Multilineage differentiation of human mesenchymal stem cells in a three-dimensional nanofibrous scaffold
ABSTRACT Functional engineering of musculoskeletal tissues generally involves the use of differentiated or progenitor cells seeded with specific growth factors in biomaterial scaffolds. Ideally, the scaffold should be a functional and structural biomimetic of the native extracellular matrix and support multiple tissue morphogenesis. We have previously shown that electrospun, three-dimensional nanofibrous scaffolds that morphologically resemble collagen fibrils are capable of promoting favorable biological responses from seeded cells, indicative of their potential application for tissue engineering. In this study, we tested a three-dimensional nanofibrous scaffold fabricated from poly(epsilon-caprolactone) (PCL) for its ability to support and maintain multilineage differentiation of bone marrow-derived human mesenchymal stem cells (hMSCs) in vitro. hMSCs were seeded onto pre-fabricated nanofibrous scaffolds, and were induced to differentiate along adipogenic, chondrogenic, or osteogenic lineages by culturing in specific differentiation media. Histological and scanning electron microscopy observations, gene expression analysis, and immunohistochemical detection of lineage-specific marker molecules confirmed the formation of three-dimensional constructs containing cells differentiated into the specified cell types. These results suggest that the PCL-based nanofibrous scaffold is a promising candidate scaffold for cell-based, multiphasic tissue engineering.
- SourceAvailable from: Kazuhiko Ishihara
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- "During the cell fate determination process, stem cells sense and react to physical properties of their microenvironment. Accordingly , some cell fates are reached only in three-dimensional (3-D) cell cultures  . To date, in spite of extensive research efforts to control stem cell differentiation, the efficiency achieved in lineagerestricted differentiation is often poor. "
ABSTRACT: A large number of lineage-committed progenitor cells are required for advanced regenerative medicine based on cell engineering. Due to their ability to differentiate into multiple cells lines, multipotent stem cells have emerged as a vital source for generating transplantable cells for use in regenerative medicine. Increment in differentiation efficiency of the mesenchymal stem cell was obtained by using hydrogel to adjust the proliferation cycle of encapsulated cells to signal sensitive phase. Three dimensional (3-D) polymer networks composed of poly(2-methacyloyloxyethyl phosphorylcholine (MPC)-co-n-butyl methacrylate (BMA)-co-p-vinylphenylboronic acid (VPBA)) (PMBV) and poly(vinyl alcohol) (PVA) were prepared as a hydrogel. The proliferation of cells encapsulated in the PMBV/PVA hydrogel was highly sensitive to the storage modulus (G') of the hydrogel. That is, when the G' value of the hydrogel was higher than 1.0 kPa, the cell proliferation was ceased and the proliferation cycle of cells was converged to G1 phase, whereas when the G' value was below 1.0 kPa, cell proliferation proceeded. By changing the G' value of hydrogels under encapsulation the cells, proliferation cycle of encapsulated mesenchymal stem cells was regulated to G1 phase and thus signal sensitivity were increased. 3-D polymer networks as hydrogels with tunable physical properties can be effectively used to control proliferation and lineage-restricted differentiation of stem cells. Copyright © 2015 Elsevier Ltd. All rights reserved.Biomaterials 07/2015; 56. DOI:10.1016/j.biomaterials.2015.03.051 · 8.56 Impact Factor
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- "MSCs are a heterogeneous population of plastic-adherent, fibroblast-like cells, from which the progenitor cells in culture are able to self-renew and differentiate into multiple lineages  . Recent studies showed that combining human MSCs and biomaterials with controlled properties, or by adding certain growth factors, differentiation towards chondrogenic  , osteogenic  , myogenic , adipogenic   endothelial , and neurogenic  lineage can be achieved. Distribution and adherence of cells in scaffolds play a crucial role in the efficiency of tissue engineering approaches. "
ABSTRACT: In regenerative medicine studies, cell seeding efficiency is not only optimized by changing the chemistry of the biomaterials used as cell culture substrates, but also by altering scaffold geometry, culture and seeding conditions. In this study, the importance of seeding parameters, such as initial cell number, seeding volume, seeding concentration and seeding condition is shown. Human mesenchymal stem cells (hMSCs) were seeded into cylindrically shaped 4 × 3 mm polymeric scaffolds, fabricated by fused deposition modelling. The initial cell number ranged from 5 × 104 to 8 × 105 cells, in volumes varying from 50 µl to 400 µl. To study the effect of seeding conditions, a dynamic system, by means of an agitation plate, was compared with static culture for both scaffolds placed in a well plate or in a confined agarose moulded well. Cell seeding efficiency decreased when seeded with high initial cell numbers, whereas 2 × 105 cells seemed to be an optimal initial cell number in the scaffolds used here. The influence of seeding volume was shown to be dependent on the initial cell number used. By optimizing seeding parameters for each specific culture system, a more efficient use of donor cells can be achieved. Copyright © 2013 John Wiley & Sons, Ltd.Journal of Tissue Engineering and Regenerative Medicine 11/2013; DOI:10.1002/term.1842 · 5.20 Impact Factor
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- "In this study, the scaffolds used have both a polymer phase and a ceramic phase. As mentioned earlier, PLGA scaffolds release acidic degradation products, which may cause inflammatory responses in vivo (Li et al., 2005). The lactic and glycolic acidosis created within the engineered tissue may promote cell hypertrophy. "
ABSTRACT: It is now widely acknowledged that implants that have been designed with an effort towards reconstructing the transition between tissues might improve their functionality and integration in vivo. This paper contributes to the development of improved treatment for articular cartilage repair by exploring the potential of the combination of electrospinning technology and cell sheet engineering to create cartilage tissue. Poly(lactic-co-glycolic acid) (PLGA) was used to create the electrospun membranes. The focus being on the cartilage-bone transition, collagen type I and hydroxyapatite (HA) were also added to the scaffolds to increase the histological biocompatibility. Human mesenchymal stem cells (hMSCs) were cultured in thermoresponsive dishes to allow non-enzymatic removal of an intact cell layer after reaching confluence. The tissue constructs were created by layering electrospun membranes with sheets of hMSCs and were cultured under chondrogenic conditions for up to 21 days. High viability was found to be maintained in the multilayered construct. Under chondrogenic conditions, reverse-transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry have shown high expression levels of collagen type X, a form of collagen typically found in the calcified zone of articular cartilage, suggesting an induction of chondrocyte hypertrophy in the PLGA-based scaffolds. To conclude, this paper suggests that layering electrospun scaffolds and cell sheets is an efficient approach for the engineering of tissue transitions, and in particular the cartilage-bone transition. The use of PLGA-based scaffold might be particularly useful for the bone-cartilage reconstruction, since the differentiated tissue constructs seem to show characteristics of calcified cartilage. Copyright © 2013 John Wiley & Sons, Ltd.Journal of Tissue Engineering and Regenerative Medicine 06/2013; DOI:10.1002/term.1765 · 5.20 Impact Factor