Chondrogenic, hypertrophic, and osteochondral differentiation of human mesenchymal stem cells on three-dimensionally woven scaffolds.

The development of mechanically functional cartilage and bone tissue constructs of clinically relevant size, as well as their integration with native tissues, remains an important challenge for regenerative medicine. The objective of this study was to assess adult human mesenchymal stem cells (MSCs) in large, three-dimensionally woven poly(ε-caprolactone; PCL) scaffolds in proximity to viable bone, both in a nude rat subcutaneous pouch model and under simulated conditions in vitro. In Study I, various scaffold permutations-PCL alone, PCL-bone, "point-of-care" seeded MSC-PCL-bone, and chondrogenically precultured Ch-MSC-PCL-bone constructs-were implanted in a dorsal, ectopic pouch in a nude rat.

Anisotropic architecture and electrical stimulation enhance neuron cell behaviour on a tough graphene embedded PVA: alginate fibrous scaffold.

Tough scaffolds comprised of aligned and conductive fibers are promising for peripheral nerve regeneration due to their unique mechanical and electrical properties. Several studies have confirmed that electrical stimulation can control the axonal extension . However, the stimulatory effects of scaffold architecture and electrical stimulation have not yet been investigated in detail.

Comprehensive proteomic characterization of stem cell-derived extracellular matrices.

In the stem-cell niche, the extracellular matrix (ECM) serves as a structural support that additionally provides stem cells with signals that contribute to the regulation of stem-cell function, via reciprocal interactions between cells and components of the ECM. Recently, cell-derived ECMs have emerged as in vitro cell culture substrates to better recapitulate the native stem-cell microenvironment outside the body. Significant changes in cell number, morphology and function have been observed when mesenchymal stem cells (MSC) were cultured on ECM substrates as compared to standard tissue-culture polystyrene (TCPS).

Non-cell-adhesive substrates for printing of arrayed biomaterials.

Cellular microarrays have become extremely useful in expediting the investigation of large libraries of (bio)materials for both in vitro and in vivo biomedical applications. An exceedingly simple strategy is developed for the fabrication of non-cell-adhesive substrates supporting the immobilization of diverse (bio)material features, including both monomeric and polymeric adhesion molecules (e.g.

Tri-layered elastomeric scaffolds for engineering heart valve leaflets.

Tissue engineered heart valves (TEHVs) that can grow and remodel have the potential to serve as permanent replacements of the current non-viable prosthetic valves particularly for pediatric patients. A major challenge in designing functional TEHVs is to mimic both structural and anisotropic mechanical characteristics of the native valve leaflets. To establish a more biomimetic model of TEHV, we fabricated tri-layered scaffolds by combining electrospinning and microfabrication techniques.

Electrospun PGS:PCL microfibers align human valvular interstitial cells and provide tunable scaffold anisotropy.

Tissue engineered heart valves (TEHV) can be useful in the repair of congenital or acquired valvular diseases due to their potential for growth and remodeling. The development of biomimetic scaffolds is a major challenge in heart valve tissue engineering. One of the most important structural characteristics of mature heart valve leaflets is their intrinsic anisotropy, which is derived from the microstructure of aligned collagen fibers in the extracellular matrix (ECM).

Biomimetic scaffold combined with electrical stimulation and growth factor promotes tissue engineered cardiac development.

Toward developing biologically sound models for the study of heart regeneration and disease, we cultured heart cells on a biodegradable, microfabricated poly(glycerol sebacate) (PGS) scaffold designed with micro-structural features and anisotropic mechanical properties to promote cardiac-like tissue architecture. Using this biomimetic system, we studied individual and combined effects of supplemental insulin-like growth factor-1 (IGF-1) and electrical stimulation (ES). On culture day 8, all tissue constructs could be paced and expressed the cardiac protein troponin-T.

A biodegradable microvessel scaffold as a framework to enable vascular support of engineered tissues.

A biodegradable microvessel scaffold comprised of distinct parenchymal and vascular compartments separated by a permeable membrane interface was conceptualized, fabricated, cellularized, and implanted. The device was designed with perfusable microfluidic channels on the order of 100 μm to mimic small blood vessels, and high interfacial area to an adjacent parenchymal space to enable transport between the compartments. Poly(glycerol sebacate) (PGS) elastomer was used to construct the microvessel framework, and various assembly methods were evaluated to ensure robust mechanical integrity.

The significance of pore microarchitecture in a multi-layered elastomeric scaffold for contractile cardiac muscle constructs.

Multi-layered poly(glycerol-sebacate) (PGS) scaffolds with controlled pore microarchitectures were fabricated, combined with heart cells, and cultured with perfusion to engineer contractile cardiac muscle constructs. First, one-layered (1L) scaffolds with accordion-like honeycomb shaped pores and elastomeric mechanical properties were fabricated by laser microablation of PGS membranes. Second, two-layered (2L) scaffolds with fully interconnected three dimensional pore networks were fabricated by oxygen plasma treatment of 1L scaffolds followed by stacking with off-set laminae to produce a tightly bonded composite.

Combined technologies for microfabricating elastomeric cardiac tissue engineering scaffolds.

