BMP2 binds non-specifically to PEG-passivated biomaterials and induces substantial signaling

Abstract

Biomaterials are widely employed across diverse biomedical applications and represent an attractive strategy to explore physiologically how extracellular matrix components influence the cellular response. In this study, we aimed to use previously developed biomimetic streptavidin platforms to investigate the role of glycosaminoglycans (GAGs) in bone morphogenetic protein 2 (BMP2) signaling. However, we observed that the interpretation of our findings was skewed due to the GAG-unrelated, non-specific adsorption of BMP2 on components of our biomaterials. Non-specific adsorption of proteins is a recurrent and challenging issue for biomaterial studies. Despite the initial incorporation of anti-fouling poly(ethylene glycol) (PEG) chains within our biomaterials, the residual non-specific BMP2 adsorption still triggered BMP2 signaling within the same range as our conditions of interest. To tackle this issue, we explored various options to prevent BMP2 non-specific adsorption. Specifically, we tested alternative constructions of our biomaterials on gold or glass substrate using distinct PEG-based linkers. We identified the aggregation of BMP2 at neutral pH as a potential cause of non-specific adsorption and thus determined specific buffer conditions to prevent it. We also investigated the induced BMP2 signaling over different culture periods. Nevertheless, none of these options resulted in a viable suitable solution to reduce the non-specific BMP2 signaling. Next, we studied the effect of various blocking strategies. We identified a blocking condition involving a combination of bovine serum albumin and trehalose that successfully reduced the unspecific attachment of BMP2 and the non-specific signaling. Furthermore, the effect of this blocking step was improved when using gold platforms instead of glass, particularly with Chinese hamster ovary (CHO) cells that seemed less responsive to non-specifically bound BMP2 than C2C12 cells.