Effect of ‘in air’ freezing on post-thaw recovery of Callithrix jacchus mesenchymal stromal cells and properties of 3D collagen-hydroxyapatite scaffolds

Mutsenko, Vitalii; Knaack, Sven; Lauterboeck, Lothar; Tarusin, Dmitriy N.; Sydykov, Bulat; Cabiscol, Ramon; Ivnev, Dmitrii; Belikan, Jan; Beck, A.; Dipresa, Daniele; Lode, Anja; Khassawna, Thaqif El; Kampschulte, Marian; Scharf, R.; Petrenko, Alexander; Korossis, Sotirios; Wolkers, Willem F.; Gelinsky, Michael; Glasmacher, Birgit; Gryshkov, Oleksandr

doi:10.1016/j.cryobiol.2020.01.015

Cited by 15 publications

(14 citation statements)

References 119 publications

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“…This temporal loss of metabolic activity could be explained by the loss of cells from the scaffold granule surface, as was shown by Calcein and DAPI staining of cells one-hour post-thaw (Figure 2C, Supplementary Figure S4). These findings are in accordance with other studies, showing that Callithrix jacchus MSCs and cells of human osteoblastic cell lines sustain their activity after being frozen on hydroxyapatite or β-TCP scaffolds, respectively [18,19]. In contrast to both these studies, we employed the simplest cell culture and cryopreservation procedures in order to develop protocols which can be later easily translated to clinical application.…”

Section: Discussionsupporting

confidence: 90%

“…Cryopreservation is the use of very low temperature to maintain living cells and tissues in a quiescent status for a long period, without losing their structure and function [21]. Several works have been devoted to the cryopreservation of engineered biological constructs [18,19,[22][23][24], which could guide our experimentation on the cryopreservation of biofabricated BTE constructs.…”

Section: Discussionmentioning

confidence: 99%

“…A recent pilot study of Hernandez-Tapia showed that osteoblasts have good survival rate after cryopreservation on β-TCP scaffold [18]. The equally recent study of Mutsenko et al [19] showed that MSCs could be frozen in 3D collagen-hydroxyapatite scaffold without significant loss of cell viability. Although these studies use technically challenging methodology, they represent invaluable first steps towards the creation of a tissue-engineered constructs biobanks.…”

Section: Introductionmentioning

confidence: 99%

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A New Perspective for Bone Tissue Engineering: Human Mesenchymal Stromal Cells Well-Survive Cryopreservation on β-TCP Scaffold and Show Increased Ability for Osteogenic Differentiation

Leppik

Gempp

Kuçi

et al. 2022

IJMS

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The clinical breakthrough of bone tissue engineering (BTE) depends on the ability to provide patients routinely with BTE products of consistent pharmacological quality. The bottleneck of this approach is the availability of stem cells. To avoid this, we suggest immobilization of random-donor-derived heterologous osteoinductive MSCs onto osteoconductive matrices. Such BTE products could then be frozen and, after thawing, could be released as ready-to-use products for permanent implantation during surgery. For this purpose, we developed a simple protocol for cryopreservation of BTE constructs and evaluated the effects of this procedure on human MSC (hMSCs) metabolic and osteogenic activity in vitro. Our findings show that hMSCs can be freeze-thawed on a β-TCP scaffold through a technically simple procedure. Treated cells sustained their metabolic activity and showed favorable osteogenic potential. Mechanistically, HIF1α and YBX1 genes were activated after freeze-thawing, and supposed to be linked to enhanced osteogenesis. However, the detailed mechanisms as to how the cryopreservation procedure beneficially affects the osteogenic potential of hMSCs remains to be evaluated. Additionally, we demonstrated that our BTE products could be stored for 3 days on dry ice; this could facilitate the supply chain management of cryopreserved BTE constructs from the site of manufacture to the operating room.

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Section: Discussionsupporting

confidence: 90%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A New Perspective for Bone Tissue Engineering: Human Mesenchymal Stromal Cells Well-Survive Cryopreservation on β-TCP Scaffold and Show Increased Ability for Osteogenic Differentiation

Leppik

Gempp

Kuçi

et al. 2022

IJMS

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“…For the validation of cryopreservation of cell-laden core-shell capsules, a CPA cocktail containing 10% DMSO and 0.3 M sucrose with the pretreatment step was selected, based on the above presented results. The latter step was dictated by the need to provide intracellular cryoprotection of the cells and thus the beneficial effect of sucrose-pretreatment as we have previously shown [51,52].…”

