Dataset Information

Collagen-Hydroxyapatite Scaffolds Induce Human Adipose Derived Stem Cells Osteogenic Differentiation In Vitro.

ABSTRACT: Mesenchymal stem cells (MSCs) play a crucial role in regulating normal skeletal homeostasis and, in case of injury, in bone healing and reestablishment of skeletal integrity. Recent scientific literature is focused on the development of bone regeneration models where MSCs are combined with biomimetic three-dimensional scaffolds able to direct MSC osteogenesis. In this work the osteogenic potential of human MSCs isolated from adipose tissue (hADSCs) has been evaluated in vitro in combination with collagen/Mg doped hydroxyapatite scaffolds. Results demonstrate the high osteogenic potential of hADSCs when cultured in specific differentiation induction medium, as revealed by the Alizarin Red S staining and gene expression profile analysis. In combination with collagen/hydroxyapatite scaffold, hADSCs differentiate into mature osteoblasts even in the absence of specific inducing factors; nevertheless, the supplement of the factors markedly accelerates the osteogenic process, as confirmed by the expression of specific markers of pre-osteoblast and mature osteoblast stages, such as osterix, osteopontin (also known as bone sialoprotein I), osteocalcin and specific markers of extracellular matrix maturation and mineralization stages, such as ALPL and osteonectin. Hence, the present work demonstrates that the scaffold per se is able to induce hADSCs differentiation, while the addition of osteo-inductive factors produces a significant acceleration of the osteogenic process. This observation makes the use of our model potentially interesting in the field of regenerative medicine for the treatment of bone defects.

SUBMITTER: Calabrese G

PROVIDER: S-EPMC4794180 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Collagen-Hydroxyapatite Scaffolds Induce Human Adipose Derived Stem Cells Osteogenic Differentiation In Vitro.

Calabrese Giovanna G Giuffrida Raffaella R Fabbi Claudia C Figallo Elisa E Lo Furno Debora D Gulino Rosario R Colarossi Cristina C Fullone Francesco F Giuffrida Rosario R Parenti Rosalba R Memeo Lorenzo L Forte Stefano S

PloS one 20160316 3

Mesenchymal stem cells (MSCs) play a crucial role in regulating normal skeletal homeostasis and, in case of injury, in bone healing and reestablishment of skeletal integrity. Recent scientific literature is focused on the development of bone regeneration models where MSCs are combined with biomimetic three-dimensional scaffolds able to direct MSC osteogenesis. In this work the osteogenic potential of human MSCs isolated from adipose tissue (hADSCs) has been evaluated in vitro in combination with ...[more]

PMID: 26982592

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Project description:Osteogenic cell signaling pathway disruption varies among bone diseases. This investigation was designed to identify adipose-derived multipotent stromal cell (ASC) and bone graft scaffold combinations for local, targeted restoration of gene expression and extracellular matrix (ECM) deposition. Human ASC osteogenesis on bone graft materials was quantified following culture in stromal (S), osteogenic (O), or osteogenic for 48 h followed by stromal medium (OS) to test the two-part hypothesis: (1) identical ASC isolates on distinct bone graft scaffolds demonstrate unique viability, differentiation, ECM production, and gene expression in the same culture conditions; (2) identical ASC-bone graft scaffold combinations have different cell viability, differentiation, ECM production, and gene expression when cultured in S, O, or OS medium. Three commercially available bone graft scaffold materials, type I bovine collagen (C), hydroxyapatite + β-tricalcium phosphate + type I bovine collagen (HT), and β-tricalcium phosphate + type I bovine collagen (CT) were evaluated. Passage 3 ASCs were loaded onto scaffold blocks with a spinner flask bioreactor, and constructs were cultured up to 28 days. Cell viability, gene expression (alkaline phosphatase [ALPL], osteoprotegerin [TNFRSF11B], osteocalcin [BGLAP], cannabinoid receptors type I [CNR1] and II [CNR2], receptor activator of nuclear factor kappa β ligand [TNFSF11]), as well as ECM DNA, collagen, sulfated glycosaminoglycan, and protein content were quantified. Matrix organization was evaluated with scanning electron microscopy. Effects of scaffold, medium, or culture duration on cell viability were minimal. Significantly higher initial ALPL expression decreased with time, while BGLAP expression increased in HT constructs in O medium, and the constructs had the most abundant ECM components and ultrastructural organization. There was a similar, although delayed, pattern of gene expression and greater ECM collagen with less organization in C constructs in O medium. Higher CNR1 expression in C versus higher TNFRSF11B/TNFSF11 expression in HT constructs throughout the study support stimulation of unique osteogenic signaling pathways by identical cell isolates. These results suggest that bone scaffold composition may be used to selectively target specific osteogenic cell signaling pathways in ASC constructs to stimulate ECM deposition based on therapeutic needs.

