A 1-min Method for Homogenous Cell Seeding in Porous Scaffolds

Tan, Lijun; Ren, Yijin; Kuijer, Roel

doi:10.1177/0885328210389504

Cited by 17 publications

(20 citation statements)

References 32 publications

(61 reference statements)

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“…The tubes were then centrifuged at 400g for 5 min to force the cell suspension into the matrix [ 42 ]. A 10-ml syringe was used to create a vacuum in each tube for 10 s in order to maximize cell seeding [ 43 ]. After 1 h scaffolds were transferred to 24-well plates.…”

Section: Methodsmentioning

confidence: 99%

Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage

et al. 2016

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Articular cartilage has a limited capacity to heal itself and thus focal defects often result in the development of osteoarthritis. Current cartilage tissue engineering strategies seek to regenerate injured tissue by creating scaffolds that aim to mimic the unique structure and composition of native articular cartilage. Decellularization is a novel strategy that aims to preserve the bioactive factors and 3D biophysical environment of the native extracellular matrix while removing potentially immunogenic factors. The purpose of this study was to develop a procedure that can enable decellularization and recellularization of intact articular cartilage matrix. Full-thickness porcine articular cartilage plugs were decellularized with a series of freeze-thaw cycles and 0.1% (w/v) sodium dodecyl sulfate detergent cycles. Chondroitinase ABC (ChABC) was applied before the detergent cycles to digest glycosaminoglycans in order to enhance donor chondrocyte removal and seeded cell migration. Porcine synovium-derived mesenchymal stem cells were seeded onto the decellularized cartilage scaffolds and cultured for up to 28 days. The optimized decellularization protocol removed 94% of native DNA per sample wet weight, while collagen content and alignment were preserved. Glycosaminoglycan depletion prior to the detergent cycles increased removal of nuclear material. Seeded cells infiltrated up to 100 μm into the cartilage deep zone after 28 days in culture. ChABC treatment enhances decellularization of the relatively dense, impermeable articular cartilage by reducing glycosaminoglycan content. ChABC treatment did not appear to affect cell migration during recellularization under static, in vitro culture, highlighting the need for more dynamic seeding methods.

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Section: Methodsmentioning

confidence: 99%

Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage

et al. 2016

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“…15,16,22 Therefore, in the present study, wet electrospinning technique and ultrasonic treatment were employed to achieve highly porous PLGA/PCL scaffolds. Furthermore, nHA was incorporated into the scaffolds to enhance dental cell differentiation.…”

Section: Discussionmentioning

confidence: 99%

“…For instance, the scaffolds were rotated in cell suspensions, or slight vacuum was used to enhance cell infiltration throughout the scaffold. 18,22 In a pilot study used to test this approach, a dynamic seeding method was found to create a rotational turbulence that over time proved to be too harsh for the DE cells (data not shown). Therefore, a static seeding method was used in the current study.…”

Section: Discussionmentioning

confidence: 99%

“…Although electrospun scaffolds have the potential to resemble natural ECM, a common problem encountered with these scaffolds is an inherently low porosity, which inhibits adequate cellular infiltration. 15,16,22 Therefore, in the present study, wet electrospinning technique and ultrasonic treatment were employed to achieve highly porous PLGA/PCL scaffolds. Furthermore, nHA was incorporated into the scaffolds to enhance dental cell differentiation.…”

Section: Discussionmentioning

confidence: 99%

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Influence of highly porous electrospun PLGA/PCL/nHA fibrous scaffolds on the differentiation of tooth bud cells in vitro

Cai

Hoopen

Zhang

et al. 2017

J Biomedical Materials Res

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In this study, we investigated the use of three-dimensional electrospun poly(lactic-co-glycolic acid)/poly(ε-caprolactone) (PLGA/PCL) scaffolds seeded and cultured with postnatal dental cells, for improved dental tissue regeneration. Wet-electrospinning combined with ultrasonic treatment was studied as a method to enhance scaffold porosity and to promote cell-cell interactions. We also investigated whether nano-hydroxyapatite (nHA) incorporation could enhance dental cell differentiation. All scaffolds were seeded with human tooth pulp-derived dental mesenchymal (hDM) cells, or a combination of hDM and pig dental epithelial (pDE) cells, cultured for up to 28 days. Developmentally staged samples were assessed using scanning electron microscopy, histological, immunohistochemical, DNA and alkaline phosphatase activity assays, and quantitative-PCR for ameloblastic, odontoblastic, and osteogenic related gene expression. Results showed that electrospun scaffolds exhibited sufficient porosity to support robust cell ingrowth. Additional ultrasonic treatment led to a less homogeneous scaffold porosity, resulting in evident cell clustering and enhanced hDM-pDE cell-cell interactions. Finally, nHA incorporation was found to enhance dental cell differentiation. However, it also resulted in smaller fibre diameter and reduced scaffold porosity, and inhibited cell ingrowth and proliferation. In conclusion, ultrasonically treated wet-electrospun PLGA/PCL scaffolds are a suitable material for dental tissue engineering, and support future in vivo evaluations of this model.

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“…The passive seeding technique is based on dropping cells onto the scaffold surface followed by the cell infiltration through the scaffold microstructure. On the other hand, the active cell seeding utilises a certain external factor, which improves the penetration of cells into the interior of the substrate (rotation, centrifugation, magnetic field) and the results have demonstrated improved cell distribution and ECM formation by differentiated cells by this method [2,9,30].…”

Section: Introductionmentioning

confidence: 99%

Effects of Cell Seeding Methods on Chondrogenic Differentiation of Rat Mesenchymal Stem Cells in Polyhydroxybutyrate/Chitosan Scaffolds

et al. 2019

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The aim of our study was to examine the effects of passive and active cell seeding techniques on in vitro chondrogenic differentiation of mesenchymal stem cells (MSC) isolated from rat bone marrow and seeded on porous biopolymer scaffolds based on polyhydroxybutyrate/chitosan (PCH) blends. This paper is focused on the distribution of the cells on and in the scaffolds, since it influences the uniformity of the created extracellular matrix (ECM), as well as the homogenity of the distribution of chondrogenic markers in vitro which ultimately affects the quality of the newly created tissue after in vivo implantation. The three types of cell-scaffold constructs were examined by: fluorescence microscopy, SEM, histology and quantitative analysis of the glycosaminoglycans after chondrogenic cultivation. The results demonstrated that the active cells seeded via the centrifugation of the cell suspension onto the scaffold guaranteed an even distribution of cells on the bulk of the scaffold and the uniform secretion of the ECM products by the differentiated cells.

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A 1-min Method for Homogenous Cell Seeding in Porous Scaffolds

Cited by 17 publications

References 32 publications

Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage

Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage

Influence of highly porous electrospun PLGA/PCL/nHA fibrous scaffolds on the differentiation of tooth bud cells in vitro

Effects of Cell Seeding Methods on Chondrogenic Differentiation of Rat Mesenchymal Stem Cells in Polyhydroxybutyrate/Chitosan Scaffolds

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