Hydrogel Effects on Bone Marrow Stromal Cell Response to Chondrogenic Growth Factors
Rhima M. Colemana, b, Natasha D. Casea and Robert E. Guldberg
Biomaterials Volume 28, Issue 12, April 2007, Pages 2077-2086
The use of chondrocytes, a type of cells that secretes cartilage matrix, for cartilage tissue engineering has presented several challenges. It is difficult to harvest enough number of chondrocytes without dedifferentiation. Passaging chondrocytes usually results in decreased production of desirable cartilage matrix components such as of type II collagen, sulfated glycosaminoglycans (sGAGs), and increased production of undesirable type I collagen. Alternatively, bone marrow stromal cells (BMSCs), which have a similar characteristic to stem cells of being able to differentiate into bone, cartilage, and fat under proper conditions, has been successfully cultured in 3D and shown to possess chondrogenesis. The purpose of this paper is to investigate the effects of biochemical factors (i.e. growth factors) and focus on choices of hydrogels as scaffold materials on chondrogenic differentialtion of rat BMCs.
The main growth factors being investigated in this study include transforming growth factor-β1 (TGF-β1), fibroblastic growth factor-2 (FGF-2), and Dexamethasone (Dex). TGF-β1 enhances differentiation of stromal cells at late passages whereas FGF-2 treatment enhances TGF-β1. While Dex upregulates the production of TGF-β1 mediated collagen II and is required for chondrogenesis, it is also shown to have negative effects on cells viability. Initial data from this study showed that the treatment of FGF-2 during monolayer expansion and TGF-β1 in the presence of Dex during 3D culture in hydrogel culture of P2 yielded the greatest production of sGAG, a cartilaginous matrix. This optimal condition was then used to study the effect of hydrogels-alginate and agarose. Although 2% alginate and agarose used in this study have similar macroscopic property such as pore size, they were shown to have different effects on cell behaviors in response to biochemical stimulations. In alginate, the presence of FGF-2 enhances the effect of TGF-β1 and results in more production of a cartilaginous matrix than the presence of TGT- β1 alone. The opposite result was found when BMSCs were cultured in agarose. Moreover, Dex did not show any negative effect on cell death when being used in alginate cultures.
The techniques used in this study were mostly the same techniques we have learned in BE115 class such as cell passaging, 3D cell culturing, cell lysis, live/dead assays. Measuring cell’s production of excretion and cartilaginous protein was a good example of cell characterization based on cell’s responses to different stimuli and environments. Live/dead assay using green and red dyes as molecular probes with a confocal microscope was used to determine cell death in gel samples. The use of software such as ImagePro, in this case, to count the number of cells in live/dead assay was proved to be necessary and useful.
3 comments:
I think it would also be important to understand what characteristics of the alginate and the agarose in conjugation with the growth factors that prevented matrix formation. This could be useful not only for understanding chondrogenesis but for other tissue engineering applications as well. And furthermore very interesting. (i should probably read the paper).
The study is interesting as it provides some explanations regarding to the interaction between alginate and positive-charged proteins,which explains the role of alginate in cartilage tissue culture.
For other cell lines of cartilage, are we using the same growth factors if we want to conduct the same experiment?
Will different combination of alginate and agarose give different results of the growth factors?
i was wondering if the paper discussed whether the differences in the amount of matrix produced between alginate hydrogels and agarose are due to the release kinetics?
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