The integrin α5β1 regulates chondrocyte hypertrophic differentiation induced by GTP-bound transglutaminase 2

Soluble GTP-bound transglutaminasc 2 (TG2) induces hypertrophic differentiation in chondrocyte cultures in a beta 1 integrin-dependent fashion. beta 1 integrin subfamily consists of 12 heterodimers with 12 different alpha subunits and a beta 1 subunit. To identify the specific integrin heterodimer(s) responsible for this process, we specifically blocked individual beta 1 integrins on the CH-8 immortalized human chondrocytes during hypertrophic differentiation. Blockade of alpha 5 beta 1 inhibited matrix metalloprotemase 13 (MMP- 13), type X collagen expression, alkaline phosphatase activity and matrix calcification by 30-50% associated with weak effects of anti-alpha 3 beta 1 and -alpha 4 beta 1. Anti-alpha 1 beta 1, -alpha 2 beta 1 and -alpha 6 beta 1 had no effect. To examine whether the dominant effect of integrin alpha 5 beta 1 was due to a direct interaction with TG2, we incubated the chondrocytic cells on plates coated with GTP-bound TG2. The immobilized GTP-bound TG2 induced hypertrophic differentiation to the same extent as the soluble GTP-bound TG2, which was also inhibited by anti-alpha 5 beta 1. CH-8 cells grown on plates coated with GTP-bound TG2 demonstrated adherence associated with focal adhesion kinase phosphorylation. These properties were inhibited by anti-alpha 5 beta 1. Furthermore, engagement of alpha 5 beta 1 on CH-8 cells via anti-alpha 5 beta 1 antibody did, in fact, induce differentiation. Although CH-8 cells adhered to GTP-free TG2 via integrin alpha 5 beta 1 1, the cells failed to undergo hypertrophic differentiation. Thus, integrin alpha 5 beta 1 is critical for the chondrocyte hypertrophic differentiation induced by GTP-bound TG2, and this induction is ligand dependent. (c) 2007 Elsevier B.V/International Society of Matrix Biology. All rights reserved.