Section: Effect Of the Cryopreservation On Cell-encapsulated Core-shementioning

confidence: 99%

Coaxial Alginate Hydrogels: From Self-Assembled 3D Cellular Constructs to Long-Term Storage

Gryshkov

Mutsenko

Tarusin

et al. 2021

IJMS

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Alginate as a versatile naturally occurring biomaterial has found widespread use in the biomedical field due to its unique features such as biocompatibility and biodegradability. The ability of its semipermeable hydrogels to provide a favourable microenvironment for clinically relevant cells made alginate encapsulation a leading technology for immunoisolation, 3D culture, cryopreservation as well as cell and drug delivery. The aim of this work is the evaluation of structural properties and swelling behaviour of the core-shell capsules for the encapsulation of multipotent stromal cells (MSCs), their 3D culture and cryopreservation using slow freezing. The cells were encapsulated in core-shell capsules using coaxial electrospraying, cultured for 35 days and cryopreserved. Cell viability, metabolic activity and cell–cell interactions were analysed. Cryopreservation of MSCs-laden core-shell capsules was performed according to parameters pre-selected on cell-free capsules. The results suggest that core-shell capsules produced from the low viscosity high-G alginate are superior to high-M ones in terms of stability during in vitro culture, as well as to solid beads in terms of promoting formation of viable self-assembled cellular structures and maintenance of MSCs functionality on a long-term basis. The application of 0.3 M sucrose demonstrated a beneficial effect on the integrity of capsules and viability of formed 3D cell assemblies, as compared to 10% dimethyl sulfoxide (DMSO) alone. The proposed workflow from the preparation of core-shell capsules with self-assembled cellular structures to the cryopreservation appears to be a promising strategy for their off-the-shelf availability.

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“…SEM was used to observe the ultrastructural morphology of the hAM, according to the previously established protocol with modifications [64]. For these purposes, the non-frozen and frozen hAM (1× and 2× freeze-thaw cycles) were first washed two times with a 0.1 M cacodylate buffer (CAC, pH 7.4, Carl Roth, Karlsruhe, Germany) for 10 min and fixed with 2.5% glutaraldehyde (Carl Roth) in 0.1 M CAC for 90 min.…”

Section: Scanning Electron Microscopymentioning

confidence: 99%

Repeated Freezing Procedures Preserve Structural and Functional Properties of Amniotic Membrane for Application in Ophthalmology

Pogozhykh

Hofmann

Gryshkov

et al. 2020

IJMS

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For decades, the unique regenerative properties of the human amniotic membrane (hAM) have been successfully utilized in ophthalmology. As a directly applied biomaterial, the hAM should be available in a ready to use manner in clinical settings. However, an extended period of time is obligatory for performing quality and safety tests. Hence, the low temperature storage of the hAM is a virtually inevitable step in the chain from donor retrieval to patient application. At the same time, the impact of subzero temperatures carries an increased risk of irreversible alterations of the structure and composition of biological objects. In the present study, we performed a comprehensive analysis of the hAM as a medicinal product; this is intended for a novel strategy of application in ophthalmology requiring a GMP production protocol including double freezing–thawing cycles. We compared clinically relevant parameters, such as levels of growth factors and extracellular matrix proteins content, morphology, ultrastructure and mechanical properties, before and after one and two freezing cycles. It was found that epidermal growth factor (EGF), transforming growth factor beta 1 (TGF-β1), hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), hyaluronic acid, and laminin could be detected in all studied conditions without significant differences. Additionally, histological and ultrastructure analysis, as well as transparency and mechanical tests, demonstrated that properties of the hAM required to support therapeutic efficacy in ophthalmology are not impaired by dual freezing.

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Effect of ‘in air’ freezing on post-thaw recovery of Callithrix jacchus mesenchymal stromal cells and properties of 3D collagen-hydroxyapatite scaffolds

Cited by 15 publications

References 119 publications

A New Perspective for Bone Tissue Engineering: Human Mesenchymal Stromal Cells Well-Survive Cryopreservation on β-TCP Scaffold and Show Increased Ability for Osteogenic Differentiation

A New Perspective for Bone Tissue Engineering: Human Mesenchymal Stromal Cells Well-Survive Cryopreservation on β-TCP Scaffold and Show Increased Ability for Osteogenic Differentiation

Coaxial Alginate Hydrogels: From Self-Assembled 3D Cellular Constructs to Long-Term Storage

Repeated Freezing Procedures Preserve Structural and Functional Properties of Amniotic Membrane for Application in Ophthalmology

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