Biomedical Engineering Reference
In-Depth Information
spreading and proliferation. This parameter of course can also be negatively
influenced by test material constituents that are (sub-)toxic for the seeded cell
type (=[Answer to questions 5 and 7).
• Level 4—cell on-growth test: In this level, material integration in tissues is tested.
Normally tissues are in direct contact with an implant material. Cell aggregates
containing around 15,000 cells of one or more typical cell types of the targeted site
of the implantation niche are prepared and placed on top of the samples. The cells
have the choice to stay within the tissue or to leave the tissue-like environment and
contact and migrate onto the test material. Cell outgrowth is analysed after five
days in culture. The area covered by cells is determined and compared with
reference surfaces. A reduced coverage gives an indication of the cells intrinsic
competitive strength to cover the test material surface and, if more cell types are
included, also relative to each other (=[Answer to question 6).
• Level 5—cell differentiation test: In this test level, cells are seeded on top of the
samples at a density ensuring sub-confluency after five days in culture. There-
after, mRNA is isolated and the relative concentration of mRNA of specific
genes determined. The outcome yields information on how far the material
supports cell differentiation and allows statements on how far differentiation is
influenced by the material. Statements regarding the extent and time evolution
of cell differentiation can be made if more than one time-point is investigated
(=[Answer to question 9).
• Level 6—cell-cell competition test: The final level would allow for assessment
of cell affinity to the substratum in a cell-cell competitive manner. Labelled
cells of different cell types in a defined ratio and amount are seeded on top of the
material. After five days in culture, the cell number of each cell type is assessed
as well as the expression of cell-type-specific differentiation markers. The latter
and final test has the potential to predict which cell type finally covers the
implant surface (=[Answer to questions 8 and 9).
The idea behind this suggested set-up is that each test is more stringent, reducing
the number of promising materials from one level to the next. Materials that perform
adequately in all test levels are considered to be optimal and ready for subsequent in
vivo testing. The proposed test battery has the potential to reduce the time to market
and cost per new developed implant that will positively pass the human trials.
Acknowledgments The present work was supported by the European Community's 7th
Framework Programme under grant agreement nos. NMP3-LA-2008-214685 (project Magister)
and NMP3-LA-2008-213939 (project POCO).
References
1. 10993-12/TC194 I (2007) ISO 10993-12:2007
2. Alves CM, Reis RL, Hunt JA (2010) The competitive adsorption of human proteins onto
natural-based biomaterials. J R Soc Interface 7:1367-1377
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