We hypothesized that a crosstalk between osteoblast and fibroblast (FB) exists, which contributes to bone as a dynamic tissue. Cell-free supernatants were harvested from fibroblast cultures and later subject pre-osteoblasts to investigate there capacity to modulate cell viability and differentiation mechanisms, reporting the possible involvement of Shh signaling as a paracrine mechanism. By exploring immunoblotting technology, we have shown that FB-released factors interfere with osteoblast metabolism by up-regulating the phosphorylation of FAK and Rac-1 proteins at the early stage and later contribute to osteoblast differentiation by up-modulating alkaline phosphatase (ALP) and in vitro mineralization. We also found that Shh signaling was not required during osteoblastic differentiation promoted by the FB-released factors as well as MAPK-ERK phosphorylation, while pre-osteoblast cultures subjected to osteogenic medium (O.M.) require downstream transducers of Shh, such as Patched and Gli-1, and MAPK-ERK. Altogether, our results indicate for the first time a possible mechanism involved in the crosstalk between fibroblasts and osteoblasts, as it was possible to observe trophic factors released by fibroblasts interfering decisively in osteoblast metabolism in a Shh-independent manner. This study collaborates the body of work that indicates paracrine signaling molecules participate in the crosstalk among bone-resident cells and explains, at least partially, the biological mechanisms responsible for bone tissue dynamism, opening new avenues to understand etiologies of bone diseases.
Bone Fibroblast Osteoblast Crosstalk Cell signaling
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The authors are grateful to FAPESP (#2015/00581-9; #2014/22689-3) and CNPq (#477452/2012-4) for the financial support. W.F.Z. is a PQ-2 fellow from CNPq (#301966/2015-0).
This study was funded by the Fundação de Amparo à Pesquisa do Estado de São Paulo - FAPESP (grants #2015/00581-9; #2014/22689-3) and the Conselho Nacional de Desenvolvimento Científico e Tecnológico - CNPq (grant #477452/2012-4).
Compliance with ethical standards
Conflict of interest
All the authors declare to have no conflict of interest with the materials used in the present study.
Gemini-Piperni S, Milani R, Bertazzo S, Al Et (2014) Kinome profiling of osteoblasts on hydroxyapatite opens new avenues on biomaterial cell signaling. Biotechnol Bioeng 111:1900–1905. doi:10.1002/Bit.25246CrossRefPubMedGoogle Scholar
Ducy P, Schinke T, Karsenty G (2000) The osteoblast: a sophisticated fibroblast under central surveillance. Science 289:1501–1504CrossRefPubMedGoogle Scholar
Lee B, Thirunavukkarasu K, Zhou L, Al Et (1997) Missense mutations abolishing dna binding of the osteoblast-specific transcription factor Osf2/Cbfa1 in cleidocranial dysplasia. Nat Genet 16:307–310. doi:10.1038/Ng0797-307CrossRefPubMedGoogle Scholar
Lee J, Sk Madhurakkat Perikamana, Ahmad T, Al Et (2017) Controlled retention of Bmp-2-derived peptide on nanofibers based on mussel-inspired adhesion for bone formation. Tissue Eng Part A. doi:10.1089/Ten.Tea.2016.0363Google Scholar
Wf Zambuzzi, Bruni-Cardoso A, Jm Granjeiro, Al Et (2009) On the road to understanding of the osteoblast adhesion: cytoskeleton organization is rearranged by distinct signaling pathways. J Cell Biochem 108:134–144. doi:10.1002/Jcb.22236CrossRefGoogle Scholar
Cavagis A, Takamori E, Granjeiro J, Al Et (2014) Tnfalpha contributes for attenuating both Y397fak And Y416src phosphorylations in osteoblasts. Oral Dis 20:780–786. doi:10.1111/Odi.12202PubMedGoogle Scholar
Milani R, Ferreira CV, Jm Granjeiro, Al Et (2010) Phosphoproteome reveals an atlas of protein signaling networks during osteoblast adhesion. J Cell Biochem 109:957–966. doi:10.1002/Jcb.22479PubMedGoogle Scholar
Wf Zambuzzi, Ferreira CV, Jm Granjeiro, Aoyama H (2011) Biological behavior of pre-osteoblasts on natural hydroxyapatite: a study of signaling molecules from attachment to differentiation. J Biomed Mater Res A 97:193–200. doi:10.1002/Jbm.A.32933Google Scholar
Rutault K, Ca Hazzalin, Lc Mahadevan (2001) Combinations of Erk and P38 Mapk inhibitors ablate tumor necrosis factor-alpha (Tnf-Alpha) Mrna induction. Evidence for selective destabilization of Tnf-Alpha transcripts. J Biol Chem 276:6666–6674. doi:10.1074/Jbc.M005486200CrossRefPubMedGoogle Scholar
Liu X, Zeng L, Zhao Z, Al Et (2017) Pbmc activation via the Erk and Stat signaling pathways enhances the anti-tumor activity of Staphylococcal enterotoxin A. Mol Cell Biochem. doi:10.1007/S11010-017-3038-5Google Scholar
Mc Moorer, Hebert C, Re Tomlinson, Al Et (2017) Defective signaling, osteoblastogenesis and bone remodeling in a mouse model of connexin 43 C-terminal truncation. J Cell Sci 130:531–540. doi:10.1242/Jcs.197285CrossRefGoogle Scholar
Wang M, Shu Z-J, Wang Y, Peng W (2017) Stachydrine hydrochloride inhibits proliferation and induces apoptosis of breast cancer cells via inhibition of Akt and Erk pathways. Am J Transl Res 9:1834–1844PubMedPubMedCentralGoogle Scholar