Towards biobanking technologies for natural and bioengineered multicellular placental constructs

Pogozhykh, Denys; Prokopіuk, Volodymyr; Prokopyuk, O. S.; Kuleshova, Larisa; Goltsev, Anatoliy; Figueiredo, Constança

doi:10.1016/j.biomaterials.2018.08.060

Cited by 23 publications

(20 citation statements)

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“…Cryopreservation is the use of very low temperature to preserve living cells and tissues in a quiescent status for a long period, without losing their viability, activity, and function [14]. Several works have been devoted to the cryopreservation of engineered biological substitutes [15][16][17][18][19][20][21][22][23][24][25], but there is still the need for information on the cryopreservation of biofabricated osteoblast constructs, a paramount goal addressed in this work. Pioneer studies demonstrate that post-thawing osteoblast viability was better maintained (40-50%) when cryopreserved in dimethyl sulfoxide (DMSO) than in other cryoprotectant additives [26,27].…”

Section: Introductionmentioning

confidence: 99%

Effects of Cryopreservation on Cell Metabolic Activity and Function of Biofabricated Structures Laden with Osteoblasts

et al. 2020

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Biofabrication and maturation of bone constructs is a long-term task that requires a high degree of specialization. This specialization falls onto the hierarchy complexity of the bone tissue that limits the transfer of this technology to the clinic. This work studied the effects of the short-term cryopreservation on biofabricated osteoblast-containing structures, with the final aim to make them steadily available in biobanks. The biological responses studied include the osteoblast post-thawing metabolic activity and the recovery of the osteoblastic function of 3D-bioprinted osteoblastic structures and beta tricalcium phosphate (β-TCP) scaffolds infiltrated with osteoblasts encapsulated in a hydrogel. The obtained structures were cryopreserved at −80 °C for 7 days using dimethyl sulfoxide (DMSO) as cryoprotectant additive. After thawing the structures were cultured up to 14 days. The results revealed fundamental biological aspects for the successful cryopreservation of osteoblast constructs. In summary, immature osteoblasts take longer to recover than mature osteoblasts. The pre-cryopreservation culture period had an important effect on the metabolic activity and function maintain, faster recovering normal values when cryopreserved after longer-term culture (7 days). The use of β-TCP scaffolds further improved the osteoblast survival after cryopreservation, resulting in similar levels of alkaline phosphatase activity in comparison with the non-preserved structures. These results contribute to the understanding of the biology of cryopreserved osteoblast constructs, approaching biofabrication to the clinical practice.

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Section: Introductionmentioning

confidence: 99%

Effects of Cryopreservation on Cell Metabolic Activity and Function of Biofabricated Structures Laden with Osteoblasts

et al. 2020

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“…Previously, we developed efficient biobanking technologies for mesenchymal stem cells, blood cells, as well as tissues and tissue engineered constructs [ 26 , 32 , 35 , 63 , 64 ]. Results of these studies show substantial differences in cryopreservation outcomes and necessity to optimize the process for each particular object.…”

Section: Discussionmentioning

confidence: 99%

Towards Reduction or Substitution of Cytotoxic DMSO in Biobanking of Functional Bioengineered Megakaryocytes

Pogozhykh

Eicke

Gryshkov

et al. 2020

IJMS

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Donor platelet transfusion is currently the only efficient treatment of life-threatening thrombocytopenia, but it is highly challenged by immunological, quality, and contamination issues, as well as short shelf life of the donor material. Ex vivo produced megakaryocytes and platelets represent a promising alternative strategy to the conventional platelet transfusion. However, practical implementation of such strategy demands availability of reliable biobanking techniques, which would permit eliminating continuous cell culture maintenance, ensure time for quality testing, enable stock management and logistics, as well as availability in a ready-to-use manner. At the same time, protocols applying DMSO-based cryopreservation media were associated with increased risks of adverse long-term side effects after patient use. Here, we show the possibility to develop cryopreservation techniques for iPSC-derived megakaryocytes under defined xeno-free conditions with significant reduction or complete elimination of DMSO. Comprehensive phenotypic and functional in vitro characterization of megakaryocytes has been performed before and after cryopreservation. Megakaryocytes cryopreserved DMSO-free, or using low DMSO concentrations, showed the capability to produce platelets in vivo after transfusion in a mouse model. These findings propose biobanking approaches essential for development of megakaryocyte-based replacement and regenerative therapies.