Polymer scaffolds that direct elongation and orientation of cultured cells can enable tissue engineered muscle to act as a mechanically functional unit. We combined micromolding and microablation technologies to create muscle tissue engineering scaffolds from the biodegradable elastomer poly(glycerol sebacate). These scaffolds exhibited well defined surface patterns and pores and robust elastomeric tensile mechanical properties.

Sox11 is expressed in early progenitor human multipotent stromal cells and decreases with extensive expansion of the cells.

There has been considerable interest in developing new therapies with adult multipotent progenitor stromal cells or mesenchymal stem cells (MSCs) in organ replacement and repair. To be effectively seeded into scaffolds for therapy, large numbers of cells are needed, but concerns remain regarding their chromatin stability in long-term culture. We therefore expanded four donors of human MSCs (hMSCs) from bone marrow aspirates with a protocol that maintains the cells at low density.

Intravenous hMSCs improve myocardial infarction in mice because cells embolized in lung are activated to secrete the anti-inflammatory protein TSG-6.

Quantitative assays for human DNA and mRNA were used to examine the paradox that intravenously (i.v.) infused human multipotent stromal cells (hMSCs) can enhance tissue repair without significant engraftment.

Human multipotent stromal cells undergo sharp transition from division to development in culture.

Human mesenchymal stem cells, or multipotent stromal cells (MSCs), are of interest for clinical therapy, in part because of their capacity for proliferation and differentiation. However, results from clinical trials and in vitro models have been variable, possibly because of MSC heterogeneity and a lack of standardization between MSC in vitro expansion protocols. Here we defined changes in MSCs during expansion in vitro.

Use of differentiating adult stem cells (marrow stromal cells) to identify new downstream target genes for transcription factors.

We developed a strategy for use of microarray data to rapidly identify new downstream targets of transcription factors known to drive differentiation by following the time courses of gene expression as a relatively homogeneous population of stem/progenitor cells are differentiated to multiple phenotypes. Microarray assays were used to follow the differentiation of human marrow stromal cells (MSCs) into chondrocytes or adipocytes in three different experimental conditions. The steps of the analysis were the following: (a) hierarchical clustering was used to define groups of similarly behaving genes in each experiment, (b) candidates for new downstream targets of transcription factors that drive differentiation were then identified as genes that were consistently co-expressed with known downstream target genes of the transcription factors, and (c) the list of candidate new target genes was refined by identifying genes whose signal intensities showed a highly significant linear regression with the signal intensities of the known targets in all the data sets.

Comparison of effect of BMP-2, -4, and -6 on in vitro cartilage formation of human adult stem cells from bone marrow stroma.

The human adult stem cells from bone marrow stroma referred to as mesenchymal stem cells or marrow stromal cells (MSCs) are of interest because they are easily isolated and expanded and are capable of multipotential differentiation. Here, we examined the ability of recombinant human bone morphogenetic protein (BMP)-2, -4, and -6 to enhance in vitro cartilage formation of MSCs. Human MSCs were isolated from bone marrow taken from normal adult donors.

Adipogenic differentiation of human adult stem cells from bone marrow stroma (MSCs).

Unlabelled: We assayed gene expressions during adipogenesis of human MSCs. Microarray assays demonstrated time-dependent increases in expression of 67 genes, including 2 genes for transcription factors that were not previously shown to be expressed during adipogenesis. INTRODUCTION Increased numbers of bone marrow adipocytes have been observed in patients with osteoporosis and aplastic anemia, but the pathological mechanisms remain unknown.

Adult stem cells from bone marrow (MSCs) isolated from different strains of inbred mice vary in surface epitopes, rates of proliferation, and differentiation potential.

For reasons that are not apparent, it has been difficult to isolate and expand the adult stem cells referred to as mesenchymal stem cells or marrow stromal cells (MSCs) from murine bone marrow. We developed a protocol that provides rapidly expanding MSCs from 5 strains of inbred mice. The MSCs obtained from 5 different strains of mice were similar to human and rat MSCs in that they expanded more rapidly if plated at very low density, formed single-cell-derived colonies, and readily differentiated into either adipocytes, chondrocytes, or mineralizing cells.

Expansion of human adult stem cells from bone marrow stroma: conditions that maximize the yields of early progenitors and evaluate their quality.

There is considerable interest in the biology and therapeutic potential of adult stem cells from bone marrow stroma, variously referred to as mesenchymal stem cells or marrow stromal cells (MSCs). Human MSCs can expand rapidly in culture, but the rate of expansion and the yields of multipotential progenitors are inversely related to the plating density and incubation time of each passage. We have defined conditions for optimizing the yields of cultures enriched for early progenitors.

In vitro cartilage formation by human adult stem cells from bone marrow stroma defines the sequence of cellular and molecular events during chondrogenesis.

One approach to resolving the complexities of chondrogenesis is to examine simplified systems in vitro. We analyzed cartilage differentiation by human adult stem cells from bone marrow stroma. Marrow stromal cells were cultured as micromass pellets for 21 days in serum-free medium containing transforming growth factor (TGF)-beta3, dexamethasone, and bone morphogenetic protein (BMP)-6.