Osteogenic Differentiation is Selectively Promoted by Morphogenetic Signals from Chondrocytes and Synergized by a Nutrient Rich Growth Environment

Abstract

Cartilage formation always precedes that of bone during endochondral skeletal development. To determine if chondrocytes provide inductive signals for osteogenesis, C3H10T(1/2) mesenchymal stem cells were co-cultured in membrane separated transwell culture chambers with chondrocytes, osteoblasts, or fibroblasts. Osteogenesis, as assessed by the expression of osteocalcin mRNAs, was strongly induced in the C3H10T(1/2) cells co-cultured with chondrocytes but not induced by co-culture with either osteoblasts or fibroblasts. Interestingly, while only osteogenic differentiation was observed in the C3H10T(1/2) cells co-cultured with chondrocytes, bone morphogenetic protein (BMP)-7 treatment induced an ordered endochondral progression of skeletal cell differentiation in which chondrogenic differentiation preceded osteogenesis by 2 to 4 days. A nutrient enriched growth environment enhanced osteogenic differentiation induced by either co-culture or BMP-7 treatment 2- to 5-fold. Nutrient enhanced osteogenic differentiation was associated with an activation of the retinoblastoma-mediated signal transduction pathways. In summary, these results show that osteogenesis is selectively induced by morphogenetic signals produced by chondrocytes and that a nutrient rich environment enhances both BMP-7- and co-culture-induced osteogenic differentiation.

Mesenchymal Stem Cell Stimulation of Tissue Growth Depends on Differentiation State

“…Some studies have utilized mixed populations of cells plated together in monolayer [39], whereas others have employed cell culture insert culture systems [40] or conditioned medium [41] to study the interaction of cells without direct contact. Biomaterials can provide the added dimension of a 3D environment for cell and tissue coculture.…”

Controlled differentiation of stem cells

“…For example, during endochondral ossification, cartilage serves as a morphological template for blood vessel invasion and MSC recruitment for future bone [88]. Recent studies have indicated that morphogenetic signals from chondrocytes regulate multiple steps of chondrogenesis and endochondral ossification during skeletal differentiation [98][99][100]. These studies have implicated that mechanisms of endochondral ossification of condensing mesenchyme can be regulated by cell-secreted morphogenetic factors.…”

Coculture Strategies in Bone Tissue Engineering: The Impact of Culture Conditions on Pluripotent Stem Cell Populations

“…1 Later studies investigating cell differentiation identified a number of factors that could play a role in the communication between chondrocytes and osteoblasts in during differentiation. Cell communication through paracrine signaling has demonstrated that it plays a role in the selective osteogenic differentiation of MSC when exposed to soluble factors secreted by nonhypertrophic chondrocytes cultured in a transwell membrane system 22,23 (Fig. 2B).…”

“…The physically distinct locations of the cell populations help track individual phenotype changes and gene expression characteristics. [21][22][23]27,35 The shared media volume also aids in accounting for factors that may be constitutively expressed by one or more of the cell populations, as well as for any temporal changes in the secretory protein expression profile that is typical of many primary cell populations involved in tissue regeneration.…”

“…8 The indirect coculture of similar cells has been shown to yield different results in a number of other studies, raising the need for a consensus on the role of factors in the engineering of mineralized bone tissue and their occurrence as a function of the method used. [21][22][23] Through the course of this review, we attempt to present a synthesis of outcomes of coculture strategies in bone tissue engineering and how recent advancements can be applied to further our understanding of molecular interactions, and, finally, to succeed in the ultimate goal of engineering viable bone tissue (Fig. 1).…”