Influence of a Cyclic RGD Peptide on MAP Kinases Signaling Pathways

Influence of a Cyclic RGD Peptide on MAP Kinases Signaling Pathways
S. PALLU1, M. DARD2, A. JONCZYK3, and J. AM?‚D?‚E1, 1 INSERM U-443, Universit?© Bordeaux 2, France, 2 Biomet-Merck Biomaterials, Darmstadt, Germany, 3 Merck Preclinical Research, Darmstadt, Germany
2003
IADR
Objectives: Osseointegration of bone substitutes materials can be improved if the implants are supplemented with autologuous osteoblastic cells. RGD-containing peptides contribute to stimulate pro-adhesive properties of materials towards osteogenic cells. Modification of materials by grafting RGD-containing peptides is based on the affinity of avb3-avb5 integrins to extracellular matrix proteins. Previous promising results have been obtained on different osteoblastic cell populations with a cyclo-DfKRG peptide. The aim of this work is to confirm these results by using of Human Bone Marrow Stromal Cells (HBMSC).
Methods: HBMSC have been cultured during 15, 30, 60, 90 min in IMDM alone onto different coatings ; cyclo-DfKRG (100 ¨µM), poly-L-lysine (0,005 %), fibronectin (10 ¨µg/ml). Total proteins were extracted using a RIPA buffer. Tyrosine kinase (PTK) activities were determined by using a kit (Promega, France). Tyrosine phosphorylation of p125FAK and MAPK have been determined by Western blotting and results have been quantified using NIH 1.62 analyser (Clark Lab Mirco Junker, USA).
Results: Culture onto cyclo-DfKRG peptide induced an increase of PTK activities which reached a maximum after 30 min contrary to poly-L-lysine. Adhesion to cyclo-DfKRG promoted early tyrosine phosphorylation of p125FAK and Erk 1 after 15 min of cell seeding, whereas Erk 2 and p38 were activated after 30 min.
Conclusions: Adhesion of HBMSC onto cyclo-DfKRG peptide coating should contribute to activate transcription factors like cbfa / osf2 responsible for stimulation of osteocalcin expression. This peptide seems to be a good candidate to modify biomaterials and to increase their osseointegration.