Skip to main content

Spatial nanoscale control of growth and adhesion factors to enhance the osteogenic differentiation of mesenchymal stem cells

Periodic Reporting for period 1 - OsteoNano (Spatial nanoscale control of growth and adhesion factors to enhance the osteogenic differentiation of mesenchymal stem cells)

Reporting period: 2015-05-01 to 2017-04-30

The main challenge of regenerative medicine is to understand the molecular basis of tissue-specific differentiation and then to apply the appropriate chemical and physical cues to guide stem cell fate.
In particular, bone tissue engineering is one of the major societal challenges in orthopedics, cranio-maxillo-facial surgery and dentistry, partially due to the aging of the population.
OsteoNano project focused on improving the efficiency of a biochemical factor, named bone morphogenetic protein 2 (BMP-2), to drive stem cells osteogenic differentiation. BMP-2 is a growth factor already used in clinics for spine fusion and tibial fractures. However, this practice presents important sides effects such as ectopic bone formation and eventually cancer.
The objectives of OsteoNano project are initially to design a 2D cellular support (biomimetic platforms) able to mimic the natural presentation of BMP-2 to cell surface. In vivo, BMP-2 is bound to extracellular matrix (ECM) components, in particular to the glycosaminoglycan heparan sulphate (HS), a sugar ubiquitously present on cellular surface and in the ECM. In normal conditions HS is not soluble but grafted on core-protein. For this reason the biomimetic platforms here developed present HS immobilized and oriented via its reducing end to a Streptavidin monolayer.
The second objective is to investigate the molecular interaction between BMP-2 and HS by the characterization of their binding affinity and the kinetics of release of BMP-2 from HS film.
Finally stem cells can be plated on these surfaces and cellular mechanism, in particular osteogenic differentiation driven by BMP-2 signalling pathway, analysed.
I proved that HS, once immobilized on the biomimetic platforms, enhances the biological effect of BMP-2 promoting osteogenic differentiation of mesenchymal stem cells and inhibiting the effect of noggin, the natural BMP-2 antagonists.
As conclusion, the immobilization of HS on 3D scaffold could be a future strategy to improve the effect of BMP-2 for bone repair.
During the first year of the OsteNano project biomimetic platforms have been characterized with surface sensitive techniques, used to obtain information on the exact amount of HS and on BMP-2 present on the surface. The control over BMP-2 surface molecular density is necessary to interpret cellular responses once interacting with the platforms.
Indeed, these surfaces have been used as support for cellular studies. In particular, murine cell model, which responds to BMP-2 (C2C12) and human mesenchymal stem cells, have been plated on these surfaces and their responses analyzed.
One of the earlier events of BMP-2 signaling pathway is the phosphorylation of SMAD protein, which is transported into cellular nucleus and activates genes responsible for osteogenic differentiation.
Cells plated on the biomimetic platforms show an enhanced SMAD phosphorylation level, only when BMP-2 is presented via HS.
Moreover, HS prevents BMP-2 recognition by noggin, one of the known BMPs antagonists. This result demonstrates that grafted HS improves the biological effect of BMP-2 and suggests that grafted HS can be a good candidate for future regenerative medicine applications.
In the last period I proved the versatility of these biomimetic platforms, which can be functionalized by multiple ligands, such as BMP-2 and Integrin ligands (RGD peptides).
During OsteoNano project I first wrote a review titled “Tuning cellular responses to BMP-2 with material surfaces” published in Cytokine & Growth Factor Reviews in 2015, which shows the open questions and the new challenges on BMP-2 presentation through engineered surfaces. The main results of the OsteoNano project have been published in a paper titled “Enhanced biological activity of BMP-2 bound to surface-grafted heparan sulfate” published in Advanced Biosystems in 2017.
This project has been also presented to several international conferences such as:
1st ERC BIOMIM Meeting, Grenoble, April 10-12, 2015; FEBS-MPST, Rhodes, September 24-29, 2015 and 11th international BMP Conference, Boston, October 26-30, 2016.
OsteoNano project clarified not only the interaction between HS and BMP-2 but also revealed the potency of immobilized and well-oriented HS to enhance the biological effect of BMP-2 on osteogenic differentiation.
OsteNano project proposed advantages and breakthroughs for future regenerative applications such as (i) reduced use of bound BMP-2 on the surfaces, reducing therefore cost and side effects (ii) possible enhancement of the osteogenic differentiation by surface co-presentation of BMP-2 and ligands targeting integrin receptors (RGD peptide) (iii) versatility of the biomimetic platforms potentially usable for other cellular lineage such as cartilage and neurons.
Graphic abstract: CGFR journal
Graphic abstract: Advanced biomaterial journal