Bone morphogenetic protein‐2 modulation of chondrogenic differentiation in vitro involves gap junction‐mediated intercellular communication

Abstract

Undifferentiated mesenchymal cells in the limb bud integrate a complex array of local and systemic signals during the process of cell condensation and chondrogenic differentiation. To address the relationship between bone morphogenetic protein (BMP) signaling and gap junction‐mediated intercellular communication, we examined the effects of BMP‐2 and a gap junction blocker 18 alpha glycyrrhetinic acid (18α‐GCA) on mesenchymal cell condensation and chondrogenic differentiation in an in vitro chondrogenic model. We find that connexin43 protein expression significantly correlates with early mesenchymal cellular condensation and chondrogenesis in high‐density limb bud cell culture. The level of connexin43 mRNA is maximally upregulated 48 h after treatment with recombinant human BMP‐2 with corresponding changes in protein expression. Inhibition of gap junction‐mediated intercellular communication with 2.5 μM 18α‐GCA decreases chondrogenic differentiation by 50% at 96 h without effects on housekeeping genes. Exposure to 18α‐GCA for only the first 24–48 h after plating does not affect condensation or later chondrogenic differentiation suggesting that gap junction‐mediated intercellular communication is not critical for the initial phase of condensation but is important for the onset of differentiation. 18α‐GCA can also block the chondrogenic effects of BMP‐2 without effects on cell number or connexin43 expression. These observations demonstrate 18α‐GCA‐sensitive regulation of intercellular communication in limb mesenchymal cells undergoing chondrogenic differentiation and suggest that BMP‐2 induced chondrogenic differentiation may be mediated in part through the modulation of connexin43 expression and gap junction‐mediated intercellular communication. J. Cell. Physiol. 193: 233–243, 2002.