Regulation of Human Mesenchymal Stem Cell Osteogenesis by Specific Surface Density of Fibronectin: a Gradient Study (original) (raw)
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Tissue Engineering Part A, 2010
In this study, we evaluated the role of fiber size scale in the adhesion and spreading potential of human mesenchymal stem cells (hMSCs) on electrospun poly(caprolactone) (PCL) nanofibrous and microfibrous scaffolds. The effect of in vivo regulators in inducing osteogenic differentiation of hMSCs on PCL nanofibrous scaffolds was investigated using osteogenic differentiation marker gene expression and matrix mineralization. Here, we report for the first time the influence of in vivo regulators in an in vitro setting with hMSCs for bone tissue engineering on PCL nanofibrous matrices. Our results indicated that hMSCs attached and spread rapidly on nanofibrous scaffolds in comparison to microfibrous PCL. Further, hMSCs proliferated well on the nanofibrous scaffolds. The cells on the nanofibrous PCL were found to differentiate into the osteoblast lineage and subsequently mineralize upon addition of in vivo osteogenic regulators. The attachment and spreading of hMSCs were more effective on the nanofibers compared with the microfibers despite the lower protein surface coverage (total adsorbed protein per unit fiber surface area) on nanofibers.
Journal of Materials Science, 2016
We demonstrated an approach that modifies the scaffold surface with a range of molecules, simultaneously conjugated to the scaffold by a single treatment with concentrated conditioned medium (CM), inducing mesenchymal stem cells (MSC) to differentiate into osteogenic lineages. We first show that the CM from MG63 cells is capable of inducing the desired MSC differentiation over 7 days. We then analyze how the biodegradable polymer polycaprolactone (PCL) can be used as the scaffold. Using a CO 2 plasma treatment, it is possible to conjugate MG63 CM proteins onto the PCL film surface, and we show a gradual release of protein from such a modified PCL scaffold. Finally, we verified cell differentiation and marker expression of MSCs grown on the modified PCL and show that osteogenic markers, including alkaline phosphatase and Runx2 mRNA, are significantly upregulated. Immunostaining also shows a strong expression of the Runx2 protein. Our study shows that the differentiation effects of a condition medium can be preserved when its content is used to modify the surface of polymer scaffolds. This approach may be further applied for the differentiation of various cell lines, and it provided a first step toward growing MSCs on more complex scaffold shapes aimed at therapeutic uses.
Macromolecular Bioscience, 2009
Developing biomaterial scaffolds to elicit specific cell responses is important in many tissue engineering applications. We hypothesized that the chemical composition of the scaffold may be a key determinant for the effective induction of differentiation in human mesenchymal stem cells (hMSCs). In this study, electrospun nanofibers with different chemical compositions were fabricated using poly[(L-lactide)-co-(e-caprolactone)] (PLCL) and gelatin. Scanning electron microscopy (SEM) images showed a randomly arranged structure of nanofibers with diameters ranging from 400 nm to 600 nm. The incorporation of gelatin in the nanofibers stimulated the adhesion and osteogenic differentiation of hMSCs. For example, the well-stretched and polygonal morphology of hMSCs was observed on the gelatin-containing nanofibers, while the cells cultured on the PLCL nanofibers were contracted. The DNA content and alkaline phosphatase activity were significantly increased on the PLCL/gelatin blended nanofibers. Expression of osteogenic genes including alkaline phosphatase (ALP), osteocalcin (OCN), and collagen type I-a2 (Col I-a2) were also upregulated in cells cultured on nanofibers with gelatin. Mineralization of hMSCs was analyzed by von Kossa staining and the amount of calcium was significantly enhanced on the gelatin-incorporated nanofibers. These results suggest that the chemical composition of the underlying scaffolds play a key role in regulating the osteogenic differentiation of hMSCs.
