Bone Conditioned Medium: Preparation and Bioassay.
Autologous bone grafts are widely used in oral and maxillofacial surgery, orthopedics, and traumatology. Autologous bone grafts not only replace missing bone, they also support the complex process of bone regeneration. This favorable behavior of autografts is attributed to the three characteristics: osteoconductivity, osteogenicity, and osteoinductivity. However, there is another aspect: Bone grafts release a myriad of molecules, including growth factors, which can target mesenchymal cells involved in bone regeneration. The paracrine properties of bone grafts can be studied in vitro by the use of bone-conditioned medium (BCM). Here we present a protocol on how to prepare bone-conditioned medium from native pig cortical bone, and bone that underwent thermal processing or demineralization. Cells can be directly exposed to BCM or seeded onto biomaterials, such as collagen membranes, previously soaked with BCM. We give examples for in vitro bioassays with mesenchymal cells on the expression of TGF-β regulated genes. The presented protocols should encourage to further reveal the paracrine effects of bone grafts during bone regeneration and open a path for translational research in the broad field of reconstructive surgery.
Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern; Laboratory of Oral Cell Biology, School of Dental Medicine, University of Bern; Department of Oral and Maxillofacial Surgery, School of Dental Medicine, Universitat Internacional de Catalunya.
Jordi
J
Caballé-Serrano
Laboratory of Oral Cell Biology, School of Dental Medicine, University of Bern; Department of Cranio Maxillofacial Surgery, Inselspital, University of Bern.
Kosaku
K
Sawada
Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern; Laboratory of Oral Cell Biology, School of Dental Medicine, University of Bern; Department of Implant Dentistry, School of Dentistry, Universidade Federal de Santa Catarina.
Guenther
G
Schuldt Filho
Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern; Robert K. Schenk Laboratory of Oral Histology, School of Dental Medicine, University of Bern.
Dieter D
DD
Bosshardt
Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern.
Daniel
D
Buser
Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern; Laboratory of Oral Cell Biology, School of Dental Medicine, University of Bern; reinhard.gruber@zmk.unibe.ch.
Reinhard
R
Gruber
D000818:Animals; D001672:Biocompatible Materials / Q000494:pharmacology; D001681:Biological Assay / Q000379:methods; D001861:Bone Regeneration / Q000502:physiology; D016025:Bone Transplantation / Q000379:methods; D001842:Bone and Bones / Q000166:cytology*; D018929:Cell Culture Techniques / Q000379:methods*; D017077:Culture Media, Conditioned; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism; D013552:Swine; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics; D014182:Transplantation, Autologous
D001672:Biocompatible Materials; D017077:Culture Media, Conditioned; D036341:Intercellular Signaling Peptides and Proteins; D016212:Transforming Growth Factor beta