Project description:Craniomaxillofacial bone defects can occur as a result of congenital, post-oncologic, and high-energy impact conditions. The scale and irregularity of such defects motivate new biomaterials to promote regeneration of the damaged bone. We have recently described a mineralized collagen scaffold capable of instructing stem cell osteogenic differentiation and new bone infill in the absence of traditional osteogenic supplements. Herein, we report the integration of a millimeter-scale reinforcing poly (lactic acid) frame fabricated via 3D-printing into the mineralized collagen scaffold with micron-scale porosity to form a multi-scale mineralized collagen-PLA composite. We describe modifications to the PLA frame design to increase the compressive strength (Young's Modulus, ultimate stress and strain) of the composite. A critical challenge beyond increasing the compressive strength of the collagen scaffold is addressing challenges inherent with the irregularity of clinical defects. As a result, we examined the potential for modifying the frame architecture to render the composite with increased compressive strength in one axis or radial compressibility and shape-fitting capacity in an orthogonal axis. A library of mineralized collagen-PLA composites was mechanically characterized via compression testing and push-out test to describe mechanical performance and shape-fitting capacity. We also report in vitro comparison of the bioactivity of porcine adipose derived stem cells in the mineralized collagen-PLA composite versus the mineralized collagen scaffold via metabolic activity, gene expression, and functional matrix synthesis. The results suggest that incorporation of the PLA reinforcing frame does not negatively influence the osteoinductive nature of the mineralized collagen scaffold. Together, these findings suggest a strategy to address often competing bioactivity, mechanical strength, and shape-fitting design requirements for biomaterials for craniomaxillofacial bone regeneration.

Project description:Objectives: Repairing articular cartilage damage is challenging. Clinically, tissue engineering technology is used to induce stem cell differentiation and proliferation on biological scaffolds to repair defective joints. However, no ideal biological scaffolds have been identified. This study investigated the effects of amniotic membrane/collagen scaffolds on the differentiation of adipose-derived stem cells (ADSCs) and articular cartilage repair. Methods: Adipose tissue of New Zealand rabbits was excised, and ADSCs were isolated and induced for differentiation. An articular cartilage defect model was constructed to identify the effect of amniotic membrane/collagen scaffolds on cartilage repair. Cartilage formation was analyzed by imaging and toluene blue staining. Knee joint recovery in rabbits was examined using hematoxylin and eosin, toluidine, safranine, and immunohistochemistry at 12 weeks post-operation. Gene expression was examined using ELISA, RT-PCR, Western blotting, and immunofluorescence. Results: The adipose tissue was effectively differentiated into ADSCs, which further differentiated into chondrogenic, osteogenic, and lipogenic lineages after 3 weeks' culture in vitro. Compared with platelet-rich plasmon (PRP) scaffolds, the amniotic membrane scaffolds better promoted the growth and differentiation of ADSCs. Additionally, scaffolds containing the PRP and amniotic membrane efficiently enhanced the osteogenic differentiation of ADSCs. The levels of COL1A1, COL2A1, COL10A1, SOX9, and ACAN in ADSCs + amniotic membrane + PRP group were significantly higher than the other groups both in vitro and in vivo. The Wakitani scores of the ADSC + amniotic membrane + PRP group were lower than that in ADSC + PRP (4.4 ± 0.44**), ADSC + amniotic membrane (2.63 ± 0.38**), and control groups (6.733 ± 0.21) at week 12 post-operation. Osteogenesis in rabbits of the ADSC + amniotic membrane + PRP group was significantly upregulated when compared with other groups. Amniotic membranes significantly promoted the expression of cartilage regeneration-related factors (SOX6, SOX9, RUNX2, NKX3-2, MEF2C, and GATA4). The ADSC + PRP + amniotic membrane group exhibited the highest levels of TGF-β, PDGF, and FGF while exhibiting the lowest level of IL-1β, IL6, and TNF-α in articular cavity. Conclusion: Amniotic membrane/collagen combination-based scaffolds promoted the proliferation and cartilage differentiation of ADSCs, and may provide a new treatment paradigm for patients with cartilage injury.

Dataset Information

Collagen-Hydroxyapatite Scaffolds Induce Human Adipose Derived Stem Cells Osteogenic Differentiation In Vitro.

Publications

Collagen-Hydroxyapatite Scaffolds Induce Human Adipose Derived Stem Cells Osteogenic Differentiation In Vitro.

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