Annals of Biomedical Engineering, 2005
In this study, the osteoinductive and cell-binding properties of three different resorbable polymers were evaluated by human mesenchymal stem cells (MSCs). MSCs were isolated, expanded, and cultivated onto resorbable D,D,L,L-polylactide (PLLA), collagen I/III, and polygalactin-910/polydioxanone (PGPD) scaffolds in vitro. To evaluate the influence of dexamethasone, ascorbic acid, and beta-glycerolphosphate (DAG) on osteoblast differentiation, MSCs were incubated in a DAG-enriched medium. After a 28-day period in vitro, the cellular loaded polymers were digested enzymatically by papain and HCl. The Ca(2+) content of the biomembranes was evaluated by an o-kresolphthalein-complexon reaction via photometer. A PicoGreen assay was performed for dsDNA quantification. Significant differences between the number of adherent MSCs were documented (collagen > PLLA > PGPD). Compared to the initial number of adherent cells, all biomaterials induced a significant decrease in cellular adherence after 28 days in vitro. The presence of DAG-enriched culture medium stimulated the cellular proliferation for PLLA and slightly for PGPD, whereas cell proliferation was inhibited when MSCs were cultivated onto collagen I/III. In comparison with the control groups, all biomaterials (PLLA, PGPD, and collagen I/III) showed a significant increase in local Ca(2+) accumulation under DAG stimulation after 28 days in vitro. Furthermore, collagen I/III and PLLA scaffolds showed osteoinductive properties without DAG stimulation. These results were verified by immunocytochemical stainings against osteoblast-typical markers (osteopontin and alkaline phosphatase) and completed by calcified matrix detection (von Kossa staining). MSCs were identified by CD105 and CD13 antigen expression. Corresponding to an absence of CD34, CD45, and collagen II expression, we found no chondrogenic or hematopoietic cell differentiation. The results indicate significant differences for the proliferation, differentiation, adherence, and Ca(2+) accumulation between the tested polymers in a MSC culture.
Artificial Cells, Nanomedicine, and Biotechnology
Stem cells can be obtained from a variety of sources. To compare the effect of cell source on the osteogenic differentiation potential, buccal fat pad-derived mesenchymal stem cells (BFP-MSCs), bone marrow-derived MSCs (BM-MSCs) and unrestricted somatic stem cells (USSCs) with different accessibility in time and region, were cultured on bioceramic (Bio-Oss V R) coated electrospun polycaprolactone (PCL) scaffold (PCL-Bio). After scaffold characterization, stem cells proliferation and osteogenic differentiation were investigated by MTT and Alizarin red staining, alkaline phosphatase activity, calcium content and gene expression assays. Proliferation rate of the stem cells was not significantly different with each other, only USSCs showed significantly lower proliferation rate while cultured on PCL-Bio; although, PCL-Bio showed better proliferation support in comparison with tissue culture plate and PCL. Mineralization of the BM-MSCs was significantly higher than others, while BFP-MSCs were close to it. Highest ALP activity was detected in BFP-MSCs cultured on PCL-Bio. USSCs demonstrated higher gene expression level in three genes, although differences were not huge compared to others. According to the results and due to the availability, facilitated preparation procedure and less patients suffering, BFP-MSCs have a better choice than BM-MSCs and USSCs for use in bone tissue engineering.
Tissue Engineering Part A, 2010
The design of porous scaffolds able to promote and guide cell proliferation, colonization, and biosynthesis in three dimensions is key determinant in bone tissue engineering (bTE). The aim of this study was to assess the role of the micro-architecture of poly(e-caprolactone) scaffolds in affecting human mesenchymal stem cells' (hMSCs) spatial organization, proliferation, and osteogenic differentiation in vitro. Poly(e-caprolactone) scaffolds for bTE and characterized by mono-modal and bi-modal pore size distributions were prepared by the combination of gas foaming and selective polymer extraction from co-continuous blends. The topological properties of the pore structure of the scaffolds were analyzed and the results correlated with the ability of hMSCs to proliferate, infiltrate, and differentiate in vitro in three dimensions. Results showed that the micro-architecture of the pore structure of the scaffolds plays a crucial role in defining cell seeding efficiency as well as hMSCs' threedimensional colonization, proliferation, and osteogenic differentiation. Taken all together, our results indicated that process technologies able to allow a fine-tune of the topological properties of biodegradable porous scaffolds are essential for bTE strategies.
