Characterization of pannexin 3 as a regulator of chondrocyte differentiation

Objectives: Through bioinformatical analysis, we identified pannexin 3 (Panx3) as a new member of the pannexin family of gap junction proteins that is preferentially expressed in cartilage. However, the physiological function of Panx3 was unknown. In this study, our objectives are to analyze the expression of Panx3 in developing cartilage and in a chondrogenic cell line and to identify the role of Panx3 in chondrocyte differentiation.

Methods: RT-PCR, Northern blotting, and in situ hybridization were performed for the expression analysis of Panx3 mRNA in cartilage. To analyze the role of Panx3 in chondrocyte differentiation, we used chondrogenic cell line ATDC5 to create stable transfectants for forced Panx3 expression by an expression vector and for inhibition of endogenous Panx3 expression by a shRNA vector.

Results: Panx3 mRNA was strongly expressed in prehypertrophic chondrocytes in the growth plate cartilage. Expression of Panx3 was progressively induced in differentiating ATDC5 cells after stimulation with insulin. Further, Panx3-transfected ATDC5 cells promoted the expression of chondrocyte marker genes, including type II collagen, aggrecan, and type X collagen, whereas suppression of endogenous Panx3 inhibited the expression of those genes.

Conclusion: Our results indicate that Panx3 promotes ATDC5 cell differentiation and suggest that Panx3 plays a role in cartilage development. [supported by NIDCR Intramural Program]