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Methods Mol Biol ; 2206: 47-56, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754810


The development and maintenance of a functioning vascular system is a critical function for many aspects of tissue growth and regeneration. Vascular endothelial cell in vitro co-culture spheroids are self-organized cell composites that have the capacity to recapitulate the three-dimensional tissue microenvironment. These spheroid testing platforms aim to better understand the mechanisms of functional tissue and how new therapeutic agents can drive these 3D co-culture processes. Here we describe direct cell-cell 3D endothelial co-culture spheroid methods, to examine the physiological spatial growth and cell-cell interaction of vascular cells and surrounding native tissue cells in the formation of vascular networks within spheroids and the potential to regenerate tissue.

PLoS One ; 10(12): e0145080, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26675008


The current study has investigated the use of decellularised, demineralised bone extracellular matrix (ECM) hydrogel constructs for in vivo tissue mineralisation and bone formation. Stro-1-enriched human bone marrow stromal cells were incorporated together with select growth factors including VEGF, TGF-ß3, BMP-2, PTHrP and VitD3, to augment bone formation, and mixed with alginate for structural support. Growth factors were delivered through fast (non-osteogenic factors) and slow (osteogenic factors) release PLGA microparticles. Constructs of 5 mm length were implanted in vivo for 28 days within mice. Dense tissue assessed by micro-CT correlated with histologically assessed mineralised bone formation in all constructs. Exogenous growth factor addition did not enhance bone formation further compared to alginate/bone ECM (ALG/ECM) hydrogels alone. UV irradiation reduced bone formation through degradation of intrinsic growth factors within the bone ECM component and possibly also ECM cross-linking. BMP-2 and VitD3 rescued osteogenic induction. ALG/ECM hydrogels appeared highly osteoinductive and delivery of angiogenic or chondrogenic growth factors led to altered bone formation. All constructs demonstrated extensive host tissue invasion and vascularisation aiding integration and implant longevity. The proposed hydrogel system functioned without the need for growth factor incorporation or an exogenous inducible cell source. Optimal growth factor concentrations and spatiotemporal release profiles require further assessment, as the bone ECM component may suffer batch variability between donor materials. In summary, ALG/ECM hydrogels provide a versatile biomaterial scaffold for utilisation within regenerative medicine which may be tailored, ultimately, to form the tissue of choice through incorporation of select growth factors.

Regeneração Óssea , Matriz Extracelular , Hidrogéis/química , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Osteoblastos/citologia , Alginatos/efeitos adversos , Alginatos/química , Animais , Condrogênese , Ácido Glucurônico/efeitos adversos , Ácido Glucurônico/química , Ácidos Hexurônicos/efeitos adversos , Ácidos Hexurônicos/química , Humanos , Hidrogéis/efeitos adversos , Ácido Láctico/efeitos adversos , Ácido Láctico/química , Camundongos , Pessoa de Meia-Idade , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoblastos/transplante , Osteogênese , Ácido Poliglicólico/efeitos adversos , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Tecidos Suporte/efeitos adversos , Tecidos Suporte/química