Role of culture conditions on in vitro transformation and cellular colonization of biomimetic HA-Col scaffolds

Abstract

We have recently developed new 3D hydroxyapatite/collagen (50/50 wt%) scaffolds using a biomimetic synthesis approach. The first in vitro tests performed in static culture showed a limited cell colonization and survival inside the scaffolds. The current study evaluated in dynamic culture the scaffold changes and colonization by human immortalized osteoprogenitor STRO-1A cells. The stability of our scaffolds in the different culture conditions (static, low flow, high flow) was validated by the maintenance of the pore diameter and interconnectivity over 21 d. The colonization and the viability of STRO-1A cells inside the scaffolds were further evaluated on histological sections. It was demonstrated that only the high flow-rate allowed cell survival after 7 d and a complete scaffold colonization. Moreover, the colonization and viability were different in function of the scaffold position inside the perfusion container. The differentiation markers (alkaline phosphatase activity, type I procollagen and osteocalcin synthesis) of STRO-1A cells were analyzed in the culture medium after 7, 14 and 21 d. The low flow-rate increased significantly the three markers compared with static conditions. In contrast, markers were reduced in high flow-rate compared with low flow-rate. To explain this surprising result, we hypothesized that the different molecules were actually adsorbed on the scaffold because of the closed circuit used in the high flow-rate conditions. In summary, this study provides original results on the influence of flow rate but mostly of the circuit used (open/closed) on the structural modifications and cell colonization of collagen-HA scaffolds.

The effect of purmorphamine on differentiation of endometrial stem cells into osteoblast-like cells on collagen/hydroxyapatite scaffolds

“…ALP increase in both control and PMA-treated groups extended up to day 21 of the culture. Even at this stage, the ALP level was considerably higher in the control group than in the PMA-treated group (Arpornmaeklong et al 2010, Campos et al 2013. This finding echoes with that of Campos, where increased ALP activity was reported in 2D cultures containing osteoprogenitor cells' osteogenic inductor.…”

Bioengineered constructs combined with exercise enhance stem cell-mediated treatment of volumetric muscle loss

“…Culturing cells within 3D substrates results in a loss of cell viability 9 39 . During the process of generating bioconstructs with Luc + MuSCs, we observed substantial bioluminescent signal decay in vitro within two hours of reconstitution, continuing up to 24 h ( Fig.…”

“…3a ). Perfusion systems have proven a practical tool for remedying the loss of cell viability in 3D culture conditions 35 39 40 . We thus sought to establish a perfusing bioreactor to be used immediately after curing of the hydrogel to test whether this would enhance MuSC viability.…”

Cellular Nutrition in Complex Three-Dimensional Scaffolds: A Comparison between Experiments and Computer Simulations

“…They showed enhanced cell proliferation at a flow rate of 10 μ L/min and induced cell differentiation at 200 μ L/min in bone constructs seeded with mouse osteoblast precursor cells. Campos et al also determined cell differentiation dependent on fluid flow rates in a perfusion culture [ 24 ]. On hydroxyapatite/collagen scaffolds colonized with STRO-1A cells they found stimulated proliferation at flow rates of 300 μ L/min, but osteogenic marker gene expression was enhanced with a low flow of only 30 μ L/min.…”