Macromolecular Bioscience, 2010
Layer-by-layer assembly of biogenic polyelectrolytes (PEL) was carried out on the surface of poly (L-lactide) to generate polyelectrolyte multilayers (PEM) that foster osteogenic differentiation of human mesenchymal stem cell (hMSC). Gelatin (GEL), hyaluronic acid (HA) and heparin (HEP) were chosen as polyanions, while chitosan (CHI) was employed as polycation. Multilayer formation was monitored by surface plasmon resonance and water contact angle measurements showing that layer formation process and surface wetting properties depended on the type of polyanions. While HEP as strong PEL led to thicker and more hydrophilic PEM, layer mass was lower for weak polyanions GEL and HA. Short-term adhesion studies with hMSC showed strong adhesion and spreading of cells on PEM composed of GEL/ CHI and low spreading, motile phenotype and aggregation of hMSC on HEP/CHI or HA/CHI. Long term studies over three weeks were carried out to follow growth and differentiation of hMSC on the PEM. Weak osteogenic differentiation of hMSC was observed on GEL/CHI if cells were cultured in normal medium while no osteogenic phenotypes were observed on HEP/CHI or HA/CHI. When cells were cultured in osteogenic differentiation medium, however, PEM composed of HEP/CHI or HA/CHI promoted differentiation of hMSC towards osteoblasts, while PEM composed of GEL/CHI failed to do so. Overall, the composition of PEMs can be used as additional tool to control osteogenic differentiation of hMSC.
Journal of Materials Science: Materials in Medicine
Bioresorbable biomaterials can fill bone defects and act as temporary scaffold to recruit MSCs to stimulate their differentiation. Among the different bioresorbable polymers studied, this work focuses on poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL). Were prepared blends of PHBV and PCL to obtain PHBV based biomaterials with good tenacity, important for bone tissue repair, associated with biocompatible properties of PCL. This study assesses the viability of Vero cells on scaffolds of PHBV, PCL, and their blends and the osteogenic differentiation of mesenchymal stem cells (MSCs). Materials were characterized in surface morphology, DSC and Impact Strength (IS). Vero cells and MSCs were assessed by MTT assay, cytochemical and SEM analysis. MSC osteogenic differentiation was evaluated through alizarin red staining and ALP activity. We found some roughness onto surface materials. DSC showed that the blends presented two distinct melting peaks, characteris...
BioImpacts
Introduction: Biocompatible and biodegradable scaffolds have gained tremendous attention because of their potential in tissue engineering. In this study, the aim was to reach a feasible setup from a ternary hybrid of polyaniline (PANI), gelatin (GEL), and polycaprolactone (PCL) to fabricate aligned and random nanofibrous scaffolds by electrospinning for tissue engineering purposes. Methods: Different setups of PANI, PCL, and GEL were electrospun. Then, the best aligned and random scaffolds were chosen. SEM imaging was done to observe nanoscaffolds before and after stem cell differentiation. Mechanical properties of the fibers were tested. Their hydrophilicity was measured using the sessile drop method. SNL Cells were then seeded onto the fiber, and MTT was performed to assess its toxicity. The cells were then differentiated. After osteogenic differentiation, alkaline phosphatase activity, calcium content assay, and alizarin red staining were done to check the validity of osteogenic ...
PloS one, 2011
The development of a new family of implantable bioinspired materials is a focal point of bone tissue engineering. Implant surfaces that better mimic the natural bone extracellular matrix, a naturally nano-composite tissue, can stimulate stem cell differentiation towards osteogenic lineages in the absence of specific chemical treatments. Herein we describe a bioactive composite nanofibrous scaffold, composed of poly-caprolactone (PCL) and nano-sized hydroxyapatite (HA) or beta-tricalcium phosphate (TCP), which was able to support the growth of human bone marrow mesenchymal stem cells (hMSCs) and guide their osteogenic differentiation at the same time. Morphological and physical/chemical investigations were carried out by scanning, transmission electron microscopy, Fourier-transform infrared (FTIR) spectroscopy, mechanical and wettability analysis. Upon culturing hMSCs on composite nanofibers, we found that the incorporation of either HA or TCP into the PCL nanofibers did not affect c...