rusanova A.V., Vasileva T.V., Strukova S.M., Markvicheva E.A. (2008). Entrapment of herbal extracts into biodegradable microcapsules. Biochemistry (Moscow) Supplemental Series B: Biomedical Chemistry 2 (2), 176–182 [+]

    The microcapsules with entrapped herbal water-soluble extracts of plantain Plantago major and calendula Calendula officinalis L. (PCE) were prepared by layer-by-layer (LbL)-adsorption of carrageenan and  oligochitosan onto CaCO3 microparticles with their subsequent dissolving after the treatment of EDTA. Entrapment of PCE was performed by using adsorption and co-precipitation techniques. The co-precipitation provided better entrapment of PCE into the carbonate matrix compared to adsorption. In vitro release kinetics (AGJ) was studied using artificial gastric juice. Using the model of acetate ulcer in rats it has been demonstrated that PCE released from the microcapsules accelerates gastric tissue repair.

    ID:384
  14. Markvicheva E., Stashevskaya K., Strukova S., Prudchenko I., Rusanova A., Makarova A., Vasilieva T., Bespalova J., Grandfils C.h. (2006). Biodegradable microparticles loaded with thrombin receptor agonist peptide for gastric ulcer treatment in rats. J. Drug Del. Sci. Tech 16 (4), 321–325 [+]

     

     

    The aim of the current paper was to elaborate an immobilization method of thrombin receptor agonist peptide (TRAP-6) in biodegradable biocompatible poly(d,l)-lactide-co-glycolide (PLGA) microparticles and to demonstrate the effect of the entrapped peptide for tissue repair, namely for a gastric ulcer treatment in rats. TRAP-6 was entrapped in polymer using w/o/w double emulsion-evaporation technique. The morphology of empty and TRAP-6 loaded microparticles was evaluated by light and scanning electron microscopy (SEM). In vitro release kinetics profile of TRAP-6 from microparticles was studied by HPLC. To investigate gastric mucosal protection effect in vivo, TRAP-6-loaded microparticles were administered in a rat stomach after a previous mucosal injury (a gastric ulcer). Microparticles with entrapped TRAP-6 were found to reduce both an inflammation and proliferation phases of wound healing, and thus accelerated tissue repair in rats.

     

    ID:402

Марквичева Елена Арнольдовна

  • Москва, ул. Миклухо-Маклая, 16/10 — На карте
  • ИБХ РАН, корп. 34, комн. 422
  • Тел.: +7(422)336-06-00
  • Эл. почта: lemark@ibch.ru