Integrin-mediated interactions with a laminin- presenting substrate modulate biosynthesis and phenotypic expression for cells of the human nucleus pulposus

With aging and pathology, cells of the nucleus pulposus (NP) de-differentiate towards a fibroblast-like phenotype, a change that contributes to degeneration of the intervertebral disc (IVD). Laminin isoforms are a component of the NP extracellular matrix during development but largely disappear in the adult NP tissue. Exposing human adult NP cells to hydrogels made from PEGylated-laminin-111 (PEGLM) has been shown to regulate NP cell behaviors and promote cells to assume a biosynthetically active state with gene/protein expression and morphology consistent with those observed in juvenile NP cells. However, the mechanism regulating this effect has remained unknown. In the present study, the integrin subunits that promote adult degenerative NP cell interactions with laminin-111 are identified by performing integrin blocking studies along with assays of intracellular signaling and cell phenotype. The findings indicate that integrin a3 is a primary regulator of cell attachment to laminin and is associated with phosphorylation of signaling molecules downstream of integrin engagement (ERK 1/2 and GSK3p). Sustained effects of blocking integrin a3 were also demonstrated including decreased expression of phenotypic markers, reduced biosynthesis, and altered cytoskeletal organization. Furthermore, blocking both integrin a3 and additional integrin subunits elicited changes in cell clustering, but did not alter the phenotype of single cells. These findings reveal that integrin- mediated interactions through integrin a3 are critical in the process by which NP cells sense and alter phenotype in response to culture upon laminin and further suggest that targeting integrin a3 has potential for reversing or slowing degenerative changes to the NP cell.