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Construction of vascular tissues with macro-porous nano-fibrous scaffolds and smooth muscle cells enriched from differentiated embryonic stem cells.

ABSTRACT: Vascular smooth muscle cells (SMCs) have been broadly used for constructing tissue-engineered blood vessels. However, the availability of mature SMCs from donors or patients is very limited. Derivation of SMCs by differentiating embryonic stem cells (ESCs) has been reported, but not widely utilized in vascular tissue engineering due to low induction efficiency and, hence, low SMC purity. To address these problems, SMCs were enriched from retinoic acid induced mouse ESCs with LacZ genetic labeling under the control of SM22? promoter as the positive sorting marker in the present study. The sorted SMCs were characterized and then cultured on three-dimensional macro-porous nano-fibrous scaffolds in vitro or implanted subcutaneously into nude mice after being seeded on the scaffolds. Our data showed that the LacZ staining, which reflected the corresponding SMC marker SM22? expression level, was efficient as a positive selection marker to dramatically enrich SMCs and eliminate other cell types. After the sorted cells were seeded into the three-dimensional nano-fibrous scaffolds, continuous retinoic acid treatment further enhanced the SMC marker gene expression level while inhibited pluripotent maker gene expression level during the in vitro culture. Meanwhile, after being implanted subcutaneously into nude mice, the implanted cells maintained the positive LacZ staining within the constructs and no teratoma formation was observed. In conclusion, our results demonstrated the potential of SMCs derived from ESCs as a promising cell source for therapeutic vascular tissue engineering and disease model applications.

SUBMITTER: Hu J

PROVIDER: S-EPMC3335865 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Construction of vascular tissues with macro-porous nano-fibrous scaffolds and smooth muscle cells enriched from differentiated embryonic stem cells.

Hu Jiang J Xie Changqing C Ma Haiyun H Yang Bo B Ma Peter X PX Chen Y Eugene YE

PloS one 20120424 4

Vascular smooth muscle cells (SMCs) have been broadly used for constructing tissue-engineered blood vessels. However, the availability of mature SMCs from donors or patients is very limited. Derivation of SMCs by differentiating embryonic stem cells (ESCs) has been reported, but not widely utilized in vascular tissue engineering due to low induction efficiency and, hence, low SMC purity. To address these problems, SMCs were enriched from retinoic acid induced mouse ESCs with LacZ genetic labelin ...[more]

PMID: 22545119

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Project description:Adequate bony support is the key to re-establish both function and esthetics in the craniofacial region. Autologous bone grafting has been the gold standard for regeneration of problematic large bone defects. However, poor graft availability and donor-site complications have led to alternative bone tissue-engineering approaches combining osteoinductive biomaterials and three-dimensional cell aggregates in scaffolds or constructs. The goal of the present study was to generate novel cell aggregate-loaded macroporous scaffolds combining the osteoinductive properties of titanium dioxide (TiO2) with hydroxyapatite-gelatin nanocomposites (HAP-GEL) for regeneration of craniofacial defects. Here we investigated the in vivo applicability of macroporous (TiO2)-enriched HAP-GEL scaffolds with undifferentiated and osteogenically differentiated multipotent adult progenitor cell (MAPC and OD-MAPC, respectively) aggregates for calvaria bone regeneration. The silane-coated HAP-GEL with and without TiO2 additives were polymerized and molded to produce macroporous scaffolds. Aggregates of the rat MAPC were precultured, loaded into each scaffold, and implanted to rat calvaria critical-size defects to study bone regeneration. Bone autografts were used as positive controls and a poly(lactic-co-glycolic acid) (PLGA) scaffold for comparison purposes. Preimplanted scaffolds and calvaria bone from pig were tested for ultimate compressive strength with an Instron 4411(®) and for porosity with microcomputerized tomography (?CT). Osteointegration and newly formed bone (NFB) were assessed by ?CT and nondecalcified histology, and quantified by calcium fluorescence labeling. Results showed that the macroporous TiO2-HAP-GEL scaffold had a comparable strength relative to the natural calvaria bone (13.8±4.5?MPa and 24.5±8.3?MPa, respectively). Porosity was 1.52±0.8?mm and 0.64±0.4?mm for TiO2-HAP-GEL and calvaria bone, respectively. At 8 and 12 weeks postimplantation into rat calvaria defects, greater osteointegration and NFB were significantly present in the TiO2-enriched HAP-GEL constructs with OD-MAPCs, compared to the undifferentiated MAPC-loaded constructs, cell-free HAP-GEL with and without titanium, and PLGA scaffolds. The tissue-engineered TiO2-enriched HAP-GEL constructs with OD-MAPC aggregates present a potential useful therapeutic approach for calvaria bone regeneration.

Project description:BackgroundTissue engineering of the annulus fibrosus (AF) shows promise as a treatment for patients with degenerative disc disease (DDD). However, it remains challenging due to the intrinsic heterogeneity of AF tissue. Fabrication of scaffolds recapitulating the specific cellular, componential, and microstructural features of AF, therefore, is critical to successful AF tissue regeneration.MethodsPoly-L-lactic acid (PLLA) fibrous scaffolds with various fiber diameters and orientation were prepared to mimic the microstructural characteristics of AF tissue using electrospinning technique. AF-derived stem cells (AFSCs) were cultured on the PLLA fibrous scaffolds for 7 days.ResultsThe morphology of AFSCs significantly varied when cultured on the scaffolds with various fiber diameters and orientation. AFSCs were nearly round on scaffolds with small fibers. However, they became spindle-shaped on scaffolds with large fibers. Meanwhile, upregulated expression of collagen-I gene happened in cells cultured on scaffolds with large fibers, while enhanced expression of collagen-II and aggrecan genes was seen on scaffolds with small fibers. The production of related proteins also showed similar trends. Further, culturing AFSCs on a heterogeneous scaffold by overlaying membranes with different fiber sizes led to the formation of a hierarchical structure approximating native AF tissue.ConclusionFindings from this study demonstrate that fibrous scaffolds with different fiber sizes effectively promoted the differentiation of AFSCs into specific cells similar to the types of cells at various AF zones. It also provides a valuable reference for regulation of cell differentiation and fabrication of engineered tissues with complex hierarchical structures using the physical cues of scaffolds.The translational potential of this articleEffective AF repair is an essential need for treating degenerative disc disease. Tissue engineering is a promising approach to achieving tissue regeneration and restoring normal functions of tissues. By mimicking the key structural features of native AF tissue, including fiber size and alignment, this study deciphered the effect of scaffold materials on the cell differentiation and extracellular matrix deposition, which provides a solid basis for designing new strategies toward more effective AF repair and regeneration.

Dataset Information

Construction of vascular tissues with macro-porous nano-fibrous scaffolds and smooth muscle cells enriched from differentiated embryonic stem cells.

Publications

Construction of vascular tissues with macro-porous nano-fibrous scaffolds and smooth muscle cells enriched from differentiated embryonic stem cells.

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