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“…An optical microscopy study with phasecontrast methods, neutral red, trypan blue vital staining, confocal laser scanning microscopy with fluorescein diacetate and propidium iodide (FDA/PI) staining according to previously described protocols [6,8] were done.…”

Section: Methodsmentioning

confidence: 99%

“…For effective clinical application, it is important to characterize the properties of spheroids after generation and short-term storage, which is necessary for its transportation from the biotechnological laboratory to the clinic [5]. In our previous studies, we have proved inappropriate of cryopreservation for spheroids storage [6]. In our view, subnormothermic (20 С) and hypothermic (4 С) regimes are promising for the spheroids short-term storage.…”

Section: Introductionmentioning

confidence: 93%

“…Для потреб медицини оптимально застосовувати субнормотермічне (20 С) зберігання сфероїдів МСК плаценти тривалістю до 24 год, подальше зберігання призводить до різкого зниження їх метаболічної активності та розпаду на окремі клітини. 6 МСК плаценты клеток получено около 825,3 ± 56,2 сфероидов размером 50-150 мкм и 62,3 ± 5,1 сфероидов размером 150-300 мкм, которые имеют сферическую форму, не окрашиваются трипановым синим, окрашиваются нейтральным красным и флюоресцеин диацетатом. Полученные сфероиды способны к адгезии к пластику и разрушению при кратковременном центрифугировании.…”

Section: характеристика сфероїдів із мезенхімальних стовбурових клітиunclassified

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Characterization of Placental Mesenchymal Stem Cells Spheroids after Generation, Hypothermic and Subnormothermic Storage

Prokopіuk¹,

Globa²,

Prokopiuk³

et al. 2019

Innov Biosyst Bioeng

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Background. Placental mesenchymal stem cells application is a promising area of cell therapy for number diseases. Their using in the form of spheroids results in enhancement of the therapeutic potential, improve the cells survival after transplantation. Medical purposes require the characterization and development of spheroids storage technology. Objective. The aim of the paper is to characterize placental mesenchymal stem cells spheroids after generation and low-temperature storage. Methods. Spheroids were produced by a hanging drop method from placental mesenchymal stem cells, characterized after production and after subnormothermic (20 C) and hypothermic (4 C) storage for up to 96 h. The structure, metabolic activity of spheroids according to the data of the MTT test and the resazurin restoration test were studied. Results. The hanging drop method makes it possible to produce from 110 6 placental mesenchymal stem cells about 825.3 ± 56.2 spheroids in the size of 50-150 microns and 62.3 ± 5.1 spheroids of 150-300 microns in size, having a spherical shape, not stained with trypan blue, stained with neutral red and fluorescein diacetate. The spheroids adhere to the plastic and destroying through short-time centrifugation. During subnormothermic storage spheroids, structural characteristics and metabolic activity change after 48 h, during hypothermicafter 24 h of storage. Conclusions. The hanging drop method makes it possible to produce about 900 spheroids from 1 million placental MSCs with stable metabolic activity. For medical purposes, subornorthomeric (20 C) storage not more than 24 h is recommended for placental MSC spheroid. Longer storage leads to a sharp decreasing of their metabolic activity and destroying to the separated cells.

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Towards biobanking technologies for natural and bioengineered multicellular placental constructs

Cited by 23 publications

References 50 publications

Effects of Cryopreservation on Cell Metabolic Activity and Function of Biofabricated Structures Laden with Osteoblasts

Effects of Cryopreservation on Cell Metabolic Activity and Function of Biofabricated Structures Laden with Osteoblasts

Towards Reduction or Substitution of Cytotoxic DMSO in Biobanking of Functional Bioengineered Megakaryocytes

Characterization of Placental Mesenchymal Stem Cells Spheroids after Generation, Hypothermic and Subnormothermic